162
ABSTRACT
POSTERS
ABSTRACTS ARRANGED ALPHABETICALLY BY
FIRST AUTHOR
Paper No.: 3125
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
EFFECT OF CHRONIC MILD STRESS ‘CMS’ ON CYP1A2
ENZYME ACTIVITY AND INDUCIBILITY IN WISTAR RATS
Ahmed M-D Abdel-tawab(1), AM Hassan(1), AN-E Hassan
Ain Shams University, Faculty of Medicine, Department of
Pharmacology, Cairo, Egypt
Cytochrome P450-1A2 enzyme activity is reported to be affected by
inflammatory cytokines and psychological stress. These are also, associated with the pathogenesis of depression. In this work we investigated
the effect of ‘CMS’ model of depression on CYP1A2 enzyme activity
and inducibility by o-toluidine ‘T’. 32 Wistar rats were exposed to CMS
for 4 months then randomly allocated into 3 groups: CMS, CMS-T
(received single injection of i.p. 10 mg/kg o-toluidine 24 hours before
decapitation) and a group left for additional 8 weeks to recover from
CMS, then received ‘T’(CMS-RT) Control rats were allocated into two
groups, naı̈ve (received vehicle) ‘T’ groups. CYP1A2 was assessed by
measuring the ratios of caffeine ‘CA’ and 3 its metabolites; theobromine
‘TB’, paraxanthine ‘PX’ and theophylline ‘TP’ in serum samples collected 3 hours after i.p. injection of 20 mg/kg CA. Results showed significant differences in the behavioral tests assessing induction of
depressive-like behaviors. The PX/CA metabolic ratio was the only metabolic ratio that showed significant increase in CMS-T compared to C-T
group. This ratio was still higher in CMS-RT group (P < 0.01 in both).
Medians of PX/CA were 0.08, 0.25, 0.17 for T, CMS-T, CMS-RT groups
respectively. Other metabolic ratios medians for these 3 groups were
0.06, 0.09, 0.13 for TB/CA, 0.06, 0.16, 0.1 for TP/CA and 0.2, 0.5, 0.39
for (PX+TB+TP)/CA, respectively. The CMS group showed no significant difference from naı̈ve group. These results suggested that CMS can
be associated with increased CYP1A2 inducibility by o-toluidine.
Paper No.: 1007
FOCUSED CONFERENCE GROUP: PW01 - PHARMACOLOGY
OF ADRENOCEPTORS: EIGHTH SATELLITE MEETING
IN VIVO AND EX VIVO RECEPTOR OCCUPANCY ASSAYS
FOR THE RAT A2A/D RECEPTOR
RS79948 in non-specific binding wells) in 50 mM glycylglycine buffer
for 45 minutes at 4 degrees. Harvesting was carried out using a Brandel
harvester through GF/B filters soaked in 0.5% polyethylimine. For the in
vivo assay, cortex homogenate from rats dosed with RS79948 followed
by intravenous [3H]-RX821002 was filtered as above using a manifold.
Radioactivity was quantified using a scintillation counter. In cortex of
rats (4 per treatment group) dosed with 0.1, 1, and 5 mg/kg RS79948,
31.2, 83.0, and 92.3% occupancy was measured in the ex vivo assay.
These values were consistent with 66 and 93% occupancy measured at
0.3 and 1 mg/kg in the in vivo assay. In conclusion, ex vivo and in vivo
occupancy assays for the measurement of a2 occupancy have been developed. Conducting the assay in the cortex region provides confidence that
measurements represent occupancy predominantly at the a2A/D subtype.
Paper No.: 1008
FOCUSED CONFERENCE GROUP:
P11 - G PROTEIN-COUPLED 7TM RECEPTORS: FROM
MOLECULAR TO PHYSIOLOGICAL FUNCTION
DEVELOPMENT OF A P2X7 BINDING AND EX VIVO
OCCUPANCY ASSAY IN RAT BRAIN
Sarah Able, R Fish, L Booth, S Katugampola
Pfizer Global Research and Development, Department of Discovery
Biology, Sandwich, UK
The aim of this study was to characterise the binding properties of a
P2X7 antagonist radioligand ([3H]2-cyano-1-[(1S)-1-phenylethyl]-3quinolin-5-ylguanidine) in native tissue, and optimise the binding assay
for use in measuring ex vivo occupancy of P2X7 antagonists. For the in
vitro assay, rat cortex homogenate was incubated with 5nM radioligand
(plus 10 lM of in-house P2X7 antagonist in non-specific binding wells)
in 50 mM Tris/0.1% BSA for 2 hours at 4 degrees. Harvesting was carried out using a Brandel harvester through GF/B filters soaked in 0.5%
polyethylimine. The radioligand concentration was increased (10 nM)
and incubation time reduced (15 minutes) for the ex vivo occupancy
assay. Approximately 60% specific binding was achieved in the cortex
binding assay. The association and dissociation rate of the radioligand
were 0.0021 min-1 nM-1 and0.007 min-1 (half times 45.1 and 88.2 minutes). The Kd derived from kinetic parameters was 3.4 nM, which was
consistent with that derived from saturation experiments of 3.1 nM. In the
brain of rats (4 per treatment group) dosed with3, 10, 30, and 100 mg/kg
of an in-house P2X7 antagonist, 21.5, 27.9, 58.9 and107.4% P2X7 receptor occupancy was measured. In conclusion, an in vitro binding assay
using the P2X7 radioligand has been developed in native tissue and the
ligand properties have been characterised. This assay has been successfully modified for measurement of ex vivo P2X7 receptor occupancy.
Sarah Able, R Fish, C Powell, S Katugampola
Pfizer Global Research and Development, Department of Discovery
Biology, Sandwich, UK
The binding of the a2 radioligand [3H]-RX821002 to rat brain has been
well characterised in the literature. The aim of this study was to use this
ligand to develop binding assays to measure ex vivo and in vivo occupancy of a2A/D antagonists in rat brain. Initial autoradiography studies
using subtype selective antagonists confirmed that there was a strong
radioligand binding signal in the cortex region, and that binding to a2A/D
receptors constitued the majority of this signal. For the ex vivo occupancy assay, cortex homogenate from rats dosed with the a2 antagonist
RS79948 was incubated with 1 nM [3H]-RX821002 (plus 100 nM
Paper No.: 1629
FOCUSED CONFERENCE GROUP: P06 - THE HEART GONE
WRONG; STABILIZATION OF CARDIAC FUNCTION - BASIC
EFFECT OF VITAMIN E ON RENAL ISCHEMIC
PRECONDITIONING IN MALE AND FEMALE RATS
Nahid Abotaleb(1), S Aryamanesh(2), SM Ebrahimi(2), M Nobakht(3)
(1) Iran University of Medical Sciences, Department of Physiology,
Tehran, Iran
(2) Tehran University of Medical Sciences, Nursing and Midwifery
Faculty, Tehran, Iran
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
163
(3) Iran University of Medical Sciences, Department of Histolology and
Neuroscience, Tehran, Iran
Vitamins E play important roles in the protective effect of ischemic preconditioning (IPC). In this study, these antioxidants have been investigated after the induction of ischemia reperfusion (IR) and (IPC) of
kidney. Forty-eight Wistar rats were divided randomly into six groups:
group A (8 male controls), group B (8 female controls), group C (8 male
IR cases) and group D (8 female IR cases). Ischemia was induced by
clamping of left renal arteries for 45 minutes. Rats in group E (8 male
IPC cases) and group F (8 female IPC cases) underwent four cycles of
4-min arterial clamping and 11 min of declamping prior to the final
45 min ischemia induction. Then, serum was collected to assess the
blood urea nitrogen, creatinine and vitamin E. Also, renal tissues were
obtained for the histological assessments. Vitamin E was significantly
increased in both male and female rats in IPC group compared with IR
group. Also, its levels showed significant increase in female IPC group
compared with male IPC group. Histological evaluation showed that in
female rats, injuries induced by IR were significantly less than male rats
and also protective effects of IPC in female rats were significantly more
than male rats. Results of the present study show that in female rats,
after renal IPC, an increase of endogenous vitamin E along with a
decrease in tissue injuries is apparent. This might indicate the protective
effects of preconditioning might be due to an increase in vitamin E in
body.
Paper No.: 506
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
THE BLOOD GLUCOSE LEVELS IN HEALTHY VOLUNTEERS
ADMINISTERED WITH REPAGLINIDE AND GENETIC
POLYMORPHISMS OF CYP3A4 AND CYP2C8
Ruzilawati Abu Bakar, MS Ab Wahab, G Siew Hua
University Sains Malaysia, Department of Pharmacology, Kota Bharu,
Kelantan, Malaysia
Repaglinide is a novel prandial glucose regulator (PGR) for the treatment
of type 2 diabetes mellitus. It is mainly metabolized in the liver by
CYP3A4 and CYP2C8. The objective of the present study is to investigate the effects of the CYP3A4 and CYP2C8 genotypes on the repaglinide’s (Novo Nordisk) blood glucose levels in healthy Malaysian
subjects. Overall, 121 healthy volunteers consisting of 61 men and 60
women enrolled in the study. Each participant received oral 4 mg of repaglinide. Six blood glucose levels (0 min, 30 min, 60 min, 120 min,
180 min and 240 min) per individual were taken according to protocol.
Subjects were genotyped by polymerase chain reaction-restriction fragment length polymorphism for CYP3A4*4,*5 and *18 and by allelesspecific multiplex PCR for CYP2C8*2,*3,*4 and *5. In the group of
121 volunteers, the allele frequency of CYP2C8*1 was 99.17%. The
allele frequency of CYP2C8*2 and CYP2C8*3 were 0.41% respectively.
The allele frequency of CYP2C8*5 was 4.02%. CYP2C8*4 was not
detected in any individual. The frequency of the CYP3A4*18 allele in
the Malaysian population is 2.07%. No mutations were detected for
CYP3A4*4 and CYP3A4*5 alleles. No statistically significant were
found in the blood glucose response to repaglinide between the genotypes. The present study shows that genetic polymorphisms in CYP3A4
and CYP2C8 are not play a role in the blood glucose response to repaglinide.
Paper No.: 507
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING BENEFITS AND MINIMIZING HARMS FROM DRUGS
POPULATION PHARMACOKINETICS OF REPAGLINIDE IN
HEALTHY MALAYSIAN SUBJECTS AND GENETIC POLYMORPHISMS OF CYP3A4 AND CYP2C8
Ruzilawati Abu Bakar, G Siew Hua, MS Ab Wahab
University Sains Malaysia, Department of Pharmacology, Kota Bharu,
Kelantan, Malaysia
This study investigated the effects of CYP3A4 and CYP2C8 genetic
polymorphisms on repaglinide’s (Novo Nordisk) pharmacokinetics in
healthy Malaysian subjects (n = 121) who had received oral repaglinide
(4 mg). Blood samplings were done for repaglinide’s serum concentration determination at 0, 30, 60, 120, 180 and 240 min using highperformance liquid chromatography. Subjects were also genotyped by
PCR-RFLP for CYP3A4*4,*5 and *18 and by an allele-specific multiplex PCR for CYP2C8*2,*3,*4 and *5 alleles. The allele frequencies of
CYP2C8*1, *2, *3, *4 and *5 were 99.17%, 0.41%, 0.41%, 0% and
4.02% respectively. The frequencies of the CYP3A4*4, *5 and *18
alleles were 0%, 0% and 2.07% respectively. CYP2C8 and CYP3A4
genotypes were not significantly associated with repaglinide’s blood
glucose lowering effect. On the other hand, the CYP3A4 genotype
significantly influenced repaglinide’s pharmacokinetics. Subjects having
the CYP3A4*1/*18 genotype has a 34% lower mean elimination rate
constant (p = 0.04) and 133% longer elimination half-lives (p = 0.04)
when compared to the normal types (CYP3A4*1/*1). This confirms that
CYP3A4 plays a larger role in metabolizing repaglinide. In conclusion,
genetic polymorphisms of CYP3A4 specifically CYP3A4*18 plays a
main role in contributing to the interindividual variability in repaglinide’s
pharmacokinetics.
Paper No.: 3113
FOCUSED CONFERENCE GROUP: P04 - PHARMACOEPIDEMIOLOGY, CURRENT CONTROVERSIES AND OPPORTUNITIES
THE EFFECT OF ADVERSE DRUG REACTIONS (ADR) ON DRUG
PRESCRIPTIONS DURING ACUTE HOSPITAL ADMISSIONS
Paul Acheampong(1), M Doshi(2), C Doig(2)
(1) North Tyneside General Hospital / Royal Victoria Infirmary, Department ofClinical Pharmacology and General Internal Medicin, Newcastle
Upon Tyne, UK
(2) North Tyneside General Hospital Rake Lane, North Shields, Tyne
and Wear, UK
adr are undesirable effects of drugs beyond their anticipated therapeutic
effects occurring during proper clinical use of drugs (Asscher AW et al,
bmj 1995; 311: 1003-1005). adr are reported to account for 5% of all
hospital admissions, occur in 10-20% of hospital in-patients and are
responsible for 0.1% of medical in-patient deaths (Pirmohamed M et al,
bmj 1998; 316: 1295-1298). adr result in substantial costs – directly and
indirectly - to patients and healthcare systems. Data was randomly collected from patients during acute admission to a district general hospital
to assess: 1) the incidence of adr in patients admitted to hospital 2) the
severity of adr using standard definitions [adr resulting in hospitalisation
or prolonged admission, physical disability or malformation to an unborn
foetus (Lazarou J et al, jama 1998; 279: 1200-1205)] and patient perspective, and 3) the effect of previous adr on prescribed medications during acute hospitalisation. In all, 322 acute admissions were screened and
of these 22.4% had adr to previously prescribed medications prior to
admission. Whereas 66.3% of patients considered their adr to be severe,
only 25% of previous adr were severe by standard definition. 18.4% of
adr affected choice of prescribed medications during acute admission to
hospital. The incidence of adr remains high and has enormous effects on
healthcare delivery by way of choice of drugs used during acute hospitalisation. There is an attendant financial implication in view of the fact that
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
164
most adr are to first-line drugs and second-line agents are usually more
expensive.
Paper No.: 1134
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
ANATOMICAL REGIONAL DIFFERENCES IN AORTIC TISSUE
REACTIVITY: THE ROLE OF PERIVASCULAR ADIPOSE
TISSUE IN NORMAL AND DIABETIC RATS
Francis I Achike(1), CB Chua(1), CY Kwan(2)
(1) International Medical University, Department of Clinical Sciences,
Kuala Lumpur, Malaysia
(2) China Medical College, Taichung, Taiwan
The perivascular adipose tissue (PVAT), until recently, was treated as a
non-physiologic structural tissue. It’s now known to exert vasorelaxant
and contractile effects. Anatomical regional variations in rat vascular tissue function and in the contribution of human adipose tissue to pathophysiology have been reported. We, therefore, explored possible regional
differences in the effect of PVAT on the contractile responses of windkessel, thoracic and abdominal aortic rings from euglycaemic and streptozotocin-induced diabetic rats. Weighed endothelium-intact or denuded rings
with or without PVAT were dissected from male Sprague-Dawley rats
and mounted in Krebs solution for isometric tension recording. The tissues were contracted with either phenylephrine (PE, 0.1nM - 0.1mM) or
angiontensin-II (Ang-II, 0.01nM - 100nM) under the various experimental conditions. PVAT had no effect on endothelium-intact tissues but it
exerted relaxant, no effect, or contractile (in diabetic tissues) effects,
respectively in the windkessel, thoracic or abdominal endotheliumdenuded tissues. Endothelium-intact or denuded diabetic tissues, respectively, contracted equally or higher than their corresponding euglycaemic
rings. We speculate that PVAT acts like a ‘buffer endothelium’ with its
relaxant function unmasked once the endothelium is damaged, and like
the damaged endothelium it also promotes vascular contractility in the
diabetic state.
Paper No.: 1135
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
POTASSIUM CHANNEL MODULATION OF ACIDOSISINDUCED VASODILATATION OF NORMAL AND DIABETIC
RAT AORTA
dent vasodilatation. 4-aminopyridine or glibenclamide inhibited acidotic
relaxation in diabetic but not euglycaemic ED+ tissues indicating KV
and KATP channel activation in diabetes and their involvement in acidosis-induced endothelium-dependent vasodilatation. In conclusion, acidosis causes endothelium-dependent and -independent vasodilatation in
normal and diabetic tissues via mechanisms differentially modulated by
NO-cGMP and selected K+-channels, some of which, altered in diabetes,
are potential therapeutic targets.
Paper No.: 3248
FOCUSED CONFERENCE GROUP: P02 - TRANSMEMBRANE
TRANSPORT: PERSPECTIVES FOR DISEASE AND DRUG
DISCOVERY
CONSEQUENCES OF CHRONIC POSTNATAL STRESS ON
GLUTAMATERGIC TRANSPORTER AND ON BEHAVIOURAL
TEST IN THE RAT
Gabriela Acosta, MM Odeon, ML Orta, A Salatino
Institute of Pharmacological Research (ININFA), CONICET-UBA, Buenos Aires, Argentina
Early life events have profound consequences in growth and development. It is well known that animals exposed to stressful stimuli during
their early life develop different neurological disorders when they
become adults. We evaluated the consequences of chronic early life environmental manipulations on glutamate transporter (GluT), evaluating the
uptake Glu by synaptosomes isolated from frontal cortex (FC) and hippocampus (Hic) by kinetic parameters, corticosterone levels and GLT-1
expression by western blot. Also we investigated the possiblility of relations between GluT and behaviour. In repeated stress the pups were separated from their mothers and exposed to cold stress (4C) for 1 h at
postnatal days during 20 days. These animals were allowed to a 30 days
recovery period until adulthood. FC and HIC were dissected to study
GluT and trunk blood samples were collected to determinate corticosterone levels. Repeated chronic stress did not change uptake levels in both
areas whereas kinetics parameters were modified. While GLT-1 expression increased on FC and Hic. The levels of corticosterone decreased. In
chronic stress we found an increment in time spent in the illuminated site
of the dark/light transition test. In summary, we have observed regional
changes in GluT produced by chronic stress. These results suggest that a
exposure to postnatal stress at different periods after birth modifies GluT,
affects hypothalamic-pituitary-adrenal axis, which could be relevant to
function of GluT in the adult rat brain and induces anxiolytic-like action
in an animal model of anxiety, and alter the glutamatergic neurons.
Francis I Achike, JL Yeo
International Medical University, Department of Clinical Sciences, Kuala
Lumpur, Malaysia
We showed that acidosis-induced vasodilatation does not involve a
cyclooxygenase product, but is via endothelium-dependent and -independent mechanisms, the former modulated by the EDNO-cGMP cascade.
This study explored K+-channel modulation of acidosis-induced vasodilatation of aortic rings from normal male Sprague-Dawley and streptozotocin-induced diabetic rats. Contractile responses to phenylephrine in
endothelium-intact (ED+) and -denuded (ED-) aortic rings in normal (pH
7.40) or acidotic (pH 7.20) Krebs solution were recorded in the presense
or absence of various potassium channel blockers. Barium chloride (Kir
blocker) significantly inhibited acidotic relaxation in euglycaemic and
diabetic endothelium-intact but not -denuded tissues indicating Kir channel involvement in endothelium-dependent vasodilatation. TEA inhibited
acidotic relaxation in ED+ but not ED- euglycaemic tissues whereas iberiotoxin had no effect in both tissues, indicating involvement of a TEAsensitive (non-BKCa) mechanism in endothelium-dependent relaxation.
In diabetic tissues, however, TEA or iberiotoxin attenuated acidotic
relaxation (ED+ and ED-), although not significantly in iberiotoxin-treated ED- tissues, indicating BKCa channel activation in diabetes and its
involvement in acidosis-induced endothelium-dependent and -indepen-
Paper No.: 427
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
MECHANISM OF ANTIPROLIFERATIVE ACTION OF
XYLOPIA AETHIOPICA (ANNONACEAE) DRIED FRUITS
EXTRACT ON HUMAN CERVICAL CANCER CELLS
Oluwatosin Adaramoye(1), N Singh(2), S Meena(2), B Changkija(3),
R Konwar(3), S Sinha(2), J Sarkar(2)
(1) Department of Biochemistry, College of Medicine, University of
Ibadan, Nigeria
(2) Drug Target Discovery and Development Division, Central Drug
Research Institute, Lucknow, India
(3) Endocrinology Division, Central Drug Research Institute, Lucknow,
India
Introduction: African guinea pepper Xylopia aethiopica is commonly
used in traditional medicine for treatment of several diseases. In this
study, we investigated the anti-cancer potential and mechanism of cell
death elicited by Xylopia aethiopica dried fruit extract (XAFE) on human
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
165
cervical cancer cellline, C33A. Materials: Human cancer cell line-C33A
(cervical) was obtained from ATCC. The cell was grown in recommended media supplemented with 10% FBS, 50 lg/ml gentamicin and
2.5 lg/ml amphotericin B in a 5% CO2 humidified atmosphere at 37C.
Results: Our study demonstrated that XAFE treatment led to dose-dependent growth inhibition in various cell lines with selective cytotoxicity
towards cancer cells. Apoptosis was confirmed by nuclear fragmentation
and sub-G0/G1 phase accumulation. Cell cycle was also arrested at G2/
M phase with decreased G0/G1 population. Semi-quantitative gene
expression studies revealed dose-dependent increase of Bax, p53 and
p21 transcripts with decrease in Bcl-2 gene expression. Conclusion:
Taken together, XAFE inhibited cell proliferation, induced apoptosis and
cell cycle arrest in C33A cells, indicating that XAFE may be a potential
therapeutic agent for cancer.
Paper No.: 3230
FOCUSED CONFERENCE GROUP: P19 - GENERAL SESSION
ANALGESIC ACTIVITY OF THE AQUEOUS SEED EXTRACT
OF HUNTERIA UMBELLATA (K. SCHUM.) HALLIER F.
(APOCYNACEAE) IN RODENTS
Olufunmilayo Adeyemi(1), A Adeneye(1,2), T Alabi(1)
(1) University of Lagos, College of Medicine, Department of Pharmacology, Idi-Araba, Lagos State, Nigeria.
(2) Lagos State University College of Medicine, Department of Pharmacology, Ikeja, Lagos State, Nigeria
Different parts of the tropical rainforest tree, Hunteria umbellata
(K. Schum.) Hallier f. (family: Apocynaceae), are highly valued for the
local management of human and veterinary diseases, including genital
infections, diabetes mellitus, obesity, gastric ulcers and pain by African
traditional healers [Falodun A, et al., Pakistani Journal of Pharmaceutical
Sciences 2006; 19(3):256-258]. Recent studies have reported the analgesic and antipyretic effects of the fruit pulp extract of the plant [Igbe I,
et al., Tropical Journal of Pharmaceutical Research 2009;8(4): 331-336].
The present study was designed at investigating the potential analgesic
property and possible mechanism(s) of action 50-200 mg/kg of the aqueous seed extract of Hunteria umbellata (HU) in different experimental
models of algesia using the tail flick, tail immersion, acetic acid-induced
writhing tests and formalin-induced algesia. Results showed that pretreatment with HU produced significant and dose related (p < 0.05,
P < 0.001) analgesic effect in the treated rats which were mediated via
central and peripheral mechanisms. Overall, results of this study showed
that HU possesses analgesic effect which lends support to its folkloric
use in the local management of pain.
Paper No.: 3392
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
THE ANALGESIC AND ANTIINFLAMMATORY ACTIVITIES
OF THE AQUEOUS LEAF AND STEM EXTRACT OF
ASYSTASIA GANGETICA
Olufunmilayo O Adeyemi, FR Aigbe, GN Uyaiabasi
in pain threshold comparable to morphine (10mg/kg) in tail flick test and
peak analgesia at 200mg/kg in the hot plate test. The extract (25-200mg/
kg) also produced significant (p < 0.05) inhibition of oedema comparable
to indomethacin (10mg/kg) in the carrageenan induced paw oedema
model. The extract (200 mg/kg) produced a significant inhibitory effect
(p < 0.05) comparable to that produced by 1mg/kg dexamethasone in the
xylene induced mouse ear oedema model. Preliminary phytochemical
screening showed that the extract contains alkaloids, tannins, cardiac glycosides and flavonoids. It did not produce mortality nor any visible sign
of lethality 24 hours after single oral administration of 10g/kg. The
results show that the extract possesses analgesic and antiinflammatory
potentials. Further studies to determine the active component(s) responsible for these activities with the view of isolating a lead compound that
may become a novel drug will be carried out.
Paper No.: 1857
FOCUSED CONFERENCE GROUP: PW23 - APPLYING
PHARMACOGENOMICS (PGX) FROM RESEARCH INTO
CLINICAL PRACTICE: PRESENT AND FUTURE
DPYD POLYMORPHISMS AND ADJUVANT 5-FU/LV
TREATMENT OUTCOME IN COLORECTAL CANCER
PATIENTS
Shoaib Afzal(1), SA Jensen(1), B Vainer(1), U Vogel(2), JB Sørensen(1),
HE Poulsen(1)
(1) Copenhagen University Hospital, Rigshospitalet, Laboratory of
Clinical Pharmacology, Copenhagen, Denmark
(2) National Food Institute, Technical University of Denmark,
Copenhagen, Denmark
Dihydropyrimidine Dehydrogenase (DPYD) is the main catabolic
enzyme of 5-Fluorouracil. The objective of this study was to test whether
selected single nucleotide polymorphisms or specific haplotypes in
DPYD influenced treatment outcome in colorectal cancer patients treated
with adjuvant 5-FU/LV. We studied 302 patients treated with adjuvant 5FU/LV after intended curative resection of stage II-III colorectal cancer.
DNA was extracted from FFPE tumor tissue and analyzed for 4 coding
SNPs in the DPYD gene (DPYD cys29arg, DPYD val166met, ile543val
and the exon 14 deletion variant DPYD 2a) using endpoint reads from
real-time PCR. Furthermore DPYD immunohistochemistry was carried
out. Endpoints were relapse free survival (RFS) and overall survival
(OS). Haplotypes containing 2 polymorphisms on different alleles (40
patients) were associated with improved treatment outcome (OS: HR
0.47, p = 0.02; RFS: HR 0.46, p = 0.01). Furthermore, the presence of
haplotypes containing DPYD val166met polymorphisms were also associated with improved outcome (OS: HR 0.51, p = 0.05; RFS: HR 0.43,
p = 0.01). These associations remained unchanged after adjusting for
known clinical prognostic markers. The two delineated patient groups
contained 40 and 36 patients respectively indicating that 85% of patients
could have a potential for an improved response to 5-FU based treatment. DPYD9a TT and haplotypes containing the DPYD 166 polymorphisms (p = 0.04) were associated with increased protein levels as
determined by immunochemistry. We were able to identify two DPYD
profiles that may be useful for allocating colorectal cancer patients to
adjuvant 5-FU based treatment. Our results need confirmation in larger
prospective studies under different treatment regimes.
University of Lagos, Department of Pharmacology, Lagos, Nigeria
The analgesic and antiinflammatory activities of the aqueous leaf and
stem extract of Asystasia gangetica (Linn.) T. Anders. [family Acanthaceae] used commonly in sub tropics and tropics for the management of
asthma and pain were evaluated. The analgesic effect of the extract (25,
50, 100 and 200mg/kg) was evaluated in rats and mice using the acetic
acid induced writhing, cold water tail flick and hot plate models. Its antiinflammatory effect was evaluated using carragenaan induced rat paw
oedema and xylene induced mouse ear oedema models. The extract (25200mg/kg) significantly (p < 0.05) reduced the number of writhes in the
acetic acid induced writhing test. At 100 mg/kg, it produced an increase
Paper No.: 2345
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
MICROPARTICLES FROM PATIENTS WITH METABOLIC
SYNDROME INDUCE VASCULAR HYPO-REACTIVITY VIA
FAS/FAS-LIGAND PATHWAY IN MICE
A Agouni(1), P-H Ducluzeau(2), T Benameur(1), M Sladkova(0), G Leftheriotis(1), O Pechanova(3), Carmen Martinez(1), R Andriantsitohaina(1)
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
166
(1) University of Angers Faculty of Medicine, INSERM, U771, CNRS
UMR, 6214, Angers, France
(2) CHU d’Angers, Département d’Endocrinologie et Diabétologie,
Angers, France
(3) Institute of Normal and Pathological Physiology, Slovak Academy of
Sciences, Bratislava, Slovak Republic
Microparticles are small membrane vesicles released during cell activation and apoptosis. Elevated circulating levels of microparticles have
been detected in several cardiovascular diseases including the metabolic
syndrome (MS). The increased number of microparticles was associated
in many of these diseases with vascular dysfunction. In the present study,
we evaluated the effects of in vivo treatment of circulating microparticles
from both patients with MS and healthy subjects on vascular function.
Microparticles obtained from whole blood either from MS patients or
healthy subjects, or a vehicle control were injected intravenously to mice
and vascular reactivity was then evaluated in aorta. Injection of microparticles from MS patients into mice induced vascular hypo-reactivity in
response to serotonin in aorta. Interestingly, hypo-reactivity was reversed
by a nitric oxide (NO)-synthase (NOS) inhibitor, and was associated with
up-regulation of inducible NOS (iNOS) protein expression and an
increased production of NO. The selective cyclo-oxygenase-2 inhibitor
reduced serotonin-induced contraction in vessels from vehicle and
healthy subject microparticles but did not affect response to the same
agonist in MS microparticle-treated mice. MS microparticles also
enhanced prostacyclin production in aorta. In addition, MS microparticles
increased reactive oxygen species production via enhanced expression of
the NADPH oxidase subunits, gp91phox and p47phox. Importantly, the
silencing of the pro-inflammatory pathway Fas/Fas-ligand, completely
prevented the vascular hypo-reactivity. These data provide evidence that
circulating microparticles from MS patients induce in vivo vascular dysfunction by increasing both oxidative and nitrosative stresses and by
altering the release of cyclo-oxygenase metabolites through the Fas/FasL
pathway.
Paper No.: 2346
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
RED WINE POLYPHENOLS PREVENT METABOLIC AND
CARDIOVASCULAR ALTERATIONS ASSOCIATED WITH
OBESITY IN ZUCKER FATTY RATS (FA/FA)
A Agouni(1), A-H Lagrue-Lak-Hal(1), HA Mostefai(1), A Tesse(1), P
Mulder(2), P Rouet(3,4), F Desmoulins(3,4), C Heymes(5), MC Martinez(1), Ramaroson Andriantsitohaina(1)
(1) University of Angers Faculty of Medicine, INSERM, U771, CNRS
UMR, 6214, Angers, France
(2) INSERM, U644, Rouen, France
(3) INSERM, U858, Toulouse, France
(4) Université Paul Sabatier, Institut Fédératif de Recherche 31,
Toulouse, France
(5) INSERM, U689, Paris, France
Obesity is associated with increased risks for development of cardiovascular diseases. Epidemiological studies report an inverse association
between dietary flavonoid consumption and mortality from cardiovascular
diseases. We studied the potential beneficial effects of dietary supplementation of red wine polyphenol extract, Provinols, on obesity-associated
alterations with respect to metabolic disturbances and cardiovascular
functions in Zucker fatty (ZF) rats. ZF rats or their lean littermates
received normal diet or supplemented with Provinols for 8 weeks. Provinols improved glucose metabolism by reducing plasma glucose and
fructosamine in ZF rats. Moreover, it reduced circulating triglycerides and
total cholesterol as well as LDL-cholesterol in ZF rats. Echocardiography
measurements demonstrated that ProvinolsTM improved cardiac performance as evidenced by an increase in left ventricular fractional shortening
and cardiac output associated with decreased peripheral arterial resistances in ZF rats. Regarding vascular function, ProvinolsTM corrected
endothelial dysfunction in aortas from ZF rats by improving endothelium-
dependent relaxation in response to acetylcholine (Ach). ProvinolsTM
enhanced NO bioavailability resulting from increased nitric oxide (NO)
production through enhanced endothelial NO-synthase (eNOS) activity
and reduced superoxide anion release via decreased expression of
NADPH oxidase membrane sub-unit, Nox-1. In small mesenteric arteries,
although ProvinolsTM did not affect the endothelium-dependent response
to Ach; it enhanced the endothelial-derived hyperpolarizing factor component of the response. Use of red wine polyphenols may be a potential
mechanism for prevention of cardiovascular and metabolic alterations
associated with obesity.
Paper No.: 744
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
PATTERNS OF INFLUENZA INFECTION IN SYLHET
DIVISION OF BANGLADESH
Salim Ahmed, ATMA Jalil, SS Ahmed, MM Siddika
Jalalabad Ragib-Rabeya Medical College, Department of Pharmacology
& Therapeutics, Sylhet, Bangladesh
The influenza virus infection varies globally with the genomic diversity
or and clinical variations and regional detection of the patients or and
viral agents is influenced by the variations of geographic, demographic
condition and patient risk factors. This study is carried out to clarify
about the pattern and seasonality of influenza infection during two consecutive years from July 2007 to April 2009 in Jalalabad Ragib-Rabeya
Medical College Hospital, Sylhet with active collaboration of ICDDR’B
and IEDCR. Three hundred eighty five (N = 385) patients with acute
respiratory infection were selected as case following WHO selection criteria of avian influenza A (H5N1) virus infection. In this study throat
swab and anterior nasal swab were collected as clinical specimen from
the cases for detection of Influenza A (H5N1) by molecular or and
by any other diagnostic methods in BSL-III laboratory of ICDDR’B;
Mohakhali, Dhaka. Overall influenza positivity was 48 (12%), in where
Flu A (n = 27), Flu B (n = 21) and subtypes of Flu A: H1 (n = 16), H3
(n = 11) & H5 (n = 0). This study also suggests that peak seasonality of
influenza infection is more marked in the month of July, August &
September.
Paper No.: 1851
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
CEREBROVASCULAR CONTRACTILE RECEPTORS
UP-REGULATE VIA A RAF SENSITIVE SIGNALLING PATHWAY
Hilda Ahnstedt, L Edvinsson
Lund University, Division of Clinical Sciences, Department of Experimental Vascular Research, Lund, Sweden
Cerebral ischemia results in a rapid increase in contractile receptors in
the ischemic region, which further impairs the local blood flow and
aggravates the tissue damage. Cerebral ischemia is associated with
enhanced expression of several G-protein coupled receptors (GPCRs)
with strong vasoactive properties such as the endothelin ETA and ETB
receptors, the angiotensin II (Ang II) type 1 receptor (AT1) and the serotonin type 1B receptor (5-HT1B). This receptor up-regulation occurs via
activation of the MEK/ERK1/2 pathway. The present study aimed to
examine the role of Raf, the first upstream signalling molecule in this
pathway. Cerebral arteries were obtained from patients undergoing neurological surgery (n = 7). The vessels were organ cultured for 48 h in the
absence or presence of Raf inhibitors: SB-386023 or SB-590885. Contractile properties were evaluated in a sensitive myograph by cumulative
addition of 5-carboxamidotryptamine (5-CT, 5-HT1B agonist), Ang II
and endothelin-1 (ET-1). Vessels treated with SB-386023 and
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
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SB-590885 showed attenuated contractile responses to 5-CT, and the
effect was most prominent after treatment with SB-590885 (p < 0.05).
The maximum contraction to Ang II was attenuated by SB-590885
(P < 0.01) and only slightly by SB-386023. ET-1 gave a biphasic concentration-dependent response. The high affinity phase corresponding to
the ETB receptor-mediated contraction was decreased in the presence of
SB-590885(p < 0.05). This study shows that Raf plays an important role
in the altered expression of several GPCRs seen after cerebral ischemia
and its inhibition might be a novel approach to reduce tissue damage
after a stroke.
Paper No.: 1480
FOCUSED CONFERENCE GROUP: P17 - NEW APPROACHES
AND TARGETS IN PSYCHIATRY
EFFECTS OF SELECTED GLUTAMATERGIC LIGANDS ON
NOVELTY-INDUCED HYPERACTIVITY OF
GLUA1-DEFICIENT MOUSE LINE
Teemu Aitta-aho, C Procaccini, ER Korpi, AM Linden
Institute of Biomedicine, Department of Pharmacology, Helsinki, Finland
Glutamate AMPA receptors have been implicated in schizophrenia,
which is a major psychiatric illness with unknown etiology. Putative
mouse model of schizophrenia, glutamate AMPA-receptor GluA1-subunit deficient mouse line (GluA1-/-mice), displays various symptoms
related to schizophrenia. Among them, a robust spatial novelty-induced
hyperactivity has been shown to be sensitive to a prototypic antipsychotic haloperidol. In our studies, male and female GluA1-/-; mice and
their littermate control wild type mice (GluA1-/-mice & -/- were injected
intraperitoneally with AMPA-receptor antagonist NBQX (10, 30 and
100 mg/kg), AMPA-receptor potentiator CX546 (10, 30 and 80 mg/kg)
or metabotropic glutamate receptor 2/3 agonist LY354740 (15 mg/kg),
placed in a novel cage and thereafter their locomotor activities measured
for 2 h. Vehicle-treated GluA1-/- mice displayed clear hyperactivity in all
experiments. NBQX reduced locomotor activity of both mouse lines at
100 mg/kg dose only. However, the reduction was greater in GluA1-/mice. No effect of CX546 was found in either mouse line. LY354740
reduced locomotor activity in the GluA1-/- males but not in the females,
while the locomotor activity of GluA1-/- mice was unchanged. These
results suggest sensitivity of a tentative antipsychotic LY354740 in a
mouse model of schizophrenia, but in a gender-specific manner.
Paper No.: 1086
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
INHIBITORY EFFECT OF CARBON MONOXIDE-RELEASING
MOLECULE CORM-2 ON TNF-A RELEASE FROM
ACTIVATED-RBL-2H3 CELLS
Masaaki Akagi(1), Y Yasui(1), M Watanabe(1), A Maruyama(1), N
Matsui(1), N Fukuishi(1), R Akagi(2)
(1) Tokushima University Hospital, Faculty of Pharmaceutical Sciences,
Tokushima,Japan
(2) Yasuda Women’s University, Japan
We recently reported that HO-1 inhibited TNF-a release from activatedRBL-2H3 cells, and Zn-protoporphyrin, which was an inhibitor of heme
oxygenase, inverted its inhibition. These results suggest that heme
metabolites can inhibit TNF-a production from the activated-RBL-2H3
cells. In the present study, we examined which heme metabolites had
involved in the inhibition of TNF-a release from activated-RBL-2H3
cells. Heme metabolites (bilirubin, iron, CORM-2 as CO donor) were
co-incubated with anti-ovalbumin(OA) rat serum(IgE) for 2hr, and then
the cells were challenged with OA. Gene expression was analyzed by
RT-PCR. Protein expression was investigated by western blot analysis
and ELISA. TNF-a release was significantly reduced by CORM-2, not
iron or bilirubin. Inactivated-CORM-2 didnhft reduce TNF-a release.
TNF-a mRNA expression was also significantly inhibited by CORM-2.
Moreover, CORM-2 reduced phosphorylation of ERK. CORM-2 can
regulate TNF-a production through inhibition of ERK. These findings
provide new insights into the anti-allergic activities.
Paper No.: 3270
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
BIOEQUIVALENCE STUDY OF 1500 MG GLUCOSAMINE
SULFATE IN THAI HEALTHY VOLUNTEERS
Pravit Akarasereenont(1), S Chatsiricharoenkul(1), P Pongnarin(1), K
Sathirakul(2), A Wongkajornsilp(1), S Kongpatanakul(1)
(1) Siriraj Hospital, Faculty of Medicine, Department of Pharmacology,
Bangkok, Thailand
(2) Mahidol University, Bangkok, Thailand
Glucosamine sulfate is widely used to relieve symptoms from osteoarthritis. This study was conducted in order to determine pharmacokinetic
and assessed the in-vivo bioequivalence of two different hard capsule
formulations of glucosamine sulfate when administered as equal dose of
1,500 mg. The two formulations contain different salt form where reference product is NaCl and test product is KCl. A randomized, single dose,
two-treatment, two-period, two-sequence crossover study was conducted.
Twenty-six healthy volunteers were recruited at Siriraj Clinical Research
Unit. Each subject received a dose of 1,500 mg glucosamine sulfate of
both formulations with at least a week washout period. Blood samples
were collected over 24 h after the oral administration. The plasma fractions were analyzed for glucosamine using LC-MS/MS. Twenty-six volunteers enrolled in the present study. Pharmacokinetic parameters were
determined using the non-compartment model. The 90 percent confidence intervals of the mean ratios (test/reference) of Cmax (111.19%;
ranged from 93.01%-132.92%) and AUC0-t (107.24; ranged from
87.16%-131.93%) was not contained within the equivalence criteria of
80.00-125.00% (US FDA, 2003). However, this study showed the high
intra-individual CV calculated from ANOVA for Cmax and AUC0-24
(30%). Thus, based on equivalence limits of US FDA (2003), the test
product is not bioequivalent to the reference product in terms of rate and
extent of absorption. However, concerning the wider equivalence criteria
for highly variable drug (EMEA, 2008), the test product is bioequivalent
to the reference formulation in terms of rate and extent of absorption.
Paper No.: 3373
FOCUSED CONFERENCE GROUP: P04
- PHARMACOEPIDEMIOLOGY, CURRENT CONTROVERSIES
AND OPPORTUNITIES
PRESCRIPTION PATTERNS OF CONTROLLED MEDICINES
IN ISTANBUL
Ahmet Akici(1), D Demircan(1), I Topcu(2), H Yilmaz(2), B Donertas(1),
Kemal K Berkman(1)
(1) Marmara University School of Medicine, Department of Pharmacology, Istanbul, turkey
(2) Provincial Health Directorate of Istanbul, Istanbul, Turkey
Objective: Drug utilization study of controlled medicines (CMs) which
are prescribed as red or green color scripts in Turkey, are important to
evaluate rational pharmacotherapy. It was planned to investigate the
details of CMs utilization in Istanbul. Method: A total 12000 prescriptions, containing CMs were collected retrospectively from Provincial
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
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Health Directorate of Istanbul medical records between January-March in
2009. CMs use was stratified by some socio-demographic characteristics
of patients, and morbidity patterns among users were investigated.
Results: The average age was 46,6 ± 23,4 and half of them were women.
CMs were prescribed by physicians mainly from psychiatry (34.9%),
internal medicine (10.6%), surgery (9.9%), anesthesia (9.1%), and neurology (7.3%) departments. Only 2.5% of scripts were prescribed by general practitioners and family physicians. The majority of scripts were
prescribed at hospitals (74.4%), mainly contained single drug (94.3%),
and also many of them (77.3%) were green color scripts. The most frequently prescribed drug was alprazolam (27.9%), followed by tramadol
(13.9%), clonazepam (12.5%), biperiden (8.8%), methylfenidate (7.8%),
respectively. The most commonly recorded indications were anxiety
(19.5%), cancer (14.8%), attention deficiency-hyperactivity disorders
(12.9%), behavioral disorders (12.5%), depression (10.9%). Conclusions:
This is the first study in this context which reflects CMs utilization in
Istanbul and reveals that CMs were mainly prescribed by specialists in
the treatment of psychiatric disorders in both sexes.
Paper No.: 580
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
IN VIVO ANTIOXIDANT ACTIVITY OF BYRSOCARPUS
COCCINEUS SCHUM. AND THONN. (CONNARACEAE)
EXTRACT IN CARBON TETRACHLORIDE-INDUCED
HEPATOTOXICITY IN RATS
(1) Kaunas University of Medicine, Kaunas, Lithuania
(2) University of Aarhus, Department of Pharmacology, Aarhus, Denmark
(3) Vilnius University, Institute of Oncology, Vilnius, Lithuania
Photodynamic therapy (PDT) is a treatment method where the combination of laser light and a photosensitizer are combined to produce a
cytotoxic or modifying effect to cancerous tissue by producing reactive oxygen species. Damage of different cellular compartments
induces activation of the cellular DNA damage repair system, which
is important factor modulating tumor sensitivity to this treatment. bGlucans is not present in the membrane of tumor cells and killing of
iC3b-opsonized tumor cells via complement receptor 3 cannot be
induced during PDT. The aim of the present study was to evaluate
the effect of PDT in combination with b-glucans on DNA damage
repair system and cell proliferation and determine whether coadministration of b-glucans produces more effective killing of PDT-treated
tumor cells. C57 Bl/6 female mice bearing Lewis lung carcinoma
were exposed to the treatment with PDT (Photofrin, 10 mg/kg, 24 h
prior to laser irradiation) or/and b-glucans (400 lg/d/mouse, 5 days).
PDT in combination with b-glucans significantly reduced tumor
growth (P < 0.05, n = 10), proliferating cell nuclear antigen (PCNA)
expression (P < 0.001, n = 9), and increased necrosis in tumor tissue
(P < 0.001, n = 9). Our data demonstrates that b-glucans enhances
efficacy of photodynamic therapy by suppressing activity of DNA
damage repair system and cell proliferation, and inducing more efficient cell killing.
A Akindele(1), K Ezenwanebe(1), C Anunobi(2), Olufunmilayo O Adeyemi(1)
(1) Department of Pharmacology, College of Medicine, University of Lagos, Lagos,Nigeria
(2) Department of Morbid Anatomy, College of Medicine, University of
Lagos, Lagos,Nigeria
The leaf decoction of Byrsocarpus coccineus (Connaraceae) is drunk for
the treatment of jaundice in West African traditional medicine (WATM).
This study was designed to investigate the in vivo antioxidant activity of
B. coccineus aqueous leaf extract (BC) in carbon tetrachloride (CCl4)induced hepatotoxicity in rats. Rats were randomly divided into different
groups (vehicle; CCl4; BC 1000 mg/kg; BC 200, 400 and 1000 mg/kg +
CCl4; and livolin 20 mg/kg + CCl4) and treatment was carried out
accordingly. On the 7th day, rats were sacrificed and livers were isolated
to assay for activity of catalase (CAT), superoxide dismutase (SOD),
reduced glutathione (GSH), peroxidase (PX) and level of malondialdehyde (MDA) using standard procedures. Histopathological assessment of
liver samples was also carried out. CCl4 produced significant (p < 0.05)
reductions in the activity of CAT, SOD, PX and GSH, and conversely
increased MDA level. BC (200, 400 and 1000 mg/kg) produced significant and dose-dependent reversal of CCl4-diminished activity of the antioxidant enzymes and reduced CCl4-elevated level of MDA. The effect of
livolin on the parameters was generally comparable to those of BC at
the most effective dose of 1000 mg/kg. Histological observation of liver
samples supported the results obtained in this study. BC (+ CCl4) treated
rats generally showed normal hepatocytes with moderate inflammation
and livolin treated rats also showed normal hepatocytes but with mild
inflammation. The results obtained in this study suggest that BC possesses hepatoprotective in vivo antioxidant activity, thus justifying its use
in WATM for the treatment of liver disease.
Paper No.: 2499
FOCUSED CONFERENCE GROUP: PW04 - ANTIOXIDANTS AS
THERAPEUTIC TARGETS
EFFECT OF B-GLUCANS ON PHOTODYNAMIC THERAPY OF
LEWIS LUNG CARCINOMA
Dalia Akramiene(1), C Aleksandraviciene(3), K Suziedelis(3), U Simonsen(2), E Stankevicius(0)
Paper No.: 751
FOCUSED CONFERENCE GROUP: P14 - ADDICTION AND
DOPING: NEUROBIOLOGICAL AND CLINICAL
NEUROPROTECTIVE EFFECT OF ADENOSINE A1 AND A2A
RECEPTORS IN CHRONIC ALCOHOL WITHDRAWAL
SYNDROME IN MICE
Kiran Kumar Akula, K Chopra, SK Kulkarni
Panjab University, University Institute of Pharmaceutical Sciences,
Chandigarh, India
Alcohol withdrawal following chronic alcohol intake results in moderate to severe withdrawal symptoms. Adenosine, an endogenous neuromodulator, has been reported to reverse the behavioral symptoms of
alcohol withdrawal. The present study was conducted to evaluate the
neuroprotective actions of selective adenosine A1 and A2A receptor
ligands against chronic alcohol withdrawal syndrome in mice. Mice
were made physically dependent on alcohol by chronic intragastric
administration of 35% v/v ethanol (2 g/kg of 10% v/v) twice daily for
10 days. On day 11, animals received only adenosine receptor ligands
twice. After 12 hours of the last adenosine ligand treatment, the withdrawal symptoms were precipitated with RO15-4513 (4 mg/kg, i.p.,
ethanol antagonist) and the mice were tested for withdrawal. Ethanoltreated animals exhibited hyperlocomotor activity and anxiogenic
response. Also, alcohol withdrawal decreased pentylenetetrazol i.v. seizure threshold for the onset of generalized clonus and tonic extensor.
Oxidative-nitrosative stress, caspase-3 activity in cytoplasmic lysate,
and NFjb/p65 unit were significantly increased in nuclear lysate of the
brain samples of ethanol-treated mice. CHA (0.1 or 0.2 mg/kg, i.p.,
selective A1 agonist), but not CPCA (0.1 or 0.2 mg/kg, i.p., selective
A2A agonist), reduced all the behavioral, biochemical and molecular
changes induced by alcohol withdrawal. On the other hand, the effect
of CHA was reversed by pretreatment with DPCPX (5 mg/kg, i.p.,
selective A1 antagonist). Moreover, A2A antagonist [8-(3-cholorostryl)
caffeine, 4 mg/kg, i.p.] showed neuroprotective action in alcohol withdrawal mice. The results emphasize neuroprotective potential of adenosine A1 receptor agonists or A2A receptor antagonists in attenuating
ethanol withdrawal syndrome.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
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Paper No.: 3380
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
COMPARATIVE STUDY OF INITIAL AND AQUIRED DRUG
RESISTANCE IN PULMONARY TUBERCULOSIS
Seyed Yousef Agha Alaie(1), SD Mansoori(2), S Khazali(1), AA Hashtroodi(1), S Arami(2)
(1) Artesh University of Medical Sciences, Department of Pharmacology,
Tehran, Iran
(2) National Research Instute of Tuberculosis and Lung Disease, Tehran,
Iran
Resistance to anti-tuberculosis agents particulary multiple drug resistant(MDR) mycobacterium tuberculosis is an important obstacle in the
treatment and control of tuberculosis in the world. Between four years
for 273 smear and culture positive pulmonary tuberculosis patients (both
old = 86/273 and new = 187/273)pretreatment susceptibility tests of isolated bacilli to INH,RIF,EMB and STM were performed by standard proportional method and the result were classified in 3 groups: 1)Newly
diagnosed without any history of treatment 2)patiente with history of
treatment with one course 3)patients with history of treatment for two or
more courses supposed to be MDR cases.The result were collected for
each drug individuall and different combination of two,three and four
medications. Resistance to single drug,two drugs,three drugs and four
drugs wase significantly increased in group 3 in comparison to group 2
and 1,also significantly increase in group 2 when compared to group
1.We observed a high rate of primary resistance to INH and STM in
group 1and 2 and a high rate of MDR(INH and RIF resistance)in group
2 and 3. The duration of bacilli exposure to anti –tuberculosis agents in
the past is a mager factor in developing resistance .In contrast to WHO’s
guideline,due to high rate of primary resistance especially to STM in our
area,we don’t recommended addition of STM for treatment of patient
whose initial four drug regimens have been faild (group2).
Paper No.: 3381
FOCUSED CONFERENCE GROUP: P08 - DEVELOPMENTS IN
THE TREATMENT OF SEXUAL DYSFUNCTION AND
DISEASES OF THE LOWER URINARY TRACT
FINASTERIDE AND PRAZOSIN MIXED THERAPY IN
PATIENT WITH BENING PROSTATIC HYPERPLASIA
Paper No.: 2708
FOCUSED CONFERENCE GROUP: P11 - G PROTEINCOUPLED 7TM RECEPTORS: FROM MOLECULAR TO
PHYSIOLOGICAL FUNCTION
PRE-VERSUS POSTSYNAPTIC A2A-ADRENOCEPTORS
FUNCTIONS MEDIATED VIA GAI-ISOFORMS
Julian Albarran-Juarez(1), R Gilsbach(1), R Piekorz(2), K Pexa(2), B
Nürnberg(2,3), L Hein(1)
(1) University of Freiburg, Institute of Experimental and Clinical Pharmacology and Toxicology, Freiburg im Breisgau, Germany
(2) Heinrich-Heine-University, Institute of Biochemistry and Molecular
Biology II, Düsseldorf, Germany
(3) Eberhard-Karls-University, Institute of Experimental and Clinical
Pharmacology and Toxicology, and Interfaculty Center of Pharmacogenomics and PharmaceuticalResearch, Tübingen, Germany
Members of the a2-adrenoceptor family are prototype Gai/o protein-coupled receptors and are involved in the regulation of a number of physiological functions including presynaptic feedback inhibition of transmitter
release, analgesia, sedation, anesthetic-sparing, and locomotor activity. In
the present study we tested whether inhibition of spontaneous locomotor
activity and anesthetic-sparing effect elicited by a2-agonists are mediated
by a2A-adrenoceptors located in adrenergic or non-adrenergic neurons and
whether these responses require specific Gai-isoforms in vivo. We evaluated mice which express a2A-adrenoceptors exclusively on adrenergic cells
under control of the dopamine b-hydroxylase promoter (Adra2AC-/-TG),
mice deficient in a2-adrenoceptors (Adra2C-/-, Adra2AC-/-), mice deficient
in Gai isoforms or wild-type mice (WT). In Adra2AC-/- animals spontaneous locomotor activity was enhanced compared with WT and Adra2C-/-,
but normalized in Adra2AC-/-TG, thus mediated by a2A-adrenoceptors
located in adrenergic neurons. After administration of the a2-agonist medetomidine locomotor activity was reduced to 9.2 ± 3.8% in WT, 7.9 ± 3.7%
in Gai1-/-, and 4.3 ± 3.3% in Gai3-/- mice; and significantly attenuated in
Gai2-/- mice to 37.2 ± 8.1%. The anesthetic-sparing effect of medetomidine was ablated in Adra2AC-/- mice and was not rescued by
Adra2AC-/-TG, thus it was mediated by a2A-adrenoceptors present on nonadrenergic cells. In WT, Gai1-/- and Gai3-/- animals medetomidine caused
a leftward shift of the isoflurane-mediated loss of righting reflex curves, but
this effect was completely absent in Gai2-/- mice. Our results suggest that
inhibition of spontaneous locomotor activity and the anesthetic-sparing
effect of a2-agonists may be elicited by a specific coupling of a2A-adrenoceptors via Gai2 proteins.
Seyed Yousef Agha Alaie, SS Kashaf, B Najafian, P Musavi
Artesh University of Medical Sciences, Department of Pharmacology,
Tehran, Iran
Bening prostatic hyperplasia (BPH) is the most common cause of male
urinary obstruction. Medical management of BPH includes anti-androgens and a adrenergic receptor blockers. In this study are compared the
efficacy of mixed therapy of finasteride as a 5-a reductase inhibitor in
combination with prazosin as a specific a 1-adrenergic receptor blocker
(F+P) and finasteride alone (F) on obstructive/stimilative symptom of
BPH.Forty men aging 60 + /-6 (mean+/)SD) years old were divided into
two groups.prazosin 1mg and finasteride 5mg daily and similar dose of
finasteride alone were prescribed to group (F+P) and group (F) for
3 months respectively. afterwhile, mean prostativ volume reduced by
16.5% (p < 0.05) in group (F+P) and 11.3% (p=ns) in group (F).
Obstructive and stimulative sympthom scores reduced by 65.3%
(p < 0.0007). in groups (F+P) and (F), respectively.there was no correlation between change in mean prostatic volume and obstructive/stimulative sympthom scores (r=-0.74). The most complication were impotence
(n = 17), loss of libido (n = 17), reduced ejaculate volume (n = 16) and
dry mouth (n = 15) in which their incidences were not different between
two groups. We concluded that combination of finasteride and prazocin
might accestuate the process of prostatic volume reduction in comparison
to finasteride alone in BPH. This measure did not increase the incidence
of complicatin.So,it may be a suitable alternative for more invasive therapeutic approaches.
Paper No.: 2728
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
PALMITOYL COA ANTAGONISM OF ADP-INDUCED
RELAXATION OF RAT THORACIC AORTA
Eman Alefishat, S Alexander, V Ralevic
University of Nottingham, Department of Biomedical Science, Nottingham, UK
Palmitoyl CoA has been described to antagonise human P2Y1 receptor
responses in recombinant systems (Coddou C et al. FEBS Lett 2003;
536: 145-150) and in isolated platelets (Manolopoulos P et al. Platelets
2008; 19: 134-145). We have noted that palmitoyl CoA antagonises
P2Y1 receptor-evoked relaxations in rat isolated thoracic aorta (Alexander SPH et al. Proc BPS 2009; 6: 150P). The present study investigated
the reversibility of this interaction in the same tissue from male Wistar
rats (200-250 g), as described previously (Bultmann R et al. Eur J Pharmacol 1998; 359: 95-101). ADP evoked concentration-dependent relaxations of the methoxamine-pre-contracted aorta, 30 lM ADP elicited a
relaxation of 71 ± 4% (n = 8). In the presence of palmitoyl CoA
(10 lM), the response at this concentration of ADP was
18 ± 4%(n = 8). Following washout of palmitoyl CoA for an hour,
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
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30 lM ADP evoked a relaxation of 73 ± 4% (n = 8) of the methoxamine contraction. Application of 10 lM palmitoyl CoA in the absence
of ADP elicited a modest relaxation (5 ± 2%), which failed to reach statistical significance. These results indicate that the antagonism of palmitoyl CoA for the ADP evoked relaxation in rat isolated thoracic aorta
through the P2Y1 receptor is reversible with no significant direct effect
of pamitoyl CoA on the rat isolated thoracic aorta. Acknowledgement:
We gratefully acknowledge and appreciate the Jordanian Government,
who sponsored this research.
Paper No.: 3104
FOCUSED CONFERENCE GROUP: PW04 - ANTIOXIDANTS AS
THERAPEUTIC TARGETS
HYDROGEN SULPHIDE GENERATION IN PORCINE TISSUES
Stephen Alexander(1), M Doe(1), M Garle(1)
University of Nottingham, Department of Biomedical Sciences, Nottingham, UK
Hydrogen sulphide is a gasotransmitter candidate with a suggested physiological role in the regulation of smooth muscle (Li et al Pharmacol Ther
2009 123: 386-400). We have analysed hydrogen sulphide generation in
cell-free extracts using the methylene blue formation method (Stipanuk
MH, Beck PW Biochem J 1982;206: 267-277). In rat liver cytosol, hydrogen sulphide generation in the presence of 5 mM L-cysteine (19 nmol/hr
mg) was inhibited by 95% in the presence of either 10 lM propargylglycine or 100 lM aminooxyacetic acid. In contrast, rat kidney cytosol
hydrogen sulphide generation (4.0 nmol/hr/mg) was not significantly
inhibited by propargylglycine, but was almost completely inhibited by
aminooxyacetic acid. Porcine liver and kidney cytosolic fractions exhibited much reduced hydrogen sulphide generation (0.6 and 3.9 nmol/hr/
mg, respectively). Cytosolic fractions from porcine smooth muscle
showed a rank order of hydrogen sulphide generation of bronchioles (12)
= splenic artery (10) > bladder (7 nmol/hr/mg). Omission of pyridoxal
phosphate inhibited hydrogen sulphide generation in the bronchioles only.
In the porcine brain, midbrain activity was greater than hippocampus (3.1
and 1.0 nmol/hr/mg, respectively). Bronchiole activity was inhibited by
both propargylglycine and aminooxyacetic acid (20% control), while kidney activity was partly inhibited by propargylglycine (40% control) and
completely by aminooxyacetic acid (3% control). These studies show the
viability of using porcine tissues to assess tissues not readily assayed for
hydrogen sulphide synthesis in rodents. However, the effects of the common inhibitors don’t allow identification of the enzymes responsible.
Paper No.: 2246
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
MUSCARINIC M3ACHR IS REGULATED BY CGMP, VIA A
PKG-II DEPENDENT PHOSPHORYLATION IN PLASMA
MEMBRANES FROM TRACHEAL SMOOTH MUSCLE
Marcelo J Alfonzo, M Alfonzo-Gonzalez, RG Alfonzo, A Misle, IL
Becemberg
Central University of Venezuela Faculty of Medicine, Experimental
MedicineInstitute, Biomembranes Section, Caracas, Venezuela
Muscarinic activation of airway smooth muscle, via m2/3AchR produces
two cGMP signals related to the stimulation of the NO-dependent guanylyl cyclase and the NPR-guanylcyclase. To understand the role of
cGMP on these mAChRs, we decided to evaluate the muscarinic receptor
(3HQNB binding) activity in the presence of ATP and cGMP using
plasma membrane (PM) fractions isolated from bovine tracheal smooth
muscle. We found that cGMP plus ATP change the binding properties of
the mAChRS specially the agonist binding affinity to carbamylcholine
and the antagonist properties for 4-DAMP, which is m3AchR-selective
antagonist, without changing the m2AchRs antagonist binding abilities
of gallamine, AFDX-116DS and methoctramine. These muscarinic binding modifications disappeared, after a 4-DAMP alkylation of these PM.
Moreover, activators and inhibitors of GPKG-II affected these binding
activities. This G-kinase was detected by Western blotting in these PM.
Using 32P-ATP, we found that cGMP induced a significant 32P-phosphorylation in these PM, being immunoprecipated using a m3AchR antibody,
which was obliterated by 4-DAMP, in a dose dependent manner. These
original results indicated that cGMP specifically regulated m3AchR, via
phosphorylation by a PKG-II bound to plasma membranes from bovine
tracheal smooth muscle.
This work was supported by grants from FONACIT-Venezuela S12002000411 (ILB) and CDCH-UCV-PG-09-07401-2008/1(RGA).
Paper No.: 2650
FOCUSED CONFERENCE GROUP: PW02 - SYMPOSIUM ON
ADVANCES IN GI PHARMACOLOGY
MECHANISMS UNDERLYING THE MODULATION OF GUT
PERMEABILITY BY CANNABINOIDS
Abdussalam Alhamoruni(1), K Wright(2), A Lee(1), M Larvin(1),
S O’Sullivan(1)
(1) University of Nottingham, The School of Graduate Entry Medicine
and Health in Derby, Derby, UK
(2) Lancaster University, School of Health and Medicine, Lancaster, UK
The small bowel plays a vital role in preventing ingress of bacteria and
antigens whilst absorbing nutrients. Changes in tight junction proteins
have been reported with agents that modulate gut permeability. We have
previously shown that cannabinoids modulate intestinal permeability, and
the aim of the present study was to examine the underlying mechanisms.
Confluent Caco-2 cells (37oC, 5% CO2) were treated with phytocannabinoids or endocannabinoids (10 lM), with or without AM251 (100 nM,
CB1 receptor antagonist). Transepithelial electrical resistance measurements were made as a measure of permeability. mRNA levels of tight
junction proteins (ZO-1, Occludin and Claudin-1) were determined by
Taqman real time PCR. Signal transduction pathways were investigated
using PD98059 (1 lM, MAPK inhibitor), L-NAME (100 lM, NOS
inhibitor) and KT5720 (1 lM, PKA inhibitor). Over 48 hrs, phytocannabinoids significantly decreased permeability, while endocannabinoids
increased permeability. Both effects were inhibited by AM251. Both
phyto- and endocannabinoids increased the expression of ZO-1 mRNA.
Claudin-1 was downregulated by endocannabinoids only. Cannabinoids
do not affect Occludin expression. The effects of cannabinoids on tight
junction protein expression were inhibited by AM251. Inhibition of
MAPK signalling blocked the action of phytocannabinoids, but had no
effect on endocannabinoids. Inhibition of NOS significantly reduced the
effects of endocannabinoids only. Neither phyto- nor endocannabinoid
actions were affected by PKA inhibition. This study demonstrates that
cannabinoids modulate gut permeability and tight junction protein
expression via the CB1 receptor. The effect of phytocannabinoids
appears to be mediated through MAPK signalling pathways, while the
effect of endocannabinoids is NOS-dependent.
Paper No.: 3351
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
ROLE OF RECEPTOR RECYCLING IN THE RECOVERY OF
CA2 + SIGNALLING BY RECOMBINANTLY EXPRESSED
NEUROMEDIN U 2 RECEPTORS (NMU2)
Khaled Al-Hosaini, RAJ Challiss, GB Willars
University of Leicester, Department of Cell Physiology and Pharmacology, Leicester, UK
Neuromedin U (NmU) regulates many physiological and pathological
events via two family A, G protein-coupled receptors, NMU1 and
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
171
NMU2. These receptors show ~50% homology and couple to Gaq/11 and
Gai/o leading to increases of intracellular [Ca2 + ]i and inhibition of forskolin-stimulated cyclic AMP accumulation respectively. In HEK293 cells
recombinantly expressing either receptor, NmU binds essentially irreversibly (Brighton et al., 2004). Here, we examined the consequences of irreversible binding on recovery of NmU-mediated Ca2 + signalling
following an initial exposure in HEK-NMU2 cells. Further, we employed
confocal imaging of fluorescently labelled NmU (Cy3B-NmU-8) and a
C-terminally eGFP-tagged NMU2 to investigate ligand and receptor trafficking and their relevance to recovery of signalling. We confirmed irreversible ligand binding and demonstrated co-internalization of ligand and
receptor starting within 5min of exposure. Irreversible binding prevented
repetitive Ca2 + signalling when cells were exposed to NmU (10-30nM;
5min), washed and NmU re-applied. Full restoration of the response
required 6h recovery. Inhibition of endosomal acidification by monensin
reduced receptor recycling and slowed re-sensitization, while inhibition
of protein synthesis by cycloheximide had little effect on re-sensitization.
We developed a rapid, low pH wash that removed cell-surface receptorbound NmU, but had no deleterious effects on signal transduction.
Following exposure to NmU, brief acid wash restored an appreciable
NmU-mediated Ca2 + response within 5min with full re-sensitization
within 2-3h. Thus, receptor internalization is required for ligand removal
and cellular re-sensitization to NmU is dependent on receptor internalization and recycling rather than de novo protein synthesis. Brighton P.J.,
et al. (2004) Mol Pharmacol 66, 1544-1556.
Paper No.: 425
FOCUSED CONFERENCE GROUP: P10 - DRUGS FOR HALF
THE WORLD: PAEDIATRIC CLINICAL PHARMACOLOGY
RISK FACTORS FOR ABNORMAL NEWBORN HEARING IN
THE ERA OF EXTENDED DOSE INTERVAL AMIKACIN
ADMINISTRATION
Karel Allegaert, K Kneepkens, K Vandendriessche, F Debruyne, M Rayyan, G Naulaers, C Desloovere
University Hospitals Leuven, Department of Paediatrics, Leuven, Belgium
Introduction: There are conflicting reports on the impact of aminoglycoside administration on hearing impairment after NICU admission. We
therefore wanted to assess the impact of amikacin on hearing impairment
following implementation of an extended dose interval regimen.Methods:
Retrospective cohort study. Hearing impairment defined as at least unilateral hearing loss (>40dB), documented by brain stem evoked auditory
response (BERA). Risk factors (amikacin (yes/no, days, through, duration), vancomycin, ventilation, birth weight, bilirubinaemia, CMV, congenital malformations, meningitis, intracranial haemorrhage, convulsions)
were extracted from a prospective collected database and the original
source documents. Chi square test was used to compare risk factors
between neonates with or without hearing impairment. Results: 615 neonates were evaluated. Following re-assessment in 70 (11%), hearing
impairment was documented in 25 (4%). There was no significant difference in administration, through level, days or cumulative amikacin dose
between impaired and normal hearing neonates. Syndromal/congenital
malformations (10/25) and congenital CMV (4/25) infection were the
most relevant risk factors. Conclusion: Amikacin exposure (through
level, days, cumulative dose) in an extended dose interval approach was
not a risk factor, while syndromal/congenital malformation and CMV
infection were the most relevant risk factors.
Paper No.: 544
FOCUSED CONFERENCE GROUP: P10 - DRUGS FOR HALF
THE WORLD: PAEDIATRIC CLINICAL PHARMACOLOGY
DOES THE NEED FOR DOMPERIDONE ADMINISTRATION
SHORTLY AFTER DELIVERY AFFECT THE DURATION
BREASTFEEDING AFTER PRETERM DELIVERY?
Karel Allegaert(1), G Bosmans(2), M Vanhaver(3), V Cossey(1), A Debeer(1), C Vaanhole(1), K Janssens(1)
(1) University Hospitals Leuven, Department of Paediatrics, Leuven,
Belgium
(2) KU Leuven, Department of Clinical Psychology, Leuven, Belgium
(3) KH Limburg, Campus Hasselt, Limburg, Belgium
Introduction: There are several reports (da Silva, CMAJ 2001, Wan, Br J
Clin Pharmacol 2008) on the short term effects of domperidone administration as a galactagogue, but observations on the impact of domperidone
administration on the subsequent duration of breast feeding have not
been reported. Methods: Retrospective study in mothers of preterm neonates (<35 weeks gestational age), admitted in a single NICU in a
6 month’s time interval. Compare the duration of breast feeding after discharge of the baby and investigate if domperidone was a risk factor for
early failure after discharge (unpaired t test). Results: At delivery, 71/96
mothers of 89/118 newborns had the intention to breastfeed. 58 (81%)
were still breastfeeding at discharge. Mothers who underwent a caesarean
had an increased risk to terminate breastfeeding earlier (6.6 versus
15.5 weeks, p < 0.05) before discharge of the baby. 7 (10%) of the
mothers who were still breastfeeding at discharge received domperidone
shortly after delivery as a galactagogue, but domperione was not a
risk factor for earlier termination (17, SD 7 to 16, SD 11 weeks).
Conclusions: The need to initiate domperidone to facilitate mother’s milk
production shortly after delivery does not affect the duration of breastfeeding after discharge in former preterm neonates. The extensive interindividual variability in the duration of breast feeding after discharge
warrants prospective studies.
Paper No.: 2076
FOCUSED CONFERENCE GROUP:
P12 - ION CHANNELOPATHIES: NEW WINDOWS ON
COMPLEX DISEASE AND THERAPY
NON-GENOMIC EFFECTS OF OESTROGEN DERIVATIVES ON
BK CHANNELS
M Allen, Jacqueline Maher, A Hunter, J Mabley
University of Brighton, School of Pharmacy & Biomolecular Sciences,
Brighton, UK
There is growing evidence for steroid hormones having non-genomic
actions, including direct actions on the function of ion channels. Oestrogens and xenoestrogens are no exception. Oestrogens activate the BK
channel when the alpha subunit is associated with the beta 1 subunit (De
Wet H et al, Molecular Membrane Biology 2006; 23: 420-429). Interestingly, structurally unrelated antioestrogens such as tamoxifen also activate BK channels with associating beta 1 subunits. We have synthesised
novel steroidal oestrogens incorporating some of the structural motifs of
the non-steroidal antioestrogens. Oestrone analogues were tested for their
ability to activate BK channels in rat aortic rings pre-contracted with
phenylephrine. These analogues were also tested for their ability to activate BK currents in HEK 293 cells expressing BK alpha subunits alone
or in combination with beta 1 subunits. In addition, BK channels were
incorporated in planar lipid bilayers under voltage clamp conditions and
the ability of analogues to directly activate BK channels was determined.
All analogues of oestrone could relax aortic rings, with the following
order of increasing potency quatDME-oestrone, DME-oestrone, oestrone,
Oestrone-oxime. None of the ligands appeared to activate the BK channels in HEK 293 cells expressing alpha subunits alone, but oestrone
could activate BK currents in HEK 293 cells expressing alpha and beta 1
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
172
subunits. These electrophysiological findings were mirrored in our single
channel studies conducted in bilayers. Our studies suggest that oestrone
activates the BK channel directly but Oestrone-oxime has additional
endothelium-dependent actions.
Paper No.: 795
FOCUSED CONFERENCE GROUP: PW02 - SYMPOSIUM ON
ADVANCES IN GI PHARMACOLOGY
EVALUATION OF THE PROULCEROGENIC EFFECT OF
ATORVASTATIN ON INDOMETHACIN INDUCED GASTRIC
INJURY IN RATS
Ilkay Alp(1), O Uyanik(1), H Ozyogurtcu(2), A Gurel(2), P Atukeren(3),
K Gumustas(3), A Kapucu(4), C Demirci(4), O Ozdemir(1), BSU
Dogan(1)
(1) Istanbul University. Faculty of Pharmacy, Department of Pharmacolgy,
Istanbul, Turkey
(2) Istanbul University, Faculty of Veterinary Medicine, Department of
Pathology, Istanbul, Turkey
(3) Istanbul University, Cerrahpasa Medical Faculty, Department of
Biochemistry, Istanbul, Turkey
(4) Istanbul University, Biology Division, Department of Zoology,
Istanbul, Turkey
Statins are suggested to possess healing properties due to their antioxidant
and antiinflammatory effects in animal models of inflammatory diseases
such as intestinal ulcer and colitis. In contrary, a clinical report indicated
gastric ulcer formation in a patient by use of atorvastatin for 3 months.
We previously observed that acute or subchronic (5 days)administration
of atorvastatin have no favourable influence on indomethacin-induced
gastric ulcer lesions at 0,5 and 5mg/kg doses in rats. Interestingly, 50 mg/
kg of atorvastatin significantly aggravated indomethacin-induced ulcer
lesions with mechanisms possibly involving iNOS and increased neutrophil accumulation to gastric mucosa but independently from the inhibition
of mevalonate pathway. Whereas, a direct ulcerogenic effect of atorvastatin was not determined. Herein, we investigated gastric mucosal levels of
TNF-a, myeloperoxidase (MPO), reduced glutathione (GSH), and prostaglandin E2 (PGE2) in order to furtherly enlighten the proulcerogenic effect
of atorvastatin. Gastric ulcer scores obtained in atorvastatin (0.5-50mg/
kg,p.o) administered groups were compared with controls received vehicle plus indomethacin (30mg/kg,i.p). Enhanced gastric mucosal lesions in
rats receiving atorvastatin (50mg/kg) were associated with an increase in
gastric mucosal TNF-a and MPO levels whereas with a decrease in PGE2
and GSH levels. These findings showed that proulcerogenic effect of atorvastatin is associated with decreased defense mechanisms of gastric
mucosa as well as increased neutrophil infiltration and proinflamatory
cytokines. Excessive production of NO via iNOS may be correlated with
the generation of TNF-a in the proulcerogenic effect of atorvastatin.
The present work was supported by the Research Fund of Istanbul
University. Project No. T-76 15122006.
Paper No.: 1276
FOCUSED CONFERENCE GROUP: P06 - THE HEART GONE
WRONG; STABILIZATION OF CARDIAC FUNCTION
MODULATORY EFFECTS OF ZINC SUPPLEMENTATION ON
CARDIOVASCULAR COMPLICATIONS IN
STREPTOZOTOCIN-INDUCED DIABETES IN RATS
Nouf MI AL-Rasheed, NMI AL-Rasheed, HA Attia, LM Faddah
King Saud University, College of Pharmacy, Department of Pharmacology, Riyadh, Saudi Arabia
vent or delay cardiovascular complications. Diabetes was induced in rats
by a single intraperitoneal injection of streptozotocin (STZ, 40 mg kg-1).
After 45 days, diabetic group showed cardiovascular complications manifested by significant elevations of troponin-I (0.24 ± 0.03 ng ml-1 Vs.
0.023 ± 0.001, p < 0.001) as well as cardiac enzymes. Total cholesterol,
LDL-C and triglycerides were significantly increased compared to nondiabetic group. Diabetic group showed impaired oxidative status
manifested by a significant elevation of lipid peroxides, and reductions
of glutathione and L-ascorbic acid. Serum levels of TNF-a were significantly increased compared to non-diabetic group (236.5 ± 16.2 ng ml-1
Vs. 169.7 ± 2.9, p < 0.001). Nitric oxide (NO) and vascular endothelial
growth factor (VEGF) were markedly higher indicating endothelial dysfunction and disturbance of angiogenesis. Treatment with glibenclamide
(600 lg kg-1, i.p) resulted in an improvement in most of the deviated
biochemical parameters. However, combination of glibenclamide with
zinc sulfate (20 mg kg-1, i.p) showed more pronounced hypoglycemic
effect and a significant improvement in cardiac function as compared to
glibenclamide alone treated group. Moreover, LDL-C was significantly
decreased by the combination (34.46 ± 2.7 mg dl-1 Vs. 49.35 ± 4.1,
p < 0.01). Oxidative stress markers were improved by the combination.
Serum levels of TNF-a, NO and VEGF were significantly reduced by
the combination. These findings indicate that combination of zinc with
glibenclamide may potentiate hypoglycemic effect of glibenclamide and
limit the diabetic complications.
Paper No.: 890
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
ADVERSE EVENTS OF BENZNIDAZOLE IN CHILDREN WITH
CHAGAS DISEASE - A COHORT STUDY
Jaime Altcheh(2), G Moscatelli(2), S Moroni(2), F Garcia-Bournissen(1),
H Freilij(2)
(1) Hospital for Sick Children, Division of Clinical Pharmacology, Toronto, Ontario,Canada
(2) Servicio de Parasitologia, Hospital de Niños ‘R. Gutierrez’, Buenos
Aires,Argentina
Introduction: Chronic Chagas disease (caused by infection with Trypanosome Cruzi) is associated with severe long term complications
such as cardiac disease. Treatment of children with benznidazole can
cure the disease and prevent chronic infection. Benznidazole is associated to a high incidence of adverse drug reactions (ADRs) in adults,
including neuropathy and gastrointestinal symptoms. However, treatment in children seems to be associated with lower incidence and
severity of ADRs. We aimed to describe ADRs in a cohort of children
treated with benznidazole. Materials/Patients: A prospective study of
children with Chagas disease treated with benznidazole in a tertiary
pediatric centre in Buenos Aires, Argentina. Results and Conclusion:
107 children with Chagas were enrolled between 2004 and 2007. Mean
age of the patients was 6.9 years (CI 95%: 10 days - 19 years). 62
ADRs (in 44 children) were observed, most (77.3%) in children over
7 years of age. ADRs could be classified as mild (50; 80.6%), moderate (10; 16%) and severe (2; 3.2%). 75% (32) of the ADRs occurred
in the first 10 days of treatment. Skin was the most commonly affected
organ (50%), followed by the central nervous system (22%), and gastrointestinal tract (21%). 7 patients did not complete treatment with
benznidazole due to ADRs. Conclusion: Treatment of Chagas disease
with benznidazole is well tolerated in children. Incidence of ADRs is
strongly correlated with age (most ADRs occurred in children over
7 years). Severe ADRs frequently reported in adults were not observed
in our cohort.
Diabetes is one of most prevalent diseases that is now regarded as the
strongest risk factor for cardiovascular diseases. This study aimed to
investigate whether combination of zinc sulfate with glibenclamide preJournal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
173
Paper No.: 951
FOCUSED CONFERENCE GROUP: P10 - DRUGS FOR HALF
THE WORLD: PAEDIATRIC CLINICAL PHARMACOLOGY
POPULATION PHARMACOKINETICS STUDY OF
BENZNIDAZOLE IN CHILDREN WITH CHAGAS DISEASE
Jaime Altcheh(1), G Moscatelli(1), G Mastrantonio(3), S Moroni(1), N
Giglio(1), G Koren(2), H Freilij(1), F Garcia-Bournissen(2)
(1) Hospital de Niños ‘R Gutierrez’ de Buenos Aires, Servicio de
Parasitologia y Chagas, Buenos Aires, Argentina
(2) University of Toronto, Hospital for Sick Children, Division of
Clinical Pharmacology & Toxicology, Toronto, Ontario, Canada
(3) LASEICIC, Departamento de Quimica, Universidad de La Plata,
Argentina
Introduction: Chagas disease is caused by Trypanosoma cruzi, and leads
to long term cardiac and gastrointestinal complications. Treatment of
children with benznidazole is effective, but no information on its pharmacokinetics and no pediatric formulation are available. Patients and
Methods: Population pharmacokinetics study in children with Chagas
disease, aged 2-12 years (clinical trials.gov #NCT00699387). Enrolled
children (target N = 50) were treated with oral benznidazole 5–8 mg/kg/
day BID for 60 days, as per current pediatric Chagas protocols. At least
3 blood samples were obtained after the first dose, at steady state or after
the last dose. Benznidazole was measured in blood by HPLC. Results
and Discussion: 25 children were enrolled to date. Mean age was 6 years
(range 2.2–12 years). Median benznidazole dose was 6.3 mg/kg/day.
Median steady-state (trough) benznidazole concentration was 0.99 mg/L,
and highest observed concentration (Cmax) was 11.07 mg/L. All children treated had a positive response, with negativization of PCR for T.
cruzi DNA, and marked decrease in anti T. cruzi antibody titers.
Observed benznidazole concentrations in children were markedly lower
than those reported in adults (treated with comparable mg/kg doses). In
spite of these lower concentrations children treatment was effective and
well tolerated, with few adverse drug reactions (ADRs). Unlike adults,
ADRs in children are uncommon, and severe ADR are rare. It is possible
that the lower blood concentrations, while still providing therapeutic
effect, may be responsible for this lower incidence of ADRs. If confirmed, our results would suggest that dosing modifications in adults may
be beneficial.
Paper No.: 2982
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
5-(4-METHANESULPHONYL-PHENYL)-THYAZOLE
DERIVATIVES: NEW TOOLS FOR CHRONIC
INFLAMMATORY DISEASE BIOTHERAPY
Melchor Alvarez de Mon(1,2), F Ledo(3), E SanAntonio(1), E Sanz(1),
R Corcostegui(3), A Berisa(3), V Rubio(3), ML Lucero(3), A Muñoz(3),
A de-la-Hera(1,4)
(1) Alcalá University, Madrid, Spain
(2) Principe de Asturias University Hospital, Department of Medicine,
Madrid, Spain
(3) FAES FARMA R&D and Innovation, Spain
(4) Molecular Medicine Institute (IMMPA-CSIC/UAH), Spain
Antagonists of TNF-a binding to its receptor are successful examples of
single-cytokine targeted biotherapies of chronic inflammatory diseases
such as reumathoid arthritis (RA). However, a large fraction of those
patients are non-responders to a given mono-biotherapy. It would be
highly desirable to find small molecule drugs that selectively block not
only TNF-a production but also the production of other pathogenic cytokine(s) to assess new poly-biotherapy strategies with novel single drug
tools. Using multiplex immunoassays of cytokines secreted by healthy
human peripheral blood mononuclear cells (PBMC) stimulated with
either CD3 + CD28 antibodies or LPS, we have discovered that
5-(4-Methanesulphonyl-phenyl)-thiazole derivatives are a new drug family able to selectively inhibit both prototype Th1 cytokine IFN-c and
TNF-a production, as exemplified by FAES compound 12 and other
structurally related drugs. Compound 12 inhibited TNF-a mRNA transcription in purified LPS-stimulated human monocytes in vitro, and
reduced TNF-a plasma levels in mice injected with LPS. It also ameliorated the clinical scores of arthritis in a collagen-induced arthritis mouse
model of human RA. Notably, it selectively reduced TNF-a and INF-c
production by PBMC stimulated with LPS and CD3 + CD28 respectively also when PBMC was from RA patients with active disease that
had been resistant to methotrexate treatment, and were to enter anti-TNFa biotherapies. It is now possible to examine putative advantages of
poly-biotherapy of inflammatory diseases with complex pathogenic cytokine deregulation with novel single drug tools.
Paper No.: 1403
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
CARDIAC AND RENAL EFFECTS OF TITYUS SERRULATUS
VENOM AND A NEW ISOLATED PEPTIDE
Renata S Alves(1), ARC Jorge(2), RM Ximenes(2), A Havt(2), NRF
Nascimento(3), MGR Queiroz(1), AMC Martins(1), MH Toyama(4), HSA
Monteiro(2)
(1) Federal University of Ceara, Department of Clinical and
Toxicological Analysis, Ceara, Brazil
(2) Federal University of Ceara, Department of Physiology and
Pharmacology, Ceara, Brazil
(3) Ceara State University, Superior Institute of Biomedical Sciences,
Ceara, Brazil
(4) Paulista State University (UNESP), São Paulo, Brazil
Tityus serrulatus is a very dangerous scorpion species accounting for
fatal stings, especially among children in Brazil. The aim of the present
work was to determine the effects of Tityus serrulatus venom and a new
peptide, recently isolated from scorpion venom, weighing 3KDa on cardiac function in vivo. The effects of the crude venom (TsV; 0,6lg/min)
or toxin (PNTsV; 0,2lg/min; n = 6) were studied on mean arterial pressure (MAP), heart rate (HR), urinary flow (UF), glomerular filtration rate
(GFR) and renal cortex blood flux (RCBF), estimated by the clearance of
p-aminohippurate by, in vivo experiments. Data were analyzed by ANOVA and Student t-test using *p < 0.05. Both TsV and PNTsV increased
MAP (C = 102.5 ± 3.5 vs TsV = 134.2 ± 6.3 and PNTsV = 116.0 ±
4.2 mmHg*) but only PNTsV increased HR (C = 239.8 ± 29.35 vs
PNTsV = 355.0 ± 25.30 bpm*). The peptide also increased UF with a
maximum effect at 90 minutes (C = 8.9 ± 1.8 vs PNTsV = 33.8 ±
6.2 lL/min*). While TsV was found to reduce GFR, PNTsV actually
increased GFR (C = 0.18 ± 0.08 vs TsV = 0.03 ± 0.01* vs PNTsV =
0.32 ± 0.05 lL/min/100g) and the renal cortex blood flux was reduced
after the crude venom treatment. In conclusion, In vivo experiments
showed how the crude venom and its isolated components act increasing
arterial blood pressure, heartbeat frequency and altered renal function.
Paper No.: 1140
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
EVALUATION OF THE ANTICONVULSANT ACTIVITY OF
ZANTHOXYLUM CAPENSE (THUNB.) HARV. (RUTACEAE) LEAF
METHANOL EXTRACT IN MICE
George Amabeoku, CG Kinyua
University of the Western Cape, School of Pharmacy, Cape Town, South
Africa
Zanthoxylum capense (Thunb.) Harv. (Rutaceae) is used in South Africa
by traditional medicine practitioners to treat various ailments including
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
174
epilepsy. However, the claims of therapeutic success of the plant species
in therapy have not been subjected to any scientific scrutiny. The main
aim of the study, therefore, was to investigate the anticonvulsant activity
of the leaf methanol extract of Z. capense in mice. Chemically-induced
convulsion tests were used to assess the anticonvulsant activity of Z.
capense in mice. The acute toxicity and HPLC studies and the phytochemical analysis of the plant species were also carried out using well
established protocols. Leaf methanol extract of Z. capense and not methanol alone, significantly antagonized seizures induced by pentylenetrazole (PTZ), bicuculline, picrotoxin and strychnine while only significantly
delaying the onset of N-methyl-DL-aspartic acid (NMDLA)-induced seizures. Phenobarbitone, diazepam and not phenytoin, significantly antagonized seizures induced by PTZ, bicuculline and picrotoxin but all three
drugs did not alter NMDLA-induced seizures. Phenobarbitone significantly attenuated strychnine-induced seizures. The LD50 value obtained
following oral administration of the leaf methanol extract of Z. capense
was above 3200 mg/kg. The phytochemical analysis of the plant species
revealed the presence of alkaloids, triterpene steroids, reducing sugars,
saponins, tannins and quinones while the HPLC study revealed distinct
characteristic peaks .The data obtained indicate that the leaf methanol
extract of Z. capense has anticonvulsant activity which may probably
involve both Gabaergic, glutaminergic and glycinergic mechanisms. The
relatively high LD50 value obtained following oral administration shows
that the plant extract is non-toxic in mice.
Paper No.: 2310
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
PROTECTIVE EFFECT OF LAFUTIDINE AGAINST
LOXOPROFEN-INDUCED SMALL INTESTINAL LESIONS IN
RATS
Kikuko Amagase, R Kawahara, S Takayama, T Sugihara, A Ochi, S
Kato, K Takeuchi
Kyoto Pharmaceutical University, Division of Pathological Sciences,
Department of Pharmacology & Experimental Therapeutics, Kyoto,
Japan
The effect of lafutidine, a histamine H2-receptor antagonist with the
mucosal protective action mediated by sensory neurons, on loxoprofeninduced smalli ntestinal lesions was examined in rats, in comparison with
other H2-receptor antagonists and an proton pump inhibitor. Male SD
rats were administered loxoprofen Na and killed 24 hr later. Lafutidine,
cimetidine, famotidine or 16,16-dimethyl prostaglandin E2 (dmPGE2)
was given, 0.5 hr before and 6 hr after the administration of loxoprofen,
while ampicillin was given 24 hr and 0.5 hr before. Omeprazole was
given 0.5 hr before loxoprofen treatment. Loxoprofen dose-dependently
caused severe lesions in the small intestine, with the up-regulation of
inducible nitric oxide synthase (iNOS) expression, and an increase of
myeloperoxidase (MPO) activity as well as enterobacterial invasion in
the mucosa. These events were all prevented by dmPGE2 and ampicillin.
Likewise, lafutidine also prevented these lesions with the concomitant
suppression of up-regulaton in iNOS expression, MPO activity as well as
bacterial invasion. However, neither cimetidine, famotidine nor omeprazole had any effect on these lesions. The protective effect of lafutidine
was significantly attenuated by chemical ablation of capsaicin-sensitive
sensoryneurons. This agent increased by itself intestinal mucus secretion
and significantly suppressed the decreased mucus response to loxoprofen.
These results suggest that lafutidine protects the small intestine against
loxoprofen-induced lesions mediated by capsaicin-sensitive sensory neurons. This effect may be essentially attributable to the increase of mucus
secretion, resulting in the inhibition of bacterial invasion and iNOS
expression in the intestinal mucosa.
Paper No.: 2332
FOCUSED CONFERENCE GROUP: PW04 - ANTIOXIDANTS AS
THERAPEUTIC TARGETS
AGGRAVATION BY SELECTIVE SEROTONIN RE-UPTAKE
INHIBITOR OF ANTRAL ULCERS INDUCED BY
INDOMETHACIN IN RATS: PATHOGENIC IMPORTANCE OF
IMPAIRED ANTI-OXIDATIVE SYSTEM
Kikuko Amagase, A Kojo, C Izuhara, K Nukui, A Tanaka, K Takeuchi
Kyoto Pharmaceutical University, Division of Pathological Sciences,
Department of Pharmacology & Experimental Therapeutics, Kyoto,
Japan
Clinical studies suggested a risk of gastric adverse reactions on the concomitant use of selective serotonin re-uptake inhibitors (SSRIs) with nonsteroidal anti-inflammatory drugs (NSAIDs). We demonstrated the
adverse effect of paroxetine, one of SSRIs, in the rat stomachs when
co-administered with indomethacin, and investigated the mechanism of
this adverse effect. We also examined the effects of various agents on the
antral ulcers induced by indomethacin plus paroxetine. The animals were
fed chow for 1 h after 24 h fasting, then given indomethacin and killed
6 h later. SSRIs were given 30 min before indomethacin. Indomethacin
caused antral ulcers in refed rats. Paroxetine dose-dependently aggravated the severity of these ulcers. The worsening effect of paroxetine
was attenuated by ondansetron, a selective 5HT3 antagonist. The combined treatment with indomethacin and paroxetine caused a significant
decrease in mucosal superoxide dismutase activity as well as gluthathione content in the antral mucosa. The development of these antral ulcers
was prevented by omeprazole, pepstatin as well as rebamipide. These
results suggest that SSRIs exert a harmful influence on the antral mucosa
through activation of 5HT3 receptors when given with NSAIDs, resulting in aggravation of antral ulcers, and the process of aggravation may
be modified by the corrosive action of acid/pepsin as well as the
decreased anti-oxidative system. Rebamipide, a mucosal protective drug
developed in Japan, is useful for preventing the antral ulcers caused by
the combined administration of NSAID and SSRI, probably by its
scavenging action as well as its anti-inflammatory effect.
Paper No.: 2333
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
ROLES OF COX/PGE/EP4 RECEPTORS IN HEALING OF
INDOMETHACIN- INDUCED SMALL INTESTINAL LESIONS
IN RATS: RELATION TO VEGF EXPRESSION
Kikuko Amagase, A Kojo, N Abe, R Hatazawa, M Tanigami, K Takeuchi
Kyoto Pharmaceutical University, Division of Pathological Sciences,
Department of Pharmacology & Experimental Therapeutics, Kyoto,
Japan
Endogenous prostaglandins (PGs) play an important role in the healing
of small intestinal lesions. We investigated the mechanism by which PGs
promote the healing of intestinal lesions in rats, including the determination of EP receptor subtype(s) involved in this action and the relation
with VEGF expression. Intestinal lesions of male SD rats were induced
by indomethacin (10 mg/kg). Indomethacin (2 mg/kg) was given for
6 days, starting 1 day after ulceration. AE1-329 (EP4 agonist) was given
for 6 days in the presence of indomethacin. For studying VEGF expression, indomethacin and AE3-208 were given for 2 days, while AE1-329
was given together with indomethacin. Indomethacin caused severe
lesions in the small intestine, and the lesions healed within 7 days. The
healing of these lesions was significantly impaired by the repeated treatment of indomethacin. The healing impairment effect of indomethacin
was mimicked by the EP4 antagonist AE3-208 given for 6 days and
reversed by the co-administration of the EP4 agonist. The COX-2 mRNA
expression in the small intestine was up-regulated after ulceration with
increase of PGE2 content. VEGF expression was also increased after
ulceration. The expression of VEGF and the number of microvessels in
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
175
the mucosa was down-regulated by indomethacin and AE1-208, while
these effects of indomethaicn were reversed by the co-treatment with
AE1-329. These results confirmed an important role of endogenous
PGE2 in the healing of small intestinal lesions and further demonstrated
that this action is associated with the up-regulation of VEGF expression
mediated by the activation of EP4 receptors.
Paper No.: 1358
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
SPECIES DIFFERENCE IN CALCIUM DEPENDENCY OF
CROMAKALIM INDUCED RELAXATION IN PORCINE AND
HUMAN CORONARY ARTERIES
Nora Ambrus(1), J Pataricza(1), I Krassoi(2), Z Marton(1), M Bitay(3),
A Varro(1,2), GJ Papp(1,2)
(1) University of Szeged, Department of Pharmacology and Pharmacotherapy, Szeged Hungary
(2) Hungarian Academy of Sciences, University of Szeged, Division of
Cardiovascular Pharmacology, Szeged, Hungary
(3) University of Szeged, 2nd Department of Internal Medicine and Cardiological Center Department of Cardiosurgery, Szeged, Hungary
Introduction: We have shown that relaxation induced by levosimendan, a
potassium channel opener inodilator drug, is decreased with the elevation
of extracellular calcium concentration in porcine and human coronary
arteries (Krassói I et al, Cardiovasc. Drug. Ther. 2000; 14:691-693). Our
aim was to study the effect of cromakalim, another potassium channel
opener, in isolated human and porcine coronary arteries, and the possible
modulation of cromakalim-induced vasorelaxation at low and high extracellular calcium concentrations. Materials: Isometric tensions of porcine
and human coronary arteries were measured in isolated organ baths.
Contractions were induced by potassium chloride (30 mM) and the prostaglandin analogue, U46619 (0.7 lM). Relaxations by cromakalim
(0.0125-10.0 lM) and 1-EBIO (1-80 lM), an opener of calcium activated potassium channels, at low (1.5 mM) and high (7.5 mM) extracellular calcium concentrations were investigated. Endothelium intact and
deprived arterial preparations were used, and their functions were controlled by bradykinin (1 lM, porcine) and acetylcholine (0.1 lM,
human). Results: High extracellular calcium ([Ca2 + ]o) concentration significantly enhanced the relaxing effect and the potency of cromakalim
both in porcine and human coronary arteries. In porcine coronary artery,
high [Ca2 + ]o potentiated the effect of cromakalim and 1-EBIO only in
the presence of endothelium. In human coronary artery, the elevation of
[Ca2 + ]o enhanced the relaxation of cromakalim both in the presence and
in the absence of functional endothelium. Conclusion: Results indicate
that the coronary dilating effect of cromakalim largely depends on the
species and is modulated by extracellular calcium concentration with a
partly endothelium dependent manner.
Paper No.: 502
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
IMPROVING THE ANTIBACTERIAL ACTIVITY OF
LYSOZYME BY CONJUGATION WITH POLYSACCHARIDES
Mahmoud Aminlari
Shiraz University, Department of Biochemistry, Shiraz, Iran
A widespread trend in biomedical sciences is replacing chemical antimicrobials with naturally occurring agents. Lysozyme is an enzyme that
destroys bacteria by lysis of the bacterial cell wall. Lysozyme is found
abundantly in nature and is effect against G-positive bacteria but not
against G-negatives due to the presence of the LPS layer in outer membrane
of the latter. The purposes of this research was to glycosylate lysozyme
with polysaccharides and to study its effect on G-positive and G-negative
bacteria in vitro. Lysozyme and polysaccharides were allowed to react
under mild conditions (60 0C for one week), the glycosylated products
were separated by ion-exchange and gel filtration chromatography and
the identity of the products was confirmed by electrophoresis. The antimicrobial properties of lysozyme-polysaccharide conjugates against test
microorganisms in culture media was evaluated. Under optimum conditions, 3.0 moles dextran, 1.2 moles dextran sulfate and 3.6 mmoles b-glucan were coupled to 1 mole lysozyme. Glycosylated lysozymes exhibited
increased solubility at different temperatures and pH’s. Evaluation of the
lysozyme-polysaccharide conjugates against test microorganisms (S. aureus
and E. coli) in culture media indicated an increase in antimicrobial
activity in a concentration-dependent manner such that at 400 lg/ml,
Lysozyme-dextran conjugate decreased S. aureus and E. coli by 2 and 4
log cycle, respectively. Corresponding values for dextran sulfate were 0.6
and 1.1 and for b-glucan were 1 and 3. These results might increase the
applicability of lysozyme as a natural antimicrobial ingredient against a
broader spectrum of bacteria in different pharmaceutical preparations.
Paper No.: 2478
FOCUSED CONFERENCE GROUP: P02 - TRANSMEMBRANE
TRANSPORT: PERSPECTIVES FOR DISEASE AND DRUG
DISCOVERY
CYCLOSPORINE A, BUT NOT TACROLIMUS, SHOW
RELEVANT INHIBITION OF OATP1B1 MEDIATED
TRANSPORT OF ATORVASTATIN
Rune Amundsen(1), H Christensen(1), B Zabihyan(1), A Åsberg(1)
School of Pharmacy, University of Oslo, Department of Pharmaceutical
Biosciences, Oslo, Norway
The aim of this study was to investigate and compare the potential of CsA
and Tac to inhibit cellular uptake of atorvastatin mediated by the liver specific organic anion transporting polypeptide 1B1 (OATP1B1) in vitro.
Due to a high incidence of dyslipidaemia following solid organ transplantation, HMG-CoA reductase inhibitors (statins) are now frequently used.
The calcineurin inhibitor cyclosporine A (CsA) however increases systemic exposure of atorvastatin several-fold, an effect not observed with
tacrolimus (Tac), another calcineurin inhibitor. The inhibition of atorvastatin transport via OATP1B1 by CsA and Tac was investigated in
transfected HEK293 cells. An in vitro-in vivo extrapolation (IVIVE) was
performed to estimate the potential for CsA and Tac to inhibit OATP1B1
mediated transport in vivo. CsA inhibited OATP1B1 mediated uptake of
atorvastatin about 90-fold more efficiently than Tac, with inhibition constants (Ki) of 0.014 ± 0.003 lM and 1.30 ± 0.27 lM, respectively. The
IVIVE showed that clinically obtainable concentrations of CsA, but not
Tac, inhibit OATP1B1 transport of atorvastatin to a relevant degree. The
estimated degree of inhibition by CsA ranged from 28-77% during a dosing interval while it was below 1% for Tac when considering free concentrations and assuming competitive inhibition. This does however not fully
explain the clinically observed interaction with CsA, suggesting a more
complex inhibitory mechanism may be present. This study provides
evidence that OATP1B1 inhibition is a relevant mechanism for the interaction observed between CsA and atorvastatin. Tac on the other hand does
not have the properties to induce a similar interaction in vivo.
Paper No.: 2809
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
IN-VITRO CHARACTERISATION OF THE CLOPIDOGREL
BIOACTIVATION PATHWAYS BY CYTOCHROMES P450 AND
POTENTIAL DRUG-DRUG INTERACTIONS
Virginie Ancrenaz(1), Y Daali(1), C Samer(1), M Besson(1),
P Fontana(2), P Dayer(1), J Desmeules(1)
(1) Geneva University Hospitals, Department of Clinical Pharmacology
and Toxicology, Geneva, Switzerland
(2) Geneva University Hospitals, Division of Angiology & Hemostasis,
Geneva, Switzerland
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
176
Aim: Clopidogrel is a thienopyridine widely used to prevent thrombosis
in patients with acute coronary syndrome or after a stent implantation,
often used in association with aspirin. It is a prodrug which needs to be
activated by CYP450s in two steps to form a pharmacologically active
metabolite. The aim of our work was to characterize the cytochrome
P450 isoforms involved in the two steps of the bioactivation of clopidogrel as well as the importance of each cytochrome in the complete bioactivation mechanism. Methods: Human liver microsomes with selective
CYP450s inhibitors and cDNA expressed human CYP isoforms were
used for clopidogrel metabolism assessment. The intermediate (2oxoclopidogrel) and the active metabolites were analyzed with an HPLC-MS
method. A derivatization method was needed to avoid a rapid degradation of the active metabolite before its determination. Results: The results
obtained with microsomes and human CYP isoforms indicated that
CYP2C9, CYP1A2 and CYP2C19 were the most involved in the production of 2oxo-clopidogrel. CYP3A4, CYP2B6 and CYP2C19 contributed to the formation of the pharmacologically active metabolite from
2oxo-clopidogrel. In the inhibition studies with specific chemical inhibitors, the formation of the active metabolite from clopidogrel was in
accordance with the results obtained in the separate two steps. Conclusion: CYP2C19 is involved in both steps required in the formation of
clopidogrel active metabolite. CYP1A2 and CYP2C9 contributed
substantially to the first step while CYP3A4 was the most important in
the second step. Hence, genetic polymorphisms or drug-drug interactions
involving these pathways should be considered during clopidogrel
therapy.
Paper No.: 2071
FOCUSED CONFERENCE GROUP:
P04 - PHARMACOEPIDEMIOLOGY, CURRENT
CONTROVERSIES AND OPPORTUNITIES
ASSOCIATION BETWEEN FDA DRUG CLASSIFICATION OF
FETAL RISKS AND THE RISK OF MAJORCONGENITAL
MALFORMATIONS: A REGISTER-BASED NATIONWIDE
COHORT STUDY
Jon T Andersen(1,2), M Petersen(1,2), NL Andersen(3), S Afzal(1,2), K
Broedbaek(1,2), BR Hjelvang(1,2), C Torp-Pedersen(4,5), HE Poulsen(1,2,5)
(1) Copenhagen University Hospital, Rigshospitalet, Laboratory of
ClinicalPharmacology, Copenhagen, Denmark
(2) Bispebjerg Hospital, Department of Clinical Pharmacology, Copenhagen, Denmark
(3) Psychiatric Center Bispebjerg, Copenhagen, Denmark
(4) Gentofte University Hospital, Department of Cardiology, Gentofte,
Copenhagen,Denmark
(5) University of Copenhagen, Faculty of Health Science,, Copenhagen,
Denmark
Introduction: Since 1979 United States Food and Drug Administration
(FDA) has classified drugs according to their fetal risk when used during
pregnancy. The categories are A, B, C, D, and X; X defining the highest
evidence of risk. The aim of the study was to investigate whether first trimester dispensing of one or more drugs within each of the categories
was associated with an increased risk of major malformations. Materials:
We conducted a nationwide cohort study using the Danish Fertility Database to identify every mother of children born in Denmark 1997-2005.
All prescriptions dispensed during pregnancy were identified through the
National Prescription Register and categorized according to FDAs classification. We included pregnant women with drugs dispensed only in the
first trimester and drug-use designated to only one of the FDA-categories
or a ‘not classified’ category. All diagnoses of major congenital malformations were identified through the National Hospital Register. All
groups where compared to women with no dispensing of drugs during
pregnancy. Results: The preliminary results did not show any significantly increased risk of major malformations associated with dispensing
of drugs in the six categories. The odds ratio adjusted for age and education of having a child with a major malformation when redeeming a prescription in the respective FDA categories in the first trimester where
A:1.12(CI95% 0.76-1.66), B:1.06(CI95% 0.96-1.16),
0.92-1.20), D:1.13(CI95% 0.97-1.31), X:1.01(CI95%
classified’:1.14(CI95% 0.99-1.32). Conclusion: FDAs
fetal risks due to pharmaceuticals does not seem useful
risk of major malformations.
C: 1.05(CI95%
0.76-1.35), ‘not
classification of
in predicting the
Paper No.: 2072
FOCUSED CONFERENCE GROUP:
P04 - PHARMACOEPIDEMIOLOGY, CURRENT
CONTROVERSIES AND OPPORTUNITIES
DRUG USE DURING PREGNANCY: A REGISTER-BASED
NATIONWIDE COHORT STUDY OF FDA’S DRUG
CLASSIFICATION OF FETAL RISKS
Jon Andersen(1,2), M Petersen(1,2), N Andersen(3), S Afzal(1,2), K
Broedbaek(1,2), B Hjelvang(1,2), C Torp-Pedersen(4,5),
H Poulsen(1,2,5)
(1) Copenhagen University Hospital, Rigshospitalet, Laboratory of ClinicalPharmacology, Copenhagen, Denmark
(2) Bispebjerg Hospital, Department of Clinical Pharmacology, Copenhagen, Denmark
(3) Psychiatric Center Bispebjerg, Copenhagen, Denmark
(4) Gentofte University Hospital, Department of Cardiology, Gentofte,
Copenhagen,Denmark
(5) University of Copenhagen, Faculty of Health Science, Copenhagen,
Denmark
Introduction: The knowledge of drugs used during pregnancy is limited.
The aim of the study was to describe the use of drugs during pregnancy
according to the United States Food and Drug Administration (FDA)
classification of fetal risks. Materials: We conducted a nationwide cohort
study using the Danish Fertility Database to identify every mother of
children born in Denmark 1997-2005. All prescriptions dispensed during
pregnancy were identified through the National Prescription Register and
categorized according to FDA’s classification of fetal risk. The categories
are A, B, C, D, and X; X defining the highest evidence of risk. All drugs
not classified were grouped as one. Results: We identified 588,832 children born in the study period. During pregnancy 63.6% (374,654) of the
women redeemed a prescription on any drug; 3.1% (18,438) were dispensed a prescription on an X-classified drug during the entire pregnancy
and 3.0% (17,805) during the first trimester. D-classified drugs were
redeemed by 10.8% (63,687) of the women during pregnancy and by
8.1% (47,729) during the first trimester whereas C-classified drugs were
dispensed to 30.5% (179,423) during the pregnancy. 38.4% (226,082)
redeemed a prescription on a drug classified as a potential fetal risk (category C, D or X) during the entire pregnancy and 20.4% (123,070) during
the first trimester. Conclusion: The majority of the pregnant women
redeemed prescriptions on drugs during pregnancy. Furthermore, a large
proportion of the women were dispensed drugs with potentially fetal risk
throughout the pregnancy and during the first trimester.
Paper No.: 1240
FOCUSED CONFERENCE GROUP: P17 - NEW APPROACHES
AND TARGETS IN PSYCHIATRY
THE ANTIPSYCHOTIC-LIKE EFFECT OF MYRICITRIN, A
PKC INHIBITOR IN MICE
Roberto Andreatini(1), M Pereira(1), L Pereira(1), MABF Vital(1), ARS
dos Santos(2), MG Pizzolatti(3)
(1) Universidade Federal do Paraná, Department of Pharmacology
(2) Universidade Federal de Santa Catarina, Department of Physiological
Sciences
(3) Universidade Federal de Santa Catarina, Department of Chemistry
Protein kinase C (PKC) has been associated with bipolar disorder, since
manic patients show elevated platelet membrane-bound/cytosol PKC
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
177
ratio and antimanic drugs (lithium, valproate and tamoxifen) inhibit PKC
activity (Zarate and Manji, CNS Drugs 2009; 23:569-582). Moreover,
these antimanic drugs blocked amphetamine-induced hyperlocomotion,
an animal model of mania (Einat H et al, J Psychopharm 2006;20:714722; Sabioni et al, Prog Neuropsychopharmacol Biol Psychiatry
2008;32:1927-31). However, this animal model is also employed in the
study of antipsychotic drugs and psychotic symptoms can be seen in
manic patients. Thus, the present study evaluates the effect of mirycitrin
(a flavonoid naturally occurring in several plants, e.g. genus Eugenia), a
PKC inhibitor (Meotti et al, J Pharmarcol Exp Ther 2006;316:789-96),
in animal models for antipsychotic-like action. Male Swiss mice treated
with mirycitrin (5 and 10 mg/kg), olanzapine (10 mg/kg; positive control) or their vehicles were tested in: (a) apomorphine-induced climbing;
(b)apomorphine-induced stereotypy; (c) bar catalepsy test (with and without haloperidol co-administration); (d) spontaneous locomotor activity.
Different groups of mice were used in each model. Mirycitrin (10 mg/
kg) and olanzapine blocked stereotypy and climbing induced by apomorphine (1.0 mg/kg). When administered alone, mirycitrin did not produce
catalepsy (up to 30 mg/kg), but it increased the catalepsy induced by
haloperidol (1.0 mg/kg). At the doses tested, mirycitrin did not alter
locomotor activity. These results suggest that mirycitrin has an antipsychotic-like effect at a dose that does not induce extra-pyramidal
side-effects and that PKC inhibition could be a new strategy for the
search of new antipsychotic drugs.
Paper No.: 1274
FOCUSED CONFERENCE GROUP: P17 - NEW APPROACHES
AND TARGETS IN PSYCHIATRY
MAGNESIUM SULPHATE AND SODIUM VALPROATE BLOCK
METHYLPHENIDATE-INDUCED HYPERLOCOMOTION, AN
ANIMAL MODEL OF MANIC STATE
Roberto Andreatini(1), FJ Barbosa(1,2), B Hesse(1), RB de Almeida(1),
IP Baretta(1,3), R Boerngen-Lacerda(1)
(1) Universidade Federal do Paraná, Department of Pharmacology, Curitiba, Brazil
(2) Universidade Federal do Paraná, Department of Forensic Medicine
and Psychiatry, Curitiba, Brazil
(3) Universidade Paranaense, Instituto de Ciências Biológicas, Médicas e
da Saúde Farmacologia, Umuarama, Brazil
Magnesium sulphate (MgSO4) is used to treat and prevent eclamptic
seizure and several anticonvulsants (e.g. sodium valproate) are clinically
effective antimanic drugs. Some small sample-size, open and/or add-on
clinical studies with MgSO4 found improvement of manic symptoms
(Heiden A et al, Psychiatry Res, 1999;89:239-246; Giannini AJ et al.,
Psychiatry Res 2000; 93:83-7). Thus, these studies suggest a potential
antimanic effect of MgSO4. Psychostimulant-induced hyperlocomotion
is proposed as an animal model for the study of antimanic drugs (Einat H
et al, J Psychopharm 2006;20:714-722; Sabioni et al, Prog Neuropsychopharmacol Biol Psychiatry 2008; 32:1927-31). Therefore, we
evaluated the effect of MgSO4 and sodium valproate (positive control)
on methylphenidate-induced hyperlocomotion in mice exposed to the
open-field. Acute MgSO4 (300-400 mg/kg), but not valproate (100300 mg/kg), prevents increase locomotor activity induced by methylphenidate (2.5 mg/kg). On the other hand, repeated (14 days) valproate
(300 mg/kg), but not MgSO4 (400 mg/kg), blocked methylphenidateinduced hyperlocomotion. These effects were seen at doses of valproate
or MgSO4 that did not change locomotor activity per se. In conclusion,
acute magnesium sulphate shows antimanic-like effect in this model,
which fits with add-on small clinical studies that employed a short-term
treatment. These results support further studies for investigating MgSO4
as a potential treatment for acute mania. Possible mechanisms of this
action are: NMDA non-competitive antagonism, PKC inhibition and/or
change in calcium effects (Murck H, Nutr Neurosci 2002;5:375-389).
However, this antimanic-like effect of MgSO4 disappears after repeated
treatment, which could be specific for the model used or a side-effect
of MgSO4.
Paper No.: 3114
FOCUSED CONFERENCE GROUP:
P11 - G PROTEIN-COUPLED 7TM RECEPTORS: FROM
MOLECULAR TO PHYSIOLOGICAL FUNCTION
FRET-BASED EVIDENCE FOR LIGAND-INDEPENDENT
PREASSOCIATION OF 5-HT7 SEROTONIN RECEPTORS WITH
GS
Kjetil W Andressen(1), L Hommers(2), AH Ulsund(1), MJ Lohse(2),
KA Krobert(1), M Bünemann(2), FO Levy(1)
(1) University of Oslo, Department of Pharmacology and Center for
Heart Failure Research, Oslo, Norway
(2) University of Würzburg, Institute of Pharmacology and Toxicology,
Würzburg, Germany
In classic pharmacology, G protein-coupled receptors (GPCRs), G proteins and effectors interact sequentially by random collision. Signalling
through GPCRs is both specific and very rapid, indicating that the proteins are efficiently organized. Currently, there are two prevailing views
on how receptors, G proteins and effectors are organized; either preassembled in one complex (preassociated/precoupled) or interacting after
receptor-activation (collision-coupling) in discrete microdomains. Our
previous comparison of the Gs-coupled 5-HT4 and 5-HT7 serotonin
receptors revealed fundamental differences in G protein activation.
Whereas 5-HT4 receptor function is consistent with collision-coupling,
the unusual pharmacological properties exhibited by 5-HT7 receptors can
best fit a model with receptor and Gs preassociated in the absence of
ligand. To test if 5-HT7 receptors preassociate with Gs, we used Fluorescence Resonance Energy Transfer (FRET) to compare the interaction
between YFP-labeled 5-HT4, 5-HT7 or b1-adrenergic receptors with
CFP-labeled G proteins in HEK293 cells. Agonist-activation of 5-HT4
or b1-adrenergic receptors increased FRET (indicating recruitment of
Gs). In contrast, 5-HT7 receptor activation decreased FRET in a concentration-dependent manner. The dissociation of G protein from 5-HT7
receptors had kinetics identical to G protein activation (dissociation
between Ga and Gbc determined by FRET), but slower than recruitment
of G protein to 5-HT4 and b1-adrenergic receptors. In addition, results
from fluorescence recovery after acceptor photobleaching were consistent
with a stable complex between 5-HT7 and G protein. Taken together,
these data provide strong direct support that 5-HT7 receptors preassociate
with G protein, a property that likely accounts for the atypical signalling
characteristics of 5-HT7 receptors.
Paper No.: 3275
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
SHIKONIN, A NATURAL NAPHTOQUINONE, PROMOTES
WOUND HEALING IN INTESTINAL EPITHELIAL CELLS-18
Isabel Andújar, MC Recio, JL Rı́os
University of Valencia, Departament of Pharmacology, Burjassot, Valencia, Spain
The intestinal epithelial cell (IEC) barrier plays an important role in
maintaining mucosal immune homeostasis. Dysregulated activation of
the intestinal mucosal immune system eases the penetration of endogenous luminal bacterial flora through tight junction defects in the intestinal
epithelial barrier and this appears to initiate and perpetuate inflammation
in inflammatory bowel diseases.(1) In our search for anti-inflammatory
agents from natural products we found that shikonin, a naphthoquinone
found in the root of Lithospermum erythrorhyzon Sieb. et Zucc., is able
to ameliorate dextran sulphate sodium (DSS)-induced acute colitis due to
the inhibition of the pro-inflammatory nuclear transcription factors
NF-jB and STAT3.(2) Moreover, shikonin significantly reduced the
extent and severity of the colon injury as shown by scores of macroscopic damage, and prevented body weight loss observed in DSS-treated
mice.(3) To further investigate the possible wound healing properties in
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
178
the damaged intestinal epithelial barrier, we examined effect of shikonin
on IEC-18 cells. IEC-18 monolayers were wounded with a razor blade.
Cells were incubated with or without shikonin 1lM and migration was
blocked with treatment with dexamethasone. After 5h, migration was
assessed by counting cells across the wound edge. Shikonin significantly
induced epithelial cell migration and wound healing in IEC-18 cells compared to untreated cells or to dexamethasone-treated cells.
(1.) Cario E (2008) Mucosal Immunol 1 (Suppl 1), S62–S66
(2.) Rı́os JL et al. (2009) Meth. Find. Exp. Clin. Pharmacol. 31 (suppl
A) 92
(3.) Andújar I et al. (2008) Meth. Find. Exp. Clin. Pharmacol. 30 (suppl
2) 86
Paper No.: 2862
FOCUSED CONFERENCE GROUP: P03 - ION CHANNELS IN
ANALGESIA & ANAESTHESIA
ROCURONIUM INTERFERES WITH CYTOCHROME P450 IN
VITRO
cytes (NHEK) and to investigate the preventive effect of EPC-K1: a
phosphate diester of vitamin C and vitamin E on UV-radiation-induced
NHEK injury. NHEK were cultured in EpiLife medium supplemented
with Human Keratinocyte Growth Supplement Kit. NHEK viability was
determined by crystal violet (CV) stain 48 hr after UV irradiation. The
mRNA of a transcription factor C/EBP homologous protein (Chop) and
an ER resident molecular chaperone Bip were determined by RT-PCR
analyses. UV was especially effective in the wavelength region 250280 nm in killing NHEK. The minimum exposure dose required to kill
50% of cells (LD50) was 1.64 mJ/cm2 at 269 nm. At 235 and 310 nm,
LD50 in NHEK were 6.62 and 293 mJ/cm2, respectively. After 660 mJ/
cm2 irradiation at 310 nm, the cell viability was significantly decreased
to 30% of control (without irradiation). The addition of 100 lM EPCK1 after irradiation returned the cell viability to 118%. Six hr after
660 mJ/cm2 irradiation at 310 nm, Chop and Bip mRNA levels in
NHEK were increased 215% and 370% of control and those were
decreased by EPC-K1 treatment. Chop and Bip are responsive to endoplasmic reticulum(ER) stress. These results suggest that EPC-K1 exerts
a protective effect against UV-induced NHEK injury, in part through
the decreased ER stress.
Pavel Anzenbacher(1), E Anzenbacherova(2), A Tunkova(1), M Adamus(3)
(1) Palacky University of Olomouc, Faculty of Medicine and Dentistry,
Department of Pharmacology, Olomouc, Czech Republic
(2) Palacky University of Olomouc, Faculty of Medicine, Department of
Medicinal Chemistry and Biochemistry, Olomouc, Czech Republic
(3) Palacky University of Olomouc Hospital, Department of Anaesthesiology and Resuscitation, Olomouc, Czech Republic
Rocuronium is a relatively new neuromuscular blocking agent (NMBA)
acting as a competitive antagonist of acetylcholine on the postsynaptic
nicotinic receptor of neuromuscular junction with the most quick onset
among currently used non-depolarizing NMBA, and with an intermediate
duration of its action. It is predominantly used to facilitate endotracheal
intubation, to provide skeletal muscle relaxation during surgery, or to
facilitate mechanical ventilation in intubated, critically ill patients. Rocuronium, however, exhibits a great variability in the effect and in the
course of neuromuscular block as in the time of recovery. Variability of
response to rocuronium is according to the literature primarily caused by
pharmacokinetics of this drug. In the literature there are reports indicating that an interaction of rocuronium with liver monooxygenase system
of cytochromes P450 may contribute to these differences. We have found
an inhibition of human cytochrome P450 (form CYP3A4) enzyme activity (6-b testosterone hydroxylation) down to one half of the original
value. The mechanism of this inhibition was evaluated; the results indicate a partially noncompetitive inhibition mechanism. Interactions of this
kind may however influence the pharmacokinetics of concomitantly
administered drugs.
The financial support through the MSM6198959216 project is gratefully
acknowledged.
Paper No.: 1732
FOCUSED CONFERENCE GROUP: P19 - GENERAL
SESSION - TOXICOLOGY
ULTRAVIOLET ACTION SPECTRUM AND EFFECT OF
EPC-K1 ON ULTRAVIOLET-RADIATION-INDUCED INJURY IN
CULTURED NHEK
Kayo Aoki(1), T Nakanishi-Ueda(1), M Tsuji(1), T Okuno(2), K Oguchi(1), H Yasuhara(1)
Paper No.: 2994
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
HEPATOPROTECTIVE EFFECT OF PISTACIA LENTISCUS
AGAINST ISONIAZIDE INDUCED LIVER TOXICITY IN RATS
M Aouichri(1,2,4), M Haouari(1), L Sfaxi(1), R Zermani(3), M Ben-Attia(2), M Lakhal(4), Anis Klouz(4)
(1) École Supérieure des Sciences et Techniques de la Santé de Tunis,
Tunisia
(2) Laboratoire de Biosurveillance de l’Environnement, Faculté des Sciences de Bizerte, Tunisia
(3) Laboratoire d’Anatomie et Cytopathologie, Hôpital Charles Nicolle,
Tunisia
(4) Centre National de Pharmacovigilance, Service de Pharmacologie
Clinique, Tunisia
Isoniazid (INH) is a widely used drug in association with other antibiotics for the tuberculosis treatment. INH can cause a severe hepatotoxicity in 1% of patients by increasing the production of free
radicals in liver. Plant extracts, especially certain vegetable oils appear
to have beneficial antioxidant effects. The aim of this study is to
evaluate the protective effect of Pistacia lentiscus oil (PLO) against
INH hepatotoxicity. Male Wistar Rats were subacutely treated with
INH (25 mg/kg, i.p.) and grouped into the INH group and the INHPLO group for 1 month. The diets fed to the two groups of rats were
identical except the second group (INH-PLO), which received PLO
added to the standard diet. The diet contains 15 percent of PLO by
weight. Animals were sacrificed 1 day after the last drug administration and the livers were rapidly removed. While one portion was used
for histopathology, other pieces were stored with plasma samples
immediately at -80C. PLO prevented INH-induced hepatotoxicity,
indicated by both diagnostic indicators of liver damage (alanine aminotransferase and aspartate aminotransferase) and plasma oxidative
stress as well as histopathological analysis. PLO markedly attenuated
the INH effects on the plasma enzymatic antioxidant defence system
(SOD and GPx). Altogether, these results suggest that PLO exerts its
hepatoprotective effects against INH hepatotoxicity by inhibiting the
production of free radicals in addition to its role as a scavenger. Conclusively, PLO might be credited with beneficial health aspects and
could also be an important nutritional source rich in natural antioxidants.
(1) Showa University School of Medicne, Department of Pharmacology,
Tokyo, Japan
(2) National Institute of Occupational Safety and Health, Japan
The aims of this study are to determine the ultraviolet (UV:235310 nm) action spectrum for killing normal human epidermal keratinoJournal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
179
Paper No. 1651
FOCUS GROUP: FC13 - MAXIMISING BENEFITS AND
MINIMIZING HARMS FROM DRUGS
CYNODON DACTYLON – A POTENT, ECONOMIC, NATURAL
ANTI-DIARRHOEAL HERB IN DEVELOPING COUNTRIES
Manoj Amrutkar, V Undale, D Bharti, A Bhosale
SGRS College of Pharmacy, Department of Pharmacology, Saswad,
Pune, India
The aqueous extract of aerial parts Cynodon dactylon (poaceae) was
investigated for its anti-diarrhoeal property in term of reduction in the
rate of defecation and consistency of faeces in castor oil induced diarrhea
in Wistar rats to substantiate folkloric claim. To understand the mechanism, the effect was further evaluated on intestinal transit and castor oil
induced intestinal fluid accumulation (enteropooling). At various doses
(250 & 500mg/kg body weight) the extract showed a remarkable antidiarrhoeal activity evidenced by the reduction of the rate of defection
and consistency of faecas. Results are comparable to that of standard
drug loparamide (50 mg/kg body weight).A single oral dose of Cynodon
dactylon extract of 250mg/kg body weight produced a significant
decrease in the severity of diarrhea. Extract produced profound decrease
in intestinal transit (39.66%) also significantly inhibited castor oil
induced enteropooling comparable to that of intraperitoneal injection of
standard drug atropine sulphate at a doses of 0.1mg/kg body weight and
3 mg/kg body weight respectively. Experimental findings showed that
aqueous extract of aerial parts of Cynodon dactylon possess significant
anti-diarrhoeal activity and may be a potent anti-diarrhoeal drug in
future. Furthermore the extract formulated as a syrup (85% simple syrup
& 15% extract) also produced significant anti-diarrhoeal activity.
Paper No.: 2917
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
A NEW ANTICOCCIDIAL PRODUCT DEVELOPED FROM
ARTEMISIA SIEBERI PLANT
Hossein Ali Arab(1), A Rasouli(1), J Kaboutari(2), S Rahbari(3)
(1) University of Tehran, Faculty of Veterinary Medicine, Depatment of
Pharmacology, Tehran, Iran
(2) Faculty of Veterinary Medicine, University of Tehran,
(3) University of Shahrekord, Faulty of Veterinary Medicine, Department
of Basic Sciences, Iran
(4) University of Tehran, Faculty of Veterinary Medicine, Department of
Parasitology, Tehran, Iran
Coccidiosis is an important parasitic disease of the chickens causing high
loss in the poultry industry. The emergence of resistance against convention anticoccidial drugs and the concerns of their tissue residues enforced
the attention toward the development of the new drugs. Artemisinin is
one the potential candidate which its anticoccidial effects have been
shown by some studies. This study aimed to develop a new anticoccidial
product from Artemisia sieberi. Artemisinin was extracted from Artemisia
sieberi and the amount of artemisin was determined by HPLC. A formulation of 1% feed additive was produced from Artemisia extract. Coccidiosis was experimentally induced in chickens by oral administration of
Eimeria tenella oocysts. The anticoccidial activity of the new product was
tested by administration of the compound with different doses of 1, 2.5 &
5mg/kg for 5 successive days. A group of infected birds was treated with
1mg/kg pure artemisinin, and the fifth group remained as negative control
group. The feces of each group were collected at the end of each day, and
the amount of oocyst per gram (OPG) was determined as the index of the
anticoccidial effects of the compounds. It was found that the new compound was significantly able to decease the OPG (P < 0.001) in a doseindependent manner. There was not any significant difference between
the effects of the new compound with the pure artemisinin. The results of
this study suggest that the artemisinin granule formulated form artemisia
sieberi can be introduced, potentially as a new anticoccidial drug.
Paper No.: 1803
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
VARIOUS TYPES OF POTASSIUM CHANNELS ARE
ACTIVATED BY THE NEW NITRIC OXIDE (NO) DONOR
[RU(TERPY)(BDQ)NO+]3 + (TERPY) IN MESENTERIC
RESISTANCE ARTERIES OF RENAL HYPERTENSIVE RATS
Alice Araújo(1), J Vercesi(2), J Biazzotto(2), R da Silva(2), L Bendhack(2)
(1) University of São Paulo, School of Medicine of Ribeirão Preto,
Department of Pharmacology, São Paulo, Brazil
(2) University of São Paulo, Faculty of Pharmaceutical Sciences of
Ribeirão Preto, São Paulo, Brazil
Nitrosyl ruthenium compounds such as Terpy are very attractive as NO
donors. We previously reported that NO released from Terpy does not
activate K+ channels in aortas from 2K-1C. The aim of this study was to
verify if Terpy relaxes the isolated third branch of rat mesenteric artery
isolated from 2K-1C and normotensive (2K) rats and the cellular mechanisms involved in the smooth muscle mol/L-0.01mmol/L) werel relaxation. Concentration-effect curves for Terpy (0.01 constructed in
endothelium-denuded mesenteric resistance arteries pre-contracted with
phenylephrine. We have analyzed its maximum effect (ME) and potency
(pD2). Terpy induced relaxation of mesenteric arteries from normotensive (ME: 93.9 ± 1.2%; pD2:4.45 ± 0.32; n = 7) and hypertensive (ME:
93.7 ± 1.4%; pD2: 4.51 ± 0.36; n = 8) rats in a similar way, which was
abolished by the soluble guanylyl-cyclase inhibitor, ODQ. The relaxation
was inhibited by the following K+ channel blockers: tetraethylammonium
(TEA, 1mmol/L), glibenclamide (3umol/L), apamin (1umol/L) and 4-aminopyridine (1mmol/L). Taken together, our results show that the new
NO donor Terpy induces vascular relaxation via activation of soluble
guanylyl-cyclase and various types of K+ channels such as ATP-sensitive, small conductance calcium-activated and voltage-activated K+ channels in isolated mesenteric resistance arteries from normotensive and
hypertensive rats.
This study was approved by Ethical Committee of the University of São
Paulo (CETEA-FMRP, protocol number: 044/2008).
Supported by FAPESP and CNPq.
Paper No.: 1369
FOCUSED CONFERENCE GROUP: P02 - TRANSMEMBRANE
TRANSPORT: PERSPECTIVES FOR DISEASE AND DRUG
DISCOVERY
UPTAKE OF FOLIC ACID AND METHYLTETRAFOLIC ACID
BY RAT AND HUMAN BLOOD-BRAIN BARRIER
ENDOTHELIAL CELL LINES
João R Araújo, P Goncalves, F Martel
Porto University Faculty of Medicine, Department of Biochemistry,
Porto,Portugal
The blood–brain barrier (BBB) is a selective permeability tissue essential
for central nervous system homeostasis. Folates are essential for the biosynthesis of purines, thymidylate and methionine. Our aim was to characterize 3H-folic acid (3H-FA) and 14C-methyltetrahydrofolic acid (14CMTHF) uptake by rat (RBE4) and human (hCMEC/D3) BBB endothelial
cell lines. Uptake of 3H-FA and 14C-MTHF by RBE4 cells was timedependent and linear for the first 2 min of incubation; uptake by
hCMEC/D3 cells showed a greater experimental variability. So, further
experiments were performed in RBE4 cells. Uptake of 3H-FA was found
to be stimulated at acidic and alkaline pH, Na+-dependent; stimulated
when F- substituted for Cl-, energy-independent, inhibited by premetrexed (PTX), stimulated by cytochalasin D (cyt D) and not affected by
MTHF, DIDS, SITS, methotrexate (MTX), monensin (MON) and FA.
Uptake of 14C-MTHF was found to be pH-, Na+-, Cl-- and energy-independent, inhibited by PTX and MTX, stimulated by cyt D and not
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
180
affected by FA, DIDS, SITS, MON and MTHF. RT-PCR analysis demonstrated mRNA expression of RFC (reduced folate carrier) but neither
of FRa (folate receptor a) nor of PCFT (proton-coupled folate transporter) in RBE4 cells, and mRNA expression of both RFC and PCFT,
but not of FRa, in hCMEC/D3 cells. In conclusion, both human and rat
BBB endothelial cells show little capacity for folates uptake.
Paper No.: 2958
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
EVALUATION OF THE GASTRIC ANTIULCER ACTIVITY OF
A PROTEOLYTIC FRACTION FROM CARICA
CANDAMARCENSIS LATEX
Ana Cândida Araújo e Silva(1), FO Lemos(1), C Figueiredo(1), CTR
Viana(1), CM Souza(2), GD Cassali(2), CE Salas(3), MTP Lopes(1)
(1) Federal University of Minas Gerais, Department of Pharmacology,
Belo Horizonte, Brazil
(2) Federal University of Minas Gerais, Department of Pathology, Belo
Horizonte, Brazil
(3) Federal University of Minas Gerais, Department of Biochemistry and
Immunology, Belo Horizonte, Brazil
P1G10 is a proteolytic fraction, rich in cysteine proteases, obtained from
Carica candamarcensis (South America species). Previously, we have
found that these proteases display mitogenic effect, angiogenic and skin
healing properties on mammalian systems. Furthermore, we verified the
gastric protective and healing activities of P1G10 on gastric lesions
induced by indomethacin and acetic acid. In this study, we have investigated the antiulcer activity of P1G10 on other models of lesions, as well
as some pathways involved in this effect. In acute lesions produced by
ethanol, stress and pyloric ligation, P1G10 (10 mg/kg) showed a significant gastroprotective activity, reducing the number/severity of lesions by
40%, 88% and 64%, respectively. The treatment with P1G10 promoted a
significant decrease on gastric acidity and on peptic activity, with reductions of 56% (1 mg/kg) and 69% (10 mg/kg). We also demonstrated that
P1G10 increases the gastric levels of glutathione (42%) and that its proteolytic activity is not important for the gastroprotective action. In
chronic lesions induced by acetic acid, P1G10 showed an interesting
healing activity, reducing the ulcer size by 58%. This property is dependent on its proteolytic activity. Furthermore, we showed that P1G10
stimulates about 4-fold the epithelial cell proliferation at the ulcer margins. Thus, the gastroprotective activity plus the ability to stimulate cell
proliferation, makes of P1G10 a potential antiulcer agent.
Financial support: CNPq and Fapemig.
Paper No.: 2084
FOCUSED CONFERENCE GROUP: P14 - ADDICTION AND
DOPING: NEUROBIOLOGICAL AND CLINICAL
COMPARISON OF THE NEUROCHEMICAL CHANGES
INDUCED BY ACUTE AND REPEATED ADMINISTRATION OF
MORPHINE, COCAINE AND AMPHETAMINE IN THE RAT
LATERAL SEPTUM
Katherine A Araya, MR Ibañez, V Noches, R Sotomayor-Zarate, K
Gysling,
Pontificia Universidad Catolica de Chile, Department of Cellular and
Molecular Biology, Santiago, Chile
Addictive drugs activate dopaminergic neurons in the ventral tegmental
area. These neurons project to various limbic nuclei such as nucleus
accumbens, prefrontal cortex and lateral septum (LS). We have shown
that acute intravenous administration of morphine increases LS dopamine
(DA) release in a dose-dependent manner. However, the effect of
repeated administration of morphine or other addictive drugs in LS has
not been determined. Thus, we studied the neurochemical changes in LS
induced by acute and repeated administration of morphine (10 mg/kg
i.p.), cocaine (15 mg/kg i.p) and amphetamine (1,5 mg/kg i.p.). We analyzed DA, Glutamate and GABA extracellular levels using microdialysis
in LS of control and repeatedly treated rats with morphine, cocaine and
amphetamine. Addictive state of treated rats was evaluated through the
experimental paradigm of conditioned place preference. Our results
showed that acute administration of morphine and cocaine increased
extracellular DA levels in LS, without changing extracellular Glutamate
and GABA levels. By the contrary, acute administration of amphetamine
increased extracellular GABA levels without any significant changes in
extracellular levels of DA and Glutamate. Although, the repeated administration of morphine and cocaine produced a significant increase in DA
extracellular levels in LS, the amount of DA released is significantly
lower compared to acute administration. These results suggest that LS is
involved as a relay nucleus modulating the response of other limbic
nuclei involved in addictive processes. We will evaluate the inactivation
of LS and its effects on place conditioning induced by morphine and
cocaine.
Funding: MSI P06/008-F and FONDECYT 107-0340
Paper No.: 1155
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
PUBLICATION TRENDS OF THE CYTOCHROMES P450
RESEARCH: 1977-2008
Charles-Daniel Arreto(1), C Wilson(2), C Robert(1)
(1) Université Paris Descartes, Laboratoire d’Anatomie Fonctionnelle,
Montrouge, France
(2) The University of New South Wales, Sydney, NSW, Australia
This study analyzes trends in cytochrome P450s research publications
from 1977 to 2008. Research articles and review papers with title keywords related to ‘‘cytochrome-P450’’ were downloaded from the Web of
Science for four periods: 1977-1978, 1987-1988, 1997-1998, and 20072008. Overall the literature on P450s increased nearly seven-fold (6.9)
from 342 papers in 1977-1978 to 2357 in 2007-2008. For the same literature, the number of countries involved in P450s-related research
increased from over three-fold from 23 in 1977-1978 to 76 in 20072008. Early on in 1977-1978, the leading producers of P450s research
publications consisted of a mixture of western countries (USA, 151
papers; Japan, 42; France, 21and Federal Republic of Germany, 18) and
socialist countries (Germanic Democratic Republic, 23 and the USSR,
23). However, by 2007-2008 the USA with 810 was followed by a mixture of western countries and emerging Asian countries. During the four
time periods, the number of scientific journals publishing P450s papers
increased more than seven-fold (7.7): from 94 journals in 1977-1978 to
724 in 2007-2008. With the exception of two journals (The Journal of
Biological Chemistry and Archives of Biochemistry and Biophysics),
there was a complete turnover among the top-10 most productive journals. While journals focusing on biological chemistry dominated the topranks in 1977-1978, journals focusing on pharmacological approaches
were prominent in the 2007-2008 ranking led by the journal, Drug Disposition and Metabolism. The transition of the P450s literature from biological chemistry to pharmacology shows the importance of translational
approaches for progressing in P450s research.
Paper No.: 2423
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
ANTINOCICEPTIVE ACTIVITY OF FRUITS OF METHANOLIC
EXTRACT OF CAPPARIS OVATA IN MICE
Rana Arslan(1), N Bektas(1)
Anadolu University, Faculty of Pharmacy, Department of Pharmacology,
Eskisehir, Turkey
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
181
Aim of the study: In this study, we attempted to identify the possible
antinociceptive action of methanol extract obtained from fruits of
Capparis ovata. Materials and Methods: The antinociceptive effect of
the methanol extract of fruits of Capparis ovata (MEC) was assessed
by intraperitoneal administration in tail immersion, hot plate and writhing tests in mice. Morphine sulfate (5 mg/kg; i.p.) and diclofenac
(10 mg/kg; i.p.) were used as reference analgesic agents. Naloxone
(5 mg/kg; i.p.) was also tested. Results: MEC was studied at the doses
of 50, 100, and 200 mg/kg (i.p.) and showed significantly antinociceptive activity in all assays. The extract at given dose range reduced the
writing by 32.21, 55.70, and 68.36% respectively. MPE% were
increased by 7.27, 12.07, 14.60% in the tail immersion, and 7.88,
11.71, 16.73% in the hot plate test at the tested doses respectively. Naloxone antagonized antinociceptive effect at the doses of 100 and
200mg/kg whereas partially antagonized the effect of MEC at the dose
of 50 mg/kg. Conclusions: Based on the results obtained, it can be
concluded that MEC has antinociceptive effects both at the peripheral
and central levels.
Keywords: Capparis ovata, fruits extract, antinociception
Paper No.: 2245
FOCUSED CONFERENCE GROUP: P06 - THE HEART GONE
WRONG; STABILIZATION OF CARDIAC FUNCTION
CARDIOPROTECTIVE MECHANISM OF TELMISARTAN
THROUGH THE PPAR-C PATHWAY AGAINST
ISOPROTERENOL-INDUCED MYOCARDIAL INFARCTION IN
EXPERIMENTAL DIABETES
Dharamvir Singh Arya, S Goyal
All India Institute of Medical Sciences, Department of Pharmacology,
New Delhi, India
Introduction: Telmisartan is a partial agonist of the PPAR-c. We investigated whether telmisartan improved the pathophysiology of myocardial
infarction in diabetes partially through the PPAR-c pathway by assessing
a variety of indices, e.g., hemodynamic, biochemical, histoarchitectural
changes, and apoptosis. Materials and methods: Diabetes was induced by
a single dose of streptozotocin (70 mg/kg, IP). Diabetic rats received
either telmisartan (10 mg/kg/day, orally), the PPAR-c antagonist
GW9662 (1 mg/kg/day, IP), or both for 14 days with concurrent administration of isoproterenol (85 mg/kg, SC) on days 13 and 14. Results:
Compared with diabetic controls, diabetic rats with myocardial infarction
exhibited altered hemodynamic profiles and reduction in the activities of
creatine kinase-MB isoenzyme, lactate dehydrogenase, superoxide
dismutase, catalase, and glutathione level along with increased level of
malondialdehyde in the heart. Further, diabetic animals with myocardial
infarction exhibited increased myonecrosis, edema, and apoptotic cell
death. Treatment with telmisartan significantly improved the redox status
of the myocardium with subsequent cardiac functional recovery. However, significant effects were lowered in animals treated with telmisartan
plus GW9662. Telmisartan markedly inhibited Bax expression, TUNELpositive cells, myonecrosis, and edema. On the other hand, administration of telmisartan plus GW9662 did not elicit the same effects, nor did
they increase Bcl-2 protein expression in isoproterenol-induced myocardially infarcted diabetic rats when administered concomitantly or individually. Moreover, down-regulated PPAR-c expression in myocardially
infarcted diabetic hearts was increased by telmisartan treatment. Conclusion: In addition to class effects of ARBs, telmisartan reduces oxidative
stress and apoptosis and improves cardiac function via the PPAR-c pathway.
Paper No.: 2046
FOCUSED CONFERENCE GROUP: P02 - TRANSMEMBRANE
TRANSPORT: PERSPECTIVES FOR DISEASE AND DRUG
DISCOVERY
INTERACTIONS BETWEEN VERAPAMIL AND DIGOXIN IN
LANGENDORFF - PERFUSED RAT HEARTS: THE ROLE OF
INHIBITION OF P-GLYCOPROTEIN IN HEART
M Aylin Arýcý(1), E Kilinc(2), O Demir(1), M Ates(3), A Yesilyurt(2), A
Gelal(1)
(1) Dokuz Eylul University School of Medicine, Izmir, Turkey
(2) Ege University, Faculty of Pharmacy, Izmir, Turkey
(3) Dokuz Eylul University, Advanced Professional School of Health
Sciences, Izmir, Turkey
Introduction: P-glycoprotein (P-gp) is expressed in tumor cells as well as
normal tissues including heart. Modulation of P-gp transport in vivo may
lead to increased drug penetrance to tissues with resulting increases in
toxicity. We aimed to investigate the effects of P-gp on the isolated heart
by digoxin infusion in the absence and presence of verapamil. Materials
and Methods: Study was performed in Langendorff isolated perfused rat
hearts. After 20 minute stabilization period with Tyrode Buffer, digoxin
(125 lg/5mL) was infused for 10 minutes in control group (n = 7). The
same dose of digoxin was infused during perfusion with verapamil (1
nM) containing Tyrode buffer (n = 8) in the study group. Outflow concentration and cardiac parameters were measured at frequent intervals for
40 minutes. Results: AUEC(0-40 min) for left ventricular developed
pressure (LVDP) was significantly increased in the presence of verapamil
(4260 ± 39.37 vs 4607 ± 98.09; %95 CI -587.7 to -105.8; p = 0.0083).
The significant increases in LVDP were at 20, 25, 30, 35 and 40 minutes.
AUC(0-40 min) values for digoxin was lower significantly in the presence of verapamil. Conclusion: Verapamil increased the positive inotropic effect of digoxin which could be related to the accumulation of
digoxin in the rat heart.
Keywords: P-glycoprotein, drug interaction, heart, digoxin, verapamil
This paper was sent to Basic and Clinical Pharmacology and Toxicology
to be evaluate for publication
Paper No. 1519
FOCUS GROUP: P02 - TRANSMEMBRANE TRANSPORT:
PERSPECTIVES FOR DISEASE AND DRUG DISCOVERY
IMPACTS OF EGFR-ANTISENSE AND CARBENOXOLONE ON
CANCER CELL LINES
Masoud Asadi(1,2), H Hamzeiy(0), J Barar(2), Y Omidi(2), S Mohammadi(1)
(1) Tabriz University of Medical Sciences, Department of Pharmacology
and Toxicology, Tabriz, Iran
(2) Research Center for Pharmaceutical Nanotechnology, Iran
Gap junctions play an important role in the cell proliferation in mammalian cells. However, there are controversial issues about cell communications in neoplastic states. This study was designed to evaluate the
cytotoxic effects of Carbenoxolone as a prototype of inter cellular gap
junction blocker in association with epidermal growth factor receptor
(EGFR) gene silencing using EGFR-antisense therapy. On the basis of
specific and general roles of EGFR in different kinds of cells, three cancerous cell lines(A549, BT20 and MCF7 cells) were cultivated, and at
designated confluency cell monolayers were treated with carbenoxolone
(150 lM) and anti EGFR antisense oligodeoxyneucleotide (ODN). Cellular cytotoxicity were examined using standard colorimetric assay associated with cell viability tests. MCF7 and BT20 cells were significantly
affected by carbenoxolone and EGFR antisense, but not A549 cells.
Treated MCF7 and BT20 cells with anti-EGFR ODN and carbenoxolone
(alone or as combination) showed significant toxicity. Our preliminary
examinations with DNA microarray technology confirmed such findings.
Based upon this investigation, it can be concluded that there exists a
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
182
novel relationship between cell-cell communications and intercellular
cross talking, perhaps via gap junctions, at which we propose interestingly orchestrated biochemical machinery of cells may be related to
EGFR downstream cell signalling which may play a key role in development of cancer.
Keywords: gap junctions, EGFR-antisense, Carbenoxolone, cancer cell
line
Paper No.: 713
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
THE VARIATIONS IN MEASUREMENTS OF BLOOD PRESSUREC VALUES OBTAINED BY THE CONVENTIONAL METHOD
AND THE AMBULATORY BLOOD PRESSURE MONITORING
Mensura Asceric(1), N Mulabegovic(2), E Kapic(2), F Becic(2),
S Loga-Zec(2), J Kusturica(2)
(1) University of Tuzla, Medical Faculty, Department of Pharmacology
and Toxicology, Tuzla, Bosnia & Herzegovina
(2) University of Sarajevo, Medical Faculty, Department of Pharmacology and Toxicology, Sarajevo, Bosnia & Herzegovina
Introduction: More frequent and longer blood pressure (BP) oscillations,
in other words variability, can lead to faster and severe damage of certain
organs and hypertrophy of left venticle and presence of increased risk
factors can lead to exponential growth of risks severe cardiovascular
events. The aim of this paper was to define and analyze variations in
measurements of systolic and diastolic BP values in treated hypertensive
patients obtained by conventional method during ambulatory monitoring
blood pressure (AMBP). Patients: A total of 72 treated patients have
been observed in 2 groups. The first group of patients without of echocardiography parameters of HLV and second group of patients with echocardiography diagnosed HLV. Results: It conformed that there were no
significant gender differences but that there was a significant growth considering age and artery hypertension length as well as in body mass
index (BMI) values of cholesterol and triglycerides, a genetic predisposition to hypertension, fundus and left ventricular mass index change in
group with HLV. Systolic and diastolic BP values do not show significant
differences in the groups during the day and they are significantly higher
than values measured with AMBP method. The systolic and diastolic BP
variability is more noticeable in patients with HLV.Conclusion: The
results of this research have shown that 24-hours AMBP is better indicator of BP variability than use the conventional method.
Paper No.: 575
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
EVALUATION OF HEPATOPROTECTIVE ACTIVITY OF
GARCINIA INDICA IN RODENTS: EVIDENCE OF
ANTIOXIDANT ACTIVITY
Hardik D Ashar, VS Panda
Principal K. M. Kundnani College of Pharmacy, Department of Pharmacology & Toxicology, Mumbai, India
Garcinia indica is an Indian spice, the fruit of which is widely used in
cooking and its syrup is consumed as a soft drink. The protective effects
of aqueous extracts of the fruit rind of G. indica (GIE) on carbon tetrachloride (CCl4)-induced hepatotoxicity and the probable mechanisms
involved in this protection were investigated in rats. Liver damage was
induced in Wistar rats by administering a 1:1 (v/v) mixture of CCl4 and
olive oil (1ml/kg, i.p.) once daily for 7 days. GIE at 400 mg/kg and
800 mg/kg and the reference drug silymarin (100 mg/kg) were administered orally for 10 days to CCl4 treated rats, this treatment beginning
3 days prior to the commencement of CCl4 administration. Levels of
marker enzymes (AST and ALT), albumin (Alb) and total protein (TP)
were estimated in serum. Antioxidant parameters {reduced glutathione
(GSH), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX) and glutathione reductase (GR)} and the lipid peroxidation
marker malondialdehyde (MDA) were determined in liver. GIE and silymarin elicited significant hepatoprotective activity by attenuating the
CCl4-elevated levels of AST, ALT and MDA and restored the CCl4depleted levels of GSH, SOD, CAT, GPX, GR, Alb and TP. GIE
800 mg/kg demonstrated greater hepatoprotection than GIE 400 mg/kg.
The present findings indicate that hepatoprotective effects of GIE in
CCl4-induced oxidative damage may be due to an augmentation of the
endogenous antioxidants and inhibition of lipid peroxidation in liver.
(Devasagayam TPA, Mishra A, Bapat MM et al, Curr Sci 2006; 91(1):
90-93).
Paper No.: 2227
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
EFFECTS OF FREE D-AMINO ACIDS PRESENT IN FOODS ON
HUMAN DERMAL FIBROBLASTS
Yutaka Ashida(1), S Shimada(1), C Okamura(1), M Mita(1),
K Hamase(2)
(1) Shiseido Research Center, Yokohama, Japan
(2) Kyushu UniversityGraduate School of Pharmaceutical Sciences,
Japan
The role of D-amino acids in mammals has been neglected due to their
ultra-low concentrations in tissues. Recent analytical progress has confirmed their existence and demonstrated their involvement in physiological functions, such as neurotransmission and hormone secretion. We have
shown that their contents in skin vary with age. Free D-amino acids in
mammalian tissues may derive from ingested foods, as well as enterobacteria. In this work, we examined their contents in foods and in stratum
corneum, and investigated their physiological functions in human dermal
fibroblasts. Amino acids were extracted with aqueous methanol from stratum corneum collected by tape stripping. Derivatized D-amino acids in
the extracts were determined by means of two-dimensional HPLC
(NANOSPACE, Shiseido), as reported previously. In human stratum corneum, D-serine, D-alanine and D-aspartate were detected. D-Aspartate
enhanced type I collagen production in fibroblasts, whereas L-aspartate
did not. D-Aspartate, but not L-aspartate, also showed antioxidative
effects in cells exposed to hydroperoxide or free radicals. D-Aspartate
content in different kinds of foods varied tremendously. Food processing
procedures, including fermentation, may also alter the content of
D-amino acids, and thereby influence physiological functions. In conclusion, D-amino acids derived from foods may have physiological roles in
humans, and therefore may have potential as pharmacological agents.
Paper No.: 1510
FOCUSED CONFERENCE GROUP: P07 - SIMULATION AND
DATA MODELLING IN DRUG DEVELOPMENT.
PROTECTIVE EFFECT OF AQUEOUS SAFFRON EXTRACT
(CROCUS SATIVUS L.) ON CARBON TETRACHLORIDEINDUCED HEPATOTOXICITY IN RATS
Amir Ashna, H Najafzadeh
Shahid Chamran University, Department of Pharmacology & Toxicology,
Ahvaz, Iran
Crocus sativus L., commonly known as saffron is used for different purposes such as an antispasmodic and expectorant. Recent studies have
demonstrated that saffron extracts have antitumour, radical scavenger,
hypolipaemic, anticonvulsant effects and improve activity on learning
and memory. The aim of the present study was to assess the effect of
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
183
aqueous saffron extract on carbon tetrachloride-induced hepatotoxicity in
rats. Four groups of rats were used as group1: control; group2: Ccl4;
group3: Ccl4 + vitamin E; group4: Ccl4 + saffron extract. Drugs were
administrated for 7 days. The serum of rats was collected and analyzed
biochemically. The mean of ALT, AST, LDH, GGT, conjugated and total
billirubin, total protein and albumin was compared between groups. The
results show the mean of ALT and AST was increased by ccl4. This elevation was prevented by saffron extract; but vitamin E prevented AST
elevation. The mean of conjugated and total billirubin was increased by
ccl4. This elevation also was prevented by saffron extract. Thus saffron
has protective effect on carbon tetrachloride-induced hepatotoxicity probably by antioxidant action.
Key words: Ccl4, Hepatotoxicity, Rats, Vitamin E, Saffron
Paper No.: 1468
FOCUSED CONFERENCE GROUP: P12 - ION
CHANNELOPATHIES: NEW WINDOWS ON COMPLEX
DISEASE AND THERAPY
THE ROLE OF POTASSIUM CHANNELS IN EXPERIMENTAL
SEPSIS
Jamil Assreuy(1), R Sordi(1), D Fernandes(2)
(1) Universidade Federal de Santa Catarina, Department of Pharmacology, Florianopolis, Brazil
(2) Universidade Estadual de Ponta Grossa, Brazil
This study investigates the involvement of potassium channel (KC) subtypes on sepsis-induced vascular hyporesponsiveness to phenylephrine
(VHP). Wistar rats were submitted to cecal ligation and puncture (CLP)
surgery. Six and 24 hours later, responses to phenylephrine were determined before and 30 min after injection of KC blockers. Additionally,
TEA and GLB were injected 4 hours after CLP and 24 hours after surgery several parameters were evaluated. The KC blockers were tetraethylammonium (TEA; a non-selective KC blocker), glibenclamide (GLB; an
ATP-dependent KC blocker), 4-aminopyridine (4-AP; a voltage-dependent KC blocker) and apamin (a calcium-dependent KC blocker). Results
show that: (1) CLP induced a severe VPH; (2) injection of TEA 30 minutes before phenylephrine injection completely restored the vascular
response to it, 24 hours after CLP; (3) apamin reversed VHP after 6, but
not after 24 hours after CLP; (4) GLB restored phenylephrine response
only 24 hours after CLP and (5) 4-AP was ineffective in all periods. In
addition, the early treatment with TEA reduced plasma NOx, lung MPO,
urea, creatinine and lactate levels, prevented the development of VPH
and reduced the mortality by 50%. Early treatment with GLB was without effect. Thus, at least part of the sepsis-induced vascular dysfunction
is maintained by the activity of distinct KC subtypes, calcium-dependent
KC in the first hours and ATP-dependent KC in later phases of sepsis. In
addition, blockade of TEA-sensitive KC early after the onset of sepsis
has several beneficial consequences reducing organ damage and mortality in experimental sepsis.
Financial support: CNPQ, CAPES, FAPESC
Paper No.: 434
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
ANTIMITOGENIC AND APOPTOTIC EFFECTS OF
SELECTIVE 5HT3 RECEPTOR ANTAGONIST ON HT29
COLORECTAL CANCER CELL LINE
Ramin Ataee(1), M Zarrindast(2), S Ajdary(1), S Sharyari(1)
(1) Institute Pasteur of Iran, Department of Immunology, Tehran, Iran
(2) Tehran University of Medical Sciences, Department of Pharmacology,
Tehran, Iran
Background: Serotonin has been known to be a mitogen factor in severalmalignancies. It elicits its mitogenic effect through a wide range of
5-HTreceptor subtypes and several internal cellular transcription pathways.According to wide distribution of 5HT3 and 5HT4 receptors in GI
tracts, the mainaim of this study was to investigate effect of these receptors’ agonists andantagonists in colorectal cell line. Methodology: In cell
culture, weinvestigated the effects of Serotonin (5-HT), 5-HT3 and 5-HT4
receptors agonists and antagonists on proliferation of HT29 cells. We also
testedapoptosis for receptor antagonists on HT29 cells with TUNEL apoptosis test. With flow cytometery we assayed effects of 5HT receptors
antagonist on cell cyclekinetics and with western blot we assayed the protein expression of 5HTreceptors Results: MTT proliferation assay revealed
that Phenylbiguanide (a 5HT3receptor selective agonist) increased proliferation of HT29 cells significantly and Y25130 Hydrochloride (a 5HT3
receptor antagonist) had the opposite effect; but for 5HT4 receptor antagonist, theses effects were not significant. Also potent apoptotic and cell
cycle arresting effect was found for selective 5HT3receptor antagonist but
as well we have not seen any significant effect for 5HT4receptor antagonist Also western blot study showed high protein expression for5HT3
receptor. The findings of this study provide strong evidence for the potential role of 5-HT3 receptor in colorectal cancer.
Paper No.: 1810
FOCUSED CONFERENCE GROUP: P17 - NEW APPROACHES
AND TARGETS IN PSYCHIATRY
NEUROPROTECTIVE EFFECTS OF THE POLYPHENOLIC
ANTIOXIDANT AGENT, CURCUMIN, AGAINST
HOMOCYSTEINE-INDUCED COGNITIVE IMPAIRMENT AND
OXIDATIVE STRESS IN THE RAT
Amin Ataie, M Sabetkasaei, A Haghparast
Shahid Beheshti Medical University, Department of Pharmacology, Tehran, Iran
Aging is the major risk factor for neurodegenerative diseases and oxidative stress is involved in the pathophysiology of these diseases. In this
study, we investigated the possible antioxidant and neuroprotective properties of the polyphenolic antioxidant compound, Curcumin against homocysteine (Hcy)neurotoxicity. Curcumin (5, 15 and 45 mg/kg) was
injected intraperitonealy once daily for a period of 10 days beginning
5 days prior to Hcy (0.2 lmol/ll) intrahippocampal injection in rats.
Biochemical and behavioral studies, including passive avoidance learning
and locomotor activity tests were studied. We detected Malondialdehyde
(MDA) and Super oxide anion (SOA) in rats’ hippocampi. Also Immunohistochemical studies were done on hippocampus cells. Results indicated that Hcy could induce lipid peroxidation and increase MDA and
SOA levels in rats’ hippocampi. Additionally, Hcy impaired memory
retention inpassive avoidance learning test. However, Curcumin treatment decreased MDA and SOA levels significantly as well as improved
learning and memory in rats. Histopathological analysis also indicated
that Hcy could induce cell death and apoptosis in rats’ hippocampi and
Curcumin inhibited these toxic effects. These results suggest that Hcy
may induce lipid peroxidation and cell death in rats ‘hippocampi and
polyphenol treatment (Curcumin) has the ability to improve learning and
memory deficits by protecting rats’ hippocampi against Hcy toxicity.
Paper No.: 2828
FOCUSED CONFERENCE GROUP: P18 - NUCLEAR
RECEPTOR TARGETS FOR TREATMENT OF DISEASES
STUDY OF 5HT3 AND HT4 RECEPTORS EXPRESSION IN
HT29 CELL LINE AND HUMAN COLON ADENOCARCINOMA
TISSUES
Ramin Ataee(1), S Ajdary(1), M Zarrindast(2), M Rezayat(2), MA Shokrgozar(5)
(1) Institute Pasteur of Iran, Department of Immunology, Tehran, Iran
(2) Terhran University of Medical Sciences, Department of Pharmacology, Tehran, Iran
(3) Institute Pasteur of Iran, Cell Bank, Tehran, Iran
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
184
Background: Serotonin (5HT) has been shown to be a mitogenic factor
in several carcinomas. It elicits its mitogenic effect through a wide range
of 5HT receptor subtypes. In this study, the effect of 5-HT, 5HT3 (1-Phenylbiguanide hydrochloride) and 5HT4 (Cisapride) agonists in promoting
the growth of HT29 cell line and the growth-inhibition effect of 5HT3
receptor antagonist (Y-25130 hydrochloride) and 5HT4 receptor antagonist (RS 23597-190) were investigated. The expression of 5HT3 and
5HT4 receptors in human colon cancer tissues and cell line was studied.
Materials and methods: The growth-promoting and growth-inhibition
effect of 5-HT, 5HT3 and 5HT4 agonists and antagonists on HT29 cell
line was studied using MTT assay. Receptor expression has been demonstrated by western blotting. Results: The results showed 5HT, 5HT3, and
5HT4 agonists caused significant proliferation of HT29 cells. 5HT3 and
5HT4 receptor antagonists had inhibitory effect on the growth of these
cells. Western blot analysis gave bands from colon tissue extracts and
HT29 cell line. Conclusion: The results indicate 5HT3 and 5HT4 receptors express in colon cancer tissue and cell line significantly and the
expression for 5HT3 receptor is more potent. Furthermore, 5HT plays a
mitogenic role in colon cancer cells and antagonists of 5HT3, and 5HT4
receptors can inhibit cancer cell growth. Key Words: Colon adenocarcinoma, HT29 cell, Serotonin, 5HT3 and 5HT4 receptors.
Paper No.: 634
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
SINGLE-DOSE PHARMACOKINETICS OF CEFEPIME IN
HEALTHY EGYPTIAN BUFFALO CALVES
Taha A Attia(1), MA Tohamy(2), AAM El-Gendy(2)
(1) Menofia University Faculty of Veterinary Medicine, Department of
Pharmacology, Egypt
(2) Beni-Suef University, Faculty of Veterinary Medicine, Department of
Pharmacology, Egypt
Pharmacokinetics of cefepime was studied after single dose intravenous,
intramuscular and subcutaneous administration. Five apparently healthy
buffalo calves were used in a crossover design with 15-day washout period. After intravenous (i.v) injection of cefepime, the half-lives of distribution and elimination (t0.5(a) and t0.5(b)), volume of distribution at
steady state (Vdss), mean residence time (MRT) and total body clearance
(ClB) were 0.113 and 2.32 h., 0.192 L kg-1., 3.12 h and 0.061 L kg-1 h1
., respectively. Following intramuscular (i.m) and subcutaneous (s.c)
administration of cefepime, the maximum concentration (Cmax) 29.094
and 27.271 ug ml-1 were achieved at a maximum time (tmax) 1.611 and
1.578 h., respectively. The mean values for absorption and elimination
half-lives (t0.5(ab) and t0.5(el)) and MRT were 0.549, 2.422 and 2.242,
4.286 and 4.025 h., respectively. The intramuscular and subcutaneous
bioavailabilities were 95.81 and 85.663%, respectively. The result of invitro protein-binding study indicated that 11.423% of cefepime was
bound to the serum proteins of buffalo calves.
Paper No.: 1517
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
SEVERAL FRUIT AND BERRY JUICES AND PURÉES ARE
POTENT INDUCERS OF ENDOTHELIUM-DEPENDENT
RELAXATIONS IN THE PORCINE CORONARY ARTERY VIA
ACTIVATION OF NITRIC OXIDE AND ENDOTHELIUMDERIVED HYPERPOLARIZING FACTOR PATHWAYS
C Auger, S Trinh, J-H Kim, T Chataigneau, Valerie B Schini-Kerth
University of Strasbourg, Faculty of Pharmacy, UMR CNRS 7213, Illkirch Graffenstaden, France
Polyphenol-rich sources such as red wine have been shown to protect
the vascular system by enhancing the endothelial formation of nitric
oxide (NO) and endothelium-derived hyperpolarizing factor (EDHF),
two major vasoprotective factors. This study evaluated the ability of
several non-alcoholic sources of polyphenols and in particular selected
pure fruit juices and purées to cause endothelium-dependent relaxations
of isolated arteries and identified the mechanism involved. Vascular
reactivity was assessed using porcine coronary artery rings in the presence of indomethacin to prevent the formation of vasoactive prostanoids. The 13 fruit juices and purées tested caused variable
concentration-dependent relaxations in coronary artery rings with endothelium and only small relaxations in rings without endothelium. The
strongest inducers of endothelium-dependent relaxations were blackcurrant, aronia and cranberry followed by lingonberry, blueberry and San
Gabriele grape whereas only small relaxations were observed with
strawberry, apple, raspberry, blackberry, boysenberry, and acerola; elderberry was without effect. Relaxations to aronia, blackcurrant, and lingonberry were not affected by the combination of charybdotoxin plus
apamin (inhibitors of EDHF-mediated responses), slightly reduced by
NG-nitro-L-arginine (an NO synthase inhibitor) and markedly reduced
by the combination of NG-nitro-L-arginine and charybdotoxin plus
apamin. Thus, the present findings indicate that several fruit juices and
purées are potent inducers of endothelium-dependent relaxations in coronary arteries by stimulating the endothelial formation of NO, and also
EDHF. They further suggest that regular intake of these active rich
sources of polyphenols may increase the protective effect of endothelial
cells on the vascular system.
Paper No.: 2249
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
WHOLE-BODY DISTRIBUTION AND PHARMACOKINETICS
OF RADIOIODINATED FULLY HUMANIZED ANTI-VAP-1
ANTIBODY – A PET IMAGING STUDY OF RABBITS
Anu Autio(1), PJ Vainio(2), A Mali(3), J Vainio(2), S Suilamo(1), V
Oikonen(1), P Luoto(1), M Teräs(1), T Karhi(4), A Roivainen(1,4)
(1) Turku University Hospital, Turku PET Centre, Turku, Finland
(2) Biotie Therapies Corp, Turku, Finland
(3) MAP Medical Technologies Oy, Tikkakoski, Finland
(4) University of Turku, Turku Center for Disease Modeling, Turku, Finland
We assessed the usefulness of positron emission tomography (PET) in
evaluation of iodine-124 labeled antibody pharmacokinetics and distribution in rabbits. PET is powerful, non-invasive method particularly
suitable for drug development because of its high sensitivity and ability
to provide quantitative and kinetic data without sacrificing the animal.
The antibody (BTT-1023) is first-in-class, fully human monoclonal antibody to vascular adhesion protein-1 (VAP-1). VAP-1 is an inflammation inducible endothelial glycoprotein. It plays a key role in leukocyte
trafficking and is a potential target for anti-inflammatory therapy. BTT1020 is potentially useful for the treatment of inflammatory diseases
and in vivo imaging of inflammation. BTT-1023 was labeled with
[124I] using Chloramine-T method. Its immunoreactivity was retained
after labeling as confirmed in hVAP-1 transfected cells and by timeresolved immunofluorometric assay using human recombinant VAP-1.
Ten rabbits were intravenously injected with [124I]BTT-1023. PET/CT
was obtained over the first 2 h after dosing and at 24, 48 and 72 h
post-injection. Blood samples were collected during PET study to clarify in vivo stability and pharmacokinetics. In rabbits, the radioactivity
was distributed especially to liver and thyroid. Also heart and lungs
showed some uptake whereas brain uptake was very low. The plasma
half-life of [124I]BTT-1023 was 58.3 ± 10.5 h and clearance
7.8 ± 1.8 mL/h/kg. [124I]BTT-1023 showed good in vivo stability
(80% of signal from intact antibody 72 h after injection) and uptake in
VAP-1 positive liver. In conclusion, PET study of [124I]-labeled VAP-1
targeting BTT-1023 further elucidated distribution and kinetics of the
therapeutical antibody.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
185
Paper No.: 1095
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
EFFECT OF THE ANTHRAQUINONE COMPOUND RHEIN ON
HUMAN COLON ADENOCARCINOMA CELL
PROLIFERATION
Gabriella Aviello(1), I Rowland(2), C Gill(3), AM Acquaviva(4),
R Capasso(1), AA Izzo(1), F Capasso(1), F Borrelli(1)
(1) University of Naples Federico II, Department of Experimental
Pharmacology,Naples, Italy
(2) University of Reading, Department of Food Biosciences, Reading,
UK
(3) University of Ulster, Northern Ireland Centre for Food and Health
(NICHE), Department of Biomedical Sciences, Belfast, Northern Ireland,
UK
(4) University of Naples Federico II, Department of Biology and Cellular
and Molecular Pathology, Naples, Italy
Background/Aim: Anthraquinones are the oldest drugs used as laxatives
in clinical practice and as self-medication. Recently the use of anthraquinone laxatives, such as senna (Cassia spp), has been associated with
damage of the intestinal epithelium and increased risk of developing
colorectal cancer. However, human and animal data are inconsistent. In
the present study we evaluated the cytotoxicity and genotoxicity of rhein,
the active metabolite of senna, on human colon adenocarcinoma cells
(Caco-2) and its effect on cell proliferation. Methods: Cytotoxicity studies were performed using MTT and TEER assays; cell proliferation was
investigated using 3H-thymidine incorporation and western blot analysis.
Genotoxicity studies were performed by Comet assay. Results: Rhein
had no significant cytotoxic effect on proliferating and differentiated
Caco-2 cells. Rhein, at low concentrations (0.1 and 1 lg/ml), significantly reduced cell proliferation and ERKs expression; by contrast, at
higher concentrations (10 lg/ml), it significantly increased cell proliferation and MAP kinase activation. Moreover, rhein did not adversely affect
the integrity of tight junctions - and hence the epithelial barrier function
– as well as it did not induce DNA damage. Finally, rhein significantly
inhibited the increase in malondialdehyde and ROS levels induced by
Fenton’s reagent. Conclusions: Rhein, was devoid of cytotoxic and genotoxic effects in Caco-2 cells. Moreover, at concentrations which are
expected to be yielded in the colon lumen after a human therapeutic dose
of senna, rhein (i) inhibited cell proliferation via a mechanism likely
involving the MAP kinase pathway and (ii) prevented the DNA damage
probably via an anti-oxidant mechanism.
Paper No.: 917
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
FUNCTIONAL DISTRIBUTION OF NITRERGIC NERVE
DERIVED FROM PTERYGOPARATINE GANGLION IN
MONKEY CEREBRAL ARTERY
Kazuhide Ayajiki(1), S Kobuchi(1), N Toda(2), M Hanamitsu(3),
T Okamura(4)
(1) Department of Pharmacy, School of Pharmacy, Hyogo University of
Health Sciences, Kobe, Japan
(2) Toyama Institute for Cardiovascular Pharmacology Research,
Toyama, Japan
(3) Department of Otoraryngology, Shiga University of Medical Science,
Shiga, Japan
(4) Department of Pharmacology, Shiga University of Medical Science,
Shiga, Japan
Functional roles of nitrergic nerve innervating monkey cerebral artery
were evaluated by measuring mechanical responses of isolated preparations in vitro and by cerebral angiography in vivo. Nicotine, which stimulates nerve terminals innervating cerebral arteries, produced relaxations
in isolated monkey arteries irrigating the cerebrum and cerebellum. The
relaxations induced by nicotine were abolished by NG-nitro-L-arginine.
We demonstrated, by using arterial angiography, that electrical stimulation of one side of the greater petrosal nerve connecting to the ipsilateral
pterygopalatine ganglion (PPG) with synapse, a parasympathetic ganglion, produced vasodilatation of anterior, middle and posterior cerebral
arteries only in the stimulated side, but failed to produce responses of
superior and anterior inferior cerebellar arteries in the stimulated side and
basilar artery of anesthetized monkeys. It is concluded that nitrergic
nerves innervate all arteries tested and that stimulation of nitrergic nerve
derived from the PPG dilates cerebral arteries, but not cerebellar arteries
and basilar artery. Nitrergic nerve derived from the PPG appears to regulate cerebral vasomotor function whereas nitrergic nerve from unknown
ganglion(s) may regulate the circulation in the cerebellum and brainstem.
Paper No.: 2126
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
INHIBITORY ACTIONS OF CARVACROL 1,4-CINEOLE BUT
NOT 1,8-CINEOLE ON FIELD STIMULATION OF ISOLATED
RAT VAS DEFERENS: ROLE OF ISOPROPYL GROUP AS A
PHARMACOPHORE
Suleyman Aydin, Y Cakmakci
Anadolu University Faculty of Pharmacy, Department of Pharmacology,
Eskisehir, Turkey
Carvacrol and cineole are volatile and fragrant natural compounds found
in various plants which have medicinal properties in folk medicine. Carvacrol, 1,8-cineole and 1,4-cineole have different chemical structures but
almost similar actions were reported. Actions of 1,4-cineole, 1,8-cineole
and carvacrol were studied on electrical field stimulated isolated rat vas
deferens in this study. Contractions of vas deferens were inhibited by
Carvacrol (10-4 M) and by 1,4-cineole (10-3 M) whereas 1,8-cineole
was inactive. Based on pharmacological actions and structural properties
of the carvacrol, 1,4-cineole and 1,8-cineole on isolated vas deferens, isopropyl group was proposed to play important role on the inhibitory
actions. Since isopropyl group is widely found in volatile constituents of
natural compounds, isopropyl group of these compounds is proposed to
play role as a pharmacophore.
Paper No.: 2111
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
THE EFFECTS OF CARVACROL THYMOL AND PROPOFOL
ON ISOLATED RAT ATRIUM
Yasemin Aydin(1), S Arkan(1), S Aydin(2)
(1) Eskisehir Osmangazi University, Faculty of Medicine, Department of
Physiology, Eskisehir, Turkey
(2) Anadolu University, Faculty of Pharmacy, Department of Pharmacology, Eskisehir, Turkey
Thymol and carvacrol are natural compounds widely found in nature but
propofol is a synthetic chemical used as general anaesthetic drug which
are all have chemical similarity being phenolic substances with aliphatic
side chains. Propofol is well known for its cardiovascular effects but
there is no sufficient information on the cardiovascular actions of thymol
and carvacrol. The effects of thymol, carvacrol and propofol were investigated on the contractions of electrical field stimulation induced isolated
rat atria in the present study. All the test compounds inhibited contraction
of field stimulated isolated atria. Thymol, which has more structural
similarity to the carvacrol exhibited inhibition only at the 10-4 M but
propofol and carvacrol were observed to inhibit contractions at 10-5 and
10-4 M doses. Although thymol has more structural similarity to carvacrol
than propofol, carvacrol and propofol exhibited similar actions compared
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
186
to thymol. Thus mechanism of action of carvacrol on myocardium is suggested similar to the propofol including calcium and potassium channels.
Paper No.: 1102
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
PHARMACOLOGICAL SCREENING OF MEDICINAL PLANTS
IN SWAZILAND
ADP-stimulation with C50~0.5 mM or its abolishment by SU. So
[SU]~Kd(SU) can eliminate >50% of diabetogenic currents in patients
with either A- or B-type KATP inhibited with submillimolar IC50(ATP)
in inside-out membrane fragments. Deciphered mechanisms of HA and
SU-inhibition of diabetogenic KATP tested so far argue that the majority
of HA-KATP patients need SU doses exceeding those currently recommended for treatment of T2DM and illuminate the clinical importance of
the NIDDK/NIH-funded basic analysis of mutant/polymorphic ABCC8/
KCNJ11 identified in humans.
Peter Aziba
Olabisi Onabanjo University, Department of Pharmacology, Ago-Iwoye,
Nigeria
The search for drugs in Africa has been intensified on the potential of
medicinal activities of plants in Traditional medical practice (TMP). In
Swaziland, a tiny kingdom within South Africa, traditional practice is
well accepted by the people due to poverty, cultural affinity and unaffordability of conventional drugs. Problems associated with this practice
bothers on dosage, use, efficacy & side effects of the plant decortion used
in therapy.The institute of medicinal plants and food products research
has focused on the possibility of fostering integration of orthodox and
traditional practice.We have been screening, some plants for pharmacological activities, using basic experimental in vitro technique for receptor
studies (via the organ bath method), analgesic activities (using the Hot
plate method) and inflammatory screening of the crude plant extracts (Ledebouria ovaltifolia & mormodica involucrata methanolic extracts).In the
Isolated rat stomach strip preparation, Acetylcholine (2x10-9 - 4x10-8M)
induced contractile responses were inhibited in non- competitive antagonism by extracts at concentration between 1.25.5mg/ml - 2.5mg/ml.shifting dose effect curve to the right. This pattern of inhibition excludes the
possibility of the extract interacting with muscarinic receptor. The result
in analgesic study, the extracts dose dependently evoked a prolonged
reaction time of 85%(p < 0.05) over a period of 20min observation time,
with no overt signs of toxicity. The results in these studies suggest the
extracts contained spasmodic and analgesic activities. Thus the use of
unknown agents may help to assess or evaluate the pharmacological
activities of this plant could be a prelude to drug discovery.
Paper No.: 1597
FOCUSED CONFERENCE GROUP: P12 - ION
CHANNELOPATHIES: NEW WINDOWS ON COMPLEX
DISEASE AND THERAPY
DIABETOGENIC ABCC8/KCNJ11: MECHANISMS TO
PERSONALIZED TREATMENT
Andrey Babenko
Paper No.: 2784
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
ANTIOXIDANT ACTIVITY OF PHENOLIC EXTRACTS OF
TUNISIAN PLANTS AND THEIR EFFECT ON
MITOCHONDRIAL RESPIRATION IN VITRO
S Bacha(1), H Ferchichi(1), I Salouage(1), L Ouanes(2), M Boussaı̈d(3),
M Lakhal(1), Anis Klouz(1)
(1) National Center of Pharmacovigilance, Laboratory of Clinical Pharmacology, Tunis, Tunisia
(2) University of Faculty of Medicine, Animal Experiment Unit, Tunis,
Tunisia
(3) National Institute of Applied and Technology, Tunis, Tunisia
Based on the popular knowledge on the therapeutic uses of some Tunisian
plants, we have selected four specimens (Myrtus communis, Hypericum
humifusum, Romarinus officinalis and Crataegus azarolus). The aim of
this study is to evaluate the antioxidant effect of these plants by: the determination of total polyphenols content, the Diphenyl Pycril Hydrazil
(DPPH) test and monitoring of malondialdhéyde (MDA) used for the
determination of the lipoperoxidation inhibition percentage. We evaluate
the effect of different extracts on a mitochondrial respiration model in vitro.
The concentration of different plants extracts used was 20mg/ml. Our
study shown an antioxidant potential with scavenging activity >85% for
Myrtus communis extracts and <25% for Hypericum humifusum, Romarinus officinalis and Crataegus azarolus. The total polyphenols concentration was 103lg/ml for Myrtus communis, and varied from 59.91 to
30.91lg/ml for the other plants. Lipoperoxidation inhibition percentage
was 100% for Myrtus communis and Romarinus officinalis, 60% and 35%
for Hypericum humifusum and Crataegus azarolus respectively. The mitochondrial respiration study shows an amelioration of 19.44% for Crataegus azarolus, 13.84% for Hypericum humifusum and 3.75% for
Rosmarinus officinalis, compared to no treated mitochondria. This study
we permitted to evaluate the antioxidant activity for a screening of different Tunisian plants. In the other hand, the combination of different tests
used in our model seems to be the best guarantee to select plant extracts
with high potential to be protective against ischemia reperfusion process.
University of Washington, Pacific Northwest Research Institute, Department of Molecular Physiology and Biophysics, Seattle, WA, USA
Sulfonylureas (SU) differentially inhibit different diabetogenic mutant
ABCC8(Sulfonylurea Receptor 1)/KCNJ11(Kir6.2) channels (KATP)
(Babenko et al, NEJM 2006;355:456-466 and Pearson et al, NEJM
2006; 355:467-477). Babenko (JBC 2008; 283:8778-8782) established
the first, or A-type, mechanism of diabetogenic hyperactivity (HA) of
KATP, its MgATP/ADP-hyperstimulation (HS), clarified why even a small
number of mutant channels in a heterozygous KATP population can
hyperpolarize insulin-secreting cells, and uncovered that some A-type
mutations can attenuate SU-inhibition of KATP in MgATP/ADP, thus
explaining in part why therapeutic doses of SU for HA-KATP carriers
often exceed those recommended by the FDA for treatment of type 2
diabetes mellitus (T2DM). Now, I show that insulin release-compromising mutations in domains partnering in SUR1/Kir6.2 inhibitory coupling
(Babenko et al, FEBS Letters 1999; 459:367-76) hyperactivate KATP by
changing the rates of channel transitions to and from its long-lived closed
state with the lowest Kd for inhibitory ATP, thus establishing the second,
or B-type, diabetogenic mechanism of KATP HA. B-type mutations alter
the coupling of SU-binding with the closed channel, but not the MgATP/
Paper No.: 2261
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
STRIATAL DELIVERY OF CDNF BY AAV2 VECTOR IS
NEUROPROTECTIVE IN A RAT PARTIAL LESION MODEL
OF PARKINSON’S DISEASE
Susanne Bäck(1), J Peränen(2), L Lonka(2), T Tamminen(1), MH Voutilainen(1), A Raasmaja(1), M Saarma(2), PT Männistö(1), RK Tuominen(1),
(1) University of Helsinki, Faculty of Pharmacy, Division of Pharmacology and Toxicology, Helsinki, Finland
(2) University of Helsinki, Institute of Biotechnology, Helsinki, Finland
Cerebral dopamine neurotrophic factor (CDNF) is a novel neurotrophic
factor that, together with vertebrate and invertebrate mesencephalic
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
187
astrocyte-derived neurotrophic factor (MANF), belongs to a family of
evolutionarily conserved neurotrophic factors. In our previous work, we
proved that a single injection of MANF or CDNF into rat striatum is able
to protect and restore midbrain dopaminergic neurons in partial lesion
model of Parkinson’s disease (PD) in rat (Voutilainen MH et al, J Neurosci 2009; 29: 9651-9659, Lindholm P et al, Nature 2007; 448: 73-77).
According to these results, CDNF and MANF could potentially be used
as treatments in PD. In this study, we constructed adeno-associated viral
serotype 2 (AAV2) vectors encoding CDNF and GDNF genes and
injected them into rat striatum. Two weeks later, rat nigrostriatal dopaminergic tract was lesioned by intrastriatal injection of 6-hydroxydopamine
(6-OHDA, 2x10 lg). To evaluate the progression of the neurodegeneration of the rat nigrostriatal dopamine tract, we measured amphetamineinduced rotational behaviour and tyrosine hydroxylase (TH)-immunoreactivity in rat striatum and SN. Results implies that over-expression of
CDNF in rat striatum is able to protect rat nigrostriatal dopamine neurons
from 6-OHDA-induced nerve cell toxicity with similar efficiency as
over-expression of GDNF. This study provides further evidence that
CDNF could be a potential drug candidate for the treatment of PD.
Paper No.: 514
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
EFFICACY AND SAFETY OF ADD-ON MIRTAZAPINE IN
DEPRESSION
Dinesh Kumar Badyal(1), P Matreja(2), R Deswal(3)
(1) Christian Medical College and Hospital, Department of
Pharmacology, Ludhiana, India
(2) Department Pharmacology, Gain Sagar Medical College and
Hospital, Banur, Patiala, India
(3) Department Of Psychiatry, Christian Medical College and Hospital,
Ludhiana, India
Major depressive disorder (MDD) continues to be a considerable problem, both for clinician and at the public health level. Although antidepressant medications are effective in approximately 70% of patients with
major depressive disorder, they have a delayed onset of therapeutic
effect, problems in remission and adverse effects. Mirtazapine, an atypical antidepressant has certain advantages which makes it an important
option for the treatment of major depression in patients who require additional therapy. The study was conducted in sixty patients suffering from
major depressive disorder. The study was approved by institutional ethics
committee. The duration of study was 1 year. Patients were randomized
to two groups and were given conventional selective serotonin reuptake
inhibitors (SSRI) alone or conventional SSRIs with mirtazapine 7.5 mg
once a day for 6 weeks. Efficacy was evaluated based Hamilton depression rating scale (HDRS), Montgomery and Asberg depression rating
scale (MADRS) and Amritsar Depressive Inventory (ADI) scores. Scores
were recorded at baseline and then at 1, 2, 3, 4, 5 and 6 weeks after the
treatment. The HDRS, MADRS and ADI score reduced significantly at
each visit as compared to baseline in both the groups. The response rate
and number of remitters were significantly more with add-on mirtazapine
therapy as compared to conventional therapy at the end of study. The
HDRS-17 subset scores indicate that the add-on mirtazapine therapy was
more effective in improving anxiety and somatic symptoms as compared
to conventional therapy. There was no significant difference in adverse
effects in the two groups.
Paper No.: 1575
FOCUSED CONFERENCE GROUP: P14 - ADDICTION AND
DOPING: NEUROBIOLOGICAL AND CLINICAL
PHARMACODYNAMIC EFFECTS OF
3,4-METHYLENEDIOXYAMPHETAMINE (MDA)
MJ Baggott, J Siegrist, JR Coyle, K Flower, GP Galloway, John
Mendelson
California Pacific Medical Center Research Institute, Addiction and Pharmacology Research Laboratory, San Francisco, CA, USA
Background: MDA, an analog of MDMA (3,4-methylenedioxymethamphetamine, ‘Ecstasy’), is an illicitly used drug. In vivo MDMA is
N-demethylated to MDA. This study measured MDA effects in humans
in a controlled setting. Methods: In a placebo-controlled, double-blind,
within-subjects study, 12 individuals received a single 98 mg/70 kg bw
dose of MDA. This is the molar equivalent of 105 mg/ 70 kg bw
MDMA, a well-studied dose. Hormonal (cortisol, prolactin), physiological (HR, BP), and self-report VAS measures of typical MDMA and
hallucinogen effects were obtained. Results: MDA increased cortisol by
16.39 ug/dL (95%CI: 13.03-19.74, P < 1e-3) and prolactin by
18.37 ng/mL (95%CI: 7.39-29.35, P < 1e-3). These hormonal changes
are comparable to those seen after MDMA. Heart rate increased by
9.05 bpm (95%CI: 6.10-11.99, P < 1e-5) and blood pressure increased
by 18.98 / 12.73 mm Hg (Systolic 95%CI: 16.47 - 21.49, P < 1e-7;
Diastolic 95%CI: 10.82 - 14.63, P < 1e-4). Heart rate and systolic
changes were significantly less than and greater than seen in a previous
study of MDMA (N = 16), respectively (P < 1e-5 and P = 2.42e-7,
respectively). There were robust self-report VAS changes in both
MDMA-like (e.g., ‘‘closeness to others’’) and hallucinogen-like (e.g.,
‘‘familiar things seem unfamiliar’’, time distortions, closed-eye visuals)
effects that were generally similar to those seen after MDMA. Conclusions: MDA is a psychoactive sympathomimetic phenethylamine with
effects similar to MDMA. Although differences may exist in the magnitude of physiological effects, the overall profiles appear remarkably
similar.
Supported by DA 016776
Paper No.: 931
FOCUSED CONFERENCE GROUP: P10 - DRUGS FOR HALF
THE WORLD: PAEDIATRIC CLINICAL PHARMACOLOGY
BIOAVAILABILITY AND BIOEQUIVALENCE OF THREE DIFFERENT SIRUP FORMULATIONS CEFELEKSINE AT CHILDREN IN THERAPY OF SEVERE TONSILLITIS
Adnan Bajraktarevic(1), J Musabegovic(2), T Frankic(2), H Sladic(2), D
Abduzaimovic(3)
(1) Public Health Institution of Canton Sarajevo, Department of Paediatrics,Sarajevo, Bosnia Herzegovina
(2) Pharmaceutical Faculty, Department for Clinical Pharmacology,
Sarajevo, BosniaHerzegovina
(3) Private Biochemical Laboratory, Tesanj, Bosnia Herzegovina
Introduction: Effective criteria for evaluating pharmaceutical antibiotics
drug products saves time for the Health institutions and Pediatric hospitals and ensures that drugs purchased are not associated with hidden
expenses related to drug use. The determination of bioavailability is
dependent on the reliable, precise, and accurate measurement of the
active ingredient, or its metabolites, as a function of time. Methods:
Three same formulations of cefeleksine were compared, a randomized
three-period, three-sequence crossover study were considered the therapy of choice in children with severe bacterial tonsillitis and elevated
CRP. Bioequivalence is usually established by measuring the drug or
its metabolites or both in plasma and serum at 60 children. Results:
There was no correlation between the drug levels and side effects or
initial and subsequent results of curing severe tonsillitis. The 90% confidence interval of the relative mean measured Cmax of the test to reference of all three cefaleksine formulation should be between 94% and
98%. There were no significant baseline demographic or clinical differences between the ethnics, sexual and ages groups. Conclusions: The
change in absorption of same antibiotics of three differenet European
companies was small and unlikely to give significant differences in
clinical effect in children. The drug product is, on the basis of scientific
evidence before submitted in this sirup cefeleksine application, shown
to meet an in vitro test that has been excellent correlated with in vivo
our data.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
188
Paper No.: 932
FOCUSED CONFERENCE GROUP: P14 - ADDICTION AND
DOPING: NEUROBIOLOGICAL AND CLINICAL
VITAMINS AS ANTIOXIDANTS HAVE IMPORTANT ROLE OF
ASTHMATIC ADDITIONAL THERAPY IN CHILDREN AND
PROTECTION OF PASSIVE SMOKING
Adnan Bajraktarevic(1), M Miokovic(1), D Abduzaimovic(2), J Ceman
Saric(3), T Frankic(4), J Musabegovic(4), I Tahmiscija(5), A Selimovic(5), JA Maglajlic(6), B Begovic(7)
(1) Public Health Institution of Canton Sarajevo, Department of Paediatrics,Sarajevo, Bosnia Herzegovina
(2) Private Biochemistry and Immunology Laboratory,Tesanj, Bosnia
Herzegovina
(3) Medical Faculty Sarajevo, Department for Biochemistry and Immunology, Sarajevo, Bosnia Herzegovina
(4) Pharmaceutical Faculty, Sarajevo, Bosnia Herzegovina
(5) Pediatrics Clinic, Sarajevo, Bosnia Herzegovina
(6) Pulmology Clinic, Department for Microbiology, Sarajevo, Bosnia
Herzegovina
(7) University Medical Center, Department for Clinical Pharmacology,
Sarajevo, Bosnia Herzegovina
Background: Oxygen is a highly reactive molecule that damages living
organisms by producing reactive oxygen species. Cigarette smoke is a
complex mixture of less than five thousand chemical compounds of
which free radicals and other oxidants are present in high concentrations.
Methods: Oxidation of proteins caused introduction of carbonyl groups
into the side chains of the protein, providing a convenient and relatively
specific marker of oxidative damage. Authors studied the effects of two
volatile components of cigarette smoke, acetaldehyde and acrolein,
which are present at high concentrations in cigarette smoke on lungs of
children. Vitamins A, C, and E as antioxidants led to the realization of
the importance of antioxidants in the biochemistry of living organisms.
Results: These antioxidant defence elements offer promising chemoprevention targets that have the potential to reduce the burden of asthma.
We have endeavoured to diminish the generation of oxygen free radicals
by decreasing the oxygen concentration in the resuscitating gas. The fall
in antioxidant capacity also correlates with the increased release of oxygen radicals from circulating neutrophils in patients with exacerbations
of asthma bronchale in children. Our study showed that some tocotrienol,
retinol and ascorbic acid have significant anti-oxidant properties. Conclusions: The use of antioxidants with good bioavailability or molecules that
have antioxidant enzyme activity may be treatments that not only protect
against the direct injurious effects of oxidants. Vitamins antioxidants as
therapeutic targets have important role in obstructive pulmonary disease
in children.
Paper No.: 1237
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
CUTANEOUS ADVERSE DRUG REACTIONS: ANALYSIS OF
SPONTANEOUS REPORTS FROM HEALTHCARE
PROFESSIONALS IN CROATIA
fied. Overall, most commonly reported reactions were rashes, eruptions
and exanthemas (16.7%,), urticarias (15.9%), pruritus (15.8%,) erythemas (15.1%) and angioedemas (7.0%). Antimicrobial agents (25.0%)
and vaccines (19.6%) were most frequently implicated; together with cardiovasculars and drugs for nervous system, they accounted for more than
70% of reported drugs. There were a total of 107 (9.8%) serious reactions, most common being angioedemas (72.0%), exfoliative conditions
(8.4%), photosensitive reactions (6.5%) and bullous conditions (5.6%)
with drugs implicated being antibacterials (27.1%), cardiovasculars
(19%), drugs for nervous system (13.8%) and antineoplastics and immunosuppressants (9.35%). Conclusion: Antibacterials were the most commonly implicated drugs causing both non-serious and serious cutaneous
reactions reported in Croatia. The reporting rate of serious cutaneous
reactions is not satisfying since spontaneous reporting remains the main
source of safety information in cases of rare severe drug reactions which
are unlikely to be identified in premarketing clinical trials.
Paper No.: 1238
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
SERIOUS PSYCHIATRIC ADVERSE DRUG REACTIONS
SPONTANEOUSLY REPORTED IN CROATIA FROM 2006 TILL
2009
Ana Balazin, V Macolic-Sarinic, S Arapovic, D Krnic, M Banovac,
S Tomic
Agency for Medicinal Products and Medical Devices, Zagreb, Croatia
Introduction: Recognition of psychiatric adverse drug reactions (ADRs) is
important because they may interfere with the treatment of the primary
illness and be disturbing for patients, families and medical professionals.
Materials: National pharmacovigilance database of spontaneously
reported ADRs was searched. Reports containing psychiatric reactions
that were serious according to Critical Term List of the WHO were
considered for analysis that was done by descriptive methods (SPSS ver.
17.0). Results: 182 spontaneous reports with at least one psychiatric ADR
were identified. One third of them contained at least one serious ADR
(33.0%), which comprised 70 serious spontaneously reported psychiatric
ADRs. Restlessness (37.1%), suicidal behavior (20%) and anxiety
(12.9%) were most commonly reported serious psychiatric ADRs. Antidepressants (24.3%), antipsychotic (17.1%) and vaccines (17.1%) were most
commonly suspected. More female patients (61.4%) experienced serious
psychiatric ADRs. No gender disbalances were found regarding reactions
except for hallucinations which were reported exclusively in female
patients (6 cases; antiepileptic implicated in 50%). Half of patients experiencing serious reactions were adults; elderly patients and infants were
equally represented by 20%. Out of 12 serious psychiatric ADRs reported
in infants, restlessness was most common (91.7%); implicated was either
vaccine (84.3%) or montelukast (16.7%). 11 cases of suicidal behavior
were reported; implicated were antipsychotic (6 cases), antidepressants (4
cases) and antiparkinsonians (1 fatal case). Conclusion: Serious psychiatric ADRs can be challenging to diagnose in everyday practice. When
evaluating psychiatric symptom in any patient, it is important to check
concomitant medication and bear in mind possible psychiatric ADR.
Ana Balazin, V Macolic-Sarinic, S Arapovic, D Krnic, M Banovac,
S Tomic
Agency for Medicinal Products and Medical Devices, Zagreb, Croatia
Introduction: The aim was to explore spontaneously reported serious and
non-serious cutaneous adverse drug reactions (ADRs) and find classes of
implicated drugs. Materials: We preformed a search within national database of spontaneous ADR reports submitted between January 2006 and
December 2009. Cutaneous reactions categorized as certain, probable,
possible and unlikely, based on WHO causality definitions, were
included in the analysis that was done using descriptive methods (SPSS
ver. 17.0). Seriousness was determined based on Critical Term List of the
WHO. Results: 628 unique cases comprising 1093 reactions were identi-
Paper No.: 1815
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
MAST CELL ACTIVATION ALTERS NEURONAL NITRIC
OXIDE RELEASE IN RAT MESENTERIC ARTERY
Gloria Balfagón, J Blanco-Rivero
Universidad Autónoma de Madrid, Madrid, Spain
This study examines how mast cell activation influences neuronal nitric
oxide (NO) release. Mesenteric arteries from 6-month-old Wistar rats
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
189
were incubated in the presence of mast cell stabilizers ketotifen and tranilast and mast cell degranulator compound 48/80 (C 48/80). Basal and
electrical field stimulated (EFS) NO release was measured by fluorescence, neuronal NO synthase (nNOS) expression by Western blot and
histamine release by ELISA. NO release was reduced after preincubation
with 0.1 mmol/L tranilast or with 0.1 or 1 umol/L ketotifen for 1, 2 and
3 hours, or with 10 nmol/L ketotifen for 2 and 3 hours, release was not
modified by 10nmol/L ketotifen for 1 hour or 10 umol/L tranilast, and
increased by preincubation with 15 ug/mL C 48/80 for 1, 2 and 3 hours.
nNOS expression was decreased by 3 hour’s incubation with 0.1 umol/L
ketotifen or with 0.1 mmol/L tranilast, and was increased by 1-hour incubation with 15 ug/mL C 48/80. 3 hour’s preincubation with 0.1 mmol/L
tranilast or 0.1 umol/L ketotifen decreased histamine release, while the
preincubation with 15 ug/mL C 48/80 increased it. These results indicate
the existence of an interaction between perivascular nitrergic nerve endings and mast cell activation that would influence neuronal NO release
through a modification in nNOS expression.
This study was supported by Ministerio de Ciencia e Innovación
(DEP2006-56187-C04-04; SAF2009-10374)
Paper No.: 2069
FOCUSED CONFERENCE GROUP: P14 - ADDICTION AND
DOPING: NEUROBIOLOGICAL AND CLINICAL
ARECA NUT DEPENDENCE AMONG CHEWERS IN A SOUTH
INDIAN COMMUNITY WHO DO NOT ALSO USE TOBACCO
(1) University of São Paulo - Medical School/ LIM 51, São Paulo, Brazil
(2) University of São Paulo - Medical School/ LIM 10, São Paulo, Brazil
Albumin is the most prevalent protein in blood. Analbuminemia is associated with increased risk of cardiovascular disease and endothelial dysfunction. Inflammatory stimuli such lipopolysaccharide (LPS) is related
to reduced cardiovascular contractile response. Adult (300–350g) male
rats Sprague-Dawley (SD) and Nagase Analbuminemic Rats (NAR) were
studied: both groups received LPS (E coli serotype 026:B6) 10 mg/kg,
ip, or saline i.p. Animals were sacrificed 4h after injection. Intact thoracic
aortic ring (5-6 mm) of each animal was kept in organ baths (37C 95% O2, 5% CO2) with Krebs solution. Cardiovascular function was performed by catheter in left ventricule (LV) and carotid (dP/dt and Mean
Arterial Pressure, respectively). Under basal condition NAR has an
enhanced response (1.88 fold) to noradrenaline (NE) and lower heart
contractile response (+dP/dt=1.4 fold). LPS stimuli enhanced mortality
(25%); TNF (19.3 fold) and IL10 production (101 fold) on NAR; IL6
levels were enhanced in SD and NAR (526 fold, 914 fold respectively);
LV measurement showed an increase on +dP/dt (mmHg/s) in SD (1.32 –
fold) whereas a decrease in NAR (1.53 – fold); we found a decreased of
-dP/dt (mmHg/s) in SD (1.46– fold), no differences was observed on vascular reactivity to NE, hemodynamic parameters by carotid measures and
serum lactate levels. Prevalence of higher response to NE and elevated
+dP/dt on physiological conditions are essential to NAR survival.
Robert L Balster(1), SJS Bhat(2), MD Blank(1), M Nichter(3),
M Nichter(3)
Paper No.: 883
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
TEMPORAL EFFECTS OF ATP/ADP IN A MURINE SEPSIS
MODEL
(1) Virginia Commonwealth University, Richmond, Virgnia, USA
(2) Care Management International, Pune, India
(3) University of Arizona, Tucson, Arizona, USA
Hermes Vieira Barbeiro(1), DF Barbeiro(1), FR Laurindo(2),
HP Souza(1), IT Velasco(1), FG Soriano(1)
The goal of this study was to examine evidence of areca dependence in a
large, representative sample of areca-only (i.e., no tobacco) chewers
using established measurement scales. Information was also gathered on
use patterns in this population. Daily chewers (N = 59) from Karnataka
State, India were surveyed in 2005. Questionnaires assessed chewing history, patterns of use, and dependence features. Additionally, the relationship between topography and dependence scores was evaluated.
Approximately half of respondents reported 1-3 chews/day (mean = 1.9).
The average number of chewing episodes/day was 4.4 and the average
number of nuts/day was 1.2. Users’ typical chew lasts up to 20 minutes
and includes spitting out the juices and rinsing the mouth with water.
Overall, the levels of reported dependence symptoms were low, but
approximately 44% of chewers endorsed at least one of the following
items: continued use despite illness or wounds, difficulty refraining from
chewing in forbidden places, or craving during periods of abstinence.
Approximately 15% of chewers reported at least one quit attempt, and
13.6% had scores indicative of dependence on the modified Cigarette
Dependence Scale (Score >16). Dependence scores were positively correlated with frequency of use. High levels of dependence were not
observed in this sample of regular betel-only users, but many users
reported at least one symptom and a few had several symptoms. The levels of dependence observed in a subset of informants’ warrant further
investigation as evidence for possible betel dependence
(1) University of São Paulo - Medical School, São Paulo, Brazil
(2) University of São Paulo - INCOR, São Paulo, Brazil
Sepsis is one of the main causes of death in non-coronary intensive care
units. One potentially important source on inflammatory vascular injury
is lipopolysaccharide (LPS). Adults (255 ± 15g) male Wistar rats
received LPS (E coli 026:B6) 10 mg/kg, ip or saline (control). Animals
were sacrificed 8h, 16h and 24h after stimuli. Serum cytokine level
(TNF) was measured by ELISA. Thoracic aortic ring (5-6 mm) was
homogenized in liquid nitrogen and reconstitute to measure: iNOS and
TLR4 by RT-PCR and nitrate by NO-analyser (Sievers Instruments).
Some thoracic ring was kept intact to analyze superoxide (O2-) by chemiluminescence and vasodilation in organ baths (37C - 95% O2, 5% CO2)
with Krebs solution. Studies of O2- and vasodilation were performed on
presence of ATP/ADP. LPS injection enhanced: TNF at 8h (8.2 fold),
16h (7.3 fold) and 24h (7.7 fold); TLR4 at 8h (3.4 fold) and 24h (2.5
fold); iNOS at 8h (11.7 fold); nitrate at 16h (2.8 fold); O2- at 16h (7.8
fold) and 24h (4.7 fold). At 16h of endotoxaemia, ATP provide higher
vasodilation when compared to control (106.72%±2.3; 100.15%±0.9,
respectively) and reduced O2- levels (1.9 fold); ADP provide higher
vasodilation vasodilation (EC50) sensitivity on endotoxaemic when compared to control (2.0 and 2.6 10-8M, respectively) at the same time. In
spite of inflammation maintenance in all periods, ATP/ADP presents a
relevant effect on aorta relaxation only at 16h.
Paper No.: 882
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
ANALBUMINEMIA: CARDIOVASCULAR FUNCTION OF
ENDOTOXEMIC RATS
Paper No.: 884
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
VASCULAR REACTIVITY ON NAGASE ANALBUMINEMIC
RATS CHALLENGED BY LPS
Hermes Vieira Barbeiro(1), CB Lorigados(1), DF Barbeiro(1),
S Catanozi(2), E Nakarandake(2), IT Velasco(1), FB Fusco(2), A dos
Santos Filho(1), FG Soriano(1)
Hermes Vieira Barbeiro(1), CB Lorigados(1), DF Barbeiro(1), S Catanozi(2), E Nakarandake(2), IT Velasco(1), FB Fusco(2), A dos Santos
Filho(1), FG Soriano(1)
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
190
(1) University of São Paulo - Medical School/ LIM 51, São Paulo, Brazil
(2) University of São Paulo - Medical School/ LIM 10, São Paulo, Brazil
Hypoalbuminemia is associated with endothelial dysfunction. It is
unclear whether this dysfunction is a direct result of the decreased levels of albumin or whether it is caused by factors as chronic inflammation. Adult (300–350g) male Nagase analbuminemic rats (NAR) rats
and Sprague-Dawley (SD) were studied before (control) and 4h after
received LPS (E coli serotype 026:B6) 10 mg/kg, ip. Thoracic aortic
ring (5-6 mm) with or without endothelium of each animal was kept in
organ baths (37C - 95% O2, 5% CO2) with Krebs solution. Dose
response curve to noradrenaline (NE) was performed. Serum cholesterol, triglycerides and nitrite was evaluated. Under physiological condition NAR has enhanced levels of cholesterol (2.8 fold), triglycerides
(3.8 fold) and nitrite (3.21 fold). LPS stimuli decreased cholesterol levels (1.6 fold) on NAR and increase triglycerides levels on SD (4.09
fold). Under basal conditions, aortic ring of NAR has elevated response
to NE when compared to SD with endothelium (4.3 ± 0.41g;
2.29 ± 0.29g, respectively) and without endothelium (6.16 ± 0.33g;
3.89 ± 0.25g, respectively); this response was abolished by indomethacin. Endotoxaemia keep higher response to NE in NAR when compared
to SD only in aorta without endothelium (4.82 ± 0.35g; 2.95 ± 0.25g,
respectively) and indomethacin unchanged this response. Higher
response to NE observed on NAR under physiological conditions has
involvement of cicloxygenase and the inflammation abolish this difference on aorta with endothelium.
Paper No.: 885
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
VOLEMIC REPOSITION ON VASCULAR REACTIVITY OF
ENDOTOXEMIC RATS
Hermes Vieira Barbeiro, RC Petroni, TSL Garcia, DF Barbeiro,
CB Lorigados, FG Soriano, IT Velasco
University of São Paulo - Medical School/ LIM 51, São Paulo, Brazil
Volume and vasopressor therapy are among the primary goals in the
early management of septic shock and it has been shown to reduce
morbidity and mortality. Adult (220–270g) male Wistar rats received
saline i.p. (Control) or LPS (E coli serotype 026:B6) 10 mg/kg, i.p.,
LPS group were separated in 3 groups: LPS; LPS+Saline (Sal) 0.9%
and LPS+Hypertonic 7.5%. Treatment was performed 15 min (i.v.)
and the rats were sacrified 24h after LPS stimuli. Serum cytokine
(TNF) level was measured by ELISA. Thoracic aortic ring (5-6 mm)
with and without endothelium was kept in organ baths (37C - 95%
O2, 5% CO2) with Krebs solution for noradrenaline (NE) study (10 to
3000 nM). TNF was enhanced (7.6 fold) after LPS and volemic reposition do not altered this magnification. LPS stimuli decreased maximal response on aorta with and without endothelium when compared
with Control (1.6 fold twice). Absence of endothelium modulated
maximal response on control (1.2 fold) and sensitivity (EC50) on LPS
(2.2 fold). Volemic reposition proportionate higher maximal response
on LPS+Saline (1.4 fold) and higher sensibility (EC50) to
LPS+Hypertonic (2.9 fold) on aorta with endothelium and higher sensibility to LPS+Hypertonic (3.3 fold) on aorta without endothelium
when compare to control. Saline and hypertonic solution enhances
maximal response or sensibility to noradrenaline on isolated aorta of
endotoxemic rats, smooth muscle and endothelium maybe involved in
this finding.
Paper No.: 623
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
EFFECTS OF SURFACEN IN LOW AND MAINTAINED DOSE
IN THE TREATMENT OF ARDS. PRELIMINARY RESULTS
Yinet Barrese Pérez(1), Y Avila Albuerne(1), E Dı́az Casañas(2),
O Fernández Lima(2), R Uranga Piña(1), CM Ballagas Flores(1)
(1) National Clinical Trials Coordinating Center, Havana, Cuba
(2) National Center for Animal and Plant Health, Havana, Cuba
The acute respiratory distress syndrome (ARDS) is caused by a pulmonary inflammation and destruction of pulmonary surfactant; the application of exogenous surfactant improves oxygenation. Our objective was to
evaluate the effect of SURFACEN in low dose and sustained for three
days on the oxygenation. To this end we conducted a controlled randomized, multicentric clinical trial, in 21 adult patients treated with 100 mg
SURFACEN 3 times for 3 days and 18 remained only with mechanical
ventilation. The response was assessed as positive when the ratio PaO2/
FiO2 ‡ 200 at the end of treatment. The results showed a better response
indicator PaO2/FiO2 in the treated group (p = 0.0411). There was no difference in the clinical manifestations observed among groups or between
groups (p = 0.308) in the occurrence of adverse events. There were no
differences in mortality (p = 0.296) among groups, 11 patients in
SURFACEN group and 6 in the control group, however the causes of
death were not by ARDS but because the basic pathology that led to the
pulmonary inflammation. The treated group showed a decrease
(p = 0.0341) of 7.23 in the days of endotracheal intubation, there was
not difference in the days of mechanical ventilation (p = 0.132) or stay
in the intensive care unit (p = 0.108) although there was a tendency to
be better in the group treated with SURFACEN. We conclude that the
use of surfactant in ARDS improved oxygenation in patients with lower
FiO2 requirements, reduced the time of endotracheal intubation and had
a good safety.
Paper No.: 2842
FOCUSED CONFERENCE GROUP: P14 - ADDICTION AND
DOPING: NEUROBIOLOGICAL AND CLINICAL
INTERACTION OF PROTEIN KINASE C AND ALLOSTERIC
MODULATION AT EXTRASYNAPTIC GABA-A RECEPTORS
FOLLOWING CHRONIC ETHANOL TREATMENT
I Barrett, Stephen Kelley
Kent University, Medway School of Pharmacy, Chatham Maritime, UK
Previous studies have shown that extrasynaptic GABA-A receptors are
sensitive to low ethanol doses (Wallner, M.et al., Proc Natl Acad Sci U S
A 2003, 100, 15218-23) However, others have not reproduced this effect
(Borghese, C.M. et al., J Pharmacol Exp Ther., 2006, 316, 1360-68).
This inconsistency may be due to activity of PKC isozymes regulating
GABA-A receptor sensitivity to ethanol. We attempted to address this
question by examining the effects of the PKC inhibitor, calphostin-C
upon receptor sensitivity to low ethanol doses in mouse fibroblast Ltkcells stably expressing human recombinant a4b3d GABA-A receptors.
Ethanol (0.3, 3, 10, 30 and 100 mM) was co-applied with EC20 GABA
concentrations. The lower doses of ethanol did not potentiate GABAevoked currents in cells expressing GABA-A a4b3d receptors, however
we did find a significant potentiation of GABA-induced currents by
100 mM ethanol (p < 0.05). Intracellular application of calphostin-C
(400 nM) resulted in a significant ethanol potentiation at 30 mM and
100 mM (p < 0.05). We also tested the effects of calphostin-C upon
3a5a-THP neurosteroid potentiation of GABA-induced currents. Calphostin-C reduced 3a5a-THP potentiated currents. Chronic ethanol administration (14 days, 20 mM) had no significant effect upon 3a5a-THP
potentiation; however calphostin-C application significantly attenuated
the 3a5a-THP potentiation in cells chronically treated with ethanol
(p < 0.05). These results indicate a complex interaction between PKC
activity and the allosteric modulation of GABA-A receptors.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
191
Paper No.: 2879
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
EFFECTS OF THE ENDEMIC PLANT, ERYNGIUM KOTSCHYI,
ON RAT ISOLATED ILEUM AND DETRUSSOR MUSCLE
E Baydan(1), M Kartal(2), S Aslan(2), S Ýnce(3), Begum Yurdakok(1),
H Ekici(1), H Alp(7)
(1) Ankara University Faculty of Veterinary Medicine, Department of
Pharmacology and Toxicology, Ankara, Turkey
(2) Ankara University Faculty of Pharmacy, Ankara, Turkey
(3) Afyon University Faculty of Veterinary Medicine, Turkey
(4) Harran University Faculty of Veterinary Medicine, Turkey
Erngium kotschyi, an endemic plant, have been used in folk medicine in
Turkey, to treat gastrointestinal disorders and for its diuretic, antiinflammatory and antinociceptive effects. However no pharmacological evidence for its effectiveness on ileum and bladder has been reported. This
study, was aimed to determine the contractile effect of the lyophilized
extracts of the aereal parts of E. kotschyi on isolated rat ileum and
detrussor muscle (DM) and financialized by Ankara University (AUBAP). For this purpose, ileal and detrussor muscle strips (n:40) were
taken from 20 healthy male Wistar rats weighting around 250-300 g and
suspended under a load of 1 g. The cumulative concentrations of acethylcholine (ACh) (0.5 log 10-8-10-4 M) and the plant extract (0.0783.125 mg/ml ileum, 3.125-100 mg/ml DM) were applied. The plant
showed contractility on both ileum and DM. EC50 of ACh were determined as (2.5X10-7). The potency of the plant extract with respect to
ACh (pD2: 6.6) was found as 0.37 in ileum (Emax: 0.032 mg/ml, pD2:
2.49) and 0.045 in bladder (Emax: 4.7 mg/ml, pD2: 0.3) in cumulative
applications; suggesting the extract has greater potency in ileum than
DM. It was seen that pretreatment with the plant extract (0.3 mg/ml to
ileum and 4.7 mg/ml to DM) potentiated the cumulative contractions of
ACh and the cumulative concentrations of the plant extract pretreated
with oxybutinin (10 nM) did not have much effect on the contractility.
Further analyzes are currently been undergone in our laboratory to determine the mechanism of its action and complete pharmacodynamic effect.
Paper No. 3415
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
THE PECULIARITIES OF ACTION OF PENTOXIFYLLINE ON
RAT ISOLATED AORTA IN EXPERIMENTAL HYPOKINESIA
Aram Baykov, A Manukyan
Yerevan State Medical University ‘M. Heratsi’, Department of Pharmacology, Yerevan, Armenia
It is shown by the number of publications that restriction of movement
activity (hypokinesia) is a factor, modifying sensitivity to different vasoactive substances. The aim of this study was to reveal the influence of
hypokinesia on sensitivity isolated aortic rings (IAR) to pentoxifylline.
The experiments were performed on male mongrel albino rats. The control group of rats was kept in ordinary vivarium conditions and the
experimental group in individual narrow cages for simulation of hypokinesia during 15 days. The ability of pentoxifylline (10-6M) to decrease
isotonic contractions amplitude of IAR contracted by phenylephrine (107M) and P PGF2a - (4, 10-6M) was estimated. Cyclooxygenase (COX)
inhibition was done by diclofenac-sodium (3, 10-6M), and NO-synthase
(NOS) inhibition by 7-nitroindazole (10-4M) to define changes of
PG- and NO-dependent components of action of pentoxifylline. The
prominent decrease of sensitivity of IAR to pentoxifylline in case of
phenylephrine-induced contraction was observed in hypokinesia with no
changes in case of PGF2a-induced contraction. In COX-inhibition there
was an increase of dilatory effect of pentoxifylline in case of both constrictors in comparison to the corresponding index of control group. In
NOS-inhibition there was an authentic decrease of dilator response of
pentoxifylline in phenylephrine-induced contraction in hypokinesia,
whereas significant increase of dilatory response was noted in case of
PGF2a -induced contraction in the same conditions. Thus, hypokinesia
decreases sensitivity of IAR to pentoxifylline in case of phenylephrineinduced contraction with no change in case of PGF2a -induced one.
Changes of PG- and NO-dependent components of pentoxifylline action
in hypokinesia have different directions.
Paper No.: 1446
FOCUSED CONFERENCE GROUP: P02 - TRANSMEMBRANE
TRANSPORT: PERSPECTIVES FOR DISEASE AND DRUG
DISCOVERY
RHO KINASE INHIBITOR FASUDIL MODIFIES THE ACTIN
CYTOSKELETON OF ASTROCYTES UP-REGULATING
GLUTAMATE TRANSPORT AND ANTI-INFLAMMATORY
MECHANISMS
Philip Beart(1), C Lau(1), V Perreau(1), N Cheung(2), M Chen(2), L
Bischof(3), R O’Shea(1)
(1) University of Melbourne, Florey Neuroscience Institutes, Melbourne,
Australia
(2) University of Tasmania, Menzies Institute, Hobart, Australia
(3) CSIRO Mathematical & Information Sciences, North Ryde, Australia
Inhibition of Rho kinase has benefits in a number of brain injuries by
reducing glial scarring and thus allowing neuronal plasticity. Astrocytes
exhibit ‘‘good-bad’’ responses during reactive astrogliosis and glutamate
transporters (EAATs) may be an integral part of these homeostatic
responses. EAAT2 may play key roles in astrocyte biology as an endogenous ‘‘sensor’’ due to its high abundance in astrocytes. We explored
responses of cultured murine astrocytes to Fasudil (HA-1077) by monitoring cytochemistry (GFAP, F- & G-actin), EAAT function (activity,
cell-surface expression, localization) and the transcriptome by microarray
(Illumina Sentrix MouseRef-8 v2.0). Fasudil (100 lM, 24 h) altered
astrocytic morphology to a ‘‘physiological’’ stellate phenotype by 1h
with loss of cobble-stoned morphology. Image analyses suggested a
decreased abundance of phalloidin-positive actin stress fibres since there
was a significant conversion of F-actin to monomeric G-actin. Kinetic
studies of glutamate transport revealed a doubling of Vmax and biotinylation with Western blotting showed cell surface expression of EAAT2,
but not EAAT1, was doubled at 24h. Bioinformatics of microarray analyses of RNA from Fasudil- and vehicle-treated astrocytes (2 - 24h)
revealed extensive changes to genes involved in actin cytoskeleton,
TGFb-signalling and anti-oxidant mechanisms. Quantitative RT-PCR
confirmed microarray analyses. Together these data reveal a remarkable
spectrum of responses induced in astrocytes by Fasudil. Not only were
there elevations of anti-inflammatory astrocytic responses (BDNF, antioxidant and metabolic responses) and a switch to a ‘‘physiological’’ stellate phenotype, but the substantial elevation of EAAT2 would imply an
availability to efficiently scavenge toxic extracellular L-glutamate.
Paper No.: 1953
FOCUSED CONFERENCE GROUP: P06 - THE HEART GONE
WRONG; STABILIZATION OF CARDIAC
FUNCTION -CLINICAL
MECHANICAL AND STRUCTURAL CHARACTERISTICS OF
CAROTID PLAQUES: ANALYSIS BY MULTI-ARRAY
ECHOTRACKING SYSTEM AND MRI
Hélène Beaussier, O Naggara, D Calvet, R Joannides, B Laloux,
E Bozec, E Guegan-Massardier, E Gerardin, I Masson, C Oppenheim,
P Boutouyrie, S Laurent
(1) Hôpital Européen Georges Pompidou, Department of Pharmacology,
Paris, France
(2) Centre Hospitalier Sainte-Anne, Departments of Neurology and
Neuroradiology, Paris, France
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
192
(3) CHU Rouen, Departments of Pharmacology, Neurology and Neuroradiology, Rouen,France
Introduction: Combining functional and structural approaches may
improve the predictive value for plaque rupture and ischemic events.
Two distinct patterns of bending strain (BS) were previously determined
along the common carotid artery (CCA) (Paini et al. Stroke 2007;
38:117-23): Pattern A (i.e. outward BS) and its opposite, Pattern B (i.e.
inward BS). Our aim is to correlate arterial mechanics and composition
of an atherosclerotic plaque at the site of the CCA. Materials/Patients: 27
patients with carotid stenosis and an atherosclerotic plaque on the ipsilateral CCA were included: 18 asymptomatics (AS) and 9 symptomatics (S,
i.e. with previous ischemic stroke). Mechanical parameters were measured at 128 sites on a 4 cm long CCA segment by a novel non-invasive
echotracking system (ArtLab) and plaque composition was determined
by non invasive magnetic resonance imaging (MRI). Results: Pattern A
plaques (n = 21) were more often associated with ‘‘simple’’ plaque (i.e.
AHA stage I-III) than complex plaque (AHA stage IV-VII), by contrast
to pattern B plaques (n = 25) (chi square P = 0.03). No significant difference in BS pattern was observed between S and AS carotids. Among S
carotids, plaques were characterized by an outward reModelling
(increased external diameter and no change in internal diameter) whereas
among AS carotids, plaques grew according to an inward remodeling.
Conclusion: Longitudinal mechanics of ‘‘complex’’ plaques follows a
specific pattern of inward BS and suggest that Pattern B, associated with
an outer remodeling, is a feature of vulnerable plaques.
Paper No.: 1513
FOCUSED CONFERENCE GROUP: P08 - DEVELOPMENTS IN
THE TREATMENT OF SEXUAL DYSFUNCTION AND
DISEASES OF THE LOWER URINARY TRACT
RHO-KINASE INHIBITION IMPACTS NEUROGENIC
DETRUSOR OVERACTIVITY IN CHRONIC SPINALIZED
RATS
Delphine Behr-Roussel(1), D Broquères-You(1), S Oger(1), S Compagnie(1), A Caisey(1), P Denys(2), E Chartier-Kastler(3), F Giuliano(2)
(1) PELVIPHARM, Orsay, France
(2) Raymond Poincaré Hospital, Garches, France
(3) La Pitié-Salpêtrière Hospital, Paris, France
Spinal cord injury (SCI) severely disrupts normal bladder function by
inducing neurogenic detrusor overactivity (NDO). First line treatments
i.e. antimuscarinics often associated with intermittent catheterization are
limited by a moderate clinical efficacy and a significant incidence of side
effects. Thus, the development of new drugs for the treatment of NDO is
of crucial importance. Rho-kinase has a central role in the regulation of
detrusor smooth muscle contraction since this signalling pathway is
involved in the Ca2 + -sensitization of the smooth muscle. Thus, we
aimed to evaluate the effects of a rho-kinase inhibitor (Y-27632) on urodynamic parameters in SCI rats. Complete T7-T8 spinal cord transection
was performed in 17 female adult Sprague-Dawley rats. At 3-4 weeks
post-SCI, cystometry was performed in conscious rats to determine the
effects of Y-27632 (150 lg/kg, intravenous injection, iv, n = 7) and vehicle (saline, iv, n = 10) on the following urodynamic parameters: maximal
amplitude of micturition pressure (MP); baseline intravesical pressure
(BP); pressure threshold for inducing micturition (PT); intercontraction
interval, (ICI); voided volume; amplitude of non-voiding contractions
(NVC) and volume threshold necessary to initiate NVC. Y-27632 significantly increased voided volume whereas it did not modify MP, BP, d PT
and ICI. The amplitude of NVC was significantly decreased and the volume threshold of NVC significantly increase after Y-27632 administration when compared to vehicle (P < 0.001). Inhibition of rho-kinase
enables a better bladder emptying in the rat model of SCI-induced NDO.
This supports the potential development of rho-kinase inhibitors for the
treatment of NDO.
Paper No.: 1514
FOCUSED CONFERENCE GROUP: P08 - DEVELOPMENTS IN
THE TREATMENT OF SEXUAL DYSFUNCTION AND
DISEASES OF THE LOWER URINARY TRACT
RHO-KINASE INHIBITION RELAXES DETRUSOR FROM
NEUROGENIC PATIENTS
Delphine Behr-Roussel(1), S Oger(1), D Gorny(1), J Bernabé(1), E
Chartier-Kastler(2), P Denys(3), F Giuliano(3)
(1) PELVIPHARM, Orsay, France
(2) La Pitié-Salpêtrière Hospital, Paris, France
(3) Raymond Poincaré Hospital, Garches, France
Rho-kinases have a central role in the regulation of bladder smooth muscle. In vitro or in vivo data from animal models of overactive bladder
(OAB) indicate that rho-kinases could be involved in the pathophysiology of OAB but the role of rho-kinases in patients with neurogenic
detrusor overactivity has not yet been explored. We evaluated the effect
of rho-kinase inhibition on detrusor from neurogenic patients pre-contracted with either carbachol or KCl. Bladder samples were obtained
from 13 neurogenic patients who underwent partial or total cystectomy.
Detrusor strips with or without urothelium were mounted isometrically
(resting tension 500 mg) in organ baths filled with Krebs-HEPES maintained at 37C bubbled with 95%O2-5% CO2. After equilibration,
concentration-response curves (CRC) for the rho-kinase inhibitor,
Y-27632, (from 10-8M to 3.10-5M) or vehicle were generated on either
carbachol (1lM)- or KCl (50 mM)-precontracted detrusor (N = 7 in each
condition). Y-27632 induced a significant concentration-dependent
inhibition of KCl-induced (pD2 = 5.4 ± 0.1;Emax=-31.0 ± 4.3%) and
carbachol-induced contraction of human detrusor strips without urothelium
(pD2 = 5.3 ± 0.1;Emax=-56.1 ± 8.0%). The presence of urothelium did
not modify these inhibitory effects. Thus, rho-kinase inhibition decreases
detrusor contractions from neurogenic patients. The mechanisms responsible for this inhibition are different depending on whether the contraction is induced by carbachol or KCl, involving different signalling
pathways. This supports further investigations regarding the development
of rho-kinase inhibitors for the treatment of OAB.
Paper No.: 534
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
THE EFFECT OF ACUTE AND CHRONIC ADMINISTRATION
OF NEVIRAPINE ON INTERLEUKIN-2 IN A RAT MODEL
Zanelle Bekker, A Walubo, JB Du Plessis
University of the Free State, Department of Pharmacology, Bloemfontein, South Africa
Nevirapine (NVP) induced hepatotoxicity is regarded as partly due to
hypersensitivity reactions, implying activation of the immune system.
Therefore the aim of this study was to determine the effect of acute and
chronic NVP administration on interleukin-2 (IL-2), a cytokine implicated in cell-mediated toxic-immune reactions, after subclinical immune
stimulation with bacterial lipopolysaccharide (LPS). Male Sprague-Dawley rats were used and the study was approved by the animal ethics
commitee. During acute phase, 2 groups of 10 rats received NVP
(200mg/kg) after pre-treatment with either normal-saline (control) or LPS
(test-group). Five animals were sacrificed at 6 and 24 hours after NVP
administration. For the chronic phase, 2 groups of 15 rats each received
NVP (200mg/kg) daily over 3 weeks. After every 7 days, 5 rats from
each group were pre-treated with either normal-saline (control) or LPS
(test-group) and sacrificed 24 hours later. Serum IL-2 was measured by
ELISA. Although not statistically significant, during acute phase, IL-2
concentration (pg/ml: mean±sd) was higher in the LPS-treated-group than
in the control (421.6 ± 65.7 vs. 386.1 ± 33.4 at 6 hrs; and 398.6 ± 42
vs. 368.9 ± 16.6 at 24 hrs), but during chronic administration, IL-2
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
193
concentration (pg/ml) was lower in the LPS-treated-group than in the
control at week 1 (298.8 ± 10.8 vs. 359.7 ± 62.9), week 2 (351.9 ± 41.4
vs. 378.9 ± 60.3) and week 3 (313.1 ± 23.2 vs. 469.4 ± 143.1). These
results imply that NVP is a slow-onset IL-2 stimulant as was observed
during chronic administration, and that this was augmented in the acute
phase by LPS. Further studies are needed on the toxicologic significancy
of these observations.
Paper No.: 2780
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
THE ENDOTHELIAL PATHWAY FOR CURRENT SPREAD IS
MODULATED BY SMOOTH MUSCLE K+-CHANNELS
Timea Z Beleznai, KA Dora
University of Oxford, Department of Pharmacology, Oxford, UK
The endothelium forms the conduit for distal spread of hyperpolarizing
signals along the longitudinal axis of resistance arteries. Any agonist that
stimulates smooth muscle cell hyperpolarization has the potential to be
associated with homo- and heterocellular intercellular electrical coupling
and spreading dilatation, and thereby influence blood flow. We have a
model to study spreading dilatation by isolating and triple-cannulating rat
mesenteric arteries. Arteries are mounted in a gravity-fed pressure myograph and kept at 37C, with one branch of the bifurcation connected to a
syringe pump containing agonist and a fluorescent dye. Constant downstream flow through the feed artery allows luminal perfusion of agonists
into the sidebranch without reaching the feed artery, and using confocal
microscopy we can simultaneously measure local dilatation in the branch
and spreading dilatation into the feed artery. Agonists acting selectively
at endothelial (acetylcholine) and smooth muscle (isoprenaline) K+-channels each evoked spreading dilatation with similar mechanical length
constants near 1 mm. The ATP-sensitive K+-channel blocker glibenclamide doubled the length constant for acetylcholine and effectively abolished that for isoprenaline. Combined blockade of large conductance
Ca2 + -activated K+-channels and voltage-gated K+-channels with iberiotoxin or tetraethylammonium together with 4-aminopyridine markedly
and consistently increased the magnitude of spreading dilatation to both
agonists, such that the length constant approached infinity in the 2 mm
length of artery. Changes in endothelial cell calcium could not account
for the apparently regenerating current. These data show that activated
smooth muscle cell K+-channels not only serve to modulate contraction,
but also reduce the effectiveness of spreading dilatation.
Paper No.: 1147
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
GLUTATHIONE S-TRANSFERASE M1 AND GLUTATHIONE
S-TRANSFERASE T1 POLYMORPHISMS AND LUNG CANCER
RISK IN TUNISIAN MEN
Majda-Noura Belkhiria(1), MA Jebali(1), I Harrabi(2), J Knani(3)
(1) Faculty of Medicine of Monastir, Laboratory of Pharmacology,
Monastir, Tunisia
(2) CHU Farhat Hached, Department of Epidemiology, Sousse, Tunisia
(3) CHU Tahar Sfar, Department of Pneumology, Mahdia, Tunisia
Glutathione S-transferases (GST) detoxify polycyclic aromatic hydrocarbons found in tobacco smoke by glutathione conjugaison. Polymorphisms within the GSTM1 and GSTT1 genes, coding for enzymes with
deficient or reduced activity, have been studied as potential modifiers of
lung cancer risk. In this study, we evaluated the association between the
GSTM1 and GSTT1 polymorphisms and lung cancer risk, and their
interaction with cigarette smoke. The GSTM1 and GSTT1 polymorphisms were determined by multiplex PCR in 80 lung cancer patients
and 86 controls. Associations between specific genotypes and the development of lung cancer were examined by use of logistic regression to
calculate odds ratios (OR) and 95% confidence intervals (CI). The frequencies of GSTM1 and GSTT1 null genotypes were 57.5 and 32.5% in
lung cancer patients and 52.3 and 17.4% in controls, respectively. The
GSTM1 homozyous null genotype was not associated with an increase
risk of developping lung cancer. There was a marginally significant association between lung cancer and GSTT1 null genotype (OR : 2.22, 95%
CI : 1.10-4.61). After grouping according to smoking status, GSTT1 null
genotype was associated with an increase lung cancer risk for smokers
(OR = 2.99, 95% CI = 1.01-8.80). Our findings suggest that the GSTT1
null genotype may be associated with an increased susceptibility to lung
cancer in Tunisian men.
Paper No.: 1372
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
A LINEAL MATHEMATICAL MODEL BASED ON HBA1C
LEVELS FOLLOW UP MAY PREDICT THE YEARS OF
EVOLUTION OF TYPE 2 DIABETES MELLITUS STARTING
FROM A SINGLE HBA1C LEVEL QUANTIFICATION
Inmaculada Bellido, AE Santos, L Garcı́a-Carrascal, JJ Sanchez-Luque,
E-I Marquez, E Blanco, A Gomez-Luque
University of Malaga School of Medicine, Department of Pharmacology
and Clinical Therapeutics, Malaga, Spain
Background: Glycosylated haemoglobin (HbA1c) is the best diabetes’
control and complication risk predictor. Aim: To design a mathematical
model to predict the type 2 diabetes mellitus (T2DM) evolution time
based in patient HbA1c levels. Methods: T2DM patients were prospectively studied (5 years). All the physiopathological characteristics and
antidiabetics’ drugs treatments were recorded. The population statistical
analyses was done by ANOVA test, correlations, polynomic interpolation,
regression toward the mean, uni-bivariate linear, non-linear (including
Levenberg-Marquardt algorithm) and logistic regression analysis. Results:
346 patients with T2DM, aged 67.2 ± 10.9 years, male 50.4%, weighed
79.8 ± 1.3, mean body mass index 31.1 ± 0.5, were enrolled. The main
population characteristics were: basal glycaemia 137.5 ± 7.3 mgr/dL,
HbA1c 6.75 ± 0.59, confirmed-diabetes evolution 7.16 ± 0.68 years,
hypertension 6.45%, dyslipemic diseases 6.48%, obesity 6.3%, and
treated with insulin 17.2%, oral antidiabetics 29.8%, anti-hypertensive
agents 33.4%, normolipemiants 53.2%, platelet antiaggregants 43.2%.
The mathematical model which really shown a good adjust to the majority of the patients data was a linear differential equation with a negative
constant of proportionality which may predict the diabetes years of
evolution starting from a single HbA1c level quantification with an
r-squared adjusted for DF of 63.9% and with a standard error of 0.124.
Conclusion: This mathematical model is able to adjust predict the years
of evolution of type 2 diabetes mellitus starting from a single glycosylated haemoglobin level quantification in patients with glycosylated
haemoglobin levels between 5.5% and 9.5%. It is not reliable in case of
very low and very high HbA1c levels, and more than 60 years of diabetes
evolution.
Paper No.: 1373
FOCUSED CONFERENCE GROUP: PW19 - INFLUENCE OF
DEGENERATION AND REPAIR IN THE CNS AND PERIPHERY
THE USE OF E-LEARNING IMPROVED THE
PHARMACOLOGY LEARNING IN CHIROPODIST’S
PHARMACOLOGY TRAINING
Inmaculada Bellido(1), E-I Marquez(1), JA Garcia-Arnes(1),
E Blanco(1), A Gomez-Luque(2)
(1) University of Malaga School of Medicine, Department of
Pharmacology and Clinical Therapeutics, Malaga, Spain
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
194
(2) University Hospital Virgen Victoria, Department of Anaesthesia,
Malaga, Spain
Background: The e-learning prepared teaching material may awake our
student’s abilities to think, to learn and to develop clinical skills for their
professional lives. Aim: To quantify the efficacy of a PC-supported programme (Chiropharm) added to a face-to-face teaching of Pharmacology
training for Chiropody students. Methods: Students of Pharmacology in
Chiropody training taught by the face-to-face (FtF) (2006-07) were
compared with student taught with FtF complemented with Chiropharm
(ChiropharmP) (2008-09) methodology. ChiropharmP is an interactive
PC-supported programme design using Windows XP, UMA’s Moodle
platform, Word, Power Point, Front Page, Hot Potatoes, specialized
medical webs’ free material and Explorer/Mozilla software, including
texts and slides presentation with iterative buttons linked to hyper-text to
questions, references, figures, films, iterative auto-evaluations, clinical
cases and high fidelity simulators. A quality and quantity evaluation of
the student’s knowledges, and an anonymous satisfaction questionnaire
were analyzed (unpaired-groups’ student t test). Results: 23 (51% of
n = 45) students were initially trained in ChiropharmP handling during
3 h. They spent a mean of 33 ± 5.7 h using ChiropharmP during the
course development (2.9 ± 0.7h/week/x13 weeks). We founded some
differences between the two groups (FtF vs. ChiropharmP) (p < 0.05):
time of study 6.1 ± 1.7h/week vs. 3.3 ± 0.9h/week; correct resolved
clinical cases 54.5%-91%; approved student 68.1% (46.6% with A-B)
vs. 95.6% (73.9% with A-B); students which prepared themselves for the
honours qualification special exam 5 vs. 16; percentage which consider
this teaching-learning system better than the classic methodology 95%.
Conclusion: The Pc-supported programme ChiropharmP improved the
pharmacology learning and was more pleasant for Pharmacology student
of Chiropody.
Paper No.: 2384
FOCUSED CONFERENCE GROUP: P19 - GENERAL SESSION ONCOLOGY
AN INCREASED PRECRYOPRESERVATION TIME REDUCES
THE FEASIBILITY AND THE RECOUNT OF THE CD34(+)
CELLS OF THE UMBILICAL CORD BLOOD
Inmaculada Bellido(3), L Ponce(1), MC Hernández(2), M Barrios(1),
I Prat(2), AI Heiniger(1)
(1) Carlos Haya Universitary Regional Hospital. Haematology Service.
Malaga
(2) Regional Center for Blood Transfusion-Umbilical Cord Unit. Malaga.
Spain
(3) University of Malaga School of Medicine, Department of Pharmacology and Clinical Therapeutics, Malaga, Spain
Background: Cryopreservation protocols for umbilical cord blood’s (UC)
haematopoietic cells try to approach an acceptable level of recovery
CD34(+) by optimizing the extraction, transport, processed and cryopreservation methods of the UC. It is generally accepted that UC units can
be safely cryopreserved more than 24 h after harvest, but a delay in
freezing can influence the final quality of the UC-unit. Aim: To determine if the total time from extraction to cryopreservation, which mainly
dependent on shipment variables, may modify the feasibility and the
recount of the CD34(+) cells in the umbilical cords units. Methods: The
feasibility and number of CD34(+) cells in the UC-units from 5400 UC
samples analyzed in Malaga’s UC-bank were analyzed. The samples proceeded from 27 Andalusia hospitals (processed from 2009 January to
September (female aged 30.7 ± 0.2 years, gestation weeks 39.8 ±
0.05 weeks, type of delivery eutocic 71.5%, instrumental 16.5%, caesarea 7.6%, male neonate proportion 52.8%). Mother and neonates clinical
data, procedure for UC collection, time of collection, shipment-phases
and cryopreservation-phases, and feasibility and CD34(+) cells were
recorded. Results: Feasible samples shown a lower precryopreservation
time than non-feasible samples (-11.9%, 24.4 ± 0.3h vs. 27.7 ± 0.5h,
P < 0.01). Both feasibility and recount of CD34(+) cells showed an
inverse correlation with precryopreservation time duration with Pearson
correlation coefficient of -0.214 (p = 0.001) and -0.088 (p = 0.049).
Conclusion: The transport, processes and storage time previous to cryopreservation must be reduced as much as possible to minimized the
reduction in the CD34(+) cells of umbilical cord blood feasibility related
with the increment of the precryopreservation time.
Paper No.: 2410
FOCUSED CONFERENCE GROUP: P10 - DRUGS FOR HALF
THE WORLD: PAEDIATRIC CLINICAL PHARMACOLOGY
GRAPHIC AND PICTORIAL TABLES EXPLAINING DRUG’S
ROUTES ADMINISTRATION REDUCE THE ANXIETY OF THE
HOSPITALIZED CHILDREN IN THE INTENSIVE CARE UNIT
MV Bellido(1), Inmaculada Bellido(2), A Gomez-Luque(3)
(1) Carlos Haya Universitary Hospital, S. Intensive Care Unit, Malaga,
Spain
(2) University of Malaga, School of Medicine, D. Pharmacology and
Clinical Therapeutics, Malaga, Spain
(3) Virgen de la Victoria Universitary Hospital, S. Anaesthesiology,
Malaga, Spain
Background: Art communication techniques are usefully tool to communicate with the children following the ‘‘narrative-based medicine’’. Aim:
To evaluate the impact of graphic/pictorial tables explaining drug’s routes
administration on pain and anxiety development by the children during
drugs administration in the intensive care unit (ICU). Methods: Children’s hospitalized in the ICU having elective major surgery and oncology therapy were collected prospectively. Clinical stage, surgery and
anaesthesia procedures, oncologic, analgesics, anxiolytics treatments and
therapy-related complications were recorded. Pain and anxiety were measured by a visual analogy scale (VAS), children faces pain scales, and
STAIC questionnaire after the use of a DRA-table to explain the drug’s
routes administration and were compared with a group with DRA-table’s
explication. esults: 125 children aged 3-10 years (6.5 ± 1.5 years old,
65% male) were enrolled. The procedures included cardiac (51%), digestive (6%), neurosurgery (20%) and oncologic diseases (23%). The drugs
routes administration were oral/nasogastric tube (1%), intranasal (3.1%),
inhalatory (30.9%), intramuscle (1.3%), intravenous (50.2%), intrathecal
(13.5%). Both VAS-pain and VAS-anxiety quantification were higher
(p < 0.05) in control than in DRA-table group (5.6 ± 0.9 vs. 5.1 ± 0.5,
and 7.8 ± 1.1 vs. 5.5 ± 0.6, respectively). The reduction of anxiety in
the DRA-table group with respect to the control group was age and gender-related (higher in children up to 8 years than under 5 years, higher in
female than in male). The control group needs much more analgesics,
anxiolytics and sedans than DRA-table group. Conclusion: Graphic and
pictorial tables explaining drug’s routes administration reduce the anxiety
of the hospitalized children in the intensive care unit.
Paper No.: 1897
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
ENDOTHELIUM-DERIVED HYPERPOLARIZING FACTOR
AND NITRIC OXIDE REGULATE ARTERIAL WALL
VISCOSITY IN VIVO IN HUMANS
Jeremy Bellien, M Lacob, C Thuillez, R Joannides
Xxxxx
Although the viscoelasticity of conduit arteries has been extensively
investigated, few studies have focused on arterial wall viscosity (AWV)
itself and its regulation by the endothelium in vivo. This is of particular
importance since AWV is a major source of energy dissipation through
the vascular system reducing cardiovascular coupling efficiency. We
simultaneously measured radial artery diameter and arterial pressure
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
195
(NIUS02) in healthy volunteers before and during local infusion of pharmacological inhibitors of endothelial pathways. L-NMMA (8 mmol/min)
was used as NO-synthase inhibitor, tetraethylammonium (TEA: 9 mmol/
min), as blocker of calcium-activated potassium channels, the target of
endothelium-derived hyperpolarizing factors (EDHF) and fluconazole
(0.4 mmol/min), as inhibitor of cytochrome epoxygenases which promote the synthesis of epoxyeicosatrienoic acids synthesis, identified as
EDHF in conduit arteries. AWV was estimated from the ratio of the area
of the hysteresis loop of the pressure-diameter relationship to the area
representing the whole energy exchanged during each cardiac cycle.
L-NMMA paradoxically reduced AWV (n = 5: 27.6 ± 0.7 to
23.4 ± 0.7%, P = 0.053). Conversely, AWV was increased by TEA
(n = 6: 25.5 ± 0.5 to 31.3 ± 0.7%, P = 0.040) and fluconazole (n = 5:
26.6 ± 0.6 to 30.6 ± 0.6%, P = 0.047). This increase was more marked
with the association of L-NMMA+TEA (n = 6: 27.6 ± 0.9 to
41.0 ± 0.7%, P = 0.002) and L-NMMA+fluconazole (n = 6: 26.1 ± 0.7
to 36.3 ± 0.3%, P = 0.001) showing a synergistic effect of both combinations on AWV. These results demonstrate that the endothelium contributes in vivo in humans to the regulation of AWV through an interaction
between NO and a cytochrome-related EDHF.
Paper No.: 2874
FOCUSED CONFERENCE GROUP:
P04 - PHARMACOEPIDEMIOLOGY, CURRENT
CONTROVERSIES AND OPPORTUNITIES
ADHERENCE TO ANTIHYPERTENSIVE MEDICATIONS AND
ALL-CAUSE MORTALITY AND CARDIOVASCULAR EVENTS:
A POPULATION-BASED STUDY BASED ON
ADMINISTRATIVE DATA
Silvia Benemei(1), S Saragoni(2), P Batacchi(3), P Geppetti(1),
M Di Bari(4), N Marchionni(4), S Buda(2), ED Esposti(5), LD
Esposti(2)
(1) University of Florence, CIFF, Department of Preclinical and Clinical
Pharmacology, Florence, Italy
(2) CliCon S.r.l., Health, Economics & Outcomes Research, Ravenna,
Italy
(3) University of Florence, CIFF, Pharmaceutical Department ASL10,
Florence, Italy
(4) University of Florence, CIFF, Department of Critical Care Medicine
and Surgery, Florence, Italy
(5) Umberto I Teaching Hospital, Rome, Italy
Adherence to antihypertensive treatment (AT) is limited, ranging from 30
to 70% of treated patients, depending on its definition. The association
between poor adherence to AT and cardiovascular (CV) morbidity/mortality remains largely unexplored. Using administrative databases of citizens living in Florence, Italy, a retrospective cohort study was conducted
with inclusion of records of subjects (>18years), receiving a first prescription of AT between January 1st, 2004 and December 31st, 2006.
Adherence to AT was defined as the proportion of days (PD) on which a
patient had pills available in the interval from the first purchase of AT
until all-cause death, non-fatal stroke or myocardial infarction (composite
outcome) or leaving database and classified as poor (PD?40%), moderate
(PD:41%-60%), good (PD:61%-80%), or excellent (PD>80%). Subjects
prescribed AT for secondary CV prevention, occasional users, and those
who suffered a major CV event within 6 months of enrollment were
excluded. Cox regression model was used to determine the association
between adherence and composite outcome. 31,306 records (mean age of
60.2 ± 14.5 years, 52.0% women) were included. Adherence was poor
in 25.7%, moderate in 14.8%, good in 18.1%, and excellent in 41.5% of
subjects. Compared to poorly adherent subjects, the risk of the composite
outcome was progressively lower in those with moderate (HR=0.87),
good (HR=0.69), and excellent adherence (HR=0.53). Sub-optimal
adherence to AT is associated with substantial numbers of avoidable
deaths and CV events in primary prevention. In a public health perspective, this evidence underlines the need for improving adherence to medications in clinical practice.
Paper No.: 492
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
HONEY: THE OLDEST MODERN REMEDIES FOR MASTITIS
Mokhtar El-Bachir Benhanifia, L Boukraâ
University of Ibn Khaldoun, Department of Agro-Veterinary Sciences,
Tiaret,Algeria
Honey has been used in medicine against bacterial infections for thousands of years, the objective of this study was to evaluate the efficacy of
intramammary honey administered during lactating period, to eliminate
Staphylococcus, Aureus and Pseudomonas aeruginosa infection. Infected
mammary quarters of three cows were treated once daily with 5 ml of
multifloral honey per quarter for 2 consecutive days. Milk samples were
taken from affected quarters immediately prior to treatment on d 0, 7, 14,
21, 52 and 60 for microbiological examination. The cure following
administraton of honey was 66% (2/3 cows) at days 7 and 14; 100% (3/3)
at days 21, 52 and 60 for Staphylococcus aureus infection and 66% (2/3)
at days 21, 52 and 60 for Pseudomonas aeruginosa. This preliminary
study has shown the efficacy of honey as natural therapeutic product
against S. aureus and P. aeruginosa sub clinical mastitis. Further study
may elucidate the suseptibility of other pathogens microorganisms implicated in acute clinical and subclinical bovine mastitis (number of cases,
herd size….), the dynamic of honey (distribution, diffusion….) in mammary gland and apropriate intramammary formulation of honey for mastitis prevention and therapy in cows. Finally, honey could be an
alternative for mastitis treatment in both conventional and organic farms.
This will lead to a huge economic and health benefit worldwide.
Paper No.: 2943
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
TOXICITY OF CYTOSTATIC AGENTS TO GRANULOCYTEMACROPHAGE PROGENITORS (CFU-GM) INCREASED IN
ZUCKER OBESE RATS
I Benkõ(1), Krisztina Géresi(1), K Benkõ(2), B Szabó(3), A Megyeri(1),
E Ungvári(1), B Peitl(1), Z Szilvássy(1)
(1) University of Debrecen, Medical and Health Science Center, Department of Pharmacology and Pharmacotherapy, Debrecen, Hungary
(2) University of Debrecen, Medical and Health Science Center, Euromedi cDiagnostics, Debrecen, Hungary
(3) Semmelweis University, Heart Center, Budapest, Hungary
Increased risk of anticancer chemotherapy in seriously obese patients is
well-known especially in children with acute myeloid leukemia. Obesity
maybe one of the factors that predict treatment-related toxicity during
chemotherapy. We studied whether the functions of granulopoietic cells
do not be altered by obesity. At the first sight - granulopoiesis as measured by cellularity, frequency of granulocyte-macrophage progenitors
(CFU-GM) and total CFU-GM content of the femoral bone marrow –
did not differ in obese, insulin resistant Zucker rats compared with control, Wistar rats. Nevertheless we tested the vulnerability of their CFUGM cells by culturing them in vitro in the presence of carboplatin and
5-fluorouracil, two cytotoxic drugs with different mechanisms of action.
Both drugs were more toxic on CFU-GM cells of insulin resistant Zucker
obese animals than on CFU-GM cells of the control rats. This based on
metabolic disorders at least in partly, because we could demonstrate similar increased toxicity to CFU-GM progenitors originated from the nonobese but insulin resistant Goto-Kakizaki rats in the same dose ranges.
After in vivo administration of rosiglitazone, an insulin sensitizer, the
toxicity of the studied cytostatic drugs decreased proportionally with the
improvement of the insulin sensitivities both in Zucker obese and GotoKakizaki rats. However the increased treatment-related myelotoxicity and
mortality is well-known among obese patients with malignant diseases,
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
196
only the altered half-lives, volumes of distribution and clearances of the
cytostatic drugs have been taught to be the underlying reasons. Our
results are the first observation about an impaired granulopoiesis in obese
animals.
Paper No. 3253
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
ANGIOTENSIN-(1-7) PREVENTS DIABETES-INDUCED
ATTENUATION IN PPAR- C AND CATALASE ACTIVITIES
Ibrahim Benter(1), G Dhaunsi(1), M Yousif(1), S Akhtar(1),
M Chappell(2), D Diz(2)
(1) Kuwait University Faculty of Medicine, Department of Pharmacology, Safat, Kuwait
(2) Wake Forest University. School of Medicine, Winston-Salem, NC,
USA
Angiotensin-(1-7) is a vasodilator peptide that exhibits antihypertensive,
antithrombotic and antiproliferative properties. The mechanisms by
which angiotensin-(1-7) exerts its beneficial effects on end-organ damage
associated with diabetes and hypertension are not well characterized.
This study was designed to compare the effects of apocynin with angiotensin-(1-7) on renal vascular dysfunction and NADPH oxidase activity
in a combined model of diabetes and hypertension and to further determine whether chronic treatment with angiotensin-(1-7) can modulate
renal catalase, and peroxisome proliferator activated receptor- c (PPARc) levels in streptozotocin induced-diabetes in both normotensive Wistar
Kyoto rats (WKY) and in spontaneously hypertensive rats (SHR).
Apocynin or angiotensin-(1-7) treatment for one month starting attenuated elevation of renal NADPH oxidase activity in the diabetic SHR kidney and reduced the degree of proteinuria and hyperglycemia, but had
little or modest effect on reducing mean arterial pressure. Both drugs also
attenuated the diabetes-induced increase in renal vascular reactivity to
endothelin-1. Induction of diabetes in WKY and SHR animals resulted in
significantly reduced renal catalase activity and in PPAR-c mRNA and
protein levels. Treatment with angiotensin-(1-7) significantly prevented
diabetes-induced reduction in catalase activity and the reduction in
PPAR-c mRNA and protein levels in both animal models. These data
suggest that activation of angiotensin-(1-7)-mediated signalling could be
an effective way to prevent the elevation of NADPH oxidase activity and
inhibition of PPAR-c and catalase activities.
Paper No.: 1287
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
APPARENT DIFFERENCES IN PHARMACOKINETIC
PERFORMANCE BETWEEN GENERIC MYCOPHENOLATE
MOFETIL TABLET FORMULATIONS
Darren Bentley(1), L Banken(2), R Robson(3)
(1) Roche Products Ltd, Department of Clinical Pharmacology, Welwyn
Garden City, UK
(2) F. Hoffmann-La Roche Ltd, Welwyn Garden City, UK
(3) Christchurch Clinical Studies Trust, Christchurch, New Zealnd
CellCept, mycophenolate mofetil (MMF), is an integral component of
immunosuppressive regimens for the prophylaxis of acute rejection in
patients receiving allogeneic renal, cardiac or hepatic transplants. Several
generic formulations have been approved based on bioequivalence to the
innovator product. However, differences in dissolution rates between
generic formulations observed in vitro may lead to disparities in pharmacokinetic profiles. We compared pharmacokinetic profiles of three gen-
eric MMF formulations (A] Renodapt, Biocon Ltd; B] Mycept, Panacea
Biotec Ltd; and C] Cellmune, Cipla Ltd) in a randomized, open label,
four-way crossover study that also included CellCept. In each treatment
period, subjects received a single 500 mg MMF tablet with a high fat,
high calorie meal. Plasma concentrations of mycophenolic acid (MPA),
the major active metabolite of MMF, were measured up to 48 hours after
intake. Global statistical comparisons were made between the three generic formulations for Cmax and AUCinf and mean treatment ratios were
calculated. Thirty two healthy male volunteers were enrolled and 31
completed the study. Although global tests could not reject the null
hypothesis, there were apparent differences in the peak exposures to
MPA between the generic formulations. Mean Cmax for B was 23%
lower than C, and that for A was 15% lower than C. However, total
MPA exposures were similar across the generic formulations and CellCept. In conclusion, some apparent differences in pharmacokinetic performance exist between generic formulations. The clinical consequences
(e.g. organ rejection) of these differences when switching between generic MMF formulations remain to be investigated.
Paper No.: 1092
FOCUSED CONFERENCE GROUP: P12 - ION
CHANNELOPATHIES: NEW WINDOWS ON COMPLEX
DISEASE AND THERAPY
THE KV11.1 CHANNEL OPENER NS1643 SHORTENS THE QT
IN TRANSGENIC RABBITS WITH LONG QT-SYNDROME 1 IN
VIVO AND SHORTENS THE APD90 EX VIVO, BUT IS
ASSOCIATED WITH INCREASED RISK OF ARRHYTHMIAS
Bo Bentzen(1), S Bahrke(2), K Wu(2), AP Larsen(1), J Biermann(2),
X Peng(3), G Koren(4), M Zehender(2), C Bode(2), M Grunnet(5),
M Brunner(2)
(1) University of Copenhagen, Faculty of Health Sciences, Danish
Arrhythmia Research Centre, Copenhagen, Denmark
(2) Universitätsklinik Freiburg, Innere Medizin III, Freiburg, Germany
(3) Penn State University, Department of Comparative Medicine,
Philadelphia, PA, USA
(4) Brown University Alpert Medical School, Cardiovascular Research
Center, Rhode Island Hospital, Providence, RI, USA
(5) NeuroSearch A/S, Ballerup, Denmark
Aim: Transgenic rabbits expressing pore mutants of hKv7.1(Y315S) display a Long QT-Syndrome-1(LQT1) phenotype. Recently, the compound
NS1643 has been described to increase IKr/Kv11.1 current. We hypothesized that NS1643 would hasten cardiac repolarization and shorten the
action potential duration(APD90) in LQT1 rabbits, thereby normalizing
the QT-interval. Methods: Female transgenic LQT1 rabbits aged
4-6 months were compared to littermate controls(LMC) in vivo and
ex vivo. In vivo ECG studies(n = 9 vs 6/group) in sedated animals were
performed at baseline and during 30min i.v. infusion of NS1643(1.5mg/
kg/min) or vehicle(PEG400/glucose) in a crossover design with 1 week
between treatments. Ex vivo 4 left ventricular monophasic action potentials were recorded from Langendorff-perfused hearts (n = 13 vs 10/
group) at baseline and during 45min perfusion with NS1643(5lM) at
atrial pacing (RR= 400ms). Left ventricular refractory periods(VERP)
were assessed before and after 45min NS1643 infusion. Results: In vivo
NS1643 shortened the QTc after 30min significantly in LQT1(NS1643:
8.5%±0.7; vehicle:2.5%±1.1(P < 0.001)), whereas NS1643 did not
shorten the QTc significantly in LMC. In Langendorff experiments
APD90 was shortened by NS1643(LQT1: 32.0ms ±4.3(P < 0.01);
LMC:21.0ms±5.0(P < 0.01)). Concomitantly, the VERP was significantly shortened by NS1643 (LMC:22.6ms±9.9; LQT1:23.7ms±8.3).
Before NS1643 2 LMC and 5 LQT1 had ventricular fibrillation either
spontaneously or during VERP-testing, whereas after NS1643 9/10 LMC
and 13/13 LQT1 animals had VF. Additionally, dp/dt was significantly
decreased(LMC:63%±4; LQT1:49%±3) after NS1643. Conclusions: We
demonstrate that NS1643 shortens the QT-intervals in LQT1 significantly
in vivo. Ex vivo NS1643 shortens the APD90 and VERP in both genotypes, but is associated with increased arrhythmia susceptibility and
negative inotropy.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
197
Paper No. 3305
FOCUSED CONFERENCE GROUP: FC13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
THE STUDY OF RESPONSE TO BRONCHODILATORS
ACCORDING TO POLYMORPHISM B2-ADRENERGIC
RECEPTORS
Nadyia Berdnikova
Moscow Medical Academy, Moscow City Hospital #23, Moscow,
Russian Federation
Aim: to study the influence of polymorphism 2-adrenergic receptors
(gene ADRB2) on changing of FEV1 after taken ipratropium bromide
and salbutamol in the patients with asthma in dependence of severity of
asthma. Methods: 62 patients (48.6 11.9 ages; 33 men, 29 women) were
included in the study. The genotype (Arg, Gly) was examined by polymerase chain reaction (PCR). All the patients were divided into 2 groups:
I group -FEV1-60%, II group -FEV1-60% pred. FEV1 was measured in
all the patients before and after taken ipratropium bromide 80 mcg
(40 min.) and salbutamol 400 mcg (30 min.). Results: 26 patients have
16ArgGly: FEV1-79.8% (14), FEV1-39.6% (12). Heterosigous showed
increase FEV1 (%) after taken 2 bronchodilators: I- 8.38% and 13% and
II- 10% and 17% (p < 0.05). 16 patients have 16ArgArg: FEV1-74.8%
(10), FEV1-37.6% (6). Homosigous Arg showed increase FEV1 after
taken 2 bronchodilators: I- 1.35% and 16.9% and II- 7% and 13.4%
(p < 0.05). 18 patients have 16GlyGly: FEV1-76.4% (9), FEV1-49.8%
(9). Homosigous Gly showed increase FEV1 after taken 2 bronchodilators: I- 11.7% and 18% and II- 13.3% and 15.6%. Conclusion: only the
16GlyGly patients with FEV1¡Ü60% did not show increase FEV1 after
taken salbutamol. All the other patients showed.
Paper No. 3428
FOCUSED CONFERENCE GROUP: FC16 - NATURAL
PRODUCTS: PAST AND FUTURE?
STUDY OF INFLUENCY INDOL-3-CARBINOL,
EPIGALLOCATECHIN-3-GALLATE AND SOY ON
PROLIFERATION DISEASES OF THE REPRODUCTIVE
SYSTEM IN WOMEN
Nadyia Berdnikova, L Zorina, S Serebrova
Moscow Medical Academy, Moscow City Hospital #23, Moscow,
Russian Federation
I3C/EGCG/soy are natural products used for treatment. To confirm the
influence of I3C/EGCG/soy on proliferation diseases of reproductive system in women we searched 30 patients (41.3 4.28 age): with benign breast
diseases-6 patients (20%), myoma of body of uterus and adnexitis-4
(13.3%), adenomyosis and polyp of cervix of uterus-5 (16.6%), myoma
of body of uterus and adenomyosis-8 (26.6%), myoma of body of uterus
and adenomyosis and chronic cystic mastitis-7 (23.3%). Nobody from
patients took the hormonal treatment. Diagnoses were confirmed by sonographic and mammographic. All the patients took I3C/ EGCG/soy per os
for 6 months. The women with benign breast diseases estimated their pain
according questionnaire for pain. Patients (n = 13) showed changes of
scores for pain in mammary glands: before I3C/EGCG/soy with severe
pain-46.15%, moderate-23.07%, mild-30.77%, none-0%; after 3 months:
0%, 23.07%, 15.3% (p < 0.05), 61.63% (p < 0.05) accordingly; after
6 months: 0%, 15.3%, 15.3%, 69.33% accordingly. The size of cysts in
mammary glands changed by sonographic: when before taking I3C/
EGCG/soy it was 1.1 0.3sm then in 3 months it decreased to 0.60 0.22sm
(p = 0.0001). Cysts with size 0,5sm and less weren’t found in 3 months
by sonographic. Patients (n = 19) showed changes of uterine volume/
myoma: before I3C/EGCG/soy-184.1 4.1ml/80.1 3.6ml; after 3 months168.3 3.5ml (p < 0.05)/72.4 2.4 ml (p < 0.05); after 6 months: 162.2
3.7ml/70.4 2.1ml (p < 0.05). After 3 month taking I3C/EGCG/soy the
decrease in pain, oedema and size of cysts in women with benign breast
diseases was confirmed. We did not find extra changes after 6 months.
The decrease in uterine volume and myoma was statistically significant.
Paper No.: 1352
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
PUBLICATION RATE AMONG INVESTIGATOR-INITIATED
CLINICAL DRUG TRIALS IN DENMARK
Louise Berendt(1,2), KF Bach(1), T Callreus(2), LG Petersen(2),
HE Poulsen(3), K Dalhoff(3)
(1) Bispebjerg University Hospital, GCP Unit, Copenhagen, Denmark
(2) Danish Medicines Agency, Copenhagen, Denmark
(3) Bispebjerg University Hospital, Department of Clinical Pharmacological, Copenhagen, Denmark
Publication of clinical drug trials (CDTs) is an important way to disseminate new knowledge. However, not all CDTs are published. Based on
approved CDT applications, a systematical search in PubMed demonstrated a publication rate of 30% among Danish investigator-initiated
CDTs (Berendt. BMJ 2008; 336 : 33). In order to verify this, we conducted a similar search in which the findings - negative as well as positive – were verified by the investigators. We included all approved CDT
applications submitted to the Danish Medicines Agency in 1999, 2001
and 2003 describing an investigator-initiated trial. Corresponding publications were identified by systematically searching PubMed May-Sept
2009. Only limited data such as investigator, protocol title, drug names
and sometimes a brief description of the study were available. Each submitting investigator was asked to confirm the findings (trial published/
not published, correct publications). A total of 245 eligible approved trial
applications were identified. Response was obtained from 174 investigators. Of these, PubMed search resulted in 84 published vs. 90 non-published trials, whereas response from investigators resulted in 94
published vs. 80 non-published trials (j statistic 0.771, good agreement).
The overall publication rate based on PubMed search was 46% (112/
245). Dividing trials in +/) response from investigators, the publication
rates were 48% and 39%, respectively. Average number of publications
per trial was 1.8 (median: 5, range: 1-9). Combination of PubMed search
with verification by investigators resulted in a publication rate higher
than PubMed search alone. Reasons are discussed.
Paper No.: 1923
FOCUSED CONFERENCE GROUP: P08 - DEVELOPMENTS IN
THE TREATMENT OF SEXUAL DYSFUNCTION AND
DISEASES OF THE LOWER URINARY TRACT
PREVENTION OF URINARY TRACT INFECTIONS IN
NORWEGIAN NURSING HOMES
Jenny Bergman(1), H Salvesen Blix(2), J Schjøtt(3,4)
(1) Haukeland University Hospital, Regional Drug Information Centre,
Bergen, Norway
(2) Norwegian Institute of Public Health, Department of Pharmacoepidemiology, Oslo, Norway
(3) Haukeland University Hospital, Section of Clinical Pharmacology,
Laboratory of Clinical Biochemistry, Bergen, Norway
(4) University of Bergen, Institute of Internal Medicine, Section for Pharmacology, Bergen, Norway
Urinary tract infections (UTIs) are the most common infections in nursing homes and prevention may reduce antibiotic use and patient suffering. The spectre of agents used in preventing UTIs in nursing homes is
scarcely documented. The aim of this study was to describe all agents
used to prevent UTIs in Norwegian nursing homes. We carried out a
one-day point-prevalence study in 44 Norwegian nursing homes during
May-April 2006, with registration of all agents prescribed as UTI prophylaxis. The results included 1473 residents. 18% (n = 269) of the residents
had at least one agent recorded as prophylaxis of UTI, varying between
0-50% among the nursing homes. Methenamine was used by 8.8% of all
residents, vitamin C by 5.9%, vaginal and systemic estrogens by 5.5%
and cranberry products by 1.9%. Trimethoprim and nitrofurantoin were
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
198
used as prophylaxis by 0.9% and 0.7% respectively. The dosage of
vitamin C varied between 60-2000mg a day, cranberry products were
given as stated by the manufacturer and the dosages of other drugs were
within the recommended doses stated in the summery of product characteristics. In conclusion the use of UTI prophylaxis is common but
variable in Norwegian nursing homes. Interestingly, methenamine and
vitamin C, which lack documented effectiveness, are the most frequently
used agents. In contrast, trimethoprim, nitrofurantoin and local vaginal
estrogens have been shown to be effective, but are less frequently used.
We conclude that prescribing habits for effective UTI prophylaxis are
controversial in Norwegian nursing homes.
Paper No.: 1449
FOCUSED CONFERENCE GROUP: P19 - GENERAL
SESSION - ONCOLOGY
IMPACT OF ABCB1 VARIANTS ON NEUTROPHIL
DEPRESSION: A PROSPECTIVE STUDY
Troels K Bergmann(1), C Brasch-Andersen(0), H Gréen(2), M Mirza(3),
K Skougaard(4), J Wihl(5), N Keldsen(6), P Damkier(3), C Peterson(2),
W Vach(7), K Brøsen(1)
(1) University of Southern Denmark, Department of Clinical Pharmacology, Odense, Denmark
(2) Linköping University, Sweden
(3) Odense University Hospital, Denmark
(4) Herlev Hospital, Denmark
(5) Lund Hospital, Sweden
(6) Herning Hospital, Denmark
(7) University of Freiburg, Germany
The standard treatment for ovarian cancer in advanced stages is surgery
followed by taxane-platin therapy. Despite an initial high response rate
most patients eventually relapse. The dose limiting toxicities of paclitaxel
are neutropenia and neuropathy but the inter-individual variability is
large. The purpose of this study was to prospectively investigate the
impact of genetic variants in key drug metabolizing/transporter genes on
paclitaxel toxicity. CYP2C8*3 and three ABCB1 variants were picked
for primary analysis and a host of other candidate genes were assessed in
92 prospectively recruited Scandinavian Caucasian patients with primary
ovarian cancer who were treated with paclitaxel (175 mg/m2) and carboplatin (AUC 5-6) after cytoreductive surgery. Clinical toxicity using
Common Toxicity Criteria for Adverse Events was assessed by a single
investigator for 97% of the patients and hematologic toxicity was registered. Patients carrying one or two variant alleles of ABCB1 C3435T
had progressively more pronounced neutrophil decrease at nadir (P-value
0.03). The same association was found for ABCB1 C1236T and
G2677T/A with P-values of 0.06 and 0.02. No statistically significant
correlations were found for paclitaxel compliance and sensoric neuropathy. In conclusion: variants in the drug transporter ABCB1 gene impact
on the neutrophil suppressing effect of paclitaxel in patients with ovarian
cancer. To our knowledge this has never been shown before directly in a
prospective study; but a similar result was reported in 18 patients by Sissung et al.(Eur J Cancer 2006;42:2893-6). This novelty has implications
for the understanding of myelosuppression in particular and for tailored
chemotherapy in general.
(1) Karolinska Institute, Neonatal Research Unit Q2:07, Woman and
Child Health, Stockholm, Sweden
(2) Örebro University, School of Health and Medical Sciences, Örebro,
Sweden
(3) University of Tennessee, Department of Pediatrics, Memphis, TN,
USA
(4) University of Leicester, Department of Health Sciences, Leicester,
UK
(5) Hospital for Children and Adolescents, Helsinki, Finland
(6) University of Edinburgh, Royal Infirmary of Edinburgh, Neonatal
Unit, Edinburgh, UK
(7) University Hospital Antwerp, Antwerp, Belgium
(8) Friedrich-Alexander Universitdt Erlangen-Nürnberg, Erlangen,
Germany
(9) Uppsala University, Uppsala, Sweden
(10) Lund University, Lund, Sweden
(11) Erasmus universitair Medish Centrum, Rotterdam, The Netherlands
(12) European Paediatric Medicine Company, Brussel, Belgium
(13) Karolinska Institutet, Clinical Pharmacology, Stockholm, Sweden
(14) Hôpital Trousseau, Paris, France
Alleviation of pain is a basic and human right regardless of age. Awareness that newborn infants experience pain during surgery, mechanical
ventilation and invasive procedures has increased over the last 20 years.
Despite the accumulating research and various international and national
guidelines for neonatal pain management, there is no recent information
about how this knowledge translates into clinical practice. The NeoOpioid Consortium is a research network with 12 partners supported by
European Commission’s Framework Programme 7, ‘‘Better Use of Medicines’’. That aims to improve the safety and efficacy of opioid treatment
in critically ill newborns. The EUROPAIN survey, part of the NeoOpioid-project, designed to collect data of neonatal pain management in different countries. Neonatal units in all European countries will be invited
to participate. For ventilated infants, data will be collected about administration of analgesic and sedative medications during a 2-week period.
Documented and reported strategies of pain relief measures will be compared with local and national guidelines. Computerized tutoring material
and data entry systems will facilitate high quality data collection. EUROPain survey will provide units with unique opportunities to compare their
analgesic practices with other units nationally and European basis.
Results will provide a scientific foundation for the development of evidence-based European neonatal pain management guidelines, estimate
the eligible neonatal populations for randomized trials, explore correlations between the burden of neonatal pain and long-term cognitive outcomes, and stimulate drug development specifically targeted for
analgesia in children. Research funding from the European Community’s
Seventh Framework Programme, grant agreement no. 223767.
Paper No.: 1672
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
ANTI-INFLAMMATORY AND ANTIOXIDANT ACTIVITIES OF
AN INDIAN MEDICINAL PLANT, EMBLICA OFFICINALIS, IN
RODENT MODELS OF INFLAMMATION
Jagriti Bhatia, M Golechha, DS Arya, F Tabassum
All India Institute of Medical Sciences, Department of Pharmacology,
New Delhi, India
Paper No.: 2081
FOCUSED CONFERENCE GROUP: P10 - DRUGS FOR HALF
THE WORLD: PAEDIATRIC CLINICAL PHARMACOLOGY
EUROPAIN – EUROPEAN PAIN AUDIT IN
NEONATES – NEOOPIOID CONSORTIUM
Lena Bergqvist(1), M Eriksson(2), KJS Anand(3), EM Boyle(4),
H Lagercrantz(1), S Andersson(5), G Menon(6), B van Overmeire(7),
M Keller(8), L Hellström-Westas(3) V Fellman(4) RHN van Schaik(11),
J-P Osselaere(12), A Rane(13), J Schollin(2), R Carbajal(14)
Introduction: Emblica officinalis has been reported to possess high antioxidant activity. However, the anti-inflammatory activity of its fruits has
not been studied earlier. Hence, in the present study the anti-inflammatory activity of the standardized hydroalcoholic extract of the fruits of
Emblica officinalis (HAEEO) in acute and chronic models of inflammation was evaluated. Methods: The acute inflammation in rats was induced
by subplantar injection of carrageenan, histamine, serotonin and prostaglandin E2 respectively. Further, the histopathological examination and
assessment of the biochemical markers of oxidative stress was done in
the carrageenan group. Cotton pellet granuloma model was used to
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
199
evaluate the effect of the HAEEO on chronic inflammation. Results: In
the acute model of inflammation it was observed that the administration
of HAEEO (300, 500, and 700 mg/kg, i.p.) significantly and dose-dependently inhibited paw edema induced by all the phlogistic agents. The
HAEEO also significantly increased glutathione, superoxide dismutase
and catalase activity and reduced the levels of malondialdehyde. Also,
HAEEO at the dose of 500 and 700 mg/kg produced a significant
decrease in the amount of cellular infiltrate and subcutaneous edema
induced by carrageenan. In the cotton pellet granuloma model HAEEO
at all the doses tested significantly (P < 0.001) reduced the granuloma
formation. Conclusion: The results obtained indicate the HAEEO
possesses significant anti-inflammatory activity in all the models. Thus,
HAEEO may hold therapeutic promise in the management of inflammatory conditions.
Paper No.: 1673
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
STUDY OF THE ANTIHYPERTENSIVE POTENTIAL OF
INDIAN MEDICINAL PLANT, EMBLICA OFFICINALIS, IN A
RAT MODEL OF HYPERTENSION
Jagriti Bhatia(1), F Tabassum(1), DS Arya(1), S Joshi(1), AK Srivastava(2)
We have previously shown that serum estradiol concentrations are
halved in smoking as compared to non-smoking postmenopausal
women during estrogen-progestogen therapy. These studies included
unselected smoking women. We now wished to evaluate if light smoking influences estradiol kinetics. In addition, we aimed to study acute
effects of tablet intake in smokers versus non-smokers. We used a posthoc strategy in two double-blind randomized trials of estrogen-progestogen treatments in postmenopausal women. Study I: In a 2-year study of
129 women (94 completed) who received either 1mg estradiol continuously combined with 125lg trimegestone or placebo, we compared
smokers (18) to non-smokers (76). Smoking more than 10 cigarettes
per day precluded participation. Serum estradiol and estrone were measured at 0, 3, 6, 12, 18 and 24 months. Multiple linear regression (fixed
effects model) showed that smoking women had significantly lower
serum estradiol (p = 0.0041) and estrone (p = 0.0093) concentrations
compared to non-smokers. Study II: In a subset from a study comparing
combinations of gestodene and 1 or 2mg estradiol in 278 postmenopausal women (153 completed) unselected for smoking status, we studied 2-hour serum estradiol and estrone at 1, 6 and 12 months. We
found that smoking reduced hormone concentrations on both regimens,
for example at 12 months, serum estrone was 157 + /-18 vs 98 + /17 pg/ml in non-smoking versus smoking women at 2 hours (p = 0.04).
In conclusion, we have shown that both heavy and light smoking
reduces serum estradiol and estrone in postmenopausal women undergoing estrogen-progestogen therapy. The effect is seen already within
2 hours after tablet intake.
(1) All India Institute of Medical Sciences, Department of Pharmacology,
New Delhi, India
(2) All India Institute of Medical Sciences, Department of Laboratory
Medicine, New Delhi, India
Introduction: Emblica officinalis (EO) has been reported to possess good
antidiabetic, hypolipidemic, cardioprotective and antioxidant activity.
However, the potential of EO in preventing the development of hypertension has not been studied so far. Hence, the present study was designed
to evaluate whether EO has antihypertensive potential in the DOCA-salt
induced model of hypertension in rats. Methods: The DOCA-salt
(20 mg/kg, s.c.) twice weekly, along with 1% NaCl solution substituted
for drinking water, was administered for 5 weeks to induce hypertension
in rats. EO (75, 150 and 300 mg/kg, orally) was concurrently administered with DOCA-salt in rats for 5 weeks. The effect of EO on haemodynamic and biochemical parameters as well as serum nitric oxide and
serum electrolyte (sodium and potassium) levels were determined. The
blood pressure was recorded non-invasively by the tail-cuff method.
Results: EO significantly and dose-dependently prevented the rise in systolic, diastolic and mean arterial blood pressure and also the heart rate as
compared to the DOCA-salt group. A significant reduction in malondialdehyde and increased content of glutathione and superoxide dismutase
were observed in the rat heart. Furthermore, EO maintained serum nitric
oxide, sodium and potassium levels as compared to DOCA-salt group.
The maximal significant effect on all the study parameters was observed
at the dose of 300 mg/kg. Conclusion: EO prevents the development of
hypertension in the DOCA-salt model. This beneficial effect may be due
to its antioxidant property.
Paper No.: 1010
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING BENEFITS AND MINIMIZING HARMS FROM DRUGS
BOTH HEAVY AND LIGHT SMOKING REDUCES SERUM
ESTRADIOL AND ESTRONE IN POSTMENOPAUSAL WOMEN
DURING ESTROGEN-PROGESTOGEN THERAPY
Nina Hannover Bjarnason(1), H Jorgensen(2), C Christiansen(3)
(1) Institute for Rational Pharmacotherapy, Danish Medicines Agency,
Copenhagen, Denmark
(2) Department of Clinical Biochemistry, Bispebjerg Hospital, Copenhagen, Denmark
(3) Center for Clinical & Basic Research, Ballerup, Denmark
Paper No.: 3245
FOCUSED CONFERENCE GROUP: P04 - PHARMACOEPIDEMIOLOGY, CURRENT CONTROVERSIES AND OPPORTUNITIES
CONSUMPTION OF DRUGS AGAINST ADHD IN DENMARK
MORE THAN TEN DOUBLED IN TEN YEARS
Nina Hannover Bjarnason(1), H Jorgensen(2), C Christiansen(3)
(1) Institute for Rational Pharmacotherapy, Danish Medicines Agency,
Copenhagen, Denmark
(2) Department of Clinical Biochemistry, Bispebjerg Hospital, Copenhagen, Denmark
(3) Center for Clinical & Basic Research, Ballerup, Denmark
We have previously shown that serum estradiol concentrations are
halved in smoking as compared to non-smoking postmenopausal
women during estrogen-progestogen therapy. These studies included
unselected smoking women. We now wished to evaluate if light smoking influences estradiol kinetics. In addition, we aimed to study acute
effects of tablet intake in smokers versus non-smokers. We used a posthoc strategy in two double-blind randomized trials of estrogen-progestogen treatments in postmenopausal women. Study I: In a 2-year study of
129 women (94 completed) who received either 1mg estradiol continuously combined with 125lg trimegestone or placebo, we compared
smokers (18) to non-smokers (76). Smoking more than 10 cigarettes
per day precluded participation. Serum estradiol and estrone were measured at 0, 3, 6, 12, 18 and 24 months. Multiple linear regression (fixed
effects model) showed that smoking women had significantly lower
serum estradiol (p = 0.0041) and estrone (p = 0.0093) concentrations
compared to non-smokers. Study II: In a subset from a study comparing
combinations of gestodene and 1 or 2mg estradiol in 278 postmenopausal women (153 completed) unselected for smoking status, we studied 2-hour serum estradiol and estrone at 1, 6 and 12 months. We
found that smoking reduced hormone concentrations on both regimens,
for example at 12 months, serum estrone was 157 + /-18 vs 98 + /17 pg/ml in non-smoking versus smoking women at 2 hours (p = 0.04).
In conclusion, we have shown that both heavy and light smoking
reduces serum estradiol and estrone in postmenopausal women undergoing estrogen-progestogen therapy. The effect is seen already within
2 hours after tablet intake.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
200
Paper No.: 2811
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
REEP FAMILY MEMBERS MODULATE A2C-ADRENOCEPTOR
EXPRESSION BY AN ER INTERACTION
Susann Björk(1,2), C Hurt(3), B Kobilka(2), T Angelotti(3)
(1) University of Turku, Department of Pharmacology, Drug Development and Therapeutics, Turku, Finland
(2) Stanford University, Department of Molecular and Cellular Physiology and Medicine, Stanford School of Medicine, Palo Alto, CA, USA
(3) Stanford University, Department of Anesthesia, Stanford University
Medical Center, Palo Alto, CA, USA
As regulators of sympathetic neurotransmitter release, a2A- and a2Cadrenoceptors (ARs) exhibit differential neuronal localization. In nonneuronal cell lines, a2A-ARs target to the plasma membrane (PM) as
opposed to a2C-ARs; but in neuronal cells, also a2C-ARs target to the
PM. We investigated whether accessory Receptor Expression Enhancing
Proteins (REEPs) affect subcellular localization to modulate a2A- and
a2C-AR expression. RT-PCR analysis of cell lysates revealed REEP1,
REEP2 and REEP6 mRNA expression in neuronal but not non-neuronal
cells. A developmental pattern of REEP expression was seen in cultured
sympathetic ganglion neurons. Immunocytochemical staining of surface
and total expression in HEK293 kidney cells revealed co-localization of
a2C-ARs with transfected REEPs in the endoplasmic reticulum (ER).
Transfected REEPs appeared to enhance cell surface expression, but
because of an increase in total a2C-AR expression and not by a selective
increase in PM trafficking (shown by FACS). A direct interaction of
REEPs and receptors was suggested as HA-tagged a2C-ARs
co-expressed with either Flag-REEP1, Flag-REEP2 or Flag-REEP6 were
co-immunoprecipitated with an anti-Flag antibody. Sucrose gradient analysis of REEP expression demonstrated ER fraction localization, without
expression in the Golgi or PM fractions. Finally, glycosylation analysis
revealed that the increase in a2C-AR surface expression was not associated with an increase in mature glycosylation. Thus, enhanced cell
surface expression of a2C-ARs in neuronal cells may be due to effects of
REEP proteins. However, REEP co-expression does not enhance trafficking to the PM. Instead, REEPs and receptors interact in the ER, REEPs
possibly acting as chaperones.
Paper No.: 2720
FOCUSED CONFERENCE GROUP: P19 - GENERAL SESSION HEPATOLOGY
LOVASTATIN PREVENTS CARCINOGENESIS IN A RAT
MODEL FOR LIVER CANCER. EFFECTS OF UBIQUINONE
SUPPLEMENTATION
Linda Björkhem-Bergman(1), J Acimovic(2), U-B Torndal(3), P Parini(4),
L Eriksson(3)
(1) Karolinska Institute & Karolinska University Hospital Huddinge,
Division of Clinical Pharmacology, Department of Laboratory Medicine,
Stockholm, Sweden
(2) University of Ljubljana Faculty of Medicine, Institute of Biochemistry, Ljubljana, Slovenia
(3) Karolinska Institute & Karolinska University Hospital Huddinge,
Division of Clinical Pathology, Department of Laboratory Medicine,
Stockholm, Sweden
(4) Karolinska Institute & Karolinska University Hospital Huddinge,
Division of Clinical Chemistry, Department of Laboratory Medicine,
Stockholm, Sweden
Statins (HMGCoA reductaste inhibitors) inhibit the mevalonate pathway and efficiently reduce cholesterol synthesis. Several studies have
shown that statins may inhibit carcinogenesis although the mechanisms
are not fully understood. Ubiquinone, an antioxidant synthesized in the
mevalonate pathway, is known to be important for cell proliferation
and is increased in preneoplastic liver tissue. The synthesis of ubiquinone is also inhibited by statins and we tested the hypothesis that a
statin-induced effect on carcinogenesis may be mediated by the inhibition of ubiquinone synthesis. We studied the effects of lovastatin treatment given orally with and without addition of ubiquinone in a rat
model for chemically induced hepatocarcinogenesis. The volume fraction of preneoplastic liver tissue was determined, the proliferation of
the cells within the liver tissue was measured and different components
of the mevalonate pathway were monitored. Lovastatin treatment
reduced the volume fraction of liver nodules by about 50% and the
proliferation of the cells within the liver nodules was reduced to one
third. Ubiquinone treatment reversed the statin-induced inhibition of
cell proliferation but did not affect the volume fraction of the liver
nodules. The lathosterol levels were significantly reduced in the statin
treated rats, indicating inhibition of the mevalonate pathway. In spite
of this, the cholesterol levels were not affected. In conclusion, lovastatin prevents carcinogenesis in a rat model for liver cancer, despite
no change in cholesterol levels. The statin-induced inhibition of cell
proliferation might, at least in part, be explained by the inhibition of
ubiquinone synthesis.
Paper No.: 2721
FOCUSED CONFERENCE GROUP:
P04 - PHARMACOEPIDEMIOLOGY, CURRENT
CONTROVERSIES AND OPPORTUNITIES
STATIN TREATMENT REDUCES MORTALITY IN SEVERE
INFECTIONS – A SYSTEMATIC REVIEW AND
META-ANALYSIS
Linda Björkhem-Bergman(1), P Bergman(2), J Andersson(3), J Lindh(1)
(1) Karolinska Institute & Karolinska University Hospital Huddinge,
Division of Clinical Pharmacology, Department of Laboratory Medicine,
Stockholm, Sweden
(2) Karolinska Institute & Karolinska University Hospital Huddinge,
Division of Clinical Microbiology, Department of Laboratory Medicine,
Stockholm, Sweden
(3) Karolinska Institute & Karolinska University Hospital Huddinge,
Division of Infectious Diseases, Department of Medicine, Stockholm,
Sweden
Several studies have reported improved survival in severe bacterial
infections among statin treated patients. In addition, statins have been
ascribed beneficial anti-inflammatory effects. The aim of this study was
to evaluate the effect of statin-treatment on mortality in patients with
severe bacterial infections, by means of a systematic review and a
meta-analysis. Studies investigating the association between statin use
and mortality in patients with severe bacterial disease were identified in
a systematic literature review and a meta-analysis was performed to
calculate the overall OR of mortality in statin users. The literature
search identified 552 citations from which 35 relevant studies were
extracted. In all, 17 studies comprising 114 572 patients were included
in the final analysis. Statin use was associated with a significantly
(p < 0.0001) reduced mortality in patients suffering from severe bacterial infections (OR 0.53, 95% CI 0.43-0.66). However, all studies
included were of observational design and funnel plot analyses indicated a possible influence of publication bias (Egger’s bias test
p < 0.05). When a precision estimate test was used to adjust for publication bias, the overall OR was reduced to 0.76 (95% c.i. 0.58-1.01).
In conclusion, patients that suffer from severe bacterial infections have
a better outcome if they are on current statin treatment, but the magnitude of this proposed statin effect may have been inflated by publication bias. Nevertheless, our data indicate that statin treatment safely can
be continued during severe infections. There is a need for randomised
controlled trials investigating the possible beneficial effect of statintreatment in bacterial infections.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
201
Paper No.: 1512
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
ORAL SILDENAFIL INCREASES SODIUM NITROPRUSSIDE
IONTOPHORESIS INDUCED SKIN HYPERAEMIA IN
HEALTHY VOLUNTEERS
S Blaise, M Hellmann, M Roustit, S Isnard, Jean-Luc Cracowski
Grenoble University Hospital, Inserm CIC3, Grenoble Clinical Research
Center, Grenoble, France
Sildenafil, a specific PDE5A inhibitor, is currently tested as a treatment
for severe Raynaud’s phenomenon. The main objective was to test
whether sildenafil, alone or associated with local sodium nitroprusside
(SNP) delivered through skin iontophoresis, increases forearm cutaneous
blood conductance in healthy volunteers. Ten healthy volunteers were
enrolled. Variations in cutaneous vascular conductance following oral
administration of 50 mg or 100 mg of sildenafil associated or not with
sodium nitroprusside iontophoresis were expressed as a percentage of
maximal cutaneous vascular conductance and were monitored using laser
Doppler imaging. SNP iontophoresis was performed on the ventral face
of the forearm, one hour after application of lidocaine/prilocaine cream.
Results. Sildenafil at 100 mg, but not 50 mg, increased SNP iontophoresis area under the curve (+ 44%, P = 0.03 versus without sildenafil) and
increased peak SNP iontophoresis (+29%, P = 0.05). Sildenafil at
100 mg, but not 50 mg, increased baseline cutaneous vascular conductance (+ 75%, P = 0.03). Incidence of headache was not different when
SNP iontophoresis was associated with sildenafil. One symptomatic arterial hypotension occurred in one volunteer with 50 mg sildenafil. Conclusions. Oral sildenafil at 100mg potentiates local skin SNP
iontophoresis-induced hyperaemia, with no increased incidence of headaches. The association of oral specific PDE5A inhibitor and nitrates
administered through skin iontophoresis should be further investigated in
diseases such as severe Raynaud’s phenomenon, with particular attention
to the incidence of arterial hypotension.
Clinicaltrials.gov (NCT00710099)
Paper No.: 2798
FOCUSED CONFERENCE GROUP: P10 - DRUGS FOR HALF
THE WORLD: PAEDIATRIC CLINICAL PHARMACOLOGY
DRUGS USE IN PAEDIATRIC POPULATION. EVALUATION IN
TWO AREAS: NEONATOLOGY AND INTENSIVE CARE UNITS
neonatology unit the most frequent reason for the classification of UL
was the use of preparations manufactured (home-label medications) or
modified by the hospital pharmacy (caffeine, dexamethasone or spironolactone). Conclusion: This study shows a high rate of UL and OL drug
use in pediatrics. Regulatory authorities and the pharmaceutical industry
could be the focus for improving the availability of appropriate authorized medicines for children.
Paper No.: 2799
FOCUSED CONFERENCE GROUP:
P04 - PHARMACOEPIDEMIOLOGY, CURRENT
CONTROVERSIES AND OPPORTUNITIES
OFF-LABEL VS COMPASSIONATE DRUG USE: LEVEL OF
EVIDENCE FOR APPLICATIONS
Encarnación Blanco(1), A Muñoz(2), I Bellido(1), M Salazar(2),
EI Márquez(1)
(1) University of Málaga School of Medicine, Department of Pharmacology and Clinical Therapeutics, Málaga, Spain
(2) Virgen de las Nieves General Hospital of Granada, Spain
Introduction: Some times compassionate drug use can also very confusing: permission to treat a single patient with an unapproved drug outside
of a trial versus the off label utilization. Methods: A cross-sectional study
was undertaken for the compassionate use applications in a general hospital of Granada (Spain). Results: The most commonly prescribed classes
of drug were (% of drugs): L group (50%), B group (13.1%), J group
(8%), and H group (8%), meanwhile B group dominated the number of
applications (45.5%) followed by L group (36.8%). The reasons for compassionate use requests were: access to an unapproved drug or real compassionate use (6 new drugs, 18% of medicinal products) and the
utilization of un authorised medicinal product for an indication, population or route of administration different from the one mentioned in the
SmPC or off label use (82% of drugs). The 46% of petitions was based
on somewhere randomized clinical trial meanwhile the rest had a low
level of evidence (open studies, case reports). Finally, after two years,
half the investigational new drugs are authorized (bevacizumab, trabectedine, tipranavir), while only the 21% of different indications and the
8.3% of different use forms has been approved. There are ten compassionate use petitions based on a high level of clinical evidence without
SmPC variations. Conclusions: Though the drugs selection was rational
in most of the cases, some incongruities were observed. In order to
favour a common approach, better treatment availability and more
dynamic processing, local legal basis has been recently revised.
Encarnación Blanco(1), I Bellido(1), EI Márquez(1), A Medina(2),
JA González-Correa(1)
(1) University of Málaga School of Medicine, Department of Pharmacology and Clinical Therapeutics, Málaga, Spain
(2) San Cecilio Clinical Hospital of Granada, Spain
Introduction: Paediatric insufficient drug evaluation and the lack of
adapted pharmaceutical formulations explain the importance of unlicensed (UL) and off label (OL) prescriptions. Moreover, the quality of
information available on formulation and stability is limited, thus preparation of children’s oral medicines is subjected to much variation
between hospitals. Aim: To determine the extent of use of drugs that are
either UL or OL. Patients and Methods: A cross-sectional study including 143 children hospitalised (intensive care unit: 81, neonatology unit:
62). Results: We analysed 601 prescriptions of 91 different drugs. In
intensive care a 22.3% of children received at least one UL prescription:
half not licensed for paediatrics (amiodarone), and half by modification
to a licensed drug (nifedipine). A 87.7% children received at least one
OL prescription; the most common categorie was outside dose recommendations (dexamethasone), followed by used for non-licensed indication (alimemazine for irritability), outside licensed age range
(voriconazole for < 2 years old), and used by unlicensed route. Of the
601 prescriptions a 35% were for drugs without SPC, a 28% for drugs
prescribed in agreement with the SPC, and a 37% were UL/OL. In
Paper No.: 3150
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD DISEASES
AGE-DEPENDENT EFFECTS OF DEXAMETHASONE ON
HYPOXIC-ISCHEMIC BRAIN EDEMA
Gábor Blazsó(1), M Pásztor(2), A Balogh(1), A Márki(1), G Dombi(2),
G Falkay(1)
(1) University of Szeged, Department of Pharmacodynamics and Biopharmacy, Szeged, Hungary
(2) University of Szeged, Department of Pharmaceutical Analysis, Szeged, Hungary
Objective: Therapeutic value of glucocorticoids in the prevention of hypoxic-ischemic brain edema is still a subject of debate. The aim of the
present study was to examine whether the neuroprotective effect of dexamethasone depends on the degree of maturity of the brain. Method: 1-,
2-, 4- and 12-week-old rats were subjected to unilateral carotid artery
occlusion plus hypoxia in 8% oxygen. The effects of dexamethasone, the
antiglucocorticoid mifepristone and their combination on the cytotoxic
and vasogenic phases of hypoxic-ischemic brain edema were measured
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
202
in neonate and adult rats. The effects of flunarisine and verapamil on the
survival of the animals was also examined alone or in combination with
dexamethason. All the drugs were administered prior to hypoxia-ischemia. Results: Dexamethasone inhibited both the cytotoxic and the vasogenic phases; this inhibition could be antagonized with mifepristone in
neonate rats. Opposite results were observed in 12-week-old animals,
where the glucocorticoid increase of these phases could not be prevented
with mifepristone. In adult rats, dexamethasone increased the Ca2 + content of the hypoxic-ischemic brain hemisphere and the death of the animals. This toxic effect could be antagonized with flunarisine and
verapamil. Tritiated dexamethasone was able to penetrate into the hemispheres of 1- or 2-week-old rats, but significantly less into the brain of 4or 12-week-old animals. This effect lasts until postnatal day 24, when
the brain blood barrier is fully developed. Conclusion: The present findings support the hypothesis that there is a strong dependence on age corresponding to dexamethasone’s penetration into the brain.
Paper No.: 3105
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
CRIP1A REGULATES CB1 CANNABINOID RECEPTOR
INTERNALIZATION AND SIGNAL TRANSDUCTION
Lawrence C Blume(1), CC Bass(1), GD Dalton(1), DE Selley(2), A
Howlett(1)
(1) Wake Forest University Health Sciences, Department of Physiology
and Pharmacology, Winston-Salem, USA
(2) Virginia Commonwealth University, Richmond, USA
CRIP1a (Cannabinoid Receptor Interacting Protein 1a) is an accessory
protein for the CB1 cannabinoid receptor, whose function is as yet
unclear (Neihaus JL et al.,Mol Pharmacol 2008;72:1557-1566). We prepared clones of the N18TG2 neuroblastoma cell that stably overexpress
CRIP1a at 3- to 5-fold greater mRNA levels than WT cells. In CRIP1aoverexpressing clones, CB1 receptor density on the plasma membrane is
modestly reduced compared with WT cells, when examined using immunostaining for surface receptors using an N-terminally-directed CB1 antibody with quantitative detection using the LI-COR Odyssey. In response
to the agonist (WIN55212-2)-stimulation, internalization of CB1 receptors occurs in WT cells, but not in CRIP1a-overexpressing cells. Upon
continued exposure to agonist, cell surface CB1 receptor density was
recovered within minutes in the WT perhaps by externalization, a process
that is also observed in CRIP1a-overexpressing clones. Upon exposure
to the CB1 antagonist rimonabant, the density of cell-surface CB1 receptors increased in both WT as well as CRIP1a-overexpressing cells. Under
conditions in which 2-AG synthesis is blocked, CB1-mediated ERK
phosphorylation in WT N18TG2 cells could be reduced by rimonabant,
consistent with ‘‘constitutive activation’’ of signal transduction. In the
CRIP1a-overexpressing clones, basal pERK levels were low compared
with WT, and rimonabant had no effect, suggesting the absence of the
constitutive activation. Agonist-stimulated ERK phosphorylation
occurred to the same extent in both WT and CRIP1a-overexpressing
clones. These findings suggest that the function of CRIP1a may be to
regulate internalization-associated signal transduction events.
Supported by R21-DA025321, R01-DA03690, K01-DA024763, T32DA007246, K01-DA24763
Paper No.: 667
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
VASCULAR AT1/AT2 RECEPTOR RATIO IMPACT ON
VASOCONSTRICTION IN EXPERIMENTAL GESTATIONAL
DIABETES
Pregnancy and diabetes mellitus are conditions that separately modify
the vascular response to Angiotensin II(Ang II) by changing AT1/AT2
receptor (R) ratio of expression. We reach to establish the impact of
gestational diabetes (GDM) on AT1/AT2 R and Ang II induced vasoconstriction. Using a conventional isolated-organ system, concentration
response curves to Ang II were constructed using thoracic aortas with
(e+) and without (e-) endothelium from term pregnant (day 20) and
non pregnant rats. Animals received streptozotocin 60 mg/kg ip, an
hypercaloric diet or none. Both pregnancy and experimental diabetes
reduced Ang II vasoconstriction mainly on e+ vessels. The response
was significantly increased in bboth models of GDM. Losartan
(10-8M) reduced the response to Ang II of the gestational diabetes
e- group, but did not affect e+ aorta rings. AT1R expression increased
in the GDM group. In contrast, increased levels of AT2R were found
in the pregnant and in the diabetic groups, while no change was
observed in the GDM group. These results suggest that pregnancy
and short term exposure to diabetes increase endothelium derived
vasodilators release probably through AT2R, which effects are masked
by an increased AT1R/AT2R ratio when the two conditions are present. This imbalance becomes evident mainly in endothelium intact
vessels.
Paper No.: 2919
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
THE DIFFERENCES OF ANTIULCER ACTIVITY OF PROTON
PUMP INHIBITOR (PPI) AND H2-ANTAGONIST IN DIABETIC
RATS
Teresa Bobkiewicz-Kozlowska(1), R Forjasz(1), M Nowaczyk(0)
(1) Poznan University of Medical Sciences, Department of Pharmacology, Poznan, Poland
(2) Poznan University of Medical Sciences, Department of Pharmaceutical Technology, Poznan, Poland
Diabetic patients develop a variety of gastrointestinal symptoms
(abdominal pain, vomiting, diarrhea, constipation, delayed gastric emptying). Little attention has been paid to the incidence and healing rate
of peptic ulcer in diabetics, however recent studies indicate that ulcers
are more severe and often associated with complications such as gastrointestinal bleeding. Previous studies documented that streptozotocin
(STZ)-induced diabetes in rats is an accepted model of insulin-dependent diabetes. Experiments were performed on rats with experimental
diabetes. Acute gastric stress ulcers were developed using waterimmersion restraint stress. Ranitidine or pantoprazol (40 mg/kg i.g. or
3 mg/kg i.g., respectively) were administered 30 min before stress.
Severity of gastric mucosa injury was revealed as an ulcer index (UI).
Chronic gastric ulcers were induced using Okabe and Pfeiffer method
with the Konturek modification. Starting on the ulcer induction day,
ranitidine (40 mg/kg i.g., twice a day) or pantoprazol (3 mg/kg i.g.,
twice a day) were administered for 7 days. Tissue concentration of
cytokines TNF-a and IL-1b in gastric mucosa was measured using
ELISA. We concluded that: 1. gastric mucosa is more sensitive to
ulcerogenic stress, but chronic ulcer healing is not impaired in experimental diabetes, however the increase of proinflammatory cytokines
tissue level is observed. 2. H2-antagonist and PPI (H2-antagonist
>PPI) protect the gastric mucosa against acute stress ulcer but do not
influence on self-healing chronic ulcers in diabetic rats. 3. in gastroprotective effect of H2-antagonist lowering TNF-a and IL-1b tissue
concentration can be involved.
Rosa A Bobadilla, C Toufiño, E Fernandez-Vallin, P Lopez
ESM, Instituto Politecnico Nacional, Mexico City, Mexico
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
203
Paper No.: 730
FOCUSED CONFERENCE GROUP: P06 - THE HEART GONE
WRONG; STABILIZATION OF CARDIAC FUNCTION
RATIONAL DRUG CORRECTION OF THE SYSTEMIC
INFLAMMATORY RESPONSE SYNDROME IN PATIENTS
WITH CONGESTIVE HEART FAILURE
Tamar T Bochorishvili, MA Rogava, ET Tsiwtsiwadze
N. Kipshidze National Centre of Therapy, Department of Cardiomypathy,
Tblisi, Georgia
Introduction: The SIRS which accompained severe form of CHF are
associated with activation of a number of complex and interrelated pathways that include the endotoxemia, tissue hyperoxia and activity of the
intrinsic defense system. The purpose of the present study was to test the
hypothesis that inotropic drug with pronounced antihypoxic/antiischemic
and antioxidant properties, adenocin, would significantly reduce inflammatory signalling and cardiac dysfunction. Materials/Patients: Plasma
levels of cytokines (interleukine-6, interleukine -8, TNF-a and soluble
adhesion molecules, redox potential NAD/NADH, superoxide anion production, activities of superoxide dismutase and catalase were determined
in samples obtained from 78 patients with CHF NYHA classes II-IV,
caused by ischemic heart disease in 65 and dilated cardiomyopathy in 13
patients. All patients including in the study were randomized into 2
groups, control received standard therapy and the main additionally
received adenocin. Results: All biological markers of the intensity of
endotoxemia and oxidative stress, immune inflammatory status, redoxpotential, considerably improved in patients of the main group. The signs
of SIRS disappeared in main group, but persisted in patients of control
group. The improvement of the symptoms of SIRS in main group was
coupled with the increasing of ejection fraction by 12%, and NYHA
functional class by the 37% (only 10% in the control group). Conclusion:
The treatment with adenocin cessates the systemic hypoxic and inflammatory syndrome, improves cardiac hemodynamics and symptoms of
congestion in patients with manifested form of CHF.
Paper No.: 1934
FOCUSED CONFERENCE GROUP: PW19 - INFLUENCE OF
DEGENERATION AND REPAIR IN THE CNS AND PERIPHERY
THE STUDY OF ABCB1 POLYMORPHISMS IN ROMANIAN
EPILEPTIC PATIENTS
Paper No.: 3252
FOCUSED CONFERENCE GROUP:
P04 - PHARMACOEPIDEMIOLOGY, CURRENT
CONTROVERSIES AND OPPORTUNITIES
GENDER DIFFERENCE IN THE IMPACT OF LONG-TERM
PSYCHOACTIVE DRUG USE ON COGNITIVE FUNCTIONING:
THE VISAT COHORT
O Boeuf-Cazou(1), B Bongue(2), D Ansiau(3), J-C Marquié(4), Maryse
Lapeyre-Mestre(1)
(1) Université de Toulouse; UPS, Faculté de Médecine Purpan, Unité de
Pharmacoépidémiologie EA3696, Toulouse, France
(2) CETAF, Centre technique d’Appui et de Formation des Centres
d’Examens de Santé, Saint-Etienne, France
(3) International University of Monaco, Principality of Monaco, Monte
Carlo
(4) Université de Toulouse II, Laboratoire Travail et Cognition, UMR
5263 CNRS5, Toulouse, France
Introduction: Few studies have investigated long-term effects of psychoactive drug use among a population aged less than 50. The aim of this
study was to underline the impact of psychoactive drug use on cognitive
functions in a population of workers according to gender. Materials/
Patients: This study included 1,251 French workers from the VISAT
(Aging, Health and Work) cohort whose the objective was to underline
the long-term impact of working conditions on health and aging. VISAT
study included initially 3,237 subjects (current or recently retired workers) born in 1934, 1944, 1954 or 1964 at baseline. Data were collected
through interviews during the annual medical examination by occupational physicians in 1996, 2001 and 2006. Cognitive function was
assessed through five cognitive tests (immediate free recall test, delayed
free recall test, recognition test, Digit Symbol Substitution Subtest and
visual search speed test). Cognitive scores obtained after a 10-year follow-up were investigated among three categories of psychoactive drug
(hypnotic, anxiolytic and antidepressant) users: none (82.1%), long-term
(9.5%) and occasional (8.4%) with analysis of covariance models
adjusted for several potential confounders. Results: In men, we found an
alteration in delayed recall and recognition tests among anxiolytic users,
and an alteration in the recognition test and the digit symbol substitution
subtest among antidepressants users. In women, we observed an alteration in the selective attention test among hypnotic users. Conclusion:
Long-term use of psychoactive medication could impair different cognitive functions depending on gender and type of drugs.
Ioana Corina Bocsan, AD Buzoianu, FC Militaru, R Popp, A Trifa,
LP Dumbrava, E Brusturean
University of Medicine and Pharmacy, Department of Clinical Pharmacology, Cluj Napoca, Romania
Introduction. Epilepsy is one of the most common, chronic neurological
disorders, affecting approximately 42 million patients worldwide. Even
with more than 15 antiepileptic drugs (AED’s), 30% of patients are still
resistant to AEDs. The pharmacokinetic hypothesis for the refractory epilepsy, postulates that resistance is caused by over expression of multidrug
transporter proteins at the blood brain barrier level, possibly as a result
of active efflux mediated by locally over expressed drug transporter proteins. Objective. The aim of this study is to determine the genotype distribution and the allele frequency of the ABCB1 variants in a Romanian
epileptic patients group and compare it with the control group. Methods.
Based on the PCR-RFLP technique, 4 variants of the ABCB1 gene were
studied in 64 epileptic patients and in 200 healthy Romanian volunteers.
Results and conclusions: Our results show low frequency of ABCB1 T
129C and ABCB1 G2677A polymorphisms, only 6.2% of patients being
heterozygous for each of them, the rest of the patients are homozygous
for normal allele. The other two ABCB1 polymorphisms, respectively
2677G>T and 3435C>T, have been frequently found in the patients
group. The distribution of genotypes for ABCB1 3435C>T polymorphism is similar to that of the general population (genotypes CC, CT and
TT had a frequency of 30.8%, 50.8% and 18.4% in the group of patients
compared to 24.5%, 54% and 21.5% in the general population).
Paper No.: 3124
FOCUSED CONFERENCE GROUP: PW02 - SYMPOSIUM ON
ADVANCES IN GI PHARMACOLOGY
PRO-APOPTOTIC AND ANTI-PROLIFERATIVE MECHANISMS
OF STW 5
G Bonaterra(1,2), S Zügel(2), W Hildebrandt(2), Olaf Kelber(3), D Weiser(3), J Metz(4), R Kinscherf(2)
(1) Section Macroscopic Anatomy, Medical Faculty Mannheim, University of Heidelberg, Mannheim, Germany
(2) Department of Medical Cell Biology, University Marburg, Marburg,
Germany
(3) Scientific Department, Steigerwald Arzneimittelwerk GmbH, Darmstadt, Germany
(4) Anatomy and Cell Biology III, University of Heidelberg, Heidelberg,
Germany
Introduction: STW 5 is widely used in the treatment of gastrointestinal
disorders, including functional dyspepsia and colon irritabile, diseases for
which an inflammatory etiology is discussed. Our objective was to determine anti-proliferative and pro-apoptotic effects of this combination of
nine plant extracts on colon adenocarcinoma cells (HT-29) in comparison
with aspirin (ASA) or diclofenac (Diclo), for which a reduction of colon
carcinoma risk is known. Methods: HT-29 cells were treated with Diclo,
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
204
ASA, STW 5 or its components. Anti-proliferative effects and markers
of apoptosis were measured after 72 h. Results: STW 5 (100 ı̀g/ml) and
some of its components induced an inhibition of proliferation by 50-60%
(ASA 0.1 mM or Diclo 2,5 mM 45-50%), a 3 to 4- fold increase in
apoptosis (as well as ASA or Diclo), a 20% to 30% induction of Caspase-3 or BAX expression (ASA or Diclo revealed inhibitory effects), an
inhibition in Bcl2 and p53 mRNA expression, but no change in GDF-15
level (qPCR). Conclusion: STW 5 and some of its components might
have antiproliferative and pro-apoptotic effects on HT-29 cells in vitro.
The STW 5-induced apoptosis pathway is different from the pathway initiated by ASA or Diclo.
Paper No.: 3348
FOCUSED CONFERENCE GROUP: P02 - TRANSMEMBRANE
TRANSPORT: PERSPECTIVES FOR DISEASE AND DRUG
DISCOVERY
A FAST AND SENSITIVE LC-MS/MS ASSAY FOR
QUANTIFICATION OF NORTRIPTYLINE AND ITS ACTIVE
METABOLITES E- AND Z-10-HYDROXYNORTRIPTYLINE IN
HUMAN PLASMA
Gitte Bonke(1,2)*, BP Jensen(2)
(1) * Current address: University of Southern Denmark, Department of
Forensic Toxicology, Odense, Denmark
(2) University of Otago, Department of Medicine, Christchurch, New
Zealand
A fast and sensitive validated liquid chromatography-tandem mass spectrometry assay for quantification of nortriptyline and its active metabolites, E-10-hydroxynortriptyline and Z-10-hydroxynortriptyline in human
plasma following a single oral dose of 25 mg nortriptyline was needed
to support a clinical study (Paper No. 2117). Plasma samples were prepared by protein precipitation with cold methanol:acetonitrile (1:9) and
mean recoveries of 90%, 100%, and 99% were achieved for nortriptyline, E-10-hydroxynortriptyline and Z-10-hydroxynortriptyline. The compounds were separated using a Gemini-NX C18 column, 50 x 2.00 mm
i.d., 3 lm with a mobile phase consisting of 0.1% formic acid in acetonitrile with a gradient of 5-100% acetonitrile over 6 minutes and detected
by ESI+-MS/MS. The standard curves were linear in the range LLOQ–
40 ng/ml (r2 ‡ 0.997) with lower limits of quantification (LLOQ) at
0.2 ng/ml for nortriptyline and 0.5 ng/ml for the metabolites. Intraday
precision C.V. was £6.4% and £16% at LLOQ, interday precision C.V.
was £7.1% and £11% at LLOQ and the accuracy of the method was
within 92-114%. The matrix effect (mean ± S.D.) for nortriptyline was
54% ± 6% but the internal standard, nortriptyline-D3 corrected for this.
For the two metabolites it was 87% ± 3%. The assay has been applied
successfully to a clinical study (as reported in Paper No. 2117).
Paper No.: 898
FOCUSED CONFERENCE GROUP: P06 - THE HEART GONE
WRONG; STABILIZATION OF CARDIAC FUNCTION CLINICAL
RELEVANCE OF FREE MPA MONITORING IN EARLY
PERIOD FOLLOWING HEART TRANSPLANTATION
Roselyne Boulieu(1,2), X Cussonneau(1,2), M Larger(1,2), O Bastien(2,3)
(1) University Lyon1- Faculty of Pharmacy, Department of Clinical Pharmacokinetics, Lyon, France
(2) Inserm ERI 22, Lyon, France
(3) University Lyon1, Cardiology Unit, Lyon, France
Mycophenolic acid (MPA), the active metabolite of the pro-drug mycophenolate mofetil, and the phenolic glucuronide (MPAG) are highly
bound to albumin. Scarce data are available on free MPA pharmacokinetics in heart transplantation. The aim of this study was to investigate free
MPA and free MPAG in 16 patients in the early period after heart trans-
plantation and to assess the relation between the free forms and biological data. MPA and MPAG were analyzed by HPLC. Free MPA and free
MPAG were isolated by ultrafiltration. AUC0-12, C0, free fraction and
Cmax were determined from full pharmacokinetic profile. Creatinine
clearance, blood cell count, hematocrit and hemoglobin were collected.
A wide interindividual variability in free MPA and free MPAG concentrations was observed. Mean absolute AUC0-12 were 0.484 mg.h/L and
691.5 mg.h/L for free MPA and free MPAG respectively. MPAG free
fraction was positively related to MPA free fraction (r = 0.814,
p = 0.001). Free MPA and free MPAG pharmacokinetic parameters were
negatively correlated to creatinine clearance. Moreover, absolute free
MPA C0 was negatively related to red blood cell count, haemoglobin
and hematocrit but not free MPA AUC0-12. This observation which
needs to be confirmed suggests that free MPA C0 could be a relevant
marker related to the myelotoxic effects of the drug and emphasize the
relevance of free MPA and MPAG monitoring in the early period following heart transplantation.
Paper No.: 899
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
MONITORING OF AZATHIOPRINE METABOLITES IN
PEDIATRIC PATIENTS WITH AUTOIMMUNE HEPATITIS
Roselyne Boulieu(1,2), TMH Nguyen(0), C Le Gall(3), A Lachaux(3)
(1) University Lyon1- Faculty of Pharmacy, Department of Clinical
Pharmacokinetics,Lyon, France
(2) Inserm ERI 22, Lyon, France
(3) University Lyon1, Cardiology Unit, Lyon, France
Azathioprine, an immunosuppressive drug commonly used in the treatment of autoimmune hepatitis (AIH) is converted to 6-thioguanine nucleotides (6-TGN), the active metabolites and to 6-methylmercaptopurine
nucleotides (6-MeMPN) via Thiopurine Methyl Transferase (TPMT).
Few data are available on drug monitoring of azathioprine metabolites in
pediatric patients. The purpose of this study was to investigate thiopurine
metabolites in children with AIH and the relevance of drug monitoring
in regard to efficacy and toxicity. Data from 28 patients with AIH treated
by azathioprine for at least 3 months were recorded. 6-TGN and 6MeMPN concentrations and TPMT activity were determined by HPLC.
Blood cell counts, liver function tests were also collected as well as clinical outcome. A wide interindividual variability in 6-TGN and 6-MeMPN
concentrations was observed in pediatric AIH patients with range values
of 51-1966 and 42-8189 pmol/8x108 RBC for 6-TGN for 6-MeMPN
respectively. 61.4% of the patients achieved remission and only 32.6%
of these children had 6-TGN values within the target range proposed for
IBD (250 – 450 pmol/8x108 RBC). Azathioprine dosage was significantly related to 6-TGN and 6-MeMPN levels (r = 0.308, P < 0.001 and
r = 0.405, P < 0.001, respectively). A significant negative correlation
was observed between 6-TGN concentrations and erythrocyte and lymphocyte counts. Thiopurine metabolite monitoring appears useful in differentiating myelotoxicity and hepatotoxicity due to azathioprine from
disease recurrence and help to identify non-compliant patients. Further
larger study are needed to confirm the optimal recommended target for
thiopurine metabolites to achieve remission in AIH patients.
Paper No.: 3322
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
ANTI-INFLAMMATORY AND ANTIPROLIFERATIVE
POTENTIAL OF EXTRACTS AND PURIFIED FRACTIONS
FROM MEDITERRANEAN MARINE ALGAE
Abderrahmene Bouraoui(1), W Chaouch(1), L Mhadhebi(1), A Dellai(1),
M Dridi(1), A Laroche(2), F Berrue(3), R kerr(3), J Robert(2)
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
205
(1) Research Unit URSAM, Laboratory of Pharmacology, Faculty of
Pharmacy of Monastir, Monastir, Tunisia
(2) Institute Bergonie, Laboratory of Pharmacology of Anticancer Drugs,
Bordeaux, France
(3) UPEI, Marine Natural Products Laboratory and Discovery, Charlottetown, Canada
As part of our search for new anti-inflammatory or anticancer potential
drugs, organic extracts and purified fractions of marine algae collected
from Mediterranean Tunisia coasts were screened and evaluated for their
pharmacological potential. The present study has established that the
organic extracts of the brown algae of the genus Cystoseira showed a
strong cytotoxic activity against three human cancer cell lines (A-549,
MCF-7, HCT-15), using a MTT cytotoxic assay. At concentrations of 0.1
to 1 mg/ml these extracts suppressed, dose dependently, the proliferation
of the three cell lines by more than 75%. The IC50 values ranged from
40 to 80 lg/ml. We also established that the organic extracts and purified
fractions of the brown algae of the genus Dictyopteris, cystoseira and of
the red algae of the genus Hypnea at different doses (25, 50, 100 mg/kg)
showed a significant anti-inflammatory activity, using the carrageenan
paw edema test in male Wistar rats and in comparison to reference drugs:
dexamethasone (1 mg/kg) and aspirin (300 mg/Kg). The inhibitory effect
on the rats paw oedema is dose dependently and the % inhibition of
oedema 3 h after carrageenan injection ranged from 60 to 85%. In addition to these pharmacological activities, some of purified fractions
showed a strong inhibitory activity against PTP1B: the IC50 ranged
from 5 to 10 lg/ml. In order to isolate and identify the active substances
responsible for these pharmacodiversities of marine algae extracts
and fractions, chemical studies (HPLC, LC /MS and NMR) are under
investigation.
Paper No.: 2349
FOCUSED CONFERENCE GROUP: P17 - NEW APPROACHES
AND TARGETS IN PSYCHIATRY
THE HPA AXIS, CARDIO-VASCULAR SYSTEM AND
BEHAVIOR IN RAT CMS MODEL OF DEPRESSION
Elena V Bouzinova(1), S Christiansen(1), K Henningsen(1), R Palme(2),
O Wiborg(1)
(1) Aarhus University Hospital Risskov, Center for Psychiatric Research,
Aarhus, Denmark
(2) University of Veterinary Medicine, Department of BiomedicalSciences/Biochemistry, Vienna, Austria
Introduction: The Chronic Mild Stress is a well-established and validated
model to develop depression-like symptoms in rodents. In the current
study we have used a rat CMS model to associate the function of the
stress axis with behavioral and physiological parameters. Methods: Rats
are exposed to unpredictable, mild stressors for 8 weeks. Stress
responses, like depression associated anhedonia-like symptoms, are measured by intake of palatable sucrose solution. Already after one week of
stress exposure rats divide in two groups: stress-susceptible, i.e. anhedonic-like, are declining in sucrose intake compared with baseline level
and stress-resilient rats. The activity of HPA axis was measured as faecal
corticosterone metabolites (FCM) in diurnal samples. Different kinds of
behaviors have been screened in open field, elevated O-maze, shuttle
box, and Y-maze. The tail occlusion method has been used to measure
the blood pressure and heart rate in CMS rats. Results: Significantly
higher FCM concentrations were observed in CMS animals from first up
to fourth week of stress. Afterwards FCM returned to the baseline level
and remained unchanged until the end of experiment. The Principal
Component Analysis revealed correlations between anhedonia and some
behavioral parameters. Anhedonic rats have lower anxiety levels in
o-maze and less emotional activity in both o-maze and open field. No
differences were observed in explicit memory test, but anhedonic rats
made significantly more mistakes in Y-maze indicating working memory
impairments. Slight increases in heart rate and blood pressure in anhedonic rats were insignificant.
Paper No.: 2669
FOCUSED CONFERENCE GROUP: P02 - TRANSMEMBRANE
TRANSPORT: PERSPECTIVES FOR DISEASE AND DRUG
DISCOVERY
SLCO1B1 VARIANTS AS PHARMACOGENETIC
PREDISPOSITION FOR HEPATOTOXICITY INDUCED BY
HMGCOA REDUCTASE INHIBITORS
Nada Božina(1), I Meræep(2), V Miroševic(3), T Božina(1), J Sertiæ(1),
Z Reiner(2)
(1) University Hospital Center, Department of Laboratory Diagnostics,
Zagreb,Croatia
(2) University Hospital Center, Division of Metabolic Diseases,
Department of Internal Medicine, Zagreb, Croatia
(3) Agency for Medicinal Products and Medical Devices, Zagreb,
Croatia
Studies identified a genetic risk factor for myotoxicity induced by HMGCoA reductase inhibitors (statins) (The SEARCH Collaborative Group,
N Engl J Med.;359(8):789-99, Voora et all, J Am Coll Cardiol. 2009;
54(17):1609-16). Researchers observed a strong association between
myotoxicity and variants in the SLCO1B1 gene which encodes
OATP1B1, one of the key hepatocellular uptake transporters providing
extraction of diverse compounds including statins from portal venous
blood into the liver. However, no studies have been done to investigate
genetic risk factors for developing statin induced hepatotoxicity. We performed SLCO1B1 genotyping in patients who developed hepatotoxicity
induced by statins. We analyzed 6 patients (5 male and 1 female, mean
age was 45, range 33-57 years) who developed hepatotoxicity caused by
atorvastatin (20-80 mg/day) in 5 cases and fluvastatin in 1 case (80 mg/
day). We genotyped blood of all patients for SLCO1B1 388 A>G and
521T>C and additionally for CYP2C9*2,*,3 in a patient who was taking
fluvastatin. Genotyping was performed by real-time PCR methods.
Results. Four patients (one carrier of SLCO1B1 521T/C, another of
521T/C and 388A/G, the 3rd of 388G/G and the 4th of 521C/C and
388G/G) developed hepatic lesion and among them the homozygote carrier for both low activity SLCO1B genotypes had worst clinical presentation. Two other patients developed hepatotoxicity (one patient had
hepatic lesion and another increased transaminases) together with
increased creatin phosphokinase (one was a carrier of CYP2C9*3,
SLCO1B1 388A/G, 521T/C and another of 388G/G). Our preliminary
data indicate associations between hepatotoxicity of statins and both
CYP2C9 and SLCO1B1 polymorphysms.
Paper No.: 2670
FOCUSED CONFERENCE GROUP: P17 - NEW APPROACHES
AND TARGETS IN PSYCHIATRY
THE ROLE OF CYP2D6 AND ABCB1 PHARMACOGENETICS
IN THE CLINICAL EFFICACY OF ORALLY ADMINISTERED
RISPERIDONE
Nada Božina(1), N Jovanoviæ(2), M Lovriæ(1), V Medved(2),
M Kudumija-Slijepèeviæ(3), J Grubišin(2)
(1) University Hospital Center, Department of Laboratory Diagnostics,
Zagreb, Croatia
(2) University Hospital Center, Department of Psychiatry, Zagreb,
Croatia
(3) Bjelovar General Hospital, Department of Psychiatry, Croatia
The aim was to evaluate the role of CYP2D6 and ABCB1 variants in the
efficacy and safety of patients treated with oral risperidone. The study
included 85 patients with first episode of psychosis who were treated
with risperidone and then followed-up for eight weeks. Their psychopathology was assessed using The Positive and Negative Syndrome Scale
(PANSS). The CYP2D6 genotyping was performed by PCR-RFLP (for
*3,*4,*6) and long PCR (for duplications and *5). For ABCB1 G2677T/
A and C3435T, real-time PCR method was applied. Plasma concentrations of risperidone and 9-OH risperidone were measured by HPLC. The
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
206
distribution of genotypes: CYP2D6 wt/wt, wt/mut, and mut/mut was 44,
32 and 8 respectively; for ABCB1 2677G/G, G/T, T/T was 29, 44 and
12, for 3435CC, C/T and T/T was 25, 39 and 21, respectively. The drug
was prescribed at the mean dose of 3.94 mg ± 1.47 (range 1-8). The
patients with the CYP2D6 wt/wt genotype had significantly lower dosecorrected levels of risperidone (C/Ds) (F = 20.46, P < 0.001) and active
moiety (F = 4.496, P = 0.014), while the CYP2D6 mut/mut carriers had
the lowest levels of 9-OH risperidone C/Ds (F = 3.22, P = 0.046). The
ABCB1 genotypes did not significantly influence these parameters. A
total of 23 participants (27.4%) were responders (defined as 50%
improvement from baseline in total PANSS) and the ABCB1 3435T
allele increased the likelihood of being a responder for more than seven
times (OR = 7.306, 95% CI = 2.738–19.493, P = 0.006). Conclusion:
The CYP2D6 and ABCB1 genotyping may serve for personalizing therapy with risperidone.
Paper No.: 1489
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
SIMVASTATIN DECREASES MALONDIALDEHYDE LEVEL IN
PLASMA AND TISSUES OF RATS
Vlasta Bradamante(1), M Marija(1), L Jasna(2), K Marijan(1),
K Pasko(3)
(1) University of Zagreb School of Medicine, Department of Pharmacology, Zagreb, Croatia
(2) University of Zagreb School of Medicine, Department of Chemistry
and Biochemistry, Zagreb, Croatia
(3) NMR Center, Zagreb, Croatia
Introduction. It is considered that beneficial effect of simvastatin treatment on cardiovascular morbidity is due to its antioxidant activity. Simvastatin also shows antioxidant properties in cisplatin-induced
nephrotoxicity and hepatotoxicity. The aim of this study was to compare
the effect of simvastatin on the malondialdehyde level in plasma and different tissues of rats and to establish whether antioxidant effects exist
after discontinuation of its administration. Material and methods. Two
groups of male rats were under simvastatin treatment which was given
orally (10 mg/kg/day for 3 weeks). At the end of the drug treatment, the
first group was sacrificed and blood and tissue (heart, brain, liver, kidney) samples were taken. Other group was without the for the next
9 days (recovery period) and than sacrificed. The malondialdehyde level
was measured in plasma (ı̀mol/l) and tissue (ı̀mol/g tissue) by spectrophotometric method. The results are expressed as the percentage of the
decrease of malondialdehyde level and are compared to the control
group. Data were analyzed by t-test. Results. Simvastatin decreased malondialdehyde level in plasma and all tissue by 20-40%. The same
decrease of malondialdehyde was established in the recovery period, i.e.
9 day after the last dose. Conclusion. Our results have shown that simvastatin exhibits the significant antioxidant effectiveness which persists
after therapy discontinuation.
Paper No.: 1003
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
PROTEIN KINASE C (PKC) REGULATION OF TYPE 5
METABOTROPIC GLUTAMATE (MGLU5)
RECEPTOR-STIMULATED CA2 + OSCILLATIONS IN RAT
CEREBROCORTICAL ASTROGLIA
Sophie J Bradley(1), JM Watson(2), RAJ Challiss(1)
(1) University of Leicester, Department of Cell Physiology and Pharmacology, Leicester, UK
(2) GlaxoSmithKline, UK
Astrocytes fulfil vital homeostatic and neuromodulatory roles in the
CNS. Glutamate stimulation of astrocytes generates oscillations in cytosolic Ca2 + concentration via a PKC-dependent mechanism, which has
been shown to involve the rapid and reversible phosphorylation of
mGlu5 receptor in a process referred to as ‘‘dynamic uncoupling’’ (Nash,
MS et al. J Biol Chem 2002, 277; 35947-35960). Stimulation of rat cortical astrocytes with glutamate induced robust oscillatory changes in [Ca2 +
]i at a single-cell level which could be prevented by pre-addition of the
mGlu5 receptor-selective negative allosteric modulator MPEP. Inhibition
of PKC with staurosporine (3 lM) converted glutamate-stimulated
Ca2 + oscillations into a peak-plateau response. Acute stimulation of protein kinase C with low concentrations of phorbol dibutyrate (PDBu)
caused a decrease in glutamate (100 lM)-stimulated Ca2 + oscillation frequency, with complete inhibition of this response at higher (100 nM)
PDBu. In cell populations, staurosporine or Ro31-8220 caused 2-fold
increases in the maximal agonist-stimulated [3H]-IP accumulation without significant change in EC50 value. In contrast, pre-incubation with the
Ca2 + -dependent PKC inhibitors Gö6976 or PKC20-28 did not affect
glutamate-stimulated Ca2 + -oscillation frequency, or agonist-stimulated
[3H]-IP accumulation. Furthermore, loading astrocytes with BAPTA-AM
(100 lM) had no significant effect on agonist-stimulated [3H]-IP accumulations. These data provide further evidence for the involvement of
Ca2 + -independent PKCs in the rapid cycles of receptor phosphorylation
and dephosphorylation that likely determine mGlu5 receptor regulation
of Ca2 + oscillation frequency in rat cortical astrocytes and pinpoint postreceptor mechanisms that can shape receptor-driven Ca2 + oscillatory
behaviours.
Paper No.: 1648
FOCUSED CONFERENCE GROUP: P17 - NEW APPROACHES
AND TARGETS IN PSYCHIATRY
CHOLINERGIC AND GLUTAMATE-NITRIC OXIDE
SIGNALLING PATHWAYS IN A GENETIC ANIMAL MODEL
OF DEPRESSION, THE FLINDERS SENSITIVE LINE (FSL) RAT
Linda Brand(1), EL Minnaar(1), J van Zyl(1), G Wegener(2),
BH Harvey(1)
(1) North-West University, Division of Pharmacology, Potchefstroom,
South Africa
(2) University of Aarhus, Centre for Psychiatric Research, Aarhus, Denmark
Depression presents with evidence for degenerative pathology that relates
to disturbances in GABA and glutamatergic pathways, while there is also
evidence for an involvement of cholinergic and nitrergic pathways. The
FSL rat is a well-validated genetic animal model of depression. This
study aimed to determine whether depressive-like behavior is accompanied by altered GABA, NMDA-NO/cGMP and cholinergic signalling in
the hippocampus and frontal cortex of the FSL rat. Total regional brain
nitrogen oxides, GABA, L-citrulline and acetylcholine (ACh) were analysed by different chromatographic methods. NMDA and muscarinic M1
receptor characteristics were determined by radioligand binding assays
and cGMP levels by a radio-immunoassay kit. Behavioural assessments
included open field and social interaction tests, with results suggestive of
increased anxiogenic behavior in the FSL rat. Significantly lower basal
cGMP levels were noted in frontal cortex, but not hippocampus, of FSL
versus control rats, although no other differences with respect to indices
of NOS activity, receptor characteristics or GABA levels were apparent
in either brain area. Significantly elevated frontal cortical, but significantly reduced hippocampal levels, of ACh were observed in FSL rats.
Results suggest the presence of bidirectional cholinergic dysfunction in
frontal cortex and hippocampus of the FSL rat. Reduced fronto-cortical
cGMP levels in absence of altered NO signalling supports an NO-independent mechanism of cGMP regulation in FSL rats, possibly involving
a muscarinic cholinergic-NOS directed pathway. This will have important
implications for our understanding of ACh-NO interactions in the depressive-like behaviour of the FSL rat and, indeed, in the neurobiology and
treatment of depression.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
207
Paper No.: 1848
FOCUSED CONFERENCE GROUP: PW12 - EXPERIMENTAL
PAIN METHODS IN DEVELOPMENT AND VALIDATION OF
ANALGESICS
THE PHARMACOGENOMICS OF ESCITALOPRAM IN THE
TREATMENT OF NEUROPATHIC PAIN
Charlotte Brasch-Andersen(1,2), MU Møller(1,2), SH Sindrup(3),
K Brøsen(1), M Otto(3)
(1) Odense University Hospital, Department of Clinical Pharmacology,
Odense, Denmark
(2) Odense University Hospital, Department of Clinical Genetics,
Odense, Denmark
(3) Odense University Hospital, Department of Neurology, Odense,
Denmark
Painful polyneuropathy is a common neuropathic pain condition with
painful diabetic neuropathy being a prominent example. Numerous pathophysiological mechanisms are involved in the generation and maintenance
of neuropathic pain. In spite of increasing knowledge about the mechanisms, the treatment of neuropathic pain is still unsatisfactory. The effects
of a selective serotonin reuptake inhibitor (SSRI), escitalopram, on pain in
polyneuropathy was tested in an earlier randomized, placebo-controlled,
double- blinded cross-over trial including 41 patients who were treated
with 20 mg escitalopram and placebo. The clinical study showed a moderate, good or complete pain relief for escitalopram in 11 patients (‘‘responders’’) but at only a slight or no relief in 30 patients (‘‘non-responders’’).
The difference in response to escitalopram may be caused by genetic differences between the responders and the non-responders, which led to a
pharmacogenomic study investigating genetic differences between the two
groups for functional variants in genes involved in the signalling pathway
for serotonin. We tested the serotonin receptor subunits 5-HTR2C G68C
(rs6318) and 5-HTR2A C1354T (rs6314) both giving rise to amino acid
changes. We furthermore tested a missense variant in the gene coding for
P-glycoprotein, ABCB1 G2677T (rs2032582) and a promoter polymorphism (5-HTTLPR) in the serotonin transporter gene. We were not able to
find any statistical significant association between the tested genetic polymorphisms and effect of escitalopram when dichotomizing the patients
into responders and non-responders. Using quantitative assessment of pain
might add more information to the association studies.
Paper No.: 2426
FOCUSED CONFERENCE GROUP: PW04 - ANTIOXIDANTS AS
THERAPEUTIC TARGETS
NUCLEIC ACID OXIDATION IS PREDOMINANTLY DETERMINED BY NON-GENETIC FACTORS IN ELDERLY DANISH
TWINS
Kasper Brødbæk(1,2), R Ribel-Madsen(3), T Henriksen(1,2),
A Weimann(1,2), M Petersen(1,2), J T Andersen(1,2), S Afzal(1,2),
B Hjelvang(1,2), A Vaag(3), P Poulsen(3,4), H E Poulsen(1,2,5)
(1) Laboratory of Clinical Pharmacology, Rigshospitalet, Copenhagen,
Denmark
(2) Department of Clinical Pharmacology, Bispebjerg Hospital, Denmark
(3) Steno Diabetes Center, Gentofte, Denmark
(4) Novo Nordisk A/S, Soeborg, Denmark
(5) Faculty of Health Sciences, University of Copenhagen, Copenhagen,
Denmark
DNA oxidation is thought to be important in both cancers and in many
non-cancerous conditions. To what extend nucleic acid oxidation is determined by genetic and environmental factors is unknown. The aim of this
study was to examine the contribution of genetic versus environmental
factors to the rate of nucleic acid oxidation by performing a classical
twin study. The study population included 198 elderly twins (aged 62–
83 years), 46 monozygotic (MZ) and 53 dizygotic (DZ) twin pairs. Urine
samples were collected for measurement of markers of nucleic acid oxidation: 8-oxodG and 8-oxoGuo using ultra-performance liquid chroma-
tography and tandem mass spectrometry detection. Classical twin
analyses including intraclass correlation coefficients and heritability estimates were used to determine the relative contributions of genes and
environment to the variation in nucleic acid oxidation. There were no statistically significant differences in intraclass correlations between MZ
and DZ twins neither for 8-oxodG (rMZ = 0.55, rDZ = 0.47; P = 0.43)
nor 8-oxoGuo (rMZ = 0.45, rDZ = 0.58; P = 0.21). The heritability estimates for 8-oxodG and 8-oxoGuo excretion were h2 = 0.17 and h2 ~ 0,
respectively. In conclusion, we demonstrated that in a large population of
elderly Danish twins the rate of nucleic acid oxidation is predominantly
determined by potentially modifiable non-genetic factors. (Workshop reference: SW8, Antioxidants as therapeutic agents)
Paper No.: 2575
FOCUSED CONFERENCE GROUP: PW02 - SYMPOSIUM ON
ADVANCES IN GI PHARMACOLOGY
AL-TRYPTOPHAN RICH DIET IMPROVES ULCER HEALING
VIA ENDOGENOUS MELATONIN, PROSTAGLANDINS AND
DOWNREGULATION OF HYPOXIA INDUCIBLE FACTOR 1A
(HIF-1)
Thomas Brzozowski(1), A Ptak-Belowska(1), R Pajdo(1), S Kwiecien(1),
M Pawlik(1), I Brzozowska(2), PC Konturek(3), WW Pawlik(1), SJ Konturek(1)
(1) Jagiellonian University Medical College, Department of Physiology,
Cracow, Poland
(2) Jagiellonian University Medical College, Department of Anatomy,
Cracow, Poland
(3) Thuringen Clinic, Department of Medicine, Saalfeld, Germany
Exogenous melatonin exhibits gastroprotective and ulcer healing activities but the role of endogenous melatonin derived from daily dietary
intake of L-tryptophan (L-Try) in the mechanism of ulcer healing has not
been extensively studied. We compared the effects of i.g. administration
of vehicle (saline) or melatonin in rats fed with normal, L-Try-depleted
(0% of L-Try) and L-Try rich-diet (Try+ were determined by RT-PCR.,
0.30% L-Try of total protein) of without or with blockade of melatonin
Mel2 receptors with luzindole (20 mg/kg-d i.p.) on healing of gastric
ulcers induced by acetic acid (ulcer area=28 mm2). At 9 days after ulcer
induction, gastric ulcers were measured by planimetry, gastric blood flow
(GBF) was determined by H2-gas clearance technique and plasma L-Try
and gastric mucosal PGE2 concentrations and expression of mRNA for
COX-1, COX-2 and hypoxia HIF-1 Ulcer healing was significantly
accelerated in rats fed with L-Try rich diet while raising GBF at ulcer
margin, plasma L-Try, melatonin and gastrin levels and mucosal generation of PGE2. In L-Try free diet the plasma levels of L-Try and melatonin levels were significantly diminished and ulcer healing was
significantly delayed. Indomethacin and rofecoxib or luzindole, an antagonist of Mel2 receptors, significantly attenuated L-Try rich diet-induced
acceleration of ulcer healing. was downregulated but COX-1 and COX-2
mRNAs were upregulated by L-Try rich diet. Gastric mucosal HIF-1 We
conclude that endogenous melatonin synthesized from L-Try accelerates
ulcer healing via interaction with Mel2 receptors, an enhancement of gastric microcirculation, mediated by PGE2 derived from COX-1 and COX2 overexpression and activity.
Paper No.: 1996
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
PKCB/HUR/VEGF A NEW PATHWAY IN DIABETIC
RETINOPATHY
Claudio Bucolo(1), A Pascale(2), M Amadio(2), S Govoni(2),
F Drago(1)
(1) University of Catania, Department of Experimental and Clinical
Pharmacology, Catania, Italy
(2) University of Pavia, Italy
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
208
Introduction: To investigate whether PKCbII could control VEGF levels
via the mRNA stabilizing human embryonic lethal abnormal vision
(ELAV)-like protein, HuR, in retina of streptozotocin(STZ)-induced diabetic rat. Methods: Retinal tissues, collected after 10 days from STZinjected rats, were processed for western blotting and preparation of
mRNP (ribonucleoproteic complexes). Anti-PKCbI mouse monoclonal
antibody, anti-PKCbII rabbit polyclonal antibody, anti-HuR mouse
monoclonal antibody, anti-VEGF rabbit polyclonal antibody and antialfa-tubulin rat monoclonal antibody were used for western blot analysis.
Samples were processed for immunoprecipitation using anti-HuR antibody, and VEGF mRNA was detected by real-time quantitative PCR.
Statistical analysis was performed by ANOVA by and an appropriate
post hoc comparison test Results: PKCbI and PKCbII were increased in
the retina of STZ injected rats compared to sham (+160% +113%,
respectively). We showed a PKC-mediated phosphorylation of HuR in
the retina from diabetic rats. HuR protein levels increased in STZ-treated
rats (+62% vs. sham), and this was accompanied by an higher phosphorylation in serine residues (+209% vs. sham). These effects were blunted
by selective PKCb inhibitor treatment. A specific binding between HuR
protein and VEGF mRNA in the retina was shown. Further, the PKCb/
HuR activation is accompanied by enhanced VEGF protein expression
and this effect was attenuated by PKCb inhibitor. Conclusion: These
findings demonstrated the existence of PKC/HuR/VEGF pathway in
experimental diabetic retinopathy. A better dissection of this cascade in
diabetic conditions may help to disclose new potential pharmacological
targets. Acknowledgments: This work was supported by a grant from
MIUR - PRIN 2007.
Paper No.: 2998
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
EVALUATION OF GUIDELINE-CONFORMITY OF INITIATION
OF ORAL HYPOGLYCEMIC TREATMENT FOR INCIDENT
DIABETES MELLITUS TYPE 2 IN AUSTRIA
Anna Bucsics(1), M Asslaber(1), R Bauer(1), T Burkhardt(1),
M Pogantsch(2), A Schautzer(2), PR Wieninger(1), WC Winkelmayer(3)
(1) Main Association of Austrian Social Security Institutions, Vienna,
Austria
(2) Styrian Sickness Fund, Austria
(3) Harvard University, Boston, MA, USA
Guidelines recommend metformin as first line oral hypoglycemic (OH)
treatment in patients with diabetes uncontrolled with diet and lifestyle
modifications. It is unknown what proportion of incident patients comply
with this recommendation in Austria. We used claims from 11 sickness
funds that covered >95% of the Austrian general population. We identified new users aged 19-100 years of a first OH drug, defined as a first
filled prescription after one year without any antidiabetic drug (including
insulin). Among these incident users, we described the OH drug class
used and correlates of guideline-conforming initiation with metformin
monotherapy using logistic regression. Over 18 months (1/2007-6/2008),
we identified 42882 incident users of an OH drug: 70.1% used metformin, 24.7% used a sulfonylurea, and 4.5% initiated treatment with
another class. Independent correlates of metformin initiation were younger age, female gender, waived copayment (as indicator of low socioeconomic status), more recent year, fewer hospital days in the year prior to
first OH therapy, and more non-OH prescriptions filled during that baseline period (all P < 0.001). There was significant heterogeneity across
sickness funds. Approximately 20% of metformin initiators had a second
antidiabetic drug added within 12 months. While we were unable to
ascertain specific contraindications to metformin in this dataset (renal
insufficiency, hepatic failure), <5% of the general population is expected
to have these conditions. Conclusion: 70% of new initiators of OH treatment in Austria received metformin as recommended by international
guidelines. At least 25% did not, taking into account possible contraindications, providing an opportunity for systematic intervention.
Paper No.: 3155
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
TNFA DOES NOT INHIBIT INSULIN-STIMULATED 2-DEOXYGLUCOSE UPTAKE IN IN SITU SKELETAL MUSCLE, BUT
DOES INCREASE LIPOLYSIS AND LOWER TOTAL GLUCOSE
UTILIZATION
Christian Selmer Buhl(1), CN Petersen(1), N Jessen(1), O Schmitz(1), S
Lund(2), ES Buhl(1)
(1) Aarhus University, Department of Pharmacology, Aarhus, Denmark
(2) Aarhus Sygehus, Department of Endocrinology M, Aarhus, Denmark
Elevated plasma levels of the proinflammatory cytokine TNFa is
strongly correlated to insulin resistance and obesity. However, the exact
role of TNFa in metabolism is not understood. The aim of the present
study was to investigate the effect of TNFa stimulation on insulin sensitivity in muscle using the isolated and perfused rat hindlimb. This model
allows us to investigate direct effects on different types of muscle fibres
without interactions with neuro-endocrine pathways. The hindlimbs of
male wistar-rats were mounted and perfused at a constant flow with a
heated and oxygenized Krebs-Ringer solution. Glucose uptake was measured using 2-deoxy-3H-glucose and 14C-mannitol. At the end of the
experiment muscle biopsies were taken out and analyzed for glycogen
content. Perfusion medium was analyzed for glycerol, FFA, lactate and
total glucose utilization. Insulin and TNFa was respectively added in
concentrations of 0, 75, 500 mU/L and 0, 250, 2500ng/L. Insulin
increased glucose uptake by 1.6-3.2-fold at low and 10.0-14.7-fold at
high concentrations. TNFa did not affect basal or insulin-stimulated
2-deoxy-glucose-uptake. However, TNFa did reduce the rate of total
glucose utilization (up to 33%) and did increase the release of lactate (up
to 43.3%) and the amount of glycogen deposited (up to 23.4%) (all
P < 0,05). Furthermore TNFa significantly increased the lipolysis with
the release of FFA by up to 2.5-fold along with an increase in total
oxygen uptake. The findings show that TNFa does not affect 2-deoxyglucose-uptake in muscle. However, TNFa mediates an increase in lipolysis that may be at the expense of an attenuated glucose oxidation.
Paper No.: 2468
FOCUSED CONFERENCE GROUP: P08 - DEVELOPMENTS IN
THE TREATMENT OF SEXUAL DYSFUNCTION AND
DISEASES OF THE LOWER URINARY TRACT
INNERVATED MYOFIBROBLASTS IN THE URINARY
BLADDER? FUNCTIONAL AND ULTRASTRUCTURAL
EVIDENCE
Elizabeth Burcher(1), P Sadananda(1), S Sandow(1)
University of New South Wales, Department of Pharmacology, Sydney,
Australia
We have reported that neurokinin A (NKA) can contract isolated mucosal strips from porcine bladder, with dense smooth muscle actin staining
on the suburothelial myofibroblasts – these may mediate the contraction
to NKA (Sadananda et al., Br J Pharmacol 2008; 153, 1465-73). Our
aim was to examine the ultrastructure of the porcine bladder mucosa and
characterize the relationship of the suburothelial myofibroblasts with
adjacent cells. Pig bladders obtained from an abbatoir were processed for
electron microscopy. The suburothelial population of myofibroblasts
were morphologically similar to the deep myofibroblasts of human detrusor, in that they have prominent rough endoplasmic reticulum and elongated processes. Extensive nerve bundles with varicosities were located
below the urothelium and in close proximity to myofibroblasts. These
nerve bundles contained individual varicosities with discrete regions
(bare of Schwann cells) containing synaptic vesicles, facing the potential
neuroeffector cell. This is in accordance with the definition of neuroeffector junctions, as neurotransmitter release sites. These dense-cored synaptic vesicles were 100-250 nm in diameter, consistent with a primarily
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
209
peptidergic content. In low resolution studies (n = 4), synaptophysin and
tachykinin immunoreactivity localised nerve fibres containing tachykinins within the suburothelial nerve plexus and in association with
myofibroblasts. In conclusion, the strategic location of suburothelial
myofibroblasts between the urothelium and afferent nerve plexus suggests that they are involved in sensory processing and cross talk between
cell layers. We hypothesise that peptides such as NKA are released from
these dense-cored synaptic vesicle-containing varicosities, to act directly
on suburothelial myofibroblasts, resulting in contraction of the porcine
bladder mucosa.
Paper No.: 1148
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
SEARCHING FOR NOVEL ANALGESIC AND
ANTI-INFLAMMATORY ANALOGS OF ANANDAMIDE
Sumner Burstein(1), G Li(2), R Salmonsen(1), C McQuain(1), T Al(2),
V Kattamuri(2)
(1) University of Massachusetts Medical School, Department of Biochemistry & Molecular Pharmacology, Worcester, MA, USA
(2) Texas Tech University, Department of Chemistry & Biochemistry,
Lubbock, TX, USA
Recently, data were presented supporting the possibility that analogs of
anandamide called elmiric acids are potential anti-inflammatory agents
[Burstein SH, et al. Bioorganic & medicinal chemistry 2007;15:334555]. We have developed a mechanism-based in vitro assay for screening
compounds for potential anti-inflammatory activity based on their stimulatory action on prostaglandin J2 levels and have applied this to the novel
compounds described in this report. We determined a rank order of antiinflammatory potencies for a series of ten analogs of the endocannabinoid
anandamide that were synthesized via new asymmetric reactions and
multiple-step protection-deprotection procedures. The parent molecules
are, (5Z,8Z,11Z,14Z)-N-(2-hydroxyacetyl)icosa-5,8,11,14-tetraenamide
(GL-A) and (5Z,8Z,11Z,14Z)-N-((1S,2R)-2,3-dihydroxy-1-phenylpropyl)icosa-5,8,11,14-tetraena mide (GL-B). Here we would like to report
some preliminary results for these new anti-inflammatory candidates.
GL-B and its enantiomer showed a potency difference of approximately
20 fold. This degree of stereospecificity suggests that the stimulation of
PGJ levels in RAW264.7 cells is probably receptor mediated. Because of
their similarity to anandamide, the cannabinoid receptors are a possibility.
As with the elmiric acids, palmitoyl derivatives have little or no effect on
prostaglandin production whereas arachidonoyl analogs are all relatively
active. Halogen substituents on the phenyl ring showed a range of
potencies all of which were lower than the parent GL-B.
Support was provided by NIDA (1R03DA026960).
Paper No.: 965
FOCUSED CONFERENCE GROUP: P18 - NUCLEAR
RECEPTOR TARGETS FOR TREATMENT OF DISEASES
EFFECTS OF GONADECTOMY ON HIPPOCAMPAL
AROMATASE EXPRESSION IN BOTH GENDERS
determine the potencial effects of gonadectomy on central aromatase
expression in hippocampus. Female and/or male Sprague-Dawley rats
(200-250g) were used in experiments. Ovariectomy, castration or sham
surgery was performed in anesthetized rats. At the end of 8-weeks, rats
were killed by decapitation and hippocampal tissue homogenates were
prepared and evaluated for aromatase proteins by western blot. b-actin
was used as internal control to ensure equel protein loading. Student’s
test was used for statistical analysis and P < 0.05 was considered significant. Aromatase enzyme levels were increased in ovariectomized rats,
but did not alter in castrated rats when compared with control sham operated groups. According to these results it can be suggested that loss of
endogenous peripheral aromatase through surgical gonadectomy alters
hippocampal local aromatase levels in female but not male rats. The
increased hippocampal levels of aromatase after gonadectomy might be a
compansatory mechanism in female brain and this adaptive mechanism
may not be efficient in male brain.
Experimental procedure was approved by Hacettepe University Animal
Ethics Committee (No:2006/8-6).
Paper No.: 2314
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
ALTERATIONS IN AROMATASE ENZYME LEVELS OF
OVARY, TESTIS, UTERUS AND VAS DEFERENS TISSUES OF
DIABETIC RATS
Nihan Burul-Bozkurt, C Pekiner, P Kelicen
Hacettepe University Faculty of Pharmacy, Department of Pharmacology,
Ankara, Turkey
Although diabetes mellitus (DM) is associated with increased risk of
reproductive problems in both genders, responsible molecular mechanisms under this pathophysiology are not identified yet. Aromatase catalyzes the biosynthesis of estrogens and these hormones have important
roles in reproductive processes. Insulin regulate the activity of aromatase
however there are few data about the effects of diabetes on this enzyme.
Present study was designed to investigate the effects of experimental diabetes on aromatase expression levels in ovary, testis, uterus and vas deferens tissues of rats. Rats were injected with streptozotocin to induce
diabetes. Insulin treatment was also applied. At the end of 4- and
12-weeks, rats were killed by decapitation and ovary, testis, uterus and
vas deferens tissue homogenates were prepared and evaluated for aromatase proteins by western blot. Student’s t test or one-way ANOVA was
used for statistical analysis. Aromatase expression levels in ovary were
significantly decreased both in short and long-terms of diabetes. In testis,
enzyme levels were not altered in short-term, but significantly decreased
in long-term of diabetes. In uterus and vas deferens tissues no significant
differences were observed at aromatase immunoreactivity. Insulin treatment prevented diabetes-induced changes. These results indicated for the
first time that, DM altered the expression levels of aromatase both in
ovary and testis tissues but did not affect the enzyme levels in uterus and
vas deferens tissues. According to our results, aromatase might be an
important target molecule for treatment of sexual dysfunction seen in DM.
Approved by Hacettepe University Animal Ethics Committee (No:2006/
8-6).
Nihan Burul-Bozkurt, P Kelicen, C Pekiner
Hacettepe University Faculty of Pharmacy, Department of Pharmacology,
Ankara, Turkey
Aromatase catalyzes the biosynthesis of estrogens from androgens.
Although the primary sources of estrogens are gonads they were
expressed in a variety of tissues including brain. In brain estrogens is
known to play an important role in many physiological processes and
have numerous reproductive and non-reproductive functions. They regulate gene expression, neuronal survial, neuronal and glial differantiation,
synaptic transmission in many central nervous system areas and have
neurotrophic and neuroprotective properties. This study was designed to
Paper No.: 1918
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
ANALYSIS OF NITRIC OXIDE SENSITIVE GUANYLYL
CYCLASE ACTIVATION BY CIGUATES BASED ON
MANT-GTP FLUORESCENCE
Mareike Busker, JR Kraehling, M Seeanner, S Behrends
Technical University Braunschweig, Institute of Pharmacology,
Toxicology and Clinical Pharmacy, Braunschweig, Germany
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
210
Cinaciguat and ataciguat activate nitric oxide sensitive guanylyl cyclase
(NOsGC) independently of heme. It has previously been demonstrated
that MANT-nucleotides ((N-methyl)anthraniloyl-substituted nucleotides)
inhibit nucleotide cyclases by competitive interaction with the substrate
binding site of adenylyl and guanylyl cyclases [Gille et al., J Biol Chem
2004;279:19955-19969]. In the current work we use MANT-GTP derivatives and Foerster resonance energy transfer (FRET) to probe the conformation of the active site of purified NOsGC. In direct fluorescence
experiments 2’-MANT-3’-dGTP was excited at 350 nm and fluorescence
was measured at 445 nm: Addition of the a1/b1 isoform of NOsGC
(3 lM) to 2’-MANT-3’-dGTP (3 lM) led to a small but significant
increase in fluorescence. A non-heme containing NOsGC form (a1/
b1H105A) led to a more pronounced increase in fluorescence. Addition
of the novel NOsGC activators cinaciguat or ataciguat increased 2’MANT-3’-dGTP fluorescence for the heme containing and even more for
the non-heme containing form. In a second set of experiments tyrosine
(Y) or tryptophane (W) residues of NOsGC were excited at 280 nm or
295 nm, respectively resulting in fluorescence with a maximum at
335 nm. Combination of 2’-MANT-3’-dGTP and NOsGC led to an additional peak with a maximum at 430 nm indicating the occurrence of
FRET. Preliminary evidence indicates that W669 of the a1 subunit acts
as FRET-donor. Addition of ataciguat further enhanced FRET. In summary, we show that MANT-GTP fluorescence can be used in direct
experiments and FRET experiments to detect conformational changes in
the catalytic region of NOsGC induced by novel activator drugs.
Paper No.: 1963
FOCUSED CONFERENCE GROUP: PW23 - APPLYING
PHARMACOGENOMICS (PGX) FROM RESEARCH INTO
CLINICAL PRACTICE: PRESENT AND FUTURE
THE STUDY OF CYP2C19 POLYMORPHISMS IN ROMANIAN
EPILEPTIC PATIENTS
Anca Dana Buzoianu, IC Bocsan, FC Militaru, R Popp, A Trifa,
L Perju-Dumbrava
University of Medicine and Pharmacy, Department of Clinical
Pharmacology, Cluj Napoca, Romania
Introduction. Genetic polymorphism of drug metabolizing enzymes is
defined as ability of individuals to metabolize drugs in different degrees,
due to differences in the enzyme capacity and function. CYP2C19, a
member of the cytochrome P450 enzymes family, metabolizes a range of
clinically significant drugs. Its homologous gene has been shown to be
polymorphic, two major alleles, CYP2C19*2 and *3 being responsible
for the poor metabolizer (PM) phenotype and CYP2C19*4 being associate with ultra-rapid metabolizer phenotype (pharmaco-resistant phenotype). Objective. The aim of this study is to determine the allele
frequency of CYP2C19 variants in a group of epileptic patients and to
compare it with the Romanian general population. Methods. Based on the
PCR-RFLP technique, CYP2C19*2, CYP2 C19 *3 and *4 variants were
studied in 64 epileptic patients and also in 200 healthy, unrelated Romanian volunteers. Results and conclusions: The data obtained shows that
the homozygous genotype for CYP2C19 * 2 allele, responsible for a
‘poor-metabolizer’ phenotype for CYP2C19 was met at 7.7% of patients,
compared to the general prevalence of 1.5% of the Romanian general
population. Heterozygous genotype CYP2C19 * 1 / * 2 was met in 21.5%
of patients, a frequency similar to what met in the Romanian general population (24%). CYP2C19 * 3 allele has not been seen in any patient.
Paper No.: 3201
FOCUSED CONFERENCE GROUP: P10 - DRUGS FOR HALF
THE WORLD: PAEDIATRIC CLINICAL PHARMACOLOGY
DOCUMENTATION OF RECENT DRUG HISTORY IN
CHILDREN VISITING A PEDIATRIC EMERGENCY CLINIC
Ylva Böttiger(1), E Kimland(1), S Lindemalm(2)
(1) Karolinska Institute, Department of Laboratory Medicine, Division of
Clinical Pharmacology, Stockholm, Sweden
(2) Karolinska Institute, Department of Women’s and Children’s Health,
Stockholm, Sweden
Introduction: There is a generally acknowledged lack of evidence in relation to pediatric drug treatment, and off-label treatment has been reported
to be higher in this age group, than in adults. Therefore, it should be
important to document previous use of medicines in children seeking
medical care. The aim of this study was to perform a systematic analysis
of which drugs, prescribed or OTC, and/or natural remedies children had
used prior to visiting a pediatric emergency clinic, and compare this
information with the documentation of drug use in their medical records.
Methods: A questionnaire based study was performed at a paediatric
emergency clinic during three weeks. Very urgent cases and non-Swedish
speaking patients/parents were excluded. The questionnaire was validated
through an interview with a subgroup of participants. Information concerning drug use in the medical records was later compared with the
information from the questionnaires. Results: Out of a total of 963 children who visited the clinic within the time frame, 274 children, aged
2 weeks to 18 years, mean age 3.85 years, were enrolled. Forty percent
had used prescribed drugs, and 65% of the children had used OTC drugs.
Natural remedies were used by 8.4%. In more than half of the medical
records, one or more drugs were not documented. Conclusion: Information concerning the intake of medicines or natural remedies is often missing in the medical record among paediatric patients at an emergency
clinic, which could possibly influence the safety of the patient in a negative way.
Paper No.: 1486
FOCUSED CONFERENCE GROUP: P02 - TRANSMEMBRANE
TRANSPORT: PERSPECTIVES FOR DISEASE AND DRUG
DISCOVERY
NEW NAPHTHALENE DERIVATIVES OF CITALOPRAM
DISCRIMINATE BETWEEN THE PRIMARY AND THE
ALLOSTERIC BINDING SITES AT THE HUMAN SEROTONIN
TRANSPORTER
Klaus Bøgesø(1), P Bregnedal(1), K Juhl(1), H Zhong(2), O Wiborg(3)
(1) H. Lundbeck A/S, Lundbeck Research, Valby, Copenhagen,
Denmark
(2) H. Lundbeck A/S - Paramus, New Jersey, USA
(3) Aarhus University, Centre for Psychiatric Research, Risskov,
Denmark
The human serotonin transporter (hSERT) has primary and allosteric
binding sites for escitalopram and R-citalopram. While only escitalopram
binds with nanomolar high affinity to the primary binding site the interaction of both compounds with the low affinity allosteric binding site
affect the dissociation half life of [3H]-escitalopram from hSERT. We
synthesized a series of naphthalene derivatives to explore the spatial
requirements of the binding sites and tested them for affinity to the primary binding site and for allosteric modulation of [3H]-escitalopram dissociation. The three possible derivatives with a benzene ring fused to the
isobenzofurane moiety of citalopram all showed high affinity to the primary binding site (Ki’s 4-12 nM) while their allosteric modulation were
4-11 times weaker than citalopram. The citalopram derivative with a
2-naphthylgroup replacing the 4-fluorophenylring had similar profile, and
its enantiomers showed a high stereoselectivity for the primary binding
site (Ki’s 4.1 and 113 nM, respectively) and a low stereoselectivity for
allosteric modulation (EC50’s 41 and 55 microM). The derivative with a
1-naphthylgroup was a selective allosteric modulator. The enantiomers
showed low stereoselectivity for allosteric modulation (EC50’s 14 and
19 microM, respectively). Affinities (Ki’s) for the primary site were 213
and 97 nM, respectively.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
211
Paper No.: 1293
FOCUSED CONFERENCE GROUP: P19 - GENERAL SESSION HEPATOLOGY
A STUDY ON THE PROTECTIVE MECHANISM OF TIBETAN
MEDICINE POTENTILLA ANSERINA ON EXPERIMENTAL
LIVER INJURY
Guangming Cai, Z Zhi, X Fu
Paper No.: 2900
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING BENEFITS AND MINIMIZING HARMS FROM DRUGS
INHIBITORY EFFECTS OF ORIGANUM ONITES ESSENTIAL
OIL AGAINST CONTRACTIONS OF ISOLATED RAT VAS DEFERENS INDUCED BY ANDROCTONUS CRASSICAUDA
VENOM
A Cakmak(1), F Caliskan(2), Suleyman Aydin(1)
China Military Institute of Chinese Materia Medica, 302 Military Hospital of China, Beijing, PR China
Objective: To observe the protection effects of JMS (the active ingredient
of Potentilla anserina) on the hepatic injury in mice and research the possible mechanism of the effects. Methods: Using models of hepatic injury
induced by CCL4, Con-A and DEX, the levels of AST, ALT, TNF-aIFN-b-IL-4 in blood serum and the contents of MDA and GSH-PX in
hepatic tissue were detected. Results: JMS could protect the hepatic cell
directly and reduce the activity of ALT, AST in blood serum significantly(P). Conclusion: The mechanism of JMS Protection effects on
liver-injured mice should involve influencing the liver’s metabolism,
improving capability of anti-oxidation and adjusting the immunity. JMS
could reduce the relief of free radical, cause the inhibition of apoptosis
and protect the hepatic cell by adjusting the cytokine secretion in microenvironment. It could adjust the immune function by correcting the
imbalance of Th1/Th2, so as to keep the homeostasis.
Paper No.: 1924
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
PKG MEDIATES THE TESTOSTERONE VASORELAXANT
EFFECT IN HUMAN UMBILICAL ARTERY WITHOUT
ENDOTHELIUM
Elisa Cairrão(1,2), AJ Santos-Silva(1), I Verde(1)
(1) CICS – Centro de Investigação em Ciências da Saúde, Universidade
da BeiraInterior, Covilhã, Portugal
(2) Centro Hospitalar da Cova da Beira E.P.E., Covilhã, Portugal
The cyclic nucleotides involvement in the vasorelaxation induced by
testosterone in human umbilical artery (HUA) was investigated. The
effect of this hormone on denuded HUA contracted by serotonin (5HT), histamine or KCl was analysed. Testosterone effect on potassium
current (IK) was also studied in HUA vascular smooth muscle cells
(HUASMC). The relaxant effects of Tes, sodium nitroprusside (SNP; a
soluble GC stimulator) and atrial natriuretic peptide (ANP; a particulate
GC stimulator) are similar. The Tes relaxant effect is not different to
the induced by the conjoint application of ANP and Tes. However, the
effects of SNP and Tes seems additive. The inhibition of PKG significantly reduced the Tes effect independently of the contractile stimuli.
The IK measured is mainly constituted by potassium exit through voltage sensitive potassium channels (KV) and large-conductance
Ca2 + activated potassium channels (BKCa). Testosterone significantly
activated the basal IK in HUASMC. SNP does not induce a significant
modification in basal or testosterone stimulated IK. In contrast, ANP
stimulates the basal IK, but does not increase the testosterone stimulation on IK. The activations of the IK induced by testosterone or by
ANP are not significantly affected by the PKA inhibitor, but are completely inhibited by the PKG inhibitor. Our results show that testosterone and ANP share the same pathway to increase the cGMP and to
induce vasorelaxation, and suggest that Tes increases cGMP levels by
activating particulate GC.
We thank the Gynaecology-Obstetrics Department staff of CHCB and
FCT which supports the fellowship SFRH/BDE/15532/2004.
(1) Anadolu University Faculty of Pharmacy Department of Pharmacology, Eskisehir, Turkey
(2) Eskisehir Osmangazi University Faculty of Science and Letters,
Department of Biology, Eskisehir, Turkey
Androctonus crassicauda is one of the most toxic venomous scorpion
species of the world and many people die due to the envenomation by
this scorpion. Origanum onites is a a plant used for various medicinal
purposes including as antiseptic, antimicrobial and an antidote for toxic
envonamations. The purpose of this study is to investigate the effect of
O. onites essential oil on the contracting effects of A. crassicauda venom
on isolated rat vas deferens. A.crassicauda venom was obtained from
specimens of South-east Turkey and O.onites essential oil, from the distillation of the plants collected from west Anatolia. Our experiments
showed that the contraction obtained by A. crassicauda venom on the
vas deferens was antagonized and relaxed by the application of 10-6,
10-5 and 10-4 M doses of O. onites essential oil. In our study it was
shown that there is an interesting antagonistic interaction between
Origanum onites essential oil and the venom of Androctonus crassicauda.
Our results indicate the possibility of O.onites as a natural antidote
against Androctonus crassicauda which requires new and detailed in vivo
investigations.
Paper No.: 1778
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
TODAY’S USE OF MEDICINAL NATURAL PRODUCTS BY
ITALIAN WOMEN IN SICILY
Gioacchino Calapai, A Pieratti, D Crupi, A De Sarro
University of Messina, Department of Experimental and Clinical Medicine and Pharmacology, Messina, Italy
Introduction: Women are the most consumers of medicinal natural products They use them for typical women disorders (menopausal symptoms
and menstrual disturbances) and eventually for pregnancy and lactation
problems. We propose a survey on their use by sicilian women. Methods:
Data on use of natural medicinal products (such as food supplements,
herbal medicines, omeopathics) and synthetic drugs in either gender diseases or not are collected through interview by 1000 north east sicilian
women aging 15-85 years. To get information about use of drugs and
medicinal natural products a questionnaire is administered in ten different
consulting health units for women. A second way of acting is through
phone interviews. Obtained data are collected in a data-base, and successively statistically analyzed; data on undesired reactions are also collected. Preliminary results: About one third of the interviews (314) were
collected and they show that principally synthetic drugs is used by
74.5%; principally natural medicinal products by 14.5%; we had no
answer by the remaining group of women. Natural products use increases
with the age with maximum use starting from 40 years of age women
and beyond. The major part of women use them without knowledge of
their side effects. The indications most cited are menopause, water retention, stipsis, sleep disturbances, venous insufficiency. Conclusions: Use
of medicinal natural products by women of our region is increased but
safety data (warnings, side effects and contraindications) are still poorly
known by consumers.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
212
Paper No.: 1868
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
THE INVERSE AGONIST EFFECT OF RIMONABANT IN
BRAIN MEMBRANES IS NOT MEDIATED BY THE CB1
RECEPTOR
ing low CNS penetration of darifenacin and 5-HMT (limited by PGP
efflux), and trospium (limited by permeability and PGP efflux). Consistent with these results in vitro, penetration in vivo in rat brain and CSF
was significant for oxybutynin, solifenacin and tolterodine while trospium, darifenacin and 5-HMT had no significant CNS penetration. These
differences in CNS penetration may contribute to different CNS side
effect profiles across the OAB agents.
Luis F Callado, AM Erdozain, R Diez-Alarcia, JJ Meana
University of the Basque Country, Department of Pharmacology, and
Centro de Investigación Biomédica en Red de Salud Mental
(CIBERSAM), Leioa, Spain
Previous data suggest that rimonabant behaves as an inverse agonist in
different biological systems Our aim was to characterize the inhibitory
effects of rimonabant on G-protein activation in postmortem human brain
membranes In [35S]GTPcS binding assays performed in human brain
cortical membranes, increasing concentrations of rimonabant blocked the
effect evoked by WIN55,212-2 (10lM) and inhibited the basal binding
of [35S]GTPcS O-2050 also blocked the effect mediated by WIN55,2122 (1lM) but barely affected basal binding of [35S]GTPcS Neither O2050 (10 and 100lM) nor WIN55,212-2 (10lM) were able to modulate
the inhibition of the basal binding of [35S]GTPcS produced by rimonabant [35S]GTPcS binding assays performed in mouse brain revealed that
rimonabant inhibited the basal binding of [35S]GTPcS in the same manner in wild type and CB1 knock out tissue In binding saturation assays
in postmortem human brain membranes, [3H]rimonabant showed a KD
of 58 ± 11 nM The [3H]rimonabant specific binding at a fixed concentration of 5 nM was only reduced by 60% in the presence of O-2050
(10lM) These data suggest that the apparent inverse agonist effect of
rimonabant over G-protein activation is not be mediated by the cannabinoid CB1 receptor In addition, rimonabant recognizes a second binding
site distinct from cannabinoid CB1 receptors
Paper No.: 2730
FOCUSED CONFERENCE GROUP: P08 - DEVELOPMENTS IN
THE TREATMENT OF SEXUAL DYSFUNCTION AND
DISEASES OF THE LOWER URINARY TRACT
COMPARISON OF CENTRAL NERVOUS SYSTEM
PENETRATION OF ANTIMUSCARINIC DRUGS FOR
OVERACTIVE BLADDER
E Callegari(1), R Webster(2), K Fenner(2), J LaPerle(1), B Malhotra(3),
Peter Bungay(2)
(1) Pfizer Inc, Groton, CT, USA
(2) Pfizer Inc, Sandwich, UK
(3) Pfizer Inc, New York, NY, USA
Common side effects of antimuscarinics for the treatment of overactive
bladder (OAB) symptoms are peripheral (dry mouth and constipation)
while central side effects like cognitive dysfunction may be seen depending on muscarinic receptor subtype affinity and/or central nervous system
(CNS) penetration of specific agents. CNS penetration was evaluated in
nonclinical models in silico, in vitro and in vivo for common OAB drugs:
oxybutynin, trospium, tolterodine, solifenacin, darifenacin and fesoterodine (including its active metabolite, 5-hydroxymethyl tolterodine
(5-HMT)). The molecular and physicochemical properties, cross-membrane permeability (using RRCK cells from Madin-Darby Canine Kidney cell line) and P-glycoprotein (PGP) transporter interaction of each
agent were evaluated as predictors of CNS penetration. Plasma, brain
and CSF concentrations were determined following subcutaneous administration of each agent to rats. Fesoterodine, a prodrug of 5-HMT, is not
detectable in systemic circulation, and was not included in experiments
in vivo. Oxybutynin, solifenacin and tolterodine were not substrates for
PGP, demonstrated high permeability in RRCK cells, and possessed
physicochemical properties predicting moderate-high CNS penetration.
Trospium, 5-HMT and darifenacin were each PGP substrates and demonstrated low, medium and high RRCK permeability, respectively, predict-
Paper No.: 3367
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
ANTI-INFLAMMATORY EFFECTS OF THE PENTACYCLIC
TRITERPENES A,B-AMIRYNS ON ANIMAL MODELS OF
IRRITATIVE CONTACT DERMATITIS INDUCED BY
TETRADECANOYLPHORBOL 13-ACETATE
Iana Calou(1), D Gonçalves(1), T Melo(1), T Olinda(1), GA Brito(1), V
Rao(1), M Chaves(2), F Santos(1)
(1) Federal University of Ceará, Departement of Physiology and Pharmacology, Fortaleza, Brazil
(2) Federal University of Piauı́, Departament of Organic Chemistry, Teresina, Brazil
The pentacyclic triterpenes a,b-amiryns (AB), isolated from the resin of
Protium heptaphyllium March (Burseraceae), were evaluated on acute
mouse ear dermatitis induced by a single application of tetradecanoylphorbol 13-acetate (TPA, 2.5 mg / ear). The anti-inflammatory actions
were accessed by measurements of ear edema, releasing of myeloperoxidase, tissue levels of TNF-a and histological study. Male Swiss mice,
20-25g (n = 8/group) were treated topically with vehicle (2% Tween 80
or only topical acetone), AB (0.125, 0.25 and 0.5 mg/10 microliters/ear)
or dexametasone (0.05mg/ 10 microliters/ear). The animals received
simultaneously, TPA (2.5 mg in acetone/10 microliters/ear). The ear
edema was recorded by a digital caliper (100.174B/DIGIMESS) before
and 4 hours after administration of TPA. After determining the swelling,
ear tissue was collected for determination of the myeloperoxidase’s
release, evaluation of TNF-a’s tissue levels and preparation of slides for
histological study. Tests were considered significant at p < 0.05. The
group treated with AB (0.125, 0.25 and 0.5 mg) has significantly
reduced the edema to 0,0920 ± 0,01172, 0,0500 ± 0,005375 and
0,0340 ± 0,005207 respectively, when compared to TPA (0,1110 ±
0,01303 mm). Dexametasone significantly reduced the edema induced
by TPA (0,0140 ± 0,002211 mm). The AB 0.125, 0.25 and 0.5 mg/ear
significantly reduced the release of myeloperoxidase in 51,47%, 71,28%
and 81,18% respectively, considering the TPA activity 100%. A considerable reduction (48,43%) in the levels of TNF-a was observed when
animals received a topical dose of AB (0.5 mg /ear). Reductions of dermal edema and inflammatory cell infiltration was observed on animal
treated with AB (0.5 mg /ear).
Paper No.: 3421
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
THE ANTI-INFLAMMATORY EFFECT OF THE
PENTACYCLIC TRITERPENES A AND B AMIRYNS ON
ANIMAL MODEL OF DELAYED HYPERSENSITIVITY
INDUCED BY REPEATED APPLICATIONS OF OXAZOLONE
Iana Calou(1), D Gonçalves(1), T Olinda(1), T Melo(1), I Figueiredo(1),
GA Brito(1), M Chaves(2), V Rao(1), F Santos(1)
(1) Federal University of Ceará, Department of Physiology and Pharmacology, Fortaleza, Brazil
(2) Federal University of Piaui, Department of Organic Chemistry,
Teresina, Brazil
The pentacyclic triterpenes a and b amiryns (AB), isolated from the gum
of Protium heptaphyllium March, were tested in a model of delayed
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
213
hypersensitivity induced by repeated applications of oxazolone (OXA)
1% (20 microliter/ear every 3 days for 19 days). The parameters used to
measure the anti-inflammatory activity of AB were epidermal hyperplasia, tissue levels of interferon c (INF-G) and histological studies. We
used Swiss mice, 20-25g, male (n = 8). The animals were sensitized with
oxazolone (2% in acetona/50 microliter) topically on the abdomen for
two consecutive days. After six days (considered day 1 of treatment)
solution of 1% oxazolone (20 microliter) in acetone and olive oil (4:1)
was topically applied on the ear every three days for 19 days. AB was
administered topically at doses of 0,125; 0,25 and 0,5 mg/20 microliter/
ear 30 minutes before and three hours after administration of OXA. The
ear thickness was recorded every 3 days before AB application until the
end of the protocol. The hyperplasia was recorded by a digital caliper.
For all tests were considered significant p < 0.05. On 19th day, AB
(0,125, 0,25 and 0,5) reduced the epidermal hyperplasia in a significant
way (37,800 ± 1,15300; 34,600 ± 0,80550 and 32,400 ± 1,24900,
respectively) when compared to OXA (44,430 ± 0,99660). At a dose of
0,5 mg/20 microliter/ear, AB reduced in a significant way tissue levels of
INF-G (19,37 ± 10,23) when compared to OXA (477,2 ± 81,50). Histopathological analysis revealed a marked decrease in epidermal hyperplasia and neutrophil infiltration in animals treated with AB (0,5mg/20
microliter/ear).
Paper No.: 718
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
PIROXICAM PROVIDES ADEQUATE ANALGESIA AFTER
IMPACTED LOWER THIRD MOLAR REMOVAL
REGARDLESS OF THE SURGEON’S EXPERIENCE
Adriana Calvo, T Dionı́sio, F Giglio, P Gonçalves, C Santos,
University of São Paulo Bauru School of Dentistry, Department of Pharmacology,São Paulo, Brazil
The aim of this study was to compare the postoperative pain control in
volunteers medicated with piroxicam who underwent impacted lower
third molar removal performed by three surgeons with distinct experiences (a specialist, a PhD student and a senior dental student of the
Bauru School of Dentistry). Fifty volunteers underwent impacted lower
third molar removal under local anesthesia. Piroxicam (20 mg, once
daily) was prescribed to all patients for four days after the surgical procedure, and paracetamol was provided as rescue medication. The volunteers operated by the senior dental student reported higher pain scores at
3, 4, 7, 8, 10, 12, 18, 24 and 48 h in comparison to the patients operated
by the PhD student (p<.05). The volunteers who underwent surgical procedure performed by the PhD student reported lower pain scores at 8 h
in comparison to those whose surgeries were performed by the specialist
(p<.05). All pain scores were lower than 30 mm, which represents a minimal discomfort for all volunteers. Accordingly, volunteers operated by
the three surgeons used the same small amount of rescue medication
(2,159 ± 192.60mg for the specialist; 2,300 ± 208.3mg for the PhD student and 2,506 ± 228.2mg for the senior dental student). These results
suggest that piroxicam provided adequate pain control after impacted
lower third molar removal regardless of the surgeon’s experience.
Paper No.: 803
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
RAT KIDNEY ANTIOXIDANT ENZYMES IN AN EXPERIMENTAL MODEL OF PREECLAMPSIA
Elsa Camacho, J Silva, MG Matos, M Varela, MR Garrido, A Israel
Central University of Venezuela, Caracas, Venezuela
Preeclampsia (PE) is a multifactorial disorder of pregnancy, characterized
by hypertension, proteinuria, edema, endothelial dysfunction, dyslipidemia and oxidative stress. It has been suggested that nitric oxide (NO) is
the most important mediator for the reduction of peripheral vascular
resistance in normal pregnancy, and it is know that this response is
impaired in preeclampsia. The chronic inhibition of NO synthesis in
pregnant rats causes a syndrome similar to preeclampsia which causes
hypertension, proteinuria, thrombocytopenia and intrauterine growth
retardation. In order to establish the role of oxidative stress in this model
of PE, we assessed the activity of antioxidant enzymes: catalase (CAT),
superoxide dismutase (SOD) and glutathione peroxidase (GPX) in the rat
kidney. Pregnant female Sprague-Dawley rats, with 13 days of gestation
were divided into two groups: control (NaCl 0.9%, 7 days) and
L-NAME (50 mg/kg/day, 7 days). Rats were sacrificed by decapitation,
the fetuses and the kidneys were extracted. CAT, SOD, GPX activity was
determined by spectrophotometry and nitric oxide synthase (NOS) by the
convertion of radiolabeled L-arginine to L-citruline. Our results demonstrate that treatment with L-NAME in pregnant rats increased MAP
(+20 mmHg), induced proteinuria and decreased the number and weight
of fetuses. In addition, there was a significant reduction in renal CAT,
SOD, GPX and NOS activities. These findings indicate the existence of
an alteration in the renal antioxidant enzymes activity associated with the
chronic inhibition of NOS in experimental preeclampsia, and suggest a
role of oxidative stress in PE.
Paper No.: 1365
FOCUSED CONFERENCE GROUP: P14 - ADDICTION AND
DOPING: NEUROBIOLOGICAL AND CLINICAL
MEMANTINE PREVENTS THE COGNITIVE DEFICITS
INDUCED BY METHAMPHETAMINE IN RATS
Jorge Camarasa, T Rodrigo, D Pubill, E Escubedo
University of Barcelona, Department of Pharmacology and Therapeutic
Chemistry, Barcelona, Spain
Methamphetamine (METH) is a street drug abused by young people.
This study seeks to determine whether pre-treatment with memantine
(MEM) counteracts the memory impairment induced by METH in
rodents. In mice, both METH (1 mg/Kg) and MEM (5 mg/Kg) induced
a locomotor stimulant response (AUC saline: 72887 ± 6909, MEM:
280605 ± 25130, P < 0.001, METH: 256680 ± 28431, P < 0.001).
Non-spatial memory was assessed in the Object Recognition Test. MEM
pre-treatment recovered the ability of rats to discriminate between a
familiar and a novel object, which had been abolished by the METH
treatment. METH-treated rats showed impaired learning in the Morris
water maze. The escape latency time was shorter in animals treated with
saline (F2,58 = 5.434, P < 0.01), MEM (F2,123 = 4.402, P < 0.01), or
MEM+METH (F2,213 = 7.263, P < 0.001) as trials progressed. In contrast, METH-treated rats showed a higher escape latency (P < 0.05) than
saline-treated animals. Probe trial demonstrated that animals treated with
saline or MEM spent longer in the target quadrant than in the opposite.
Moreover, MEM alone induced an improvement of learning (time spent
in the target: 25.82 ± 2.44 s and opposite: 7.60 ± 1.30 s). METH- treated
rats showed a cognitive deficit (time in the target: 17.40 ± 1.23 s and
opposite: 14.62 ± 2.10 s, not different from random: 15.00 s). MEM pretreatment prevented this deficit (23.58 ± 3.22 s, target and 11.14 ± 2.35
s, opposite). MEM constitutes the first successful approach to prevent the
cognitive deficits induced by amphetamine derivatives, irrespective of
their neurochemical profile.
Paper No.: 2209
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
MODULATION OF LIGAND BINDING TO THE ALPHA1B
ADRENERGIC RECEPTOR: A POTENTIAL ROLE FOR THE
SECOND EXTRACELLULAR LOOP
Adrian Campbell(1), I McDougal(2), R Griffith(0), A Finch(1)
(1) University of New South Wales, School of Medical Sciences, Department of Pharmacology, Kensington, NSW, Australia
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
214
(2) University of Newcastle, School of Environmental and Life Sciences,
Callaghan, NSW, Australia
Activation of the a1B adrenergic receptor (AR) has previously been attributed to the disruption of an interhelical salt-bridge between an aspartate in
transmembrane helix 3 (TMIII) and a lysine in TMVII (Porter JE et al, J.
Biol. Chem. 1996; 271: 28318-23. Our homology model of the a1B AR
based on the crystal structure of bovine rhodopsin (1U19) identified a
putative salt-bridge between the same lysine (K331) and a second extracellular loop (ECL2) aspartate (D191). We propose that the salt-bridge
between TMVII and ECL2 stabilises an active state of the a1B AR. To test
this hypothesis, we created alanine mutants at positions 191 and 331 to
characterise and compare to wild type (WT) receptor and Porter et al’s
previously described mutants. Saturation data shows no significant difference in the binding affinity of 3H-prazosin (WT: 295.9 ± 199.4; D125A:
no binding; K331A: 248.9 ± 117.4; D191A: 482.9 ± 94.5; D191A/
K331A: 372.7 ± 96.7 pM, n = 4). Competition binding experiments
show a significant increase in logKi for (-)epinephrine for D191 mutants
(WT: -5.624 ± 0.03; K331A: -5.641 ± 0.03; D191A: -4.854 ± 0.06;
D191A/K331A: -4.953 ± 0.12 M, n = 3, P < 0.001). Functionally, no
mutants have a significantly different logEC50 compared to WT (WT:5.534 ± 0.21; K331A: -5.887 ± 0.15; D191A: -5.534 ± 0.25; D191A/
K331A:-5.627 ± 0.11 M, n = 3). Further studies will determine the
effects of these mutations on antagonist inhibition of receptor activation.
The apparent conflict between binding and activation data for (-)epinephrine suggests that in the absence of the charge on D191, the receptor is
biased towards the inactive state, decreasing affinity for (-)epinephrine,
but once bound, the receptor can still adopt the active state.
Paper No.: 519
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
AN OVERVIEW ON ATTENUATION OF DRUG-INDUCED
NEPHROTOXICITY
Guoying Cao, X Hu
Beijing Hospital, Department of Clinical Pharmacology, Beijing, PR
China
Amino glycoside antibiotics, anti cancer drugs, immunosuppressant,
radio contrasts and TCM are associated with nephrotoxicity. Druginduced renal dysfunction generally induces a lysosomal phospholipidosis which is accompanied by cellular necrosis and postnecrotic cell
regeneration. Drug toxicity remains an important cause of acute kidney
injury that, in many circumstances, can be prevented or at least minimized by early intervention. Hydration therapy and structure optimization are widely used in preventing nephrotoxicity of cisplatin while preor coadministration of inhibitors are still questionable. In this article we
review the most prevalent drugs which induce nephrotoxicity, and from
the point of clinical and laboratory research view, analyze the available
evidence for preventive measures.
Paper No.: 2387
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
INCREASED SENSITIVITY OF CEREBRAL AND CORONARY
ARTERIES TO GPCR AGONISTS IN SPONTANEOUSLY
HYPERTENSIVE RATS
Lei Cao(1), Y Zhang(1), Y-X Cao(2), L Edvinsson(1), C-B Xu(1)
(1) Lund University, Institute of Clinical Science, Division of Experimental Vascular Research, Lund, Sweden
(2) Xi’an Jiaotong University College of Medicine, Department of Pharmacology, Xi’an, Shaanxi, PR China
Hypertension is a major risk factor closely associated with myocardial
infarct and stroke. However, the underlying molecular mechanisms
describing how hypertension leads to vascular damage are not fully
understood. Vascular G-protein coupled receptors (GPCR) mediate vascular smooth muscle contraction and proliferation, which contribute to
narrowing of vascular lumen. The present study was designed to examine
the hypothesis that in hypertension, there is increased sensitivity of cerebral and coronary arteries to vasoconstrictors that activate GPCR. Cerebral and coronary arteries were removed from spontaneously
hypertensive (SHR, n = 8) and normal tension (n = 14) rats. Contractile
responses to GPCR agonists (U46619, thromboxane A2 receptor agonist;
sarafotoxin 6c, endothelin type B receptor agonist and endothelin-1,
endothelin type A and B receptor agonist) were recorded by a sensitive
myograph. Real-time PCR was used to semi-quantify GPCR mRNA
expression. The results show that the contractile responses to U46619,
sarafotoxin 6c and endothelin-1 in cerebral and coronary arteries were
increased in SHR, compared with the arteries from normotensives. In
parallel, there were increased mRNA expressions for thromboxane A2,
endothelin type A and B receptors, suggesting involvement of transcriptional mechanisms. In conclusion, the present study suggests that
increased vascular GPCR expression in hypertension is a mechanism that
may in part mediate the increased sensitivity of cerebral and coronary
arteries seen in hypertension.
Paper No.: 2388
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
LONG-TERM CIGARETTE SMOKE EXPOSURE INDUCES
BRONCHIAL HYPERRESPONSIVENESS TO G-PROTEIN
COUPLED RECEPTOR AGONISTS
Lei Cao(1), Y Zhang(1), Y-X Cao(2), L Edvinsson(1), C-B Xu(1)
(1) Lund University, Institute of Clinical Science, Division of Experimental Vascular Research, Lund, Sweden
(2) Xi’an Jiaotong University College of Medicine, Department of Pharmacology, Xi’an, Shaanxi, PR China
Cigarette smoke exposure strongly associates with airway hyperreactivity (AHR) and inflammation. The present study was designed to examine if G-protein coupled receptors (GPCR), inflammatory mediators and
Raf-1 activity are involved in the smoke-induced airway pathogenesis.
SD rats (200-300g) were divided into three groups (control, smoke
exposure and GW5074 treatment; n = 10). In smoke exposure group,
rats were exposed to cigarette smoke generated from 10 cigarettes/day
for 8 weeks. Control rats received fresh air only. GW5074 (a Raf-1
inhibitor) treatment group was exposed to the same amount of smoke
and treated by intraperitioneal administration of GW5074 (0.5mg/kg)
every day for 8 weeks. At the end of the experiments, bronchi were
isolated from all groups and cut into ring segments. Contractile
responses of bronchial segments were recorded by a sensitive myogragh. The mRNA expression for GPCR and inflammatory mediators
were semi-quantified by real-time PCR. The results show that after
8 weeks of smoke exposure, the airway exhibited increased contractile
responses to carbachol, 5-HT, sarofotoxin 6c and endothelin-1, which
was parallel with enhanced mRNA expression for 5-HT2A, endothelin
type A and B receptors. In addition, there was increase in mRNA
expression of inflammatory mediators (IL-1beta, TNF-a and ICAM-1)
in smoke group. Treatment with GW5074 throughout the smoke exposure period abrogated the smoke-induced AHR and inflammation. In
conclusion, long-term cigarette smoke exposure results in airway
inflammation and AHR to GPCR agonists. Inhibition of Raf-1 activity
might provide new strategies for the treatment.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
215
Paper No. 1222
FOCUS GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD DISEASES
MITIGATION EFFECT OF ESCULENTIC ACID ON
RADIATION-INDUCED PNEUMONIA AND PULMONARY
FIBROSIS
Yong-Bing Cao(1), S-M Yang(2), Y-P Tian(1), M Zhang(2), L Zhang(3),
K-Z Zhang(2), L-J Yin(2), B-R Zhang(2), R Howell(2), Y-Y Jiang(1) P
Okunieff(2) L Zhang(2)
(1) Second Military Medical University School of Pharmacy, Department
of Pharmacology, Shanghai, PR China
(2) University of Rochester Medical Center, Department of Radiation
Oncology, Rochester, NY, USA
(3) West China Hospital of Sichuan University, Chengdu, Sichuan, PR
China
The effect of Esculentic acid (EA) on radiation-induced pneumonia and
pulmonary fibrosis and its possible molecular mechanisms were investigated in lung irradiated mouse model. C57BL/6 mice (5-10/group) were
received 15 Gy ionizing radiation (IR) on whole lung. Thirty minutes
late, the mice treated with vehicle alone (as control group) or 10 mg/kg
Esculentic Acid (as test group) orally every other day for days (pneumonia group) or up to 3 months (fibrosis group). At the different time points
post IR (2.5, 17 days for pneumonia group, and 7 months for fibrosis
group), the mice were sacrificed, the BAL (bronchial alveolar lavage),
plasma and lung tissue were collected and assess for the alterations in
BAL cell counts and differentiation, cytokines and lung fibrosis with
means of microscope, ELISA or pathological staining. The lung function
was assessed by the breath rate and compliance. The results showed that:
1) EA inhibited neutrophil in BAL 17 days post-radiation; 2) EA
reduced IR-induced P-selectin, PF-4, sTNFR-1, Lymphotaxin on day 2.5
and 17; the IR-induced IL-1b,TNF-a and VCAM-1 were also reduced
on day 2.5; 3) the EA inhibit lung acute and sub-acute inflammatory
response induced by radiation; 4) EA reduced the breath rate and
increased the lung compliance 7 months post-radiation; and 5) EA
decreased the amounts of collagen deposited in lung tissue as compared
to the vehicle alone treated mice. In conclusion, the EA mitigates
IR-induced pneumonia and pulmonary fibrosis via inhibition of the productions of several pro-inflammatory cytokines and inflammatory mediators.
Paper No.: 823
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD DISEASES
REMICADE ALLEVIATES AIRWAY INFLAMMATION AND
HYPERREACTIVITY IN ASTHMATIC E3 RATS
epithelial damage. Serum levels of IgE increased, and so did the levels
of NO, iNOS, TNF-a, and IL-4, IL-5, IL-13 in lung homogenate and
serum. Further, the contractile responses in bronchi induced by endothelin-1, sarafotoxin 6c and bradykinin increased, and isoprenalineinduced relaxations decreased. All these changes induced by the sensitization procedure were reduced by the remicade treatment. The results
suggested that Remicade prevented the development of local airway
inflammation and antagonized changes of the bronchial smooth muscle
receptor phenotype, thereby blocking the development of airway hyperreactivity of asthmatic rats.
Paper No.: 2668
FOCUSED CONFERENCE GROUP: P11 - G PROTEINCOUPLED 7TM RECEPTORS: FROM MOLECULAR TO
PHYSIOLOGICAL FUNCTION
MMLDL UPREGULATES ENDOTHELIN TYPE A AND TYPE B
RECEPTORS IN RAT BASILAR ARTERY
Yong-Xiao Cao(1), J Li(1), ZY Yuan(1), L Cao(2), CB Xu(2)
(1) Xi’an Jiaotong University College of Medicine, Department of
Pharmacology, Xi’an, PR China
(2) Lund University, University Hospital of Lund, Institute of Clinical
Science, Division of Experimental Vascular Research, Lund, Sweden
Mminimally modified low density lipoprotein (mmLDL) can induce an
inflammatory response and enhance plaque formation in arteries. Vascular endothelin type A (ETA) and type B (ETB) receptors are implicated
in the pathogenesis of cardiovascular diseases. The present study was
designed to test if mmLDL upregulates ET receptors in rat basilar artery.
Rat basilar artery ring segments were cultured with mmLDL 10 g/ml for
24 h. Tension of the ring segments was recorded by a sensitive myogragh. The mRNA expressions and proteins of ETA and ETB receotors
were semi-quantified by real-time PCR and western blot techniques,
respectively. The results showed that organ culture with mmLDL
increased the contractions induced by sarafotoxin 6c and endothelin-1
and enhanced the expression of mRNA and proteins of ETB and ETA
receptors in in rat basilar artery, suggesting mmLDL up-regulated ETB
and ETA receptors. The general PKC inhibitors, staurosporine, prevented
the upregulation. MAPK inhibitors for extracellular signal related kinases
1 and 2 (ERK1/2), SB386023 and U0126, p38 MAPK, SB203580 inhibited the upregulation of ETB and ETA receptors. C-jun terminal kinase
(JNK), SP600125 and NF-jB, wedelolactone inhibited the upregulation
of ETB. In conclusion, mmLDL up-regulates ETB and ETA receptors in
rat basilar artery. PKC, MAPK and NF-jB signalling pathways seem
involved in the up-regulations.
Yong-Xiao Cao(1), Y Cai(1), SM Lu(1), CB Xu(2), LO Cardell(3)
(1) Xi’an Jiaotong University College of Medicine, Xi’an, Shaanxi, PR
China
(2) MedDepartment of Experimental Vascular Research, Institution of
Medicine, Lund University, Sweden
(3) Department of Otorhinolaryngology, Malmö University Hospital,
Malmö, Sweden
Tumour necrosis factor-a (TNF-a) has been implicated in pathogenesis
of asthma, and neutralization of TNF-a is an effective therapy for inflammatory diseases. The present study tested the idea that a TNF-a antibody,
remicade, may be useful in the management of asthma. E3 rats were sensitized to OVA/alum and received remicade intraperitoneally. Two weeks
later OVA-PBS was intranasally instilled daily for 7 days. Bronchoalveolar lavage fluids (BALF), serum and lung homogenates were collected
for analysis of cells and inflammatory mediators. Contractile responses
of lobar-bronchus segments to agonists were tested. Pulmonary tissues
were investigated using histological examination. The results showed that
the sensitized ‘‘model E3 rats’’ exhibited an increase of the total amount
of cells, primarily eosinophils in BALF and pulmonary tissue, and
Paper No.: 420
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
INVESTIGATION OF THE RELATIONSHIP BETWEEN
PLASMA L-CARNITINE CONCENTRATION AND ITS
ANTIOXIDANT ACTIVITY AFTER SINGLE ORAL
ADMINISTRATION OF L-CARNITINE IN HEALTHY
VOLUNTEERS
Yu Cao(1), Z-W Han(1), HJ Qu(1), C-B Wang(2)
(1) The Affiliated Hospital of Medical College Qingdao University,
PharmacyDepartment, Qingdao, PR China
(2) Department of Pharmacology, Medical college Qingdao University,
Qingdao, PRChina
Introduction: This study was to investigate the plasma concentration
of L-carnitine (LC) after a single oral administration of LC solution in
healthy volunteers and its antioxidant activity. The plasma antioxidant
enzymes and total antioxidant were evaluated through the measure-
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
216
ment of SOD, GSH-Px, CAT and T-AOC activities in plasma, and
evaluate the relationship between concentration and antioxidant index
in plasma of healthy humans. Materials/Patients: Liquid LC (2.0g)
was administered orally as a single dose in 12 healthy subjects.
Plasma concentration of LC was detected by HPLC in 24h, and the
plasmas were subjected to the measurement of SOD, GSH-Px, CAT
and T-AOC activities by spectrophotometric methods to evaluate the
relationship between concentration and antioxidant activity of LC in
plasma. Results: Antioxidant enzymes,total antioxidant and plasma
concentration increased gradually from 0h to 3.5h point and reached
the peak at 3.5h,then go down gradually, and were as the same level
as the beginning(0h point) at 24h point. Correlation analysis was
studied between plasma concentration and antioxidant activity through
linear regression. The regressive equations were Y = 0.5319X+3.2429
(r = 0.9922), Y = 0.1606X+103.63(r = 0.9316), Y = 0.019X+0.3845
(r = 0.9720),Y = 0.1346X+5.4452 (r = 0.9338) respectively between
LC plasma concentration and antioxidant index (SOD, GSH-Px, CAT,
T-AOC). The antioxidant activity of LC was significantly positive
correlation with plasma concentrations of LC after single oral administration of L-carnitine in healthy volunteers. Conclusion: Administration
of liquid LC could raise the activities of antioxidant enzymes and total
antioxidant capacity in a concentration-dependent manner of LC in
plasma.
Paper No.: 2373
FOCUSED CONFERENCE GROUP: P17 - NEW APPROACHES
AND TARGETS IN PSYCHIATRY
RESCUE OF TGF-B1 SIGNALLING AS A NEW
NEUROPROTECTIVE TOOL IN ALZHEIMER’S DISEASE
Filippo Caraci(1), G Battaglia(2), ML Giuffrida(1), V Bruno(2),
P Bosco(3), MA Sortino(4), F Drago(4), F Nicoletti(2), A Copani(1)
(1) University of Catania, Department of Pharmaceutical Sciences, Catania, Italy
(2) I.N.M. Neuromed, Pozzilli, Italy
(3) IRCCS Oasi Maria S.S, Troina, Italy
(4) University of Catania, Department of Experimental and Clinical Pharmacology,Catania, Italy
Introduction: Transforming growth factor-b1 (TGF-b1) is a neurotrophic
factor that exerts neuroprotective effects in different models of neurodegeneration, including Ab toxicity (Vivien D & Ali C. Cytokine &
Growth Factor Reviews 2006; 17:121-28). Nevertheless the molecular
mechanisms underlying the neuroprotective effects of TGF-ß1 against
Ab-induced neurodegeneration are presently unknown. Materials and
results: We examined the neuroprotective activity of TGF-b1 in pure cultures of rat cortical neurons challenged with synthetic Ab. Ab induced
the apoptosis of about 30-40% of the total neuronal population. TGF-b1
was equally protective when added either in combination with, or
6 hours after Aba. Co-added TGF-b1 prevented Ab-induced cell cycle
reactivation, whereas lately added TGF-b1 rescued late b-catenin degradation and s hyperphosphorylation. Interestingly, lithium, a mood stabilizer that protects against Ab-induced neurodegeneration, induced both
the synthesis and the release of TGF-b1 from cultured astrocytes. We
also assessed the neuroprotective activity of endogenous TGF-b1 in vivo.
Ab injection into the rat dorsal hippocampus had a small neurotoxic
effect that was amplified by i.c.v. injection of SB431542, a selective
inhibitor of TGF-b1 receptor, thus suggesting that endogenous TGF-b1
acts as a factor limiting Ab toxicity. Conclusions: Our data demonstrate
that TGF-b1 is neuroprotective in vitro and in vivo against Ab-induced
neurodegeneration, and suggest that drugs able to increase TGF-bb1,
such as lithium, might represent a new neuroprotective tool for the treatment of AD.
Paper No.: 2359
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
CORRELATION BETWEEN AMINE OXIDASES KINETIC
PARAMETERS AND NITRIC OXIDE PRODUCTION IN
HUMAN MESENTERIC ARTERIES
Maria Margarida Caramona(1), S Nunes(1), J Pereira(2), C Lopes(1), I
Figueiredo(1)
(1) University of Coimbra Faculty of Pharmacy, Laboratory of Pharmacology, Coimbra, Portugal
(2) Portuguese Oncology Institute, Coimbra, Portugal
There are evidences suggesting that the enzymes monoamine oxidase
(MAO, type A and B), semicarbazide-sensitive amine oxidase (SSAO)
and nitric oxide sintase (NOS) play a broad role in the diabetes1,2. However, it is not known the correlation between NO production and the
activity of the enzymes MAO and SSAO. Therefore, the aim of this
work was to study the correlation between the nitrite and nitrate (NOx)
levels and the kinetic parameters of these enzymes in human inferior
mesenteric arteries (n = 12). The NOx concentrations were determined
by Griess reaction and the kinetic parameters were evaluated by radiochemical assay. We found that the NOx levels (57.24 ± 19.60lmol/l))
were positively correlated with MAO-B parameters: Km (r = 0.612,
p = 0.034) and Vmax (r = 0.593, p = 0.042), but it was negatively correlated with SSAO parameters: Km (r= - 0.625, p = 0.029) and Vmax (r=
-0.754, p = 0.0046). In conclusion, our results demonstrate that the correlation between NOx levels with MAO B and SSAO could be a finding
with relevance in some diseases, such as type 2 diabetes, and they are
the first step for therapeutic strategies using specific inhibitors of these
enzymes.
1 Irer SV et al, Neurotoxicology 2007; 114:811-815.
2 Nunes SF et al, Acta Diabetol 2009; 46: 135-140.
Supported by grant SFRH/BD/23606/2005 from FCT-Portugal.
Paper No.: 2360
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
PHARMACOLOGICAL TREATMENT OF POSTSTROKE
DEPRESSION
Maria Margarida Caramona(1), E Silva(2), B Oliveiros(3),
E Ponciano(3)
(1) University of Coimbra Faculty of Pharmacy, Laboratory of Pharmacology, Coimbra, Portugal
(2) Comunity Pharmacy, Coimbra, Portugal
(3) University of Coimbra Faculty of Medicine, Laboratory of Pharmacology, Coimbra, Portugal
The issue of poststroke depression has gained worldwide interest in the
past 10 years. There has been a general agreement about the prevalence
of both major and minor depression after stroke. Depression is one of the
major impediments to full physical and mental recovery from stroke. The
aim of this study was to define the prevalence of depression in stroke
survivors and assess the treatment as well as the impact on their quality
of life. It was designed a prospective study. Patients under antihypertensive therapy visiting community pharmacies all over Portugal were
recruited. Patients’ medication data, gender, age, smoking history, alcohol
intake, medical conditions, quality of life, were collected in a validated
self administered instrument. The sample included 155 patients (mean
age =66,91 + /- 10 years, 52,9% female) recruited during 2007 that had
a previous stroke and gave informed consent for a direct interview. 30%
of those assessed were depressed. 16% were taking an antidepressant
medication. Poststroke depression and low quality-of-life seem to be
more common among women. Women have less favourable functional
outcome because of higher age at stroke onset and more severe strokes.
Psychiatric disorders, mostly depression, are fairly common in stroke
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
217
survivors. Although being depressed, few patients were taking antidepressants. We consider that the prevalence of depression is relevant and
that the risk factors as well as the early diagnosis are important for the
best management and prognosis of stroke patients. Further exploration of
this low level of treatment is warranted.
Paper No.: 2931
FOCUSED CONFERENCE GROUP: P19 - GENERAL
SESSION - ONCOLOGY
IONIZING RADIATION ALTERS THE BONE MARROW
MICROENVIRONMENT BY INDUCING SENESCENCE OF
MESENCHYMAL STROMAL CELLS
Cynthia Carbonneau(1,2), G Despars(1,2), A Fortin(1,2), O Le(1,2),
T Hoang(1,3), CM Beausejour(1,2)
(1) Université de Montréal, Department of Pharmacology, Montreal,
Canada
(2) CHU Ste-Justine, Montreal, Canada
(3) IRIC, Montreal, Canada
The extent to which the integrity of the bone marrow (BM) stroma contributes to the success of hematopoietic stem cell (HSC) transplantation
is not well known. Alterations of the BM microenvironment, which has
been shown to occur during aging or following genetic manipulations of
telomere length, impair HSC functions and engraftment. These results
suggest that increased number of senescent marrow stromal cells could
affect the niche functions. Exposure to ionizing radiation (IR), by inhibiting marrow stromal cells colony forming unit (CFU) potential, is
believed to severely impair the BM microenvironment. Here, we demonstrate that IR alters the niche by inducing a p16INK4a-dependent senescence phenotype in marrow stromal cells. Intriguingly and in contrast to
p21 expression, p16INK4a expression is delayed post-IR. In accordance
with this observation, IR-induced long-term loss of BM stromal cell
CFU’s potential was partially rescued in p16INK4a deficient mice. Moreover, while the absence of p16INK4a expression did not prevent
IR-induced short-term homing impairment, it improved homing potential
long-term post-IR. In addition, IR could contribute to alter BM microenvironment by inducing a p16-independent increase in G-CSF expression.
Indeed, increased levels of G-CSF, a cytokine known to induce HSC
mobilization, were detected in the bone cavity shortly after IR. Co-culture experiments of stromal cells with hematopoietic progenitors further
confirmed that irradiated stromal cells secrete greater levels of G-CSF
than control cells. Together, our results provide a mechanistic link as to
how irradiation, by inducing senescence of the stroma and G-CSF
expression, could impair the function of the hematopoietic niche.
Paper No.: 754
FOCUSED CONFERENCE GROUP: PW19 - INFLUENCE OF
DEGENERATION AND REPAIR IN THE CNS AND PERIPHERY
THE EFFECT OF CHEMICAL ABSENCE MODEL ON THE
PROGRESSION OF KINDLING IN WISTAR RATS
Nihan Carcak(1), M Sahiner(2), E Sakallý(3), K Tezcan(3), R Gülhan(3),
F Onat(3)
(1) Istanbul University Faculty of Pharmacy Department of Pharmacology, Istanbul, Turkey
(2) Acýbadem University Faculty of Medicine, Department of Physiology, Istanbul, Turkey
(3) Marmara University Faculty of Medicine Department of Pharmacology and Clinical Pharmacology, Istanbul, Turkey
Thalamo-limbic circuits are thought to regulate limbic seizure activity
whereas thalamo-cortical circuits are involved in the expression and generation of spike-and-wave discharges (SWDs) in the absence epilepsy
models. Kindling as a model of temporal lobe epilepsies is a well known
model which causes secondary generalized seizures in rats. Recent studies showed that there is resistance in relation with SWD activity to the
kindling progress in animal models of genetic absence epilepsy. c-Butyrolactone (GBL), the prodrug of GHB (c-Hydroxybutyric acid), represent a chemical experimental model of generalized absence seizures. In
this study we aimed to determine whether the resistance to the development of kindling in absence epilepsy can be independent of the genetic
background. Electrodes were stereotaxically implanted into the basolateral amygdala. After a recovery period, the animals were stimulated at their
afterdischarge thresholds twice daily. GBL+KI animals were stimulated
20 min after the intraperitoneal (i.p) administration of GBL twice daily
until they reached stage 5 seizure state. KI animals were stimulated without GBL administration. The seizure severity was evaluated by Racine’s
scale. The KI animals had stage 5 seizures by 15 stimulations. However,
GBL-KI group reached stage 5 by 30 stimulations. Our data show a
delay in the development of kindling as well as a relation of SWD activity to the kindling progress after GBL administration. The resistance to
limbic epilepsy in absence epilepsy rats seems to be partially influenced
by the absence epilepsy itself and possibly also be the genetic background.
Paper No.: 1389
FOCUSED CONFERENCE GROUP: P19 - GENERAL 1
SESSION - ONCOLOGY
TEMOZOLOMIDE INDUCES APOPTOSIS AND AUTOPHAGY
IN GLIOMA CELLS
A Carmo(1,2), H Carvalheiro(1), Maria Celeste Lopes(1,2,3)
(1) Center for Neuroscience and Cell Biology, Department of Cellular
Immunology and Oncobiology, Coimbra, Portugal
(2) CIMAGO, University of Coimbra, Coimbra, Portugal
(3) University of Coimbra, Faculty of Pharmacy, Coimbra, Portugal
Glioblastoma is the most malignant primary brain tumor, comprising
approximately 50% of the cerebral gliomas. Despite all the efforts to
improve the efficacy of the treatment, the median survival time for GBM
patients remains approximately 12-14 months. This relative lack of success in GBM therapy is probably due to the genetic and cellular heterogeneity of these tumours and to the development of chemotherapeutic
resistance. The chemotherapeutic treatment of glioma is based on the use
of temozolamide (TMZ). Nevertheless, the median survival of glioma
patients treated with TMZ is 14.6 months. In order to understand the
effect of TMZ in glioma cells, we studied apoptosis and autophagy
induction in TMZ-treated glioma cells. For that, U-118 glioma cells were
incubated with different concentrations of TMZ for different periods of
time. Cell viability was assessed by the MTT assay and cell proliferation
was determined using BrdUrd incorporation. Apoptosis was addressed
by confocal microscopy using hoescht and propidium iodide and by flow
cytometry. Autophagy was evaluated through the LC3 expression which
was determined by western blot. The results indicated that TMZ reduced
the U-118 cell viability and proliferation. Regarding the survival, the
results showed that TMZ induced apoptosis and simultaneously autophagy. Since authophagy is a survival pathway, the low efficacy of TMZ
in the treatment of gliomas may be related with the activation of
autophagy.
Paper No.: 2992
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
CORONARY ARTERY BYPASS GRAFTING SURGERY
WORSENS OXIDATIVE STRESS AMONG PATIENTS WITH
CORONARY DISEASE
Manuel Carnero(1), J Muñoz-Marin(2), JA González Correa(2),
F Reguillo(1), J Navarro-Dorado(3), M Ramajo(3), JP de la Cruz(2),
T Tejerina(3)
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
218
(1) Hospital Clı́nico San Carlos, Service of Cardiac Surgery, Madrid,
Spain
(2) Universidad de Málaga, School of Medicine, Department of Pharmacology, Málaga,Spain
(3) University Complutense of Madrid, School of Medicine, Department
of Pharmacology, Madrid, Spain
Introduction: Oxireduction enzymatic mechanisms in human beings can
be physiologically altered due to inflammatory or infectious stimuli to
the organism. Inflammatory response to heart surgery has been widely
studied, but, up to date, little is known about its possible influence over
oxidative stress. We studied the variations of multiple oxidative stress
related products and enzymes in a cohort of patients who underwent a
coronary artery bypass grafting surgery (CABG). Methods: We measured
2 hours before and 24 after CABG the concentration of malondialdehidic
acid (MDA), nitrates, and peroxynitrites (all of them considered to be
prooxidative markers); and reducted glutation and mithocondrial superoxide dismutase (SOD-Mn) (antioxidative markers) in a cohort of
patients with coronary disease in a single center. Results: 119 patients
were included in the present study. Statistically significant differences
were detected in the mean plasmatic MDA concentration before (0.148
mmoL/L (SD 0.12)) and after (0.283 mmoL/L /SD 0.16) surgery
(P < 0.001). Higher concentrations of peroxynitrates (p = 0.443) and
nitrates (p = 0.078) were also detected, though differences did not reach
statistical significance. On the other hand, lower levels of reducted glutation and SOD-Mn were detected after surgery (p = 0.94 and p = 0.070),
though differences were not significant. Conclusions: CABG surgery
worsens the oxidative stress in patients with coronary disease higher oxidation activity and greater concentrations of its products and a lower
antioxidant activity.
Paper No.: 3143
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
ORAL PHARMACOKINETICS OF THREE
FLUOROQUINOLONES IN MEXICANS: EVIDENCE FOR
INTERETHNIC DIFFERENCES
Miriam Carrasco-Portugal(1), G Lozoya-Moreno(2),
S Patiño-Camacho(2), F Flores-Murrieta(1,2)
(1) Instituto Nacional de Enfermedade Respiratorias, Mexico City,
Mexico
(2) Escuela Superior de Medicina del Instituto Politécnico Nacional,
Mexico City, Mexico
Interethnic differences in the oral pharmacokinetics of drugs metabolized
by CYP3A4, i.e. nifedipine, cyclosporine, midazolam, sildenafil, meloxicam, have been reported, Mexicans reaching higher plasma levels than
Caucasians. It is assumed that such differences are due to a reduced
metabolism by CYP3A4 in Mexicans. However, very frequently, drugs
metabolized by CYP3A4 are also substrates of p-Glycoprotein (p-GP)
and, therefore, participation of this p-GP may contribute to such differences. In this study, we evaluated the oral pharmacokinetics of three
fluoroquinolones (ciprofloxacin, levofloxacin and gatifloxacin) that are
p-GP substrates and they are not metabolized by CYP3A4. Seventyseven male healthy subjects were enrolled in this study. The volunteers
were divided in three groups. The first group received an oral dose of
250 mg of ciprofloxacin, the second 500 mg of levofloxacin and the
third 400 mg of gatifloxacin. Blood samples were drawn at selected
times and plasma was obtained and stored frozen until analyzed by selective high-performance liquid chromatographic methods. Pharmacokinetic
parameters were obtained by non-compartmental analysis and compared
with those reported in other populations. After administration of the
drugs, plasma levels increased rapidly reaching the maximal concentration (Cmax) in about 60 to 90 minutes. Cmax and AUC values observed
in Mexicans were higher than the values described in studies conducted
in Caucausians, however, no important changes in half-life were
observed. These results suggest increased bioavailability of ciprofloxacin,
levofloxacin and gatifloxacin in Mexican subjects, maybe due to
increased absorption of these drugs, by a lower activity of p-GP.
Paper No.: 2395
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
THIOPURINE METHYLTRANSFERASE
PHENOTYPE-GENOTYPE CONCORDANCE IN CHILDREN
WITH ACUTE LYMPHOBLASTIC LEUKAEMIA
Cher Cartwright(1), C Mitchell(2), N Goulden(3), A Vora(1),
L Lennard(1)
(1) University of Sheffield, Department of Clinical Pharmacology,
Sheffield, UK
(2) University of Oxford, Oxford, UK
(3) Great Ormond Street Hospital, London, UK
The clinical importance of the thiopurine methyltransferase (TPMT)
genetic polymorphism has been illustrated by many studies which have
linked the inheritance of lower TPMT activities with thiopurine sensitivity, specifically bone marrow toxicity. The aim of this multi-centre study
was to assess TPMT phenotype-genotype concordance in children with
acute lymphoblastic leukaemia (ALL) treated according to the UKALL2003 trial. Blood samples were requested at 100% protocol mercaptopurine (75mg/sqm), or the maximum tolerated, dose. Red cell TPMT
activity and mercaptopurine derived cytotoxic thioguanine nucleotide
(TGN) metabolites were measured; samples were screened for TPMT*2
and the TPMT*3 family of low-activity variant alleles. Ranges were
compared by the Mann-Whitney test. Samples were forwarded from 800
children (median dose 75mg/sqm, range 7 to 122). TPMT activities ranged from 0 to 26.4 units, median 14.1; 733 children were wild-type, 56
TPMT*1/*3A, 7 TPMT*1/*3C, two TPMT*3A/*3C, one TPMT*3A/
*3A and one TPMT*1/*2. The sensitivity for detecting the variant allele
at 9.5units TPMT was 84% (specificity 94%), at 10.5units 95% (specificity 90%) and at 11.5units 98% (specificity 84%). At 10.5 units the overall concordance was 91%, but only 46% within the heterozygous
phenotype group <10.5units TPMT. Median TGN concentrations in children <10.5 units were 545pmol and >10.5 units 323pmol. Within the
<10.5unit cohort children with a variant allele (n = 61) had 737pmol
compared to 361pmol in wild-type children (n = 72), median difference
354pmol P < 0.001 95% CI 263 to 451. Wildtype children with TPMT
activity less than 10.5units did not exhibit the same metabolite profile as
heterozygous variant allele children.
Paper No.: 1569
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
PATIENT’S ACCESS TO ANTIRETROVIRAL MEDICINES:
HOW DO THEY VIEW THEMSELVES?
DM Carvalho, MH Magalhães, AMR Felix, ECA Costa, WA Souza,
OMGC Vilas-Boas, Marcia HMC Podestá
Federal University of Alfenas, Department of Pharmacy, Alfenas, Minas
Gerais, Brazil
The Acquired Immune Deficiency Syndrome (AIDS) shows a life impact
on the individuals, although the offer of costless treatment to the patient.
The meaning of this research is to analyze the perceptions of the quality
of life on those patients who receive medicines from a public health
unity of Alfenas, Brazil. The patients answered a questionnaire, between
September and October 2009, about specific questions of their disease
followed by the investigation SF-36 (de Soárez, PC et al Pan Am J Public Health 2009; 25: 69-76)This study was approved by the Ethic Committee in Human Research (nordm; 23087003008/2009). From the 92
patients that are in medicament treatment for about 5 years, 49% are
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
219
female. The most of them are singles (40%) with age predominantly
between 30 to 45 years old (52%), primary school instruction uncompleted (51%) and family earnings approximately US$ 375 by month
(51%). When inquired about AIDS knowledge, patients reported sexual
contamination (75%) and said that they understand, but hide this condition (65%). Concerning about life quality, the most preoccupants factors
for them are physical aspects (56%), vitality (57,8%) and emotional
aspects (60,1%), on media averages between 0 to 100. We concluded that
the AIDS treatment has interference on the self-perceptions of these individuals, already affected by social and cultural conditions.
Paper No.: 2890
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
MOLECULAR MECHANISMS OF PISTACIA LENTISCUS
ESSENTIAL OIL ANTIPROLIFERATIVE ACTIVITY
from mRNA extracted from 12 pooled samples of internal mammary
artery obtained with written informed consent from seven individuals
undergoing coronary artery bypass graft surgery. The study was
approved by the local research ethics committee. Pooled tissue samples
(4.3 g, sample weight 355 ± 302 mg SD, n = 12) yielded 608 ll total
RNA (sample RNA 51 ± 25 ll SD). A 167 lg sample of this total RNA
gave 2.48 ± 0.008 lg mRNA (SD, n = 2) and was used to prepare a
cDNA library using the OrientExpress Oligo d(T) Primer cDNA kit from
(Novagen #69992-3) from which the phage display library was prepared
using the T7 Select Phage Display System (Novagen #70550-3). Simvastatin was immobilised on a pre-prepared Magic Tag 96-well plate
and a preliminary screen against the new library revealed 25 sequences,
five of which are of interest for further study. This method has the potential to be scaled to an individual subjec’s sample, to allow Magic Tag
analysis of rare diseases and uncommon serious adverse events as well
as to improve personalising treatment to increase efficacy and reduce toxicity in more common, serious disorders.
Elisabetta Casarin(1), B Barocco(1), A Buriani(2), M Carrara(1)
(1) Padova University, Department of Pharmacology and Anesthesiology,
Padova, Italy
(2) Data Medica Research Labs, Padova, Italy
The phytocomplex from Pistacia lentiscus, a shrub of the Anacardiaceae
family, is an essential oil obtained by hydrodistillation of leaves, fruits or
from a trunk exudate (mastic gum). The mastic gum has been known to
be effective in several gastric diseases, against Helicobacter Pylori and
for its antibacterial and antifungine activities. Furthermore, Pistacia oil’s
major chemical constituents are monoterpenes with chemiopreventive
and chemiotherapic properties. We investigated the antiproliferative properties of the volatile oil from Pistacia lentiscus twigs and leaves using
human cell lines from ovarian (2008 and C13*) and colon (LoVo) adenocarcinoma, and human stable fibroblast line (HFFF2) as in vitro models.
The MTT test showed that, after 3 hour treatment, phytocomplex (about
150 lg\ml) was able to inhibit the growth of all adenocarcinoma cell
lines. After 24 hour treatment the IC50 on 2008 and LoVo cells resulted
3 times lower. On fibroblast line the phytocomplex was active only after
72 hour treatment. Western blot analysis confirmed the oil capability to
reduce carcinoma cell growth by decreasing the expression of p-ERK,
MAPKs induced by mitogenic stimuli. Using AnnexinV with propidium
iodide we observed that oil was able to stimulate apoptosis in a dosedependent manner. Analysis of mitochondrial membrane potential, with
Rhodamine123, and ROS generation, with H2DCF-DA, showed the oil
capability to activate mitochondrial apoptotic pathway. Our data also
indicate that Pistacia lentiscus oil caused programmed cell death via a
caspase-independent pathway. We also performed a flow cytometrybased cell cycle analysis, observing that the phytocomplex induced dosedependent arrest in G2M phase.
Paper No.: 1526
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
A CHEMICAL GENOMICS APPROACH TO
CARDIOVASCULAR THERAPEUTICS
Paper No.: 1527
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
PERSONALISED THERAPIES USING A CHEMICAL
GENOMICS APPROACH
Katherine Casey-Green(1), A Marsh(1), P Taylor(1), S Ladwa(2), W
Dimitri(3), D Singer(4)
(1) University of Warwick, Coventry, UK
(2) Tangent Reprofiling Ltd, Bicester, UK
(3) University Hospital, Coventry, UK
(4) Clinical Sciences Research Institute, Coventry, UK
Therapeutic agents may interact with the body in many ways apart from
the major identified mechanism, and often the range of targets through
which a clinical effect is brought about remain unclear. It is desirable to
characterise more fully the action of a pleiotropic therapeutic upon a system, identifying each of the proteins with which it interacts in the cell. In
previous studies, we have developed and tested proof of concept that the
new Magic Tag chemical genomics (Dilly, S. J., et al., Chem. Commun. 2007; 2808-2810) can enable a drug to be covalently immobilised
so that it can be probed by phage display libraries and interacting partners identified. (Ladwa, S. R., et al., ChemMedChem 2008; 3: 742-744).
In order to elucidate structure-function relationships between the immobilised drug molecule and the captured protein, it would also be useful to
be able to remove the covalently bound drug molecule from the resin in
order to observe its configuration. We present here the preparation of a
new cleavable linker immobilisation system based on ‘Wang’ methodology (Wang, S. S., J. Am. Chem. Soc. 1973; 95: 1328-1333) to allow separation of the immobilised drug - tag moiety, which will allow structural
determination and hence allow for development of further structure-activity assays to investigate new efficacy and toxicity targets. This new
approach thus has the potential to identify important new classes of therapeutic action. It may also unmask new mechanisms for toxicity thereby
allowing development of new structural chemistry approaches to reducing risk drug toxicity.
Katherine Casey-Green(1), A Marsh(1), P Taylor(1), S Ladwa(2), W
Dimitri(3), J Zhang(4), D Singer(4)
(1) University of Warwick, Coventry, UK
(2) Tangent Reprofiling Ltd, Bicester, UK
(3) University Hospital, Coventry, UK
(4) Clinical Sciences Research Institute, Coventry, UK
The Magic Tag chemical genomics approach (Dilly, S. J., et al., Chem.
Commun. 2007; 2808-2810) has identified novel drug-target interactions
(Ladwa et al. Chem Med Chem 2008;3:742-744). We now report its use
to uncover pathology-specific therapeutic targets for the pleiotropic
HMG-CoA reductase inhibitor simvastatin in patients with severe ischaemic heart disease. A new vascular phage display library was prepared
Paper No.: 1995
FOCUSED CONFERENCE GROUP: P01 - CLINICAL PHARMACOLOGY IN THE EMERGING COUNTRIES
INEQUIVALENCE OF DOXORUBICIN LIPOSOMAL FORMULATIONS COMMERCIALIZED IN LATIN AMERICA
Gilberto Castañeda-Hernández(1), L Villafaña-Godı́nez(1),
MA Valdés(2), L González-Flores(1)
(1) CINVESTAV-IPN, Department of Pharmacology, Mexico DF,
Mexico
(2) Universidad de Sonora, Hermosillo, Mexico
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
220
It is well established that liposomal formulations significantly increase
the safety of doxorubicin without compromising the antitumor efficacy
of this agent. The innovator liposomal doxorubicin is Caelyx (Doxil in
USA). A generic formulation, Doxopeg, is presently commercialized in
Latin America, although no information is available regarding its biopharmaceutical properties. Hence, we decided to comparatively study
Doxopeg and Caelyx. Six vials were studied per formulation. Each vial
contained 20 mg doxorubicin in 10 ml, according to the label. Doxorubicin content in Caelyx and Doxopeg was within the 95-105% range of the
labeled dose and no impurities were detected. In the two products, at
least 99.5% of doxorubicin content was encapsulated in liposomes. Both
formulations were similar when examined by electron microscopy and
liposomes exhibited a mean diameter of 100 nm, as determined by light
scattering. However, atomic force microscopy revealed a more stable
liposomal membrane in Caelyx, suggesting differences in lipid composition between formulations. Doxorubicin release rate in human plasma at
37 degrees in 24 h was about 1% of the liposomal content of Caelyx,
being consistent with previous reports. On the other hand, doxorubicin
release from Doxopeg was three times faster, likely due to differences in
the liposomal membrane. Since doxorubicin release rate is critical for
antitumor efficacy, it is concluded that Caelyx and Doxopeg are not
equivalent formulations. In the absence of data on the impact of faster
liposomal doxorubicin release on clinical efficacy and safety, the therapeutic use of Doxopeg is not recommended.
Paper No.: 1033
FOCUSED CONFERENCE GROUP:
P04 - PHARMACOEPIDEMIOLOGY, CURRENT
CONTROVERSIES AND OPPORTUNITIES
MANAGEMENT OF CARDIOVASCULAR MEDICATION IN
SURGICAL PATIENTS: INDICATIONS IN SUMMARY OF
PRODUCT CHARACTERISTICS AND SUBSTITUTION FOR
INTRAVENOUS FORMS
Liliana Castanheira(1), P Fresco(2), AF Macedo(1)
(1) University of Beira Interior Faculty of Health Sciences, Health Sciences Research Centre, Covilhã, Portugal
(2) Oporto University Pharmacy Faculty, Oporto, Portugal
Introduction: Each year, millions of patients around the world undergo
surgical procedures. The majority of Portuguese surgical patients
(71.5%) were taking regular medication, namely for cardiovascular disorders (53%). Recommendations for management of regular medication in
perioperative period and the availability of intravenous forms of drugs
are critically needed for an efficient iatrogenic risk management. The
purpose of this study was to identify information in summary of product
characteristics for management of cardiovascular medication in perioperative period and the availability of these medications in intravenous
form. Materials/Patients: Summary of product characteristics of cardiovascular medication was screened for information about drug management in the perioperative period. Additionally, it was verified, for the
same drugs, if an intravenous form was available to use in the perioperative period. Results: Of 46 summary of product characteristics screened,
33% had information about its use in the perioperative period and 13%
had intravenous forms available. Conclusions: For the majority of cardiovascular medications, the summary of product characteristics is not
enough to support the management of these medications in the perioperative period. Proper pharmaceutical formulations for cardiovascular drugs,
namely intravenous, are lacking to use in surgical patients.
Paper No.: 2673
FOCUSED CONFERENCE GROUP: P17 - NEW APPROACHES
AND TARGETS IN PSYCHIATRY
NEUROPLASTIC CHANGES PRECEDE CHRONIC
ANTIDEPRESSANT RESPONSES OF THE 5-HT4 RECEPTOR
AGONIST RS67333
Elena Castro, A Dı́az, EM Valdizán, R Vidal, F Pilar-Cuellar,
J Pascual-Brazo, A Pazos
Institute of Beiomedicine and Biotechnolgy- IBBTEC (UC-CSIC-IDICAN) Department of Physiology and Pharmacology, Santander, Spain
It has been recently suggested that activation of 5-HT4 receptors might
possess antidepressant properties in rats after a 3 days treatment, indicating a new strategy for developing faster-acting antidepressants. In this
regard, here we have evaluated in rat brain the expression of proteins
related to neuroplasticity after a subacute RS67333 treatment and their
correlate on behavioural paradigms. A 3-days treatment with RS67333
(1.5 mg/kg/day), previously shown to increase neurogenesis in dentate
gyrus, induced an upregulation in the expression of BDNF (% increase =
64%; p < 0.05) and pCREB/CREB ratio (% increase = 93%; p < 0.01)
in the hippocampus. A significant reduction in the forced swimming test
(immobility time) was also observed after 3 (% red= 27%; p < 0.001)
and 7 (% red=29%; P < 0.001) days of treatment. However, in the novelty-feeding suppressed test, a validated paradigm to predict chronic antidepressant efficacy, a significant reduction in the latency to feed (% red=
49%; p < 0.03) was observed only after 7 days of treatment. Short-term
treatment with RS67333 also failed to downregulate 5-HT4 receptor-coupled adenylate cyclase activity. Our data suggest that neural proliferation-related changes induced by antidepressants precede clinical
improvement. Furthermore, when compared to data regarding classical
antidepressants, our results show that the activation of 5-HT4 receptors
could represent a good strategy for developing antidepressants with a
minor onset of action.
Supported by: Ministerio de Ciencia e Innovación (SAF07-61862) and
Fundación Alicia Koplowitz
Paper No.: 1844
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
CENTRAL INJECTION OF CDP-CHOLINE SUPPRESSES
SERUM GHRELIN LEVELS WHILE IT INCREASES SERUM
LEPTIN LEVELS IN RATS
Sinan Cavun(1), S Kýyýcý(2), NF Basaran(1), V Savci(1),
S Imamoglu(2)
(1) Uludag University Faculty of Medicine Department of Medical Pharmacology, Bursa,Turkey
(2) Uludag University Faculty of Medicine Department of Endocrinology, Bursa, Turkey
Introduction: Ghrelin and leptin are two important peptides which control food intake. Recently it was shown that CDP-choline has suppressive effect on appetite. In present study we aimed to test
intracerebroventricular (icv) administration of CDP-choline on serum
ghrelin, leptin, glucose and corticosterone levels in rats. Material and
Methods: Male Wistar rats were used in all experiments. For repeated
blood withdrawal a cannula was inserted into the left carotid artery and
also a cannula was placed into the right lateral ventricule for icv injections. Rats were randomized to three treatment groups and icv CDP-choline was administered at three different doses to the groups as follows:
0.5 (n = 8), 1.0 (n = 8) and 2.0 (n = 8) lmol. Saline injected rats were
used as controls. Blood samples were withdrawn before and 5th, 15th,
30th, 60th and 120th min after icv injections. Results: Baseline serum
ghrelin and leptin levels were not different between groups. Intragroup
analysis revealed that serum ghrelin levels were suppressed significantly
at 60th min with 1lmol CDP-choline injection (p = 0.025) whereas
serum leptin levels were increased at 60th min with 0.5, 1.0 and
2.0 lmol doses of CDP-choline (p = 0.036, p = 0.012, p = 0.043,
respectively). Serum leptin levels were significantly higher at 120th min
with 1.0 and 2.0 lmol doses of CDP-choline compared with baseline
values (p = 0.017, p = 0.046, respectively). Central CDP-choline injections also induced a dose- and time dependent increase in serum glucose
and corticosterone levels. Conclusions: These results suggest that central
injection of CDP-choline suppress serum ghrelin levels while it increases
serum leptin levels.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
221
Paper No.: 590
FOCUSED CONFERENCE GROUP: PW19 - INFLUENCE OF
DEGENERATION AND REPAIR IN THE CNS AND PERIPHERY
CALCIUM ANTAGONISTIC EFFECTS OF SOME
ANTIMIGRAINE DRUGS IN RAT ISOLATED BASILAR
ARTERY
Edip Guvenc Cekic, M Tuncer
Hacettepe University Faculty of Medicine, Department of Pharmacology,
Ankara, Turkey
Vasodilator substances such as CGRP released form central perivascular
nerve endings as a result of activation of calcium channels play a role in
the pathogenesis of migraine. Propranolol, a b-adrenergic receptor
blocker, is used for chronic prophylaxis of the disease. Propranolol has
been reported to block calcium channels in the rat mesenteric artery and
aorta (Priviero FB, et al. 2006; 33: 448-455), however such kind of effect
of the drug has not yet been shown in cerebral arteries. Therefore, we
aimed to investigate calcium antagonistic effects of propranolol and two
drugs, namely pizotifen and methysergide, which are also used for
chronic prophylaxis of migraine, in the rat basilar artery. Both sexes of
Wistar rats (250-350 g) were used. After decapitation, isolated basilar
arteries were mounted in myograph system (Danish Myograph 610M)
and isometric responses were recorded. Propranolol (10 nM-30 lM) and
pizotifen (10 nM-30 lM) caused endothelium-independent relaxations in
the precontracted arteries. Methysergide (10 nM-30 lM) did not induce
any relaxation. In order to examine a calcium antagonistic activity, concentration-response curves to calcium chloride were obtained in a depolarizing Ca-free solution, in the absence and presence of propranolol (130 lM), pizotifen (1-10 lM) and methysergide (1-10 lM). Propranolol
and pizotifen but not methysergide, shifted the calcium concentration response curves to the right. The results suggest that calcium channel
blocking activities of propranolol and pizotifen may contribute to their
antimigraine effect by preventing the release of vasodilator substances
from perivascular nerve endings.
Paper No.: 3237
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
EFFECT OF CHROMIUM ENRICHED FERMENTATION
PRODUCT OF BARELEY AND BEER YEAST AND ITS
COMBINATION WITH ROSIGLITAZONE ON
EXPERIMENTALLY INDUCED HYPERGLYCEMIA IN MICE
Vlada Cekic(1), V Vasovic(2), V Jakovljevic(2), D Lalosevic(3), M Mikov(1), A Sabo(1)
(1) Hemofarm AD, Medical Department, Belgrade, Republic of Serbia
(2) University of Novi Sad, Faculty of Medicine, Department of Pharmacology, Toxicology and Clinical Pharmacology, Novi Sad, Republic of
Serbia
(3) University of Novi Sad, Faculty of Medicine, Department of Histology and Embryology, Novi Sad, Republic of Serbia
To examine the action of BBCr, rosiglitazone (R) and their combination
(BBCr + R) on glycemia in mice on an empty stomach and hyperglycemia induced with glucose, adrenaline and alloxan. The BBCr preparation
is an auxiliary medicine registered as a diabetic supplement in the treatment of insulin-independent diabetes mellitus. It is fermentation product
of barley and beer yeast enriched with organically bound chromium.
Active ingredient of this product is Glut 4 plus – insulin sensitizer with
the percentage of tablet composition of 76.92%. Animals were divided in
three groups, depending of the way of inducing hyperglycemia: I–glucose, single dose 500 mg/kg, p.o.; II–adrenaline, single dose 0.2 mg/kg,
s.c.; III–alloxan, multiple dose, 100 mg/kg, i.p. Groups were divided into
subgroups according to the preparation (BBCr, R, BBCr+R). In the first
two groups, hyperglycemia was measured on an empty stomach, 30min
after glucose load, and 45min after adrenaline injection, and at the same
time intervals after a 7-day treatment. In Group III hyperglycemia was
measured both before and after the diabetes-inducing treatment with
alloxan and tested substances (after 12 days). Histological analyses of
sections of the liver and pancreas were carried out in mice of Group III.
In the glucose load test, BBCr and BBCr+R decreased significantly glycemia value compared to control (7.4 ± 0.6mmol/l : 7.7 ± 0.7mmol/l :
8.9 ± 1.3mmol/l; p < 0.05). In the adrenaline test, R and BBCr+R
decreased significantly glycemia with respect to control (8.6 ± 1.8mmol/l
: 9.6 ± 2.4mmol/l : 15.2 ± 3.3mmol/l; P < 0.01). After treatment with
alloxan the glycemia was significantly lower in the groups treated with R
and BBCr compared to control (8.4 ± 0.8mmol/l : 8.6 ± 3.2mmol/l :
18.8 ± 5.5mmol/l, P < 0.01). The least damage to pancreas tissue caused
by alloxan was observed in the subgroups treated with BBCr and
BBCr+R. In the liver tissue, there was more or less pronounced leukocyte infiltrate in subgroups treated with BBCr and R.
Paper No.: 1411
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
DOSAGE REGIMEN OF VANCOMYCIN IN NICU/PICU
PATIENTS
O Cerná(1), Pavla Pokorná(1), J Sedivý(2)
(1) General Teaching Hospital, Department of Paediatrics, Prague, Czech
Republic
(2) General Teaching Hospital, Department of Clinical Biochemistry and
Laboratory Diagnostics, Prague, Czech Republic
Introduction: The value of vancomycin therapeutic drug monitoring and
therapeutic range of vancomycin is discussed for routine monitoring in
pediatric population. Vancomycin is a glycopeptide used in neonates and
children with staphylococcal sepsis. Methods: The goal of this prospective study is to predict a dosage regimen of vancomycin. This presented
study (2004-2009) enrolls n = 57 neonates (gestational age 25-41 weeks,birth weight 750 - 4000 g) during first month of life and n = 40 children (actual age 6 months -18 years, actual weight 12 - 40 kg) with
blood stream infection (BSI) indentified in n = 53/97 patients. Neonates
received 30 mg/kg/day i.v. divided in two or three doses according to
postnatal age, children received 20 mg/kg/dose i.v. administered by
60 minutes infusion. Trough (Ctrough) or peak (Cpeak) of vancomycin
serum concetrations were determinedbefore and after third dose of
vancomycin or later using Abbott Diagnostics Laboratories Method. Targets were Cmin concentrations between 5 and 15 mg/L and Cmax below
40 mg/L. Vancomycin data were analysed according to one- compartment model using PC- MW Pharm -version 3,30 Programme and various
dosing schemes were simulated. Results: Detected Ctrough concentrations were within targets in 27/97 patients, Cpeak levels were higher than
40 mg/l in 3/97 patients. Vancomycin dosage regimen was individualized
in 31/97 and individual prediciton revisedin 20/31 of patients. Conclusions: The use of individual therapeutic vancomycin monitoring help to
optimize target levels in NICU/PICU patients.
Paper No.: 907
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
INHIBITION OF CATECHOLAMINE SECRETION BY A
TERPENOID EXTRACTED FROM SYNEILESIS ACONITIFOLIA
(BUNGE) MAXIM IN CULTURED BOVINE ADRENAL
CHROMAFFIN CELLS
L Chai, H Mao, Y Su, Han Zhang, H Guo, Y Jin, Z Zang
Tianjin University of Traditional Chinese Medicine, Tianjin, PR China
Evidences indicated a critical relationship between activation of sympathetic nervous system by stress and chronic joint pain or inflammation.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
222
Catecholaminergic system was activated as a part of the sympathetic nervous system (CNS) response to stress. The goal of this study was to
examine the effects of a terpenoid, (6b,8b)-6-hydroxyeremophil-7(11)en-12,8-olide, extracted from Syneilesis aconitifolia (Bunge) Maxim on
CA secretion in cultured bovine adrenal chromaffin cells, a Chinese herb
which was used to treat rheumatism, arthralgia and some other diseases.
Our results showed that the terpenoid (1lM-100lM) significantly inhibited CA secretion induced by 300lM acetylcholine (ACh) and 56 mM
K+ solution in a concentration-dependent manner, which are activators
of the nicotinic acetylcholine receptor and voltage-dependent Ca2 + channels, respectively. Raise of intracellular Ca2 + concentration ([Ca2 + ]i) is
a requisite for CA secretion. We next investigated the effects of the terpenoid(1lM-100lM) on various activators induced elevation of [Ca2 + ]i.
The results indicated the terpenoid also inhibited elevation of [Ca2 + ]i
caused by ACh and high K+ in a concentration-dependent manner, which
was consistent with the inhibitory effects on CA secretion. These findings suggested that the terpenoid inhibited CA secretion in cultured
bovine adrenal Chromaffin Cells through suppression on the nicotinic
acetylcholine receptor as well as voltage-dependent Ca2 + channels,
thereby inhibition on the raise of [Ca2 + ]i.
Keywords: catecholamine secretion; calcium; Syneilesis aconitifolia
(Bunge) Maxim; joint pain; inflammation; terpenoid
Paper No.: 2344
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
ESTROGEN RECEPTOR A AS A KEY TARGET OF RED WINE
POLYPHENOLS ACTION ON THE ENDOTHELIUM
M Chalopin(1), A Tesse(1), MC Martinez(1), D Rognan(2), JF Arnal(3),
Ramaroson Andriantsitohaina(1)
(1) University of Angers Faculty of Medicine, INSERM, U771, CNRS
UMR, 6214, Angers, France
(2) University Louis Pasteur-Strasbourg I, Department of Bioinformatics
of Drugs, UMR 7175 CNRS-ULP, Illkirch, France
(3) University of Toulouse III Paul Sabatier, CHU - Centre HospitalierUniversitaire, INSERM U858, Toulouse, France
A reduction in cardiovascular risk and vascular protection associated
with diet rich in polyphenols are generally accepted; however, molecular
targets for polyphenols effects remain unknown. Meanwhile evidences in
literature have enlightened, structural similarities between estrogens and
polyphenols known as phytoestrogens, and also in their vascular effects.
We hypothesized that estrogen receptor a isoform (ERa) could be
involved in transduction of the vascular benefits of polyphenols. Here,
we used ERa deficient mice to show that endothelium-dependent vasorelaxation induced either by red wine polyphenol extract, ProvinolsTM, or
delphinidin, an anthocyanin possessing similar pharmacological profile,
is mediated by ERa. Indeed, ProvinolsTM, delphinidin and ERa agonists, 17-b-estradiol and PPT, are able to induce endothelial vasodilatation
in aorta from ERa Wild-Type but not from Knock-Out mice, by activation of nitric oxide (NO) pathway in endothelial cells. Besides, silencing
the effects of ERa completely prevented effects of ProvinolsTM and delphinidin to activate NO pathway (Src, ERK 1/2, eNOS, caveolin-1) leading to NO production. Furthermore, direct interaction between
delphinidin and ERa activator site is demonstrated using binding assay
and docking. Most interestingly, the ability of short term oral administration of ProvinolsTM to decrease response to serotonin and to enhance
sensitivity of the endothelium-dependent relaxation to acetylcholine,
associated with concomitant increased NO production and decreased
superoxide anions, was completely blunted in ERa deficient mice. This
study provides evidence that red wine polyphenols, especially delphinidin, exert their endothelial benefits via ERa activation. It is a major
breakthrough bringing new insights of the potential therapeutic of
polyphenols against cardiovascular pathologies.
Paper No.: 2470
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
THE HYPOCHOLESTEROLEMIC EFFECT OF A NOVEL
SERIES OF COMPOUNDS WITH STRUCTURAL PROPERTIES
OF ALPHA-ASARONE AND FIBRATES IN TYLOXAPOL
TREATED MICE
Germán Chamorro(1), C Zúñiga(1), L Garduño(1), MdC Cruz(1),
R Pérez(1), F Labarrios(1), J Tamariz(1)
Escuela Nacional de Ciencias Biológicas, IPN, México D.F., México
Because of the putative link between serum cholesterol and incidence of
atherosclerosis, a novel series of ten analogs of a-asarone related with
fibrates has been previously prepared. Most of them are phenoxyacetic
acids or their methyl and ethyl esters which was substituted by a single
side chain of the different positions of the benzene ring. They were evaluated in a model of hyperlipidaemia induced in ICR male mice by a single
400 mg/kg intraperitoneal injection of tyloxapol. Mice were treated with
the drugs by gavage 1 h before and 22 and 48 h after the tyloxapol injection at doses of 0, 25, 50 or 100mg/kg. The derivatives exhibited potent
hypocholesterolemic activity, lowering the mice serum cholesterol up to
64.6%, low-density protein cholesterol up to33.8% and triacylglycerols
up to 70.2%. These findings have led identification of a class of compounds that represent a promising new class of hypolipidaemic drugs.
Paper No.: 2224
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
SECRETONEURIN INDUCES ENDOTHELIUM-DEPENDENT
RELAXATION OF THE PORCINE CORONARY ARTERY
Calvin KY Chan(1), JCW Mak(0), PM Vanhoutte(1)
(1) University of Hong Kong, Department of Pharmacology and
Pharmacy, Hong Kong, PR China
(2) University of Hong Kong, Department of Medicine, Hong Kong, PR
China
Secretoneurin is a part of the peptide encoded by the Secretogranin II
gene. Secretoneurin enhances the adhesion and transendothelial migration properties of monocytes. In addition, in regenerated endothelium,
the expression of the Secretogranin II gene is up-regulated. The objective
of the present study was to examine the effects of Secretoneurin on the
vascular reactivity. Isometric tension of rings with or without endothelium from porcine coronary arteries was measured in conventional organ
chambers. Secretoneurin did not induce contraction, but in preparations
contracted by the TP-receptor agonist U46619 it caused relaxation. This
relaxation was endothelium-dependent and reduced by the nitric oxide
synthase inhibitor Nomega-nitro-L-arginine methyl ester (L-NAME). It
was abolished by combined incubation with L-NAME and the cyclooxygenase inhibitor indomethacin. By contrast the relaxation to secretoneurin was not affected by TRAM 34 and UCL 1684. These results suggest
that secretoneurin induces relaxation by activation of both endothelial
nitric oxide synthase and cyclooxygenase, with the nitric oxide pathway
playing a more dominant role.
Paper No.: 981
FOCUSED CONFERENCE GROUP: P06 - THE HEART GONE
WRONG; STABILIZATION OF CARDIAC FUNCTION - BASIC
PROSTACYCLIN IP RECEPTOR ACTIVATION,
TRANSFORMING GROWTH FACTOR SIGNALLING AND
CARDIAC FIBROSIS IN HYPERLIPIDEMIC MICE
Elsa Chan(1), G Dusting (1), N Guo(1), H Peshavariya(1), R Dilley(1),
S Narumiya(2), F Jiang(3)
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
223
(1) O’Brien O’Brien Institute and University of Melbourne, Department
of Cytorpotection Pharmacology, Fitzroy, Victoria, Australia
(2) Kyoto University, Faculty of Medicine, Department of Pharmacology,
Kyoto, Japan
(3) Chinese Ministry of Education and Chinese Ministry of Health, Key
Laboratory of Cardiovascular ReModelling and Function Research, Qilu
Hospital, Shandong University, Jinan, Shandong Province, PR China
Prostacyclin IP receptor activation has been shown to protect against cardiac fibrosis but its molecular mechanisms in the heart remain unknown.
Using mouse cardiac fibroblasts, we examined whether IP activation with
cicaprost affects collagen expression by interfering with the signalling of
a key fibrotic factor transforming growth factor (TGF)-b. In fibroblasts,
TGF-b receptor inhibition suppressed collagen I mRNA by 25 ± 3%
(n = 3-4; P < 0.05) and the phosphorylation of TGF-b signalling protein
Smad2 was detected, suggesting that TGF-b regulates basal collagen
expression. Cicaprost reduced the basal expression of TGF-b responsive
genes including collagen I and plasminogen-activator inhibitor I (PAI-1)
indicating an inhibitory effect of IP receptor activation on TGF-b signalling. cAMP-elevating agents including cicaprost induce phosphorylation
of cAMP response element binding protein (CREB), which competes
with Smad for transcription coactivators such as CBP (CREB binding
protein)/p300 to reduce collagen synthesis. In fibroblasts with a CREB
mutant protein, cicaprost did not reduce expression of collagen and PAI1. Furthermore, double knockout of apolipoprotein E and IP receptor
[ApoE(-/-)/IP(-/-); n = 4-9; P < 0.05] in mice showed greater collagen
deposition (3-fold) and mRNA (5-fold) in the heart compared to controls
[ApoE(-/-)/IP(+/+)] following two-week angiotensin II infusion (1000 ng/
kg/min, SC). Angiotensin II-induced TGF-b mRNA in the heart was similar in both genotypes, suggesting that IP receptor activation did not affect
TGF-b gene expression, but its downstream signalling mechanism. Our
data suggest the protective effect of IP signalling operates at least partly
by antagonizing TGF-b mediated profibrotic effects in cardiac fibroblasts.
Paper No.: 1554
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
NADPH OXIDASE AND TGF-BETA SIGNALLING IN
EXTRACELLULAR MATRIX PRODUCTION IN CARDIAC
FIBROBLASTS
Elsa Chan(1), H Peshavariya(1), G Nancy(1), F Jiang(2), G Dusting(1)
(1) O’Brien Institute and University of Melbourne, Department of Cytorpotection Pharmacology, Fitzroy, Victoria, Australia
(2) Key Laboratory of Cardiovascular ReModelling and Function
Research, Chinese Ministry of Education and Chinese Ministry of
Health, Qilu Hospital, ShandongUniversity, Jinan, Shandong Province,
PR China
Oxidative stress, especially activation of the superoxide generating Noxcontaining NADPH oxidase is involved in the development of cardiac
fibrosis. We hypothesized that a profibrotic factor transforming growth
factor (TGF)-b acts via NADPH oxidase to induce collagen production
in mouse cardiac fibroblasts. TGF-b (10 ng/mL; n = 3) stimulated gene
expression of collagen I (by 2.1 ± 0.4 fold) and the fibroblast differentiation marker smooth muscle a-actin in mouse cardiac fibroblasts (by
2.8 ± 0.6 fold). EUK-134 a scavenger of superoxide and hydrogen peroxide suppressed the TGF-b-induced collagen gene expression (by
46 ± 13%; n = 4). By expressing a dominant negative form of NADPH
oxidase in fibroblasts, TGF-b-induced collagen gene expression was
inhibited, suggesting that NADPH oxidase is important to TGF b signalling in extracellular matrix production. Nox2 and Nox4 are the two
major isoforms found in cardiac fibroblasts. TGF-b upregulated Nox4
expression (2.9 ± 0.8 fold; n = 3) without affecting Nox2 in mouse cardiac fibroblasts. On the other hand, Nox1 was undetected suggesting that
Nox4 regulates TGF-b mediated responses. The effect of siRNA against
Nox4 gene expression on TGF-b-induced matrix synthesis is under
investigation. In conclusion, Nox4 may act as an effector molecule in
cardiac fibroblasts mediating fibrotic reactions by acting as an intermediary in the signalling of TGF-b.
Paper No.: 1502
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
CHARACTERIZATION OF VIP AND PACAP RECEPTORS IN
THE HUMAN CORONARY AND MENINGEAL ARTERY
Kayi Chan(1), M Baun(2), J Danser(1), I Jansen-Olesen(2), AM van
denBrink(1), S Gupta(2)
(1) Erasmus Medical Center, Department of Internal Medicine, Rotterdam, The Netherlands
(2) Glostrup University Hospital, Copenhagen, Denmark
Vasoactive intestinal peptide (VIP) and the pituitary adenylate cyclase
activating polypeptides (PACAPs) are involved in various processes,
including dilation of cephalic arteries. PACAP induces migraine-like
headache in migraine patients. Interestingly, PACAPs do not only stimulate VPAC receptors, like VIP, but also PAC receptors. We characterized
responses to PACAPs and VIP, as well as their VPAC/PAC receptors in
human coronary and meningeal arteries to obtain more knowledge about
their role in migraine. Concentration response curves to the VPAC/PAC
receptor agonists PACAP38 and PACAP27, as well as the VPAC receptor agonist VIP, were constructed in human coronary and meningeal
arteries in the absence or presence of the PAC receptor antagonist
PACAP6-38 or the VPAC receptor antagonist, PG-97269. mRNA
expression was measured using RT-PCR. PACAP38 and VIP induced
relaxations, which was higher in the coronary than in the meningeal
artery. The pEC50 of PACAP38 was smaller than the pEC50 of VIP in
all arteries. The antagonist did not block vasodilation, except for
PG97269, which blocked VIP-induced vasodilation in the coronary
artery. The mRNA expression of VPAC receptors was higher than that of
PAC receptors in coronary and meningeal arteries. PACAP38 and VIP
have a low potency and efficacy in meningeal arteries. PACAP38 is less
potent than VIP in human arteries. Except for VIP-induced vasodilation
in the coronary artery, our antagonist experiments do not confirm
involvement of either VPAC or PAC receptors, although mRNA of these
receptors is present. We suggest that PACAP38 may induce migraine-like
headaches via a mechanism not involving meningeal vasodilation.
Paper No.: 1503
FOCUSED CONFERENCE GROUP: PW19 - INFLUENCE OF
DEGENERATION AND REPAIR IN THE CNS AND PERIPHERY
CHARACTERIZATION OF THE CGRP RECEPTOR
ANTAGONIST TELCAGEPANT (MK0974) IN HUMAN
ISOLATED CORONARY ARTERIES
Kayi Chan(1), L Edvinsson(2), S Eftekhari(2), R de Vries(1), S Kane(3),
R Hargreaves(3), J Danser(1), AM van denBrink(1)
(1) Erasmus Medical Center, Department of Internal Medicine, Rotterdam, The Netherlands
(2) Lund University Hospital, Malmö, Sweden
(3) Merck Research Laboratories
Calcitonin gene-related peptide (CGRP) plays a role in primary headaches and CGRP antagonists are effective in migraine treatment. Since
CGRP is also a potent vasodilator in coronary arteries, antagonism of its
receptor might cause coronary side effects. Therefore, we investigated
the effect of telcagepant (MK-0974) on human coronary arteries with different internal diameter. Human alfaCGRP induced concentration-dependent relaxations which was larger in distal (id: 600-1000lm,
Emax=83 ± 7%) than in proximal coronary arteries (id: 2-3mm,
Emax=23 ± 9%) and in coronary arterioles (id: 200-300lm, Emax=15 ±
7%). Pre-treatment with telcagepant (10 nmol/L to 1 lmol/L) antagonized the alfaCGRP-induced relaxation in a competitive manner with a
pA2 value of 8.43 ± 0.24 in the distal coronary arteries. In proximal coronary arteries and coronary arterioles, telcagepant (1 lmol/L) antagonized the alfaCGRP-induced relaxations with a pKB of 7.89 ± 0.13 and
7.78 ± 0.16 respectively. Telcagepant did not induce contraction or relaxation per se. alfaCGRP significantly increased cAMP levels and this was
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
224
abolished by pretreatment with telcagepant in distal coronary arteries.
Immunohistochemistry revealed the expression and co-localization of the
receptor elements calcitonin like receptor (CLR) and receptor activity
modifying protein 1 (RAMP1) in the smooth muscle cells in the media
layer of human coronary arteries. The findings expand our understanding
of on the peripheral vascular properties of telcagepant. When extrapolated to the clinical situation, telcagepant is unlikely to induce coronary
side effects under normal conditions in cardiovascular healthy patients.
Paper No.: 1440
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
CALCIUM CHANNEL IS INVOLVED IN NEOMYCIN-INDUCED
HAIR CELL TOXICITY IN THE ZEBRAFISH LATERAL LINE
Ming-Huan Chan, C-F Liu, H-H Chen
Tzu Chi University, Department of Pharmacology, Hualien, Taiwan
Aminoglycosides are commonly and clinically used antibiotic agents that
cause severe side effects, such as hearing loss. The extracellular divalent
cations can modulate aminoglycoside-induced sensory hair cell death.
However, the role of cytoplasmic membrane calcium channel in aminoglycoside ototoxicity is not clear. In this study, the lateral lines of zebrafish were used to investigate the effects of verapamil, a calcium channel
blocker, and Bay K8644, a calcium channel activator, on neomycininduced hair cell toxicity, which was detected in vivo by the dye FM143FX, a reliable indicator to assess hair cell viability. Results showed
that hair cells from neomycin toxicity were protected under high calcium
(3.3 mM) conditions. On the contrary, lower calcium (33 microM)
enhanced hair cell death mediated by neomycin treated for 30 minutes.
Importantly, verapamil was observed to attenuate neomycin-induced
toxic response in calcium concentrations at 33 and 330 microM. Bay
K8644, in contrast, significantly reversed the protective action of high
calcium on neomycin-induced hair cell loss. These findings indicate that
both extracellular calcium concentration and calcium channel activity
influence hair cell loss from neomycin toxicity.
Paper No.: 671
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
GASTROPROTECTIVE ACTIVITY OF ANNONA MURICATA L.
ON ETHANOL INDUCED GASTRIC LESIONS IN RODENTS
Pit Foong Chan, R Abdul Hamid
University Putra Malaysia, Faculty of Medicine and Health Sciences,
Department of Biomediacal Science, Selangor, Malaysia
Annona muricata L (AML), locally known as ‘‘durian belanda’’ belonging to family Annonaceae, is a plant been known to have various pharmacological and nutritional properties in folklore medicine (Dos Santos
et al, 2001; 8: 115-120). The antiulcerogenic effect of a crude hydroalcoholic (80%) extract obtained from AML leaves was evaluated in rodents
by employing absolute ethanol induced ulcer model. AML was administered orally (p.o.) 1h prior to ulcer induction in the dose range of 10, 30,
100 and 300mg/kg. Results of the study revealed that AML at the given
dose range significantly inhibited the ulcerative lesion in a dose-dependent manner by 22.44, 51.77, 75.43 and 92.80% respectively. Oral
administration of AML (300mg/kg) to L-NAME (a NO-synthase inhibitor)-pre-treated animals, did not reduce the antiulcerogenic activity of
AML, suggesting that the mechanism of pharmacological activity has no
relationship with the nitric oxide (NO). The pre-treatment with N-ethymaleimide, a thiol blocker, including mucosal nonprotein sulfhydryl
groups, increased the gastric lesions in comparison for control groups
(186.33 ± 18 mm2 with saline versus 379.67 ± 15.47 mm2 with NEM).
The rats treated with AML (300mg/kg) inhibited the gastric lesions as
82.02% (pre-treated with saline) and 58.21% (pre-treated with NEM).
This significant attenuation of gastric lesions showed that the crude
hydroalcoholic extract of AML has active substances that increase the
mucosal nonprotein sulfhydryl groups contents for the gastroprotection
effects.
Paper No.: 1202
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
BIOEQUIVALENCE STUDY OF TWO FORMULATIONS OF
35MG TRIMETAZIDINE MODIFIED RELEASE TABLETS IN
THAI HEALTHY VOLUNTEERS UNDER FASTING AND FED
CONDITIONS
Weerawadee Chandranipapongse, T Ruangnapa, S Ngokpol,
K Sathirakul, P Pongnarin, S Kongpattanakul
Mahidol University Faculty of Medicine & Siriraj Hospital, Department
of Pharmacology, Bangkok, Thailand
The generic product of trimetazidine, an anti-anginal agent, is now available in Thailand. Bioequivalence study is used to demonstrate that the
generic and the original drug products have comparable rate and extent
of absorption. Thus, the objective of this study was to compare the bioavailability of two modified release tablets of 35mg trimetazidine dihydrochloride (Matenol MR; generic product and Vastarel MR;
innovator’s product). This study was a single-dose, two-treatment, twoperiod, two-sequence randomized crossover design under fasting and fed
conditions with a minimum of 7 days washout period. Twenty-four
healthy Thai male and female volunteers were enrolled. For both conditions, the volunteers were received a 35mg trimetazidine modified release
tablet of both formulations. Each volunteers were obtained blood samples
16 times over a period of 24 hours after each oral administration. The trimetazidine plasma concentrations were quantified using a validated
method employing liquid chromatography with tandem mass spectrometry with the lower limit of quantification of 0.25 ng/mL. All of the pharmacokinetic parameters were investigated using non-compartmental
analysis model. The results demonstrated that the 90% CI of the geometric mean ratio of Cmax, AUC0-t and AUC0-¥ were within the equivalence criteria (80.00-125.00%) which were 105.53% (95.71%-116.36%),
104.28% (96.24%-112.98%, and 105.26% (96.61%-114.67%) under fasting condition and 110.21% (102.72%-118.25%), 101.95% (94.33%119.19%), and 99.7% (91.18%-109.02%) under fed condition, respectively. No statistical differences of median Tmax (p > 0.05) between two
formulations in both conditions were observed. Furthermore, both preparations were well tolerated and had a few non-serious adverse events. In
conclusion, these two trimetazidine products were bioequivalent.
Paper No.: 1253
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
THE EFFECT OF C825T POLYMORPHISM OF THE GNB3
GENE ON VALPROATE-INDUCED METABOLIC
ABNORMALITIES IN BIPOLAR DISORDER PATIENTS
Hui Hua Chang(1), PW Gean(2), YK Yang(3), HC Huang(3),
HC Wang(3), RB Lu(0), PS Chen(0)
(1) National Cheng Kung University, College of Medicine, Institute of
Biopharmaceutical Sciences, Tainan, Taiwan
(2) National Cheng Kung University, College of Medicine, Department
of Pharmacology, Tainan, Taiwan
(3) National Cheng Kung University, Hospital and College of Medicine,
Department of Psychiatry, Tainan, Taiwan
(4) National Cheng Kung University, College of Medicine, Institute of
Behavioral Medicine, Tainan, Taiwan
(5) National Cheng Kung University Hospital, Department of Psychiatry,
Dou-liou Branch, Taiwan
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
225
Valproate (VPA) is a mood stabilizer for treating patients with bipolar
disorder. It may cause metabolic abnormalities in certain bipolar patients.
However, the genetic factors that influcence the susceptibility remain
unclear. Genetic polymorphism of the G-protein 3 Subunit (GNB3) is
reported to be associated with metabolic phenotypes. In the current study,
we investigated the possible associations between the GNB3 variation
and VPA-induced metabolic abnormalities. Subjects (n = 96) who met
the DSM-IV criteria for bipolar disorder were recruited from the National
Cheng Kung University Hospital. Their metabolic indexes were measured. The variation of GNB3 825C/T showed an association with higher
plasma total cholesterol (p = 0.037), triglyceride (p = 0.014), and leptin
(p < 0.001) levels in bipolar disorder patients treated with VPA. After
adjusting for age, gender, types of bipolar disorders, serum concentration
of VPA, the variation of GNB3 825C/T remained significantly associated
with the levels of serum leptin and BMI (p < 0.001 and p = 0.025,
respectively). In addition, the GNB3 825C/T showed significant
drug*SNP interactions with TG levels (p = 0.013), leptin levels
(p = 0.013), and BMI (p = 0.018). These results indicated that the T
allele may be associated with lower serum leptin level and BMI in BD
patients treated with VPA.
Paper No.: 1292
FOCUSED CONFERENCE GROUP: P19 - GENERAL
SESSION - ONCOLOGY
INDOLE-3-CARBINOL ATTENUATES MIGRATION AND
INVASION OF BREAST CANCER CELLS BY BLOCKING THE
ERK/SP1-MEDIATED MMP-2 GENE TRANSCRIPTION
Hui-Chiu Chang
Kaohsiung Medical University, Graduate Institute of Medicine, Department of Medical Laboratory Science and Biotechnology, Kaohsiung,
Taiwan
Indole-3-carbinol (I3C) is the major bioactive food component of cruciferous vegetables and has been shown to exhibit inhibition of invasion
activity on various cancer cell lines. This study addressed the effect of
I3C on the expression of matrix metalloproteinases (MMPs) and clarified
the underlying mechanism. Migration, invasion, MMP-2 activity, and
MMP-2 mRNA level of MCF-7 breast cancer cells were inhibited by
I3C in a dose-dependent manner. Promoter deletion and mutation analysis suggested that I3C inhibited MMP-2 gene transcription via the -85/7 bp promoter region and the Sp1 transcription factor binding site
located within the -72/-64 bp promoter region was important for the inhibition. Chromatin immunoprecipitation assay confirmed that Sp1 proteins
constitutively bound to this consensus sequence in vivo and that the binding was attenuated by I3C. In addition, I3C inhibited the extracellular
signal-regulated kinase (ERK) signalling pathway in MCF-7 cells. The
results suggest that I3C attenuates MMP-2 expression by blocking the
ERK/Sp1-mediated gene transcription to reduce migration and invasion
of breast cancer cells.
Paper No.: 1201
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
BIOEQUIVALENCE STUDY OF 10 MG RAMIPRIL TABLETS IN
HEALTHY THAI VOLUNTEERS
Somruedee Chatsiricharoenkul, W Thangboonjit, K Sathirakul,
P Pongnarin, K Konhan, S Kongpattanakul
Mahidol University Faculty of Medicine & Siriraj Hospital, Department
of Pharmacology, Bangkok, Thailand
The objective of this study was to determine the bioequivalence of
10 mg dose of ramipril tablets between the test product (Ramtace
10 mg, Unison Laboratories, Thailand) and the reference product (Tritace 10 mg, Aventis Pharma S.P.A., Italy). The study was carried out
with a single dose, 2-treatment, 2- period, 2-sequence randomized cross-
over design under fasting condition with a minimum of 14 days washout
period in 24 healthy Thai male and female volunteers. Plasma samples
for determination of ramipril and ramiprilat were obtained pre-dose and
at frequent intervals for up to 72 h post dose. Ramipril and ramiprilat
plasma concentrations were quantified by a validated method employing
high performance liquid chromatography with tandem mass spectrometry
(HPLC-MS/MS) method with the lower limit of quantification (LLOQ)
of 0.05 ng/mL and 0.25 ng/mL for ramipril and ramiprilat, respectively.
All of the pharmacokinetic parameters were investigated using non-compartmental analysis. The result demonstrated the 90% confidence interval
(90%CI) of the geometric mean ratio (test/reference) of Cmax, AUC0-72
and AUC0-¥ of ramipril was 97.26% (84.50%-111.93%), 100.70%
(89.47%-113.34%) and 100.29% (88.90%-113.15%), respectively which
were within the equivalent criteria (80.00%-125.00%). For ramiprilat, the
90% CI for Cmax, AUC0-72 and AUC0-¥ was entirely within the equivalence criteria which were 108.87% (103.00%-115.07%), 104.93%
(100.50%-109.55%) and 103.30% (98.03%-108.85%) for Cmax, AUC0-72
and AUC0-¥, respectively. In conclusion: the 90% confidence intervals
for log-transformed geometric mean test/reference formulation ratios of
primary parameters were entirely within 80.00%-125.00%. Thus, it can
be concluded that the test formulation was bioequivalent to the reference
formulation.
Paper No.: 1492
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
A-LINOLENIC ACID PROTECTS RENAL CELLS AGAINST
PALMITIC ACID LIPOTOXICITY VIA REDUCTION OF
ENDOPLASMIC RETICULUM STRESS
Prabal Chatterjee, E Katsoulieris, J Mabley, M Samai, I Green
University of Brighton, Centre for Biomedical and Health Science
Research, Brighton, UK
Elevated levels of saturated fatty acids (FA) are associated with diabetes/
obesity and with development of diabetic nephropathy. The aim of this
study was to investigate the effects of palmitic acid (PA), a saturated FA,
on renal cells and to evaluate the potential protection afforded by a-linolenic acid (a-LA), a polyunsaturated FA. Cultures of NRK-52E cells, a
rat proximal tubular cell-line, were incubated with PA for 24 h in the
absence or presence of a-LA. Apoptotic cell death was measured using
Hoechst-propidium iodide staining. The expression of CHOP and phosphorylated eIF2a (p-eIF2a) were determined using Western blotting and
used as indicators of endoplasmic reticulum (ER) stress. PA (300 lmol/
L) increased apoptosis from 1.2 + /- 0.2% to 17.1 + /- 2.6% (n = 4,
P < 0.05). PA increased CHOP expression from 0.19 + /- 0.04 to
0.88 + /- 0.14 arbitrary units (au) (n = 6, P < 0.05) and p-eIF2a from
0.32 + /- 0.03 to 0.70 + /- 0.03 au (n = 5, P < 0.05). In the presence of
a-LA (100-300 lmol/L), PA-mediated apoptosis was reduced from
17.1 + /- 2.6% to 4.2 + /- 0.7% (n = 4, P < 0.05), CHOP expression
was reduced from 0.88 + /- 0.14 to 0.22 + /- 0.06 au (n = 6, P < 0.05)
and p-eIF2a expression from 0.70 + /- 0.03 to 0.42 + /- 0.02 au (n = 5,
P < 0.05). In conclusion, these results suggest that a-LA can reduce the
apoptosis of renal cells caused by PA and that this protection involves
reduction of ER stress.
Paper No.: 967
FOCUSED CONFERENCE GROUP: P16 - NATURAL PRODUCTS:
PAST AND FUTURE?
PHARMACOLOGICAL EXPLORATION OF HYPERICUM
PERFORATUM GROWING IN INDIA USING RODENT MODELS
Shyam Sunder Chatterjee(1), V Kumar(2)
(1) Herbal Consultant, Karlsruhe, Germany
(2) Banaras Hindu University, Institute of Technology, Pharmacology
Laboratory,Department of Pharmaceutics, Varanasi, India
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
226
All therapeutic trials with Hypericum perforatum extracts reported during
past decades have concentrated on its antidepressants like efficacy only.
Preclinical studies have revealed though, that such extracts possess
diverse other therapeutically interesting pharmacological activities as
well. Critical analyses of available preclinical information on such
extracts and their bio-active constituents lead speculate that they could as
well be medicinally used as adaptogens suitable for coping with, or ameliorating, health problems caused by environmental and/or metabolic
stress. Efforts made to experimentally verify this working hypothesis
revealed clear dose dependant anti-aggressive activity of one such extract
in a battery of five rodent behavioral models commonly used to test antistress activities of psycho-active drugs and agents. Similar was the case
also in rodent metabolic disorders models for hyperglycemia and hypercholesterolemia; two conditions often associated with environmental
stress. Effective oral daily doses of the extract in these models were similar to those needed to observe its anti-depressant like efficacy in rodent
models. Although analogous was also the case for hyperforin, quantitatively the observed effects of the extract was higher than expected by its
hyperforin content. These observations lend further experimental evidences to our conviction that pharmacologically the tested extract cannot
be defined as an antidepressant only, and that hyperforin is not its only
bio-active component.
Rasayan drugs act inside the human body by modulating the neuroendocrino-immunesystems and have been found to be a rich source of
possible therapeutic measure has become a subject of active scientific
investigations. Vajikaran is one of the eight branches that deal with
improving male sexual potency and thereby ensuring a supraja, or better
progeny. The main aim of Vajikaran besides achieving successful copulation for healthy reproduction, with sexual pleasureis an additional benefit.
The plant Curculigo orchioides, Astercanthalongifolia, Mucuna pruriens
are well known vajikaran rasayan herbs. The studywas therefore performed to effect of these plants on reproductive parameters.Following
parameters were evaluated the effect of extract on body and organweights, change in histoarchitecture of testis, fructose level in seminalvesicles and hormonal level was studied (Chauhan et al., Fitoterapia 2007;
78:530-534). Administration of ethanolic extract had pronounced anabolic andspermatogenic effect in treated animals as evidenced by weight
gains in the bodyand reproductive organs. Increase in spermatogenesis
was shown in all treatedgroup. The level of follicular stimulating hormone, leutinizing hormone andtestosterone level is significantly
increased in extract treated group andfructose content in seminal vesicles
was significantly increases in treatedgroups. Thus it was concluded that
drug was justifying the use in thetraditional system of medicine as a
vajikaran rasayana.
Paper No.: 1004
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
STRUCTURE ACTIVITY RELATIONSHIP (SAR) OF
HYPERFORIN FOR ITS ANTIDEPRESSANT-LIKE ACTIVITY
PROFILE
Paper No.: 2241
FOCUSED CONFERENCE GROUP: P06 - THE HEART GONE
WRONG; STABILIZATION OF CARDIAC FUNCTION - BASIC
EFFECT OF PRE-ISCHAEMIC CONDITIONING ON ANOXIC
DOPAMINE EFFLUX IN THE MOUSE CAUDATE
Shyam Sunder Chatterjee(1), O Kristhal(2)
(1) Herbal Consultant, Karlsruhe, Germany
(2) Bogomoletz Institute of Physiology, Kiev, Ukraine
Hyperforin is a labile, but quantitatively the major, bio-active secondary
metabolite of Hypericum perforatum with a broad spectrum of therapeutically interesting pharmacological activity profile. Initial SAR studies
have indicated that the phloroglucinol moiety of this structurally complex
molecule could be its essential pharmacophore, and that mono-acylated
hyperforin derivatives could lead to pharmaceutically stable molecules
with antidepressant-like activity profiles. Our more recent SAR studies
revealed that such molecules possess also a broad spectrum of ion channels related functions modulating activity profile of major ionotropic
mechanisms. These efforts were made to identify structurally simpler and
pharmaceutically stable molecules with activity profiles analogous to hyperforin suitable for drug development purposes. They led not only to a
library of phloroglucinol derivatives with diverse spectra of therapeutically interesting ion channels modulating activities, but also to several
symmetrically di-acylated phloroglucinols with hyperforin-like activity
profiles in a battery of in vitro pharmacological screening models. However, in animal models none of these di-acylated phloroglucinols
revealed anti-depressant-like activities. Surprisingly though, a few other
phloroglucinol derivatives with no similarity to the activity profile of hyperforin observed in vitro, were almost as active as hyperforin in
behavioural models of depression. Therefore, we conclude that observed
ion channel functions and biogenic amine reuptake modulating effects of
hyperforin are not involved in its antidepressant like efficacy.
Paper No.: 416
FOCUSED CONFERENCE GROUP: P08 - DEVELOPMENTS IN
THE TREATMENT OF SEXUAL DYSFUNCTION AND
DISEASES OF THE LOWER URINARY TRACT
ROLE OF RASAYAN HERBS ON REPRODUCTIVE
PARAMETERS OF MALE RATS
Nikky Kanti Chauhan, AMJ Young, CL Gibson, C Davidson
University of Leicester, School of Psychology, Leicester, UK
Pre-ischaemic conditioning (PIC) with brief periods of ischaemia has been
shown to be neuroprotective against a subsequent ischaemic event. Here
we examined whether short periods of PIC can affect hypoxia induced
dopamine (DA) release in the mouse caudate. Adult male mice (C57Bl/6
~30 g) were killed by cervical dislocation and their brains rapidly
removed. Brain slices (400 lm) were kept in oxygenated artificial cerebrospinal fluid (aCSF) at room temperature (22C). After at least 45 min
equilibration, slices were transferred to a brain slice chamber and perfused
with oxygenated aCSF at 33C. After a further 45 min equilibration they
were subject to PIC (0, 5 10 or 15 min) comprising hypoxia (N2/CO2)
and reduced glucose (10 to 2 mM) aCSF, then, 60 min later, a 15 min
hypoxic episode. DA efflux was measured using fast cyclic voltammetry
at carbon fibre electrodes. We recorded a) time from start of hypoxia to
start of DA release; b) time from start of DA release to peak DA release;
c) peak DA release and d) initial rate of DA release (dDA/dt). Data were
analyzed with 1-way ANOVA. The main result was that 10 and 15 min
PIC increased the time to onset of DA efflux during the second hypoxic
event (F(3, 18) = 10.7, P < 0.001) values are 349, 561, 684 and 745s for
0, 5, 10 & 15 min PIC respectively. These data suggest that PIC of 10 and
15, but not 5 min are neuroprotective in the mouse caudate.
Paper No.: 3231
FOCUSED CONFERENCE GROUP: P12 - ION
CHANNELOPATHIES: NEW WINDOWS ON COMPLEX
DISEASE AND THERAPY
GRP-78 IS A POSSIBLE TARGET IN
GABAA-CHANNELOPATHIES
Ksenia Chekina
Nagendra Chauhan, VK Dixit
Research Institute of Pharmacology, RAMS, Moscow, Russian
Federation
Natural Product Research Laboratory, Department of Pharmaceutical
Sciences, Dr. Hari Singh Gour Vishwavidyalaya, Sagar, India
The pharmacologic analysis of several nontypical anxiolytics (Afobazol,
Ladasten, Noopept, GB-115) revealed that the analyzed compounds have
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
227
number of properties in common: antioxidant activity, Ca+2-current stabilisation, antiapoptosis effect, and ability to prevent a stress induced
decrease in benzodiazepine binding. Furthermore, the literature analysis
revealed interactions between GABAAcomplex, glucose metabolism and
Ca+2-current regulation. In additional, there are literature data that some
of the mud stabilisation drugs direct effect on GRP78 expression. GRP78
(BIP) – is a glucose-regulated chaperon which has binding domains for
Ca+2, caspase, nucleotides, and glucose. For their structure, GRP78 proteins function in neurons as adjusters of Ca+2-current, anti-apoptosis regulators, and starters of antioxidant cascades. It is not clear is GRP78
involved in GABAAcomplex regulation or not, but background of this
assumption consists of many substituted data. In the present paper we
bring out an indirect evidence of interactability between GRP78 and
GABAAcomplex. Yohimbine is a standard-like anxiogenic compound
which, as it shown in literature, leads to decrease in benzodiazepine binding in intact brains and breaks normal Ca+2 current by blocking Na+-channel. If GRP-78 is involved in GABAAcomplex regulation, the
yohimbine-induced decrease in benzodiazepine binding should be levelled
by excess of glucose. In the present study we demonstrate a dose-independent yohimbine-induced decrease in benzodiazepine binding in sinaptosomas and control level of benzodiazepine binding in sinaptosomas after
glucose-enrich incubation medium either with or without yohimbine in it.
So, our present results would be a basis of availability of further research
on GRP-78 as one of possible regulators of GABAA-Cl- channel.
Paper No.: 1761
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
IN VITRO ANTIPLASMODIAL ACTIVITY OF
8-HYDROXYQUINOLINE DERIVATIVES
National Health Research Institutes, Division of Biotechnology and Pharmaceutical Research, Zhunan, Taiwan
Type II diabetes is one of the common diseases worldwide and
accounted for 90% of the total population of patients with diabetes mellitus. It is a chronic disease status with reduced insulin effectiveness and
glucose tolerability. Normal glucose homeostasis is maintained by gastrointestinal hormones such as the insulinotrpoic incretins family of two
principal members glucose-dependent insulinotropic polypeptide and glucagon-like peptide-1. Both incretins have a short circulation half-life due
to rapid inactivation by the proteolytic enzyme dipeptidyl peptidase IV
(DPP-IV). One therapeutic strategy is, therefore, to maintain the blood
levels of incretins by the inhibition of DPP-IV activities. BPR1G462 is a
novel synthetic small compound that inhibits the DPP-IV activity. We
demonstrated that BPR1G462 is orally active and dose-dependently
inhibits the plasma DPP-IV activities in Sprague-Dawley rats and in dietinduced obese (DIO) mice. Oral administration of BPR1G462 caused a
significant increase in the plasma levels of active glucagon-like peptide-1
and insulin activities in rats. BPR1G462 also increased the oral glucose
tolerability in lean and DIO mice. BPR1G462 is now a candidate for further preclinical and clinical developments for curing Type II diabetes.
Paper No.: 1695
FOCUSED CONFERENCE GROUP: P19 - GENERAL
SESSION - ONCOLOGY
PEPTIDE-CONJUGATED BIODEGRADABLE
NANOPARTICLES AS A CARRIER TO TARGET PACLITAXEL
TO TUMOR NEOVASCULATURE
Hong-Zhuan Chen, D-H Yu, Q Lu, J Xie, C Fang,
Chien-Teng Chen, R van Zyl
Shanghai JiaoTong University School of Medicine, Institute of Medical
Sciences, Department of Pharmacology, Shanghai, PR China
University of the Witwatersrand, Department of Pharmacy and Pharmacology, Johannesburg, South Africa
In present study, we developed a novel drug delivery system (DDS),
nanoparticles conjugated with K237-(HTMYYHHYQHHL) peptides for
tumor neovasculature targeting drug delivery. Paclitaxel, a chemotherapeutic agent with potent antiangiogenic activity, was used as a prototype
drug. We synthesized the aldehyde poly(ethylene glycol)–poly(lactide)
(aldehyde-PEG-PLA) block copolymer by ring opening polymerization.
The nanoparticles loading paclitaxel (PTX-NP) were fabricated using the
O/W emulsion and evaporation technique. K237 ligand, a peptide that can
bind to the KDR receptors predominantly expressed on the surface of
tumor neovasculature endothelial cells with high affinity and specificity
and inhibit the VEGF-KDR angiogenic signal pathway, was conjugated
to the aldehyde group of PEG chain using the N-terminal PEGylation
technique. The K237 conjugated paclitaxel-loaded nanoparticles (K237PTX-NP) had a hydrodynamic diameter of 150 nm. The K237 density on
nanoparticle surface was 474. The K237 conjugated nanoparticles could
be significantly internalized by human umbilical vein endothelial cells
(HUVEC) through K237-KDR interaction, and this facilitated uptake led
to the expected enhanced antiangiogenic activity shown by HUVEC proliferation, migration and tube formation compared to cells treated with
Taxol?and PTX-NP. The long-circulating property and K237 ligand of
K237-PTX-NP warranted rapid, long-term, and accurate in vivo tumor
neovasculature targeting, and thereafter the significant apoptosis of tumor
neovasculature endothelial cells and necrosis of MDA-MB-231 breast
tumors implanted in female BLAB/c nude mice. This nanoparticulate
DDS offers a new strategy for paclitaxel chemotherapy application and it
could also be used to carry other chemotherapeutic drugs, genes, and proteins with antiangiogenic activity for antiangiogenic cancer therapy.
8-Hydroxyquinoline is a derivative of heterocycle quinoline with reported
antiseptic, disinfectant, antifungal, antiprotozoal, metal chelating and anticancer properties. It is structurally related to antimalarial compounds such
as quinine and chloroquine and derivatives could inhibit the in vitro
growth of the malaria parasite, as well as the formation of the malaria pigment, haemozoin. The antimalarial properties of the 8-hydroxyquinoline
and 14 derivatives were screened using the radiolabelled hypoxanthine
uptake method over a period of 48 hours. Haemozoin formation was
observed by incubating the compounds with haemozoin crystal precursors
under acidic conditions for 24 hours, while toxicity of the compounds
was screened against human kidney epithelial (Graham) cells using the
MTT–cell viability assay. 8-Hydroxyquinoline was the most effective of
all the compounds screened (0.432 lM) as compared to quinine
(0.099 lM) against malaria. The antimalarial activity was not related to
the inhibition of haemozoin formation at a molar ratio of 5:1 (IC50:
959 lM) to haematin compared to quinine (0.2:1; IC50: 45 lM). At
1 lM, 8-hydroxquinoline exhibited 25% haemolysis compared to 1% for
quinine. 8-Hydroxyquinoline was 75 times more cytotoxic than quinine
against the kidney epithelial cells. A dichloro- substitution was more
active than dibromo-, dimethyl- and diiodide- substitutions (IC50 value:
1.709 > 4.879 > 6.083 > 7.771 lM, respectively). An amino substitution
increased the antimalarial activity compared to a nitro substitute (0.893
and 4.396 lM, respectively). Elucidating the mechanism of action of
8-hydroxyquinoline as an antimalarial is needed.
Paper No.: 2192
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
DISCOVERY OF A NOVEL DIPEPTIDYL PEPTIDASE-IV
INHIBITOR BPR1G462
Chiung-Tong Chen, K-C Yeh, C-Y Huang, W-T Jiaang, C-B Hu,
Y-W Huang, T Hsu, J-S Song, T-K Yeh, C-H Chien, S Chen, Y-S Chao
Paper No.: 2044
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
ACUTE EFFECT OF SULFAPHENAZOLE ON ISOLATED
MESENTERIC SMALL ARTERIES FROM DB/DB MICE
Hua Chen, U Simonsen, C Aalkjaer
University of Aarhus, Department of Physiology, Aarhus, Denmark
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
228
db/db mice are fat and hyperglycemic, and an established model for
human type 2 diabetes. Experiments were designed to investigate
whether treatment with sulfaphenazole, a specific cytochrome P450 2C9
inhibitor, could improve endothelial dysfunction in mesenteric small
arteries of db/db mice in vitro. Obese db/db mice and lean db/+ littermates, aged 9-12 weeks, were sacrificed after cervical dislocation. First
or second-order mesenteric small arteries were dissected and mounted
for isobaric force measurements. Intracellular calcium ([Ca2 + ]i) transients and production of reactive oxygen species (ROS) were measured
by a confocal laser scanning microscope. Impaired relaxations to acetylcholine (ACh) were observed in mesenteric resistance arteries from db/
db mice. Incubation with 10 M sulfaphenazole for 30 min significantly
improved ACh-induced relaxation in these mice. The endothelial cell
[Ca2 + ]i transients induced by ACh were significantly lower and cytosolic ROS signals significantly more pronounced in arteries from db/db
mice. Both abnormalities were normalized after sulfaphenazole incubation. The present study shows that sulfaphenazole enhances endothelial
dysfunction by decreasing ROS production and increasing [Ca2 + ]i
within endothelial cells. These observations may suggest an effect of
ROS on endothelial cell Ca2 + handling and consequently for endothelial
function in diabetes.
Paper No.: 495
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
THE ADJUVANT EFFECTS OF SPECIFIC
PHYTOCOMPOUNDS ON TUMOR CELL LYSATE-PULSED
DENDRITIC CELL-BASED VACCINE VIA INDUCTION OF
IMMUNOGENIC TUMOR CELL DEATH
Hui-Ming Chen(1,2), S-S Chen(3), N-S Yang(1)
(1) Department and Institute of Pharmacology, National Yang Ming
University, Taipei, Taiwan
(2) Agricultural Biotechnology Research Center, Academia Sinica,
Taipei, Taiwan
(3) IgE Therapeutics, Inc., Department of Allergy and Immunology, San
Diego, California, USA
An emerging concept of ‘‘a key-lock paradigm’’ between the immunogenic cell death and dendrite cell (DC) is recently proposed. Immunogenic cell death is characterized by immunogenic factors (HMGB1,
calreticulin, heat shock proteins), which are helpful for the maturation,
antigen uptake and presentation of DC and serve as powerful immunological adjuvants. Whether some specific phytocompounds claimed
to have anti-tumor effects can deliver the immunogenic signals and
keep expending the tumor-recognition factors is a challenging question. To address this possibility, we first determined the expression
profiles of immunogenic factors in tumor cell lysate (TCL) obtained
from B16 cells treated with test phytocompounds (emodin, epigallocatechin-3-gallate, curcumin, shikonin and it derivatives) and chemotherapeutic drugs (doxorubicin and pacilitaxol) at varying
concentrations. Anthracyclines-Dorxorubicin is reported to confer a
higher capacity in eliciting immunogenic cell death. Shikonin, similar
to doxorubicin, can induce tumor cell death and specific immunogenic
factors expression (Hsp70 and Hsp90) in TCL in a dose-dependent
manner, but decrease the survivin expression. The cytotoxic effects of
splenocytes on target cancer cells were much higher in TCL-shikoninDC vaccinated mice than in TCL-DMSO-DC vaccinated mice. TCLshikonin-DC vaccination can substantially decrease the tumor growth
and maintain over 70% of survival rate of tumor-bearing mice after
35 days of tumor cell injection. In conclusion, shikonin is shown here
to sensitize tumor cells to the lytic effects of DC-activated immune
effector cells, which may warrant future evaluation as adjuvant of
tumor lysate-pulsed DC vaccine.
Paper No.: 957
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
THEOPHYLLINE-SUBSTITUTED KMUP-1 INHIBITS
ALLERGEN-INDUCED PULMONARY VASCULAR AND
BRONCHIAL INFLAMMATION: ENOS-ENHANCEMENT AND
MMP-9-SUPPRESSION
Ing-Jun Chen
Kaohsiung Medical University, Department of Pharmacology,
Kaohsiung, Taiwan
KMUP-1 has been proposed to be a potential treatment for vascular and
bronchial inflammation characterized by chronic asthma and obstructive
pulmonary disease. Theophylline has been recognized as a bronchodilator in the treatment of asthmatic attacks and bronchitis related to an immunoresponse. However, a low therapeutic margin of safety has led it
hard to be used. We here investigated the effects of inhaled KMUP-1 in
the non-asthmatic mice and the asthmatic model of ovabumin (OVA)sensitized and challenged mice. Mice were allocated to either: 1) a short
term inhalation protocol, to receive KMUP-1 (1-5 mM) for 30 min in
mice; or 2) a 28-day inhalation protocol to treat asthmatic mice, sensitized with intra-peritoneal OVA on day 1 and day 8 and challenged with
OVA nebulization and treatment with KMUP-1 nebulization (5 mM/30
mins/day) on day 21-27. In both protocols, expression of endothelial
nitric oxide synthase (eNOS) was increased and matrix metalloproteinases-9 (MMP-9) was decreased in lung tissue. Pretreatment of L-NAME
(12 mM, 15 min) reversed these expressions in the short-term experiment. In 28-day experiment, OVA-sensitized decrease of sGCa1 and
PKG and increase of iNOS, ICAM-1, and VCAM-1 expression were
reversed by KMUP-1. Cell count, NO metabolite (NOx) and MMP-9activity in bronchoalveolar lavage fluid (BALF) were decreased. KMUP1 reduced OVA-sensitized vascular and bronchial wall thickening,
eNOS-immunostaining at the alveolar septa and MMP-9-immunostaining
in the bronchioles and infiltrated inflammatory cells in the peri-vascular
and peri-bronchiolar region. KMUP-1 can be useful in inhibiting pulmonary vascular and bronchial inflammation in chronic asthma and obstruction pulmonary disease.
Paper No.: 2286
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
A NOVEL ANTITUSSIVE BOTANICAL DRUG PDC-1427
Jui-Ching Chen, C-Y Lin, Y-L Ku
Medical and Pharmaceutical Industry Technology and Development Center, Natural Product Research Department, Taipei, Taiwan
Chronic cough is a common symptom with significant morbidity. This
study was aimed at evaluating the antitussive activity of PDC-1427, an
extract generated from natural plant, in citric acid-induced cough reflex
assay in guinea pigs. PDC-1427 at doses of 2000, 1000, 500, 250 and
125 mg/kg were given orally twice a day for one day. It is observed that
PDC-1427 (500~2000 mg/kg) demonstrated significant inhibitory effect
in citric acid-induced cough reflex. PDC-1427 at 2000 mg/kg possessed
cough suppression effect comparable to opioid agonist codeine (100 mg/
kg). In addition, PDC-1427 also caused significant reduction in cough
induced by citric acid challenge at 1, 2, 4 and 6 hours after final dosing.
A broad spectrum activities study on central nervous system, autonomic
system, cardiovascular function, respiratory function, gastrointestinal system, renal function, allergy, inflammation and metabolism as well as in
vitro tissue functions was also conducted for PDC-1427. In toxicological
studies, we found no obvious adverse effects in genotoxicity tests, an
acute oral toxicity study, and a repeated dose 28-day subacute oral toxicity study in rats and dogs. The NOAEL of each study are estimated to be
5000, 1500~4500 and 1500 mg/kg/day, respectively. This study provides
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
229
novel insights in the therapeutic effect of PDC-1427 against citric acidand capsaicin-induced cough, and then opens scope for further studies on
identification of chemical constituents and pharmacological mechanisms
of PDC-1427.
Paper No.: 1740
FOCUSED CONFERENCE GROUP: P19 - GENERAL
SESSION - ONCOLOGY
CRYPTOTANSHINONE HAS OPPOSITE CELL
CYCLE-ARRESTING EFFECTS ON MELANOMA CELLS OF
DIFFERENT METASTATIC CAPACITY
L Chen, S-Z Zheng, A-Y Wang, Lu Yin
(1) Nanjing University of Chinese Medicine, College of Pharmacy, Nanjing, PR China
Introduction: Cryptotanshinone (CTs) is a main active component of
Radix Salviae Miltiorrhizae, which is often used to therapy cancer as traditional Chinese herb.In this study, the effect of CTs on two melanoma
cells(B16BL6/B16) of high/low metastatic capacity has been systematically compared.Methods: We employed the effect of CTs on cell apoptosis or proliferation by Annexin V, TUNEL assay or BrdU assay etc.
Assayed the cell distributions of cell cycle by flow cytometry. The checkpoint integrity was determined by mutational analyses of B-RAF and
N-RAS, and the effect of CTs on the expression of cell cycle associated
proteins was performed by Western blotting. Results:In B16BL6 we
could found that CTs could not detect any obvious apoptosis but inhibited cell proliferation,also CTs induced cell cycle arrest via induction the
expression of checkpoint enforced by p53, Chk1 and Chk2. CTs-induced
G1 arrest of B16BL6 cells was correlated to an increase in p21. By contrast, in B16 cells, the G2/M arrest is the consequence of the induction
of Cdc25c, and the regulation of Cyclin A1, Cyclin B1 and Cdk1/cdc2
expression could be a major contributor .Conclusions: CTs, which has
opposite cell cycle-arresting effect on melanoma cells of different metastatic capacity, may offer opportunities to investigate the reasons for this
different pattern of malignancy.It will be used as a leading compound for
further studies.
Paper No.: 993
FOCUSED CONFERENCE GROUP: P19 - GENERAL
SESSION - ONCOLOGY
PULMONARY TOXICITY OF BLEOMYCIN INVOLVED IN
UPREGULATION OF LYSYL OXIDASE ACTIVITY IN
CULTURED HUMAN FETAL LUNG FIBROBLASTS
Lijun Chen, S Li, X Su, G Lin
Sun Yat-sen University Zhongshan Medical School, Department of Pharmaclogy, Guangzhou, PR China
Bleomycin (BLM) has been used as a cancer chemotherapy for many
years. Although pulmonary toxicity and pulmonary fibrosis of BLM
often occur in clinic, its mechanism is poorly understood. Lysyl oxidase
(LO), a copper (Cu)-dependent amino oxidase, plays a critical role in
morphogenesis and tissue repair of lung and other organs by catalyzing
the crosslinking of elastin and collagen and stabilizing extracellular
matrix (ECM). To better understand mechanisms of BLM-induced pulmonary fibrosis, we studied changes in LO mRNA, protein, and catalytic
activity levels in cultured human fetal lung fibroblasts (HLF) exposed to
BLM. Exposure of HLF cells to BLM at 10 lg/ml and 30 lg/ml
increased obviously LO catalytic activity in the conditioned media. The
expression of LO mRNA was enhanced significantly in HLF cells exposure to BLM at 3 lg/ml. BLM at 3 lg/ml also increased the expression
of 46 kDa preproLO, 50 kDa proLO and 32 kDa mature LO. To probe
mechanisms of upregulation of LO activity, we tested the Cu concentration in the medium of HLF cells exposed to BLM. Consistently, the Cu
concentrations in conditioned media of cultured HLF cells exposed to
BLM were increased significantly with increases of BLM ranging from
10 to 30 lg/ml. These results suggest that BLM enhanced LO activity in
association of increasing copper concentration in conditioned media of
cultured HLF cells, which may be involved in pulmonary toxicity and
pulmonary fibrosis of BLM.
Key words: bleomycin (BLM); pulmonary fibrosis; lysyl oxidase (LO);
copper (Cu).
Paper No.: 2466
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
OXYTOCIN INJECTED ON OVARIECTOMIZED FEMALE
RATS CAN AMELIORATE HEATSTROKE VIA INCREASING
CIRCULATING ENDOTHELIAL PROGENITOR CELLS
Sheng-Hsien Chen(1), W-T Huang(2), CY Kang(1)
(1) Chi Mei Medical Center, Southern Taiwain University, Department
of Gynecology & obstetrics, Department of Biotechnology, Tainan,
Taiwan
(2) Chia Nan University of Pharmacy Science, Taiwan
Background: The number of circulating endothelial progenitor cells
(EPCs) is decreased in postmenopausal women, and increased in those
on hormone replacement therapy (HRT). Some researchers have indicated that oxytocin (OT) controls differentiation of human mesenchymal
stem cell to osteoblasts (OBs) and reverse osteoporosis. Whether OT can
resuscitate heatstroke rats via increase circulating EPCs is unclear. Aim:
Our current study was designed to investigate the therapeutic effects of
OT on 2 weeks after ovariectomized (OVX) female SD rats subjected to
heatstroke insults. Measurements and Results: Female rats post OVX
2 weeks later were randomly assigned into three groups: a) vehicle 1 cc
subcutaneous injection (sc) only for 8 weeks. b) OT 1mg/kg/day sc for
8 weeks. c) then following normothermia (Urethane-anesthetized rats
exposed to 26 C or 480 min) or heatstroke (Urethane-anesthetized rats
were exposed to an ambient temperature of 43 C to induce heatstrokemean arterial pressure (MAP) decreasing more than 25 mmHg). OT pretreatment on OVX female rats significantly increased circulating EPCs
before heat stress compared with saline controls via flow cytometry
assay. After the onset of heatstroke, animals treated with saline displayed
hyperthermia, hypotension, bradycardia, hypothalamic neuronal apoptosis and degeneration, and up-regulation of systemic inflammatory
response molecules including serum tumor necrosis factor-a, soluble
intercellular adhesion molecule-1 and E-selectin. Heatstroke-induced
hypotension, bradycardia, hypothalamic neuronal apoptosis and degeneration, and increased systemic inflammatory response molecules were significantly attenuated by OT pretreatment. Conclusion: Increased
circulating EPCs produced by OT administered to OVX rats can protect
against heatstroke via attenuation of systemic inflammation, neuronal
damage and vascular endothelial damage.
Paper No.: 1459
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
ROLE OF RHOA/ROK IN VASCULAR HYPOREACTIVITY
INDUCED BY ENDOTOXIN IN EARLY AND LATE SEPSIS
Shiu-Jen Chen(1), M-H Liao(2), C-C Wu(2)
(1) Kang-Ning Junior College of Medical Care and Management,
Department of Nursing, Taipei, Taiwan
(2) National Defence Medical Centre, Taipei, Taiwan
Blood vessel tone plays an important role in the determination of tissue
perfusion and blood pressure. In severe sepsis, the vascular reactivities to
vasconstrictor and vasodilator are greatly impaired. However, the mechanism of development of vascular hyporeactivity remains unclear. Calcium
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
230
sensitization and desensitization are involved in mechanisms regulating
vascular tone. Myosin phosphatase (MYPT) and the RhoA-associated
Rho-kinase (RhoA/ROK) pathway are two major cellular targets for regulating Ca2 + sensitivity of agonist-induced contraction. In this study, we
investigated the role of RhoA/ROK in endotoxin (lipopolysaccharide,
LPS)-induced vascular hyporeactivity in early (at 1 h and 2 h after LPS)
and late (at 4 h and 6 h after LPS) sepsis. The sepsis-induced vascular hyporeactivity was performed by intravenous infusion of LPS (10 mg/kg,
10 min) to male Wistar rats. In this study, the changes of hemodynamics,
blood glucose, hepatic (GPT, GOT) and renal (CRE, BUN) function,
plasma nitrate (an indicator of NO), and the vascular reactivities to norepinephrine (NE) and acetylcholine (Ach) of thoracic aorta in vivo and ex
vivo were measured. In addition, the expression of RhoA in the thoracic
aorta was analyzed by immunohistological chemistry and.Western blot,
whereas pathological studies of lungs, livers, kidneys and thoracic aorta
were also examined. Our preliminary results showed that LPS induced
not only dysfunction of liver and kidney but also vascular hyporeactivity
at 1 h, 2 h, 4h, and 6 h in vivo and this hyporeactivity was varied ex vivo
and associated with RhoA expression in early and late sepsis.
Paper No.: 1599
FOCUSED CONFERENCE GROUP: P18 - NUCLEAR
RECEPTOR TARGETS FOR TREATMENT OF DISEASES
INVOLVEMENT OF AUTOPHAGY IN THE SEX-DIFFERENT
BRAIN INJURY CAUSED BY IRON OVERLOAD
Tzu-Yin Chen, S-I Lue, C Hsu
Kaohsiung Medical University, Department of Physiology, Kaohsiung,
Taiwan
Overproduction of iron, a degradation product of hemoglobin after hemorrhage, leads to free radical formation and oxidative stress in brain tissue. Oxidative stress and subsequent organelle damage simultaneously
induces autophagy that is an intracellular recycling protein degradation
system, while over activation of autophagy may be harmful for the acute
brain injury such as iron overload caused by hemorrhage. Our previous
study showed that infusion of ferrous citrate (FC) in caudate nucleus
(CN) caused a severer brain injury in male rats than that in females and
the estradiol pretreatment diminished the FC-induced brain injury. We
propose autophagy participates in the sex-different FC-induced brain
injury. Microtubule-associated protein 1 light chain 3 was detected to
estimate autophagy. The cleavage of a II-spectrin was quantified to estimate the severity of brain injury. The results showed that iron-induced
autophagy and DNA damage with intact nuclei implying autophagic cell
death were observed. In addition, mice of Atg5(+/)), an autophagyrelated gene, showed a less FC-induced brain deficit and injury compare
to Atg5(+/+) mice. The levels of constitutive and iron-induced autophagy
in CN were lower in female rats than in males. Levels of iron-induced
autophagy correlate with the severity of brain injury in both male and
female rats. Moreover, pretreatment of estradiol decreased both the level
of iron-induced autophagy and cleavage of a II-spectrin in females. Since
ER-a expression is higher in females than in males, the inhibition of
iron-induced autophagy by estradiol may mediate the ER-dependent neuroprotection in females from brain injury caused by iron overload.
Paper No.: 1059
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
MODULATORY EFFECTS OF THE EXTRACT OF BURDOCK
TEA ON HYPERLIPIDEMIA RAT AND RICE MODELS
Xiangguo Chen(1), Y Xu(2), Y Cao(2), Z Han(2), P Li(2), C Wang(3)
(1) Qingdao Food and Drug Administration, Department of Pharmacy,
Qingdao, PR China
(2) The Affiliated Hospital of Medical College Qingdao University,
Qingdao, PR China
(3) Qingdao University, Medical College, Qingdao, PR China
Arctium lappa Linne (AL; burdock) is a biennial member of the Asteraceae (Compositae) family, and its carrot-like root is commonly cooked
and eaten as a vegetable in parts of Asia. In traditional Brazilian and
Asian folk medicine, AL is used for the treatment of different diseases.
In the present study the effects of the extract of burdock tea on lipid profile were observed in hyperlipemic rats and mice. Seven-week-old male
Wistar rats were fed a high-fat diet to induce hyperlipemia with doses of
extract of burdock tea at 0.83 (low), 1.67 (medium), and 5.0 (high) g/kg
of body weight. The hyperlipidemia mice models were induced by intraperitoneal injection of 75% solution of egg yolk, and the Kunming mice
were intragastric administered with extract of burdock tea (25g/kg, 50g/
kg of body weight). The body weight and the concentrations of total cholesterin (TC), triglyceride (TG) and high density lipoprotein cholesterol
(HDL-C) were detected. The results showed that TG and HDL-C in rats
were decreased significantly in the two burdock tea groups (1.67g/kg.bw,
5.00g/kg.bw) and the body weight of rats was reduce remarkably. The
TG and HDL-C in mice were remarkably decreased in the burdock tea
groups (50g/kg of body weight). Taken together, these results indicated
that the extract of burdock tea has modulatory effect on blood lipid in
hyperlipidemia model rats and mice.
(Supported by the Shandong Natural Science Foundation of China, No.
Q2008C04 and Z2007c09).
Paper No.: 761
FOCUSED CONFERENCE GROUP: P19 - GENERAL SESSION ONCOLOGY
THE MECHANISM OF MNNG-INDUCED-APOPTOSIS: AMPK
COULD BE A TRANSMITTER FROM PARP ACTIVATION TO
MITOCHONDRIAL DYSFUNCTION
Yi-Cheng Chen, J-H Guh
National Taiwan University, Pharmacy Department, Taipei, Taiwan
N-Methyl-N-Nitro-N-Nitrosoguanidine (MNNG), a DNA-alkylating
agent, was well known to exert its cytotoxicity by activating poly (ADPribose) polymerase (PARP) and mitochondrial dysfunction is one of the
mechanisms to induce apoptotic cell death. However, how PARP, a
nuclear protein, conveys the death signal to mitochondria was still
unclear. Here we try to figure out what signal is activated and could be
regarded as an inducer of mitochondrial dysfunction after MNNG treatment. We found that AMP-activated protein kinase (AMPK) could be a
candidate. In PC-3 prostate cancer cells, MNNG can phosphorylate
AMPK on Thr172 and Ser385, indicating the activation. Furthermore,
among the MNNG-treated cells, we separately collected the detached
and attached parts, and observed that AMPK phosphorylation is more
obvious in detached fraction, which are judged dying or to die by flow
cytometry. Besides, MNNG can arrest cells on s/G2 phase at 6 hr, G2/M
phase at 12 hr and finally result in subG1 increase. The combination with
compound C (a specific AMPK inhibitor) can reverse cell cycle arrest
and subG1 increase induced by MNNG. These data suggest the involvement of AMPK in MNNG-induced apoptosis. So we propose that
AMPK could be a transmitter role which accepts the death message from
nucleus to mitochondria in MNNG-induced apoptosis.
Paper No.: 2298
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
APPROACH OF ACTION MECHANISM OF TCM-98 AND ITS
ACTIVE COMPONENT ON NEURO- INFLAMMATION IN IN
VIVO AND IN VITRO MODELS
Yuh-Fung Chen(1,2), Y-W Wang(2), H-Y Tsai(3)
(1) China Medical University, Department of Pharmacolgy, Taichung,
PR China
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
231
(2) China Medical University, Graduate Institute of Chinese Pharmaceutical Sciences, Taichung, PR China
(3) China Medical University Hospital, Department of Pharmay,
Taichung, PR China
Cerebral vascular diseases are the third leading cause of death in Taiwan in recent years. Neuro-inflammation and neuronal cell death occur
in the early stage of stroke. Our preliminary data showed that TCM-98
and TCM-9811 significantly reduced the infract volume of the stroke
rat brain. Furthermore, TCM-98 crude and TCM-9811 obviously
decreased both productions of NO and protein expression of iNOS and
COX2 induced by LPS, while increasing p53 protein expression. Specially, the results of TCM-98 crude and TCM-9811 reversing p53 protein attenuated by LPS is interesting for us and remain for further
investigation because p53 traditionally is thought to be involved in
apoptosis, cell cycle arrest, and DNA repair. Although previous studies
have demonstrated p53 is a suppressor of inflammatory response in different experimental types, the underlying mechanisms of anti-inflammation are poorly understood in microglia cells. In the other hand, TCM9811 was able to inhibit the effects that LPS enhanced the nuclear
translocation of p65 and p50 subunits, an effect preceded by the cytosolic decrease in IjBa and promote in phosphorylation of IjBa. Meanwhile, TCM-9811 also reversed the cytosolic p65 and p50 subunits
expression repressed by LPS. Results from our data showed that TCM98 and TCM-9811 have anti-inflammatory actions against inflammation
caused by LPS. The possible action mechanism of TCM-98 and TCM9811 involved in BV-2 cell stimulated by LPS model is investigated in
this study.
Paper No.: 3236
FOCUSED CONFERENCE GROUP: P19 - GENERAL
SESSION - ONCOLOGY
THE RELATIONSHIP BETWEEN MEKK1 AND
CARCINOGENESIS & DEVELOPMENT OF PANCREATIC
CANCER
Xiaoguang Cheng(1), F Su(1), K Li(1), Y Zhang(1), C Yan(2)
(1) Chinese Academy of Medical Sciences & Peking Union Medical
College, Institute of Materia Medica, Beijing, PR China
(2) Harbin Medical University, The Second Affiliated Hospital, Beijing,
PR China
To observe whether the MEKK1 expression is related to pancreatic cancer in vitro, we examined the MEKK1 mRNA and protein levels in different pancreatic cancer cell lines (BxPC3, Capan2, SW1990 and
PANC1). The data showed that MEKK1 mRNA level remained highly
abundant in BxPC3 and Capan2 cell lines while it had weak expression
in the SW1990 and PANC1 cell lines. A Western blot analysis and immunofluence assay further confirmed the MEKK1 protein expression in
a similar pattern in the four observed pancreatic cell lines. To identify the
detailed roles of MEKK1 in the regulation of malignant characteristics of
human pancreatic adenocarcinoma in vivo. We analyzed MEKK1 expression in patient’s samples by immunohistochemistry assay. The results
demonstrated that MEKK1 positive staining was noted in 32 samples
among the 41 pancreatic adenocarcinomas (78.1%). All results suggested
that MEKK1 expression was significantly associated with differentiation,
TNM staining and lymph node metastasis in the observed pancreatic cancer patients (P < 0.05). The MEKK1 staining did not show significantly
different among the patients of different gender, age, tumor location and
tumor size (P > 0.05). In addition, there is a mainly founding that
MEKK1 cell location verified with cancer differentiation grade. The
amounts of nuclear staining is 100% in poorly differentiated cancers,
while the amount of nuclear staining in moderate and well differentiated
cancers were 72.2% and 22.2%.In conclusion, this study demonstrates
that MEKK1 plays a major role in the metastasis of pancreatic cancer.
Our findings suggest that MEKK1 may be a potential molecule target for
pancreatic cancer therapy.
Paper No.: 562
FOCUSED CONFERENCE GROUP: P17 - NEW APPROACHES
AND TARGETS IN PSYCHIATRY
EXPRESSION OF VGLUTS IN THE PROCESS OF
DEGENERATION AND ACQUISITION OF LEARNING AND
MEMORY
Xiaorui Cheng, Y Zhao, W Zhou, Y Zhang
Beijing Institute of Pharmacology and Toxicology, Department of Neuroimmunopharmacology, Beijing, PR China
Vesicular glutamate transporters (VGLUTs) include VGLUT1, VGLUT2
and VGLUT3, and are responsible for the uploading of L-glutamate into
synaptic vesicles. The expression of VGLUTs determines the level of synaptic vesicle filling and glutamate quantal size, indicates the glutamatergic
neurons and synapse number, thus directly influences glutamate receptors
and glutamatergic synaptic transmission. In order to investigate whether
VGLUTs has correlation with the process of degeneration and acquisition
of learning and memory, we choose senescence-accelerated mouse/prone
8 (SAMP8) as the animal model of degeneration of learning and memory,
mouse through the shuttle-box test as animal model of acquisition of
learning and memory, the expression of VGLUTs and synaptophysin
(Syp) mRNA and protein in the cerebral cortex and hippocampus of the
tested subjects was investigated using QPCR and western-blot technique
respectively. Results showed that VGLUT1, VGLUT2 and Syp in the
cerebral cortex, VGLUT1 and Syp in the hippocampus of SAMP8
decreased with aging and aged senescence-accelerated mouse resistant
strains 1 (SAMR1), while VGLUT2 in the hippocampus decreased in
SAMR1 and increased in SAMP8 with aging. VGLUT3 appeared nonregular pattern. The expression of VGLUT1 mRNA and protein increased
in the process of acquisition of learning and memory in the shuttle box
test. This indicates the complementary expression of VGLUT1 and
VGLUT2 in the hippocampus of SAMP8 and VGLUTs related with learning and memory tightly. These results suggested the involvement of the
brain glutamatergic system in the degeneration of learning and memory in
SAMP8 and glutamateric neuron loss in the brain of the SAMP8.
Paper No.: 2220
FOCUSED CONFERENCE GROUP: PW12 - EXPERIMENTAL
PAIN METHODS IN DEVELOPMENT AND VALIDATION OF
ANALGESICS
SINGLE DOSE OF TRAMADOL ON NEUROTRANSMITTER
SYSTEMS IN TARGET AND NON-TARGET AREAS OF RAT
BRAIN WITHOUT NOCICEPTION
Pandanaboina Chetan(1), S Rajbanshi(2), R Wudayagiri(3)
(1) Arkansas State University, Department of Biological Sciences, Jonesboro, AR, USA
(2) Sri. Venkateswara University, Department of Zoology, Tirupati, India
(3) Mahila University, Department of Pharmacy, Tirupati, India
In the present investigation the changes in the levels of Monoamines
such as epinephrine (E), norepinephrine (NE), dopamine (DA), and serotonin (5-HT) and also levels of the major metabolites Homovanillic acid
(HVA) and Hydroxyindole acetic acid (HYAA) were estimated in different areas of the rat brain during the administration of the synthetic opioid
analgesic drug Tramadol (Ultram) without induction of pain. Following
administration of single dose of tramadol (31 mg/kg body weight), the
levels of DA, 5-HT were elevated in all the regions of brain, while NE,
EP, HIAA levels and MAO activity recorded increase in some areas and
decrease in other areas by 3 or 6 hrs. The levels of biogenic amines
returned to the respective control levels by 24 hrs after tramadol administration. The monoamine oxidase activity levels were elevated in all the
brain regions except in hypothalamus after treatment with a single dose
of tramadol. The region specific alterations in biogenic amines and their
metabolites in different areas of rat brain following administration of single dose of tramadol suggest that the analgesic actions of tramadol might
be implicated to the enhanced monoaminergic neurotransmission similar
to other opiods as well as other classes of analgesics.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
232
Paper No.: 1707
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
A GENETIC VARIANT IN THE GENE ENCODING
ADRENOMEDULLIN PREDICTS THE DEVELOPMENT OF
DYSGLYCAEMIA OVER 6.4 YEARS IN CHINESE
Bernard MY Cheung(1), AWK Tso(1,2), KL Ong(1), RYH Leung(1),
SS Cherny(3), PC Sham(3), TH Lam(4), KSL Lam(1,2)
(1) University of Hong Kong, Department of Medicine, Hong Kong, PR
China
(2) University of Hong Kong, Research Centre of Heart, Brain, Hormone
and Healthy Aging, Hong Kong, PR China
(3) University of Hong Kong, Department of Psychiatry and Genome
Research Centre, Hong Kong, PR China
(4) University of Hong Kong, Department of Community Medicine,
Hong Kong, PR China
Adrenomedullin (AM), a vasodilatory peptide, facilitates the differentiation of pre-adipocytes and affects lipolysis and glucose uptake. We investigated the association of common single nucleotide polymorphisms
(SNPs) in the gene encoding adrenomedullin (ADM) with dysglycaemia
in the Hong Kong Chinese population. Four SNPs (rs3814700,
rs11042725, rs34354539 and rs4910118) were genotyped in 1391 subjects without dysglycaemia at baseline from the Hong Kong Cardiovascular Risk Factor Prevalence Study-2 (CRISPS-2), which had a median
follow-up time of 6.4 years. Dysglycaemia included impaired fasting
glucose, impaired glucose tolerance and diabetes according to the WHO
1998 criteria. At follow-up, 382 subjects had developed dysglycaemia.
In stepwise logistic regression, the SNP rs11042725 (minor allele frequency = 28.7%) was a significant independent predictor of the development of dysglycaemia at 6.4 years (OR=1.31, P = 0.014), together with
age (P < 0.001), plasma triglycerides (P < 0.001), body mass index
(P < 0.001), 2-h glucose after oral glucose tolerance test (P < 0.001),
change in body mass index (P = 0.001) and follow-up duration
(P = 0.006). The association was more significant in women (OR=1.65,
P = 0.002) and in subjects without regular exercise (OR=1.56,
P = 0.001). In women without regular exercise, the odds ratio (95% CI)
for developing dysglycaemia was 1.93 (1.30-2.85) (P = 0.001). The other
two SNPs, rs3814700 and rs34354539 also showed nominal associations
with the development of dysglycaemia (P = 0.034 and 0.021 respectively). Our study suggests a potential role of genetic variants in the
ADM gene in the development of dysglycaemia. Further studies on the
role of adrenomedullin in glucose metabolism are warranted.
Paper No.: 1708
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD DISEASES
RELATIONSHIP OF PLASMA INTERLEUKIN-6 AND ITS
GENETIC VARIANTS WITH HYPERTENSION IN HONG KONG
CHINESE
Bernard MY Cheung(1), AWK Tso(1,2), KL Ong(1), RYH Leung(1),
SS Cherny(3), PC Sham(3), GN Thomas(4), TH Lam(5), KSL Lam(1,2)
(1) University of Hong Kong, Department of Medicine, Hong Kong, PR
China
(2) University of Hong Kong, Research Centre of Heart, Brain, Hormone
and Healthy Aging, Hong Kong, PR China
(3) University of Hong Kong,Department of Psychiatry and Genome
Research Centre, Hong Kong, PR China
(4) University of Birmingham, Department of Public Health and Epidemiology, Birmingham, UK
(5) University of Hong Kong, Department of Community Medicine,
Hong Kong, PR China
Interleukin-6 (IL-6) plays a central role in inflammation and insulin resistance as well as atherogenesis. We investigated if IL-6 is associated with
hypertension in a sample of the general population, and whether genetic
variants that might influence plasma IL-6 level could be associated with
hypertension. Plasma IL-6 was measured using a high-sensitivity
enzyme-linked immunosorbent assay in 648 normotensive and 294
hypertensive subjects from the Hong Kong Cardiovascular Risk Factor
Prevalence Study-2 and three tagging single nucleotide polymorphisms
(SNPs) (rs17147230, rs1800796 and rs2069837) in the IL-6 gene were
genotyped. Plasma IL-6 level correlated with age (r = 0.171, P < 0.001),
body mass index (r = 0.109, P < 0.001), systolic blood pressure
(r = 0.083, P < 0.001), pulse pressure (r = 0.092, P = 0.010), C-reactive
protein (r = 0.225, P < 0.001) and high-density lipoprotein cholesterol
(r=-0.100, P = 0.002). Hypertensive subjects have significantly higher
plasma IL-6 level after adjusting for age and sex (geometric mean (95%
CI) = 0.60 [0.54-0.65] vs 0.47 [0.44-0.50] pg/ml, P = 0.024).In multivariate logistic regression analysis, plasma IL-6 was independently associated with hypertension in women (P = 0.004), but not in men. The SNP
rs1800796 was independently associated with plasma IL-6 level (b=0.098, P = 0.002) in stepwise multiple linear regression. The association
was significant in both men (P = 0.042) and women (P = 0.016).However, this SNP was not associated with hypertension. Elevated plasma
IL-6 level is associated with hypertension, especially in women. Plasma
IL-6 level is influenced by the SNP rs1800796. However, this SNP is
not associated with hypertension, suggesting that hypertension is caused
by other factors that elevate plasma IL-6 level.
Paper No.: 1895
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
RESVERATROL INHIBITS 8-ISO-PROSTAGLANDIN F2ALPHA
PRODUCTION IN YOUNG AND AGED RAT BRAIN
Annalisa Chiavaroli, L Brunetti, G Orlando, L Menghini, L Recinella,
S Leone, C Ferrante, C Di Nisio, R Shohreh, P Di Michele, M Vacca
G. d’Annunzio University, Department of Drug Sciences, Chieti, Italy
Oxidative stress plays a role in the pathogenesis of neurodegeneration
typical of aging and 8-iso-prostaglandin F2a (8-iso-PGF2a), an isoprostane generated from oxygen radical peroxidation of arachidonic acid,
may represent a reliable marker of cellular oxidative damage. Resveratrol
is a naturally occurring phytoalexin that is produced in response to
injury, such as mechanical trauma, ultraviolet light and infection by pathogenic microorganisms, especially fungi, providing means for defense.
Resveratrol has been associated with numerous health benefits from
chemoprevention to cardioprotection and might prevent or reverse 8-isoPGF2a production. We have tested the possible antioxidant effects of resveratrol in rat brain synaptosomes obtained from young (3 months old)
and aged (15 months old) male Wistar rats that were perfused, in vitro,
with graded concentration of resveratrol (0.01-50 microM), both in the
basal state and after hydrogen peroxide-induced oxidative stress, evaluating 8-iso-PGF2a levels in the perfusate by radioimmunoassay. In young
rats, we observed a concentration-dependent inhibitory effect of resveratrol on brain production only after hydrogen peroxide-induced oxidative
stimulus. In aged rats, a concentration dependent antioxidant effect of
resveratrol was evidenced both in the basal state and after hydrogen peroxide-induced oxidative stimulus. In conclusion, resveratrol could be
effective in preventing brain oxidative damage both in young and aged
rats, showing a greater inhibition of isoprostane production in the latter.
Paper No.: 1321
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
INHIBITORY EFFECT OF AS1517499, A NOVEL STAT6
INHIBITOR, ON ANTIGEN-INDUCED AIRWAY
HYPERRESPONSIVENESS IN MICE
Yoshihiko Chiba, M Todoroki, Y Nishida, M Tanabe, M Misawa
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
233
Hoshi University School of Pharmacy, Department of Pharmacology,
Tokyo, Japan
Interleukin-13 (IL-13) is one of the central mediators for development of
airway hyperresponsiveness in asthma. The signal transducer and activation of transcription 6 (STAT6) is one of the major signal transducers
activated by IL-13, and a possible involvement of IL-13/STAT6 pathway
in the augmented bronchial smooth muscle (BSM) contraction has been
suggested. In the present study, the effect of a novel STAT6 inhibitor,
AS1517499, on the development of antigen-induced BSM hyperresponsiveness was investigated. In cultured human BSM cells, IL-13 (100 ng/
mL) caused a phosphorylation of STAT6 and an upregulation of RhoA, a
monomeric GTPase responsible for calcium sensitization of smooth muscle contraction: both events were inhibited by co-incubation with
AS1517499 (100 nM). In BALB/c mice that were actively sensitized and
repeatedly challenged with ovalbumin antigen, an increased IL-13 level
in bronchoalveolar lavage fluids and a phosphorylation of STAT6 in
bronchial tissues were observed after the last antigen challenge. These
mice had an augmented BSM contractility to acetylcholine together with
an upregulation of RhoA in bronchial tissues. Intraperitoneal injections
of AS1517499 (10 mg/kg) 1 h before each ovalbumin exposure inhibited
both the antigen-induced upregulation of RhoA and BSM hyperresponsiveness, almost completely. A partial but significant inhibition of antigen-induced production of IL-13 was also found. These findings suggest
that the inhibitory effects of STAT6 inhibitory agents, such as
AS1517499, both on RhoA and IL-13 upregulations might be useful for
asthma treatment.
Paper No.: 1299
FOCUSED CONFERENCE GROUP: P17 - NEW APPROACHES
AND TARGETS IN PSYCHIATRY
IMPAIRMENT OF HEME OXYGENASE-1 EXPRESSION BY
THE DEFECT OF PARKINSON-RELATED PINK1 PROTEIN
Paper No.: 3298
FOCUSED CONFERENCE GROUP: P19 - GENERAL
SESSION - TOXICOLOGY
MELAMINE MIGRATION FROM MELAMINE FOOD
CONTACT ARTICLES AVAILABLE ON THE MALAYSIAN
MARKET
Zamri Chik, DEM Haron, MA Mohamad, ED Ahmad, H Taha
University of Malaya, Faculty of Medicine, Department of Pharmacology, Kuala Lumpur, Malaysia
The migration of melamine from melamine-ware products retail in
Malaysia has been determined from 41 products. This study has been
conducted in the midst of public anxiety on the possibility of melamine
leaching into foods that come into contact with the melamine-wares.
Samples of melamine table-wares included cups and plates, forks and
spoons, tumblers, bowls, etc. were collected from various retail outlets.
The samples were exposed to the food stimulant, 3% acetic acid and also
distilled water at room temperature, 70C and 100C for 30 minutes.
The melamine detection was done using Liquid Chromatography-Tandem Mass Spectrometry (LC/MS/MS) method. The column used was a
HILIC C18 with a mobile phase consisting of a mixture of ammonium
acetate / formic acid (0.05%) in water and a mixture of ammonium acetate/ formic acid (0.05%) in acetonitrile with 95/5 (v/v). The limit of
quantitation (LOQ) was 1 ppb. The presence of melamine migration was
detected in all samples. For the articles tested with water, melamine
migration were (median (IQR)) 22.2 (32.6), 49.3 (50.9), 84.9 (89.9) ppb
at room temperature, 70 C, and 100 C, respectively. In acetic acid, melamine migration were 31.5 (35.7), 81.5 (76.2), 122.0 (126.7) ppb at
room temperature, 70 C, and 100 C, respectively. This study suggests
that excessive heat and acidity may directly affect melamine migration
from melamine-ware products. However the results showed that melamine migration in the tested items were well below the specific migration
limit at 30 mg/kg (30,000 ppb) in accordance to The European Commission Directive 2002/72/EC.
Wei-Lin Chien(1), J-R Li(1), M-J Li(3), W-M Fu(0)
(1) National Taiwan University College of Medicine, Pharmacological
Institute,Taipei, Taiwan
(2) National Taiwan University, Neurobiology and Cognition Center,
taipei, Taiwan
(3) National Taiwan University Hospital, Department of Neurology,
Taipei, Taiwan
Parkinson’s disease (PD) is one of the most common neurodegenerative
diseases. PINK1 (PTEN induced putative kinase 1) is a mitochondria
serine/threonine protein kinase. Mutation in the PINK1 gene causes an
autosomal recessive form of PD. Although PINK1 is thought to protect
cells from stress-induced mitochondria dysfunction, the etiology related
to PINK1 mutation is still not clear. We have established stable clones
of PINK1 kinase domain mutant G309D in SH-SY5Y. It was found
that ROS and apoptosis increased in G309D mutant cells following
H2O2 treatment. Heme oxygenase 1 (HO-1) increases and exerts its cell
protection when the cells are under oxidative stress. It was found here
that HO-1 increased in wilt-type cells following H2O2 treatment.
Although PINK1 mutant stable clones had a higher basal level of
HO-1, however, HO-1 induction in response to H2O2 and MPP+ treatment was impaired by PINK1 mutation. Using adenovirus infection to
overexpress HO-1 can antagonize the H2O2-induced apoptosis in
PINK1 mutant stable clones. In addition, small hairpin RNA-mediated
knockdown of TNF receptor-associated protein 1 (TRAP1), which is
the substrate of PINK1 kinase, in SH-SY5Y cells also inhibited the
expression of HO-1 in response to H2O2 treatment. TRAP1 can
increase following H2O2 treatment, whereas, the expression of TRAP1
in response to H2O2 was also impaired by PINK1 G309D mutation.
Our findings suggest a novel pathway by which the defect of PINK1
and its downstream effector TRAP1 inhibits the oxidative stressinduced HO-1 upregulation, the impairment may enhance the dopaminergic neurodegeneration in Parkinson.
Paper No.: 2244
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
ACTIVATION OF OX1, BUT NOT OX2, RECEPTORS IN THE
PERIAQUEDUCTAL GRAY PRODUCES ANTINOCICEPTION
VIA RETROGRADE ENDOCANNABINOID-INDUCED
DISINHIBITION
Lih-Chu Chiou(1,2,3), Y-C Ho(1), H-J Lee(2), Y-Y Liao(1), S-Y Fu(1),
S-F Teng(3), H-T Liao(3)
(1) National Taiwan University, College of Medicine, Graduate Institute
of Pharmacology, Department of Pharmacology, Taipei, Taiwan
(2) National Taiwan University, Graduate Institute of Zoology, Taipei,
Taiwan
(3) National Taiwan University, Neurobiology and Congnitive Science
Center, Taipei,Taiwan
Orexin A and B are endogenous agonists of a family of orphan Gq-protein coupled receptors, OX1 and OX2. The orexin system has been
implicated in arousal, metabolic, rewarding and pain regulations, and
their action mechanisms have been intensively explored except that in
pain regulation. In this study, we found orexin A and [Ala11,
D-Leu15]orexin B (AL-orexin B), an orexin B analogue selective to
OX2 receptors, increased mouse hot-plate latency when microinjected
into the ventrolateral periaqueductal gray (vlPAG), a midbrain region
mediating pain inhibition. The antinociceptive effect of orexin A, but not
AL-orexin B, was primarily mediated through OX1 receptors, opioidindependent, reversed by AM 251 (a CB1 antagonist), and mimicked by
WIN 55,212-2 (a CB1 agonist). The antinociceptive effect of AL-orexin
B was via OX2 receptors and also opioid-independent. In vlPAG slices,
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
234
orexin A, via OX1 receptors, caused membrane depolarization and neuronal firing, and depressed GABA-mediated evoked inhibitory postsynaptic currents (eIPSCs) by a presynaptic mechanism. Orexin A-induced
eIPSC depression was reversed by AM 251, mimicked by WIN 55,2122, prevented by U73122 and tetrahydrolipstatin, phospholipase C (PLC)
and diacylglycerol lipase (DAGL) inhibitors, respectively, and enhanced
by URB 602, which inhibits enzymatic degradation of 2-arachidonoylglycerol (2-AG). AL-orexin B also caused membrane depolarization via
OX2 receptors. These results suggest that activation of OX1, but not
OX2, receptors, in the vlPAG induces antinociception through the Gqprotein-PLC-DAGL cascade, yielding 2-AG, an endocannabinoid which
produces retrograde inhibition of GABA release (disinhibition). OX2
receptor activation in the vlPAG also induces antinociception by increasing neuronal activity via membrane depolarization.
Paper No.: 1116
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
HIGH-SODIUM LOADED IN RATS IMPAIRS CENTRAL
BAROREFLEX AND ACH-INDUCED VASODILATION
Akiko Chisyaki(1), M Tsuruhisa(1), M Miyakawa(1), T Masuda(1),
Y Hirabara(2), K Honda(1), R Saito(1), Y Takano(1)
(1) Fukuoka University, Department of Pharmacology, Fukuoka City,
Japan
(2) University Miyazaki Hospital, Department of Pharmacy, Japan
Recently, the enzyme 11 b-hydroxysteroid dehydrogenase type 2
(HSD2) and the mineralocorticoid receptors (MR) were identified in the
nucleus tractus solitarius (NTS), in addition to common neurotransmitters; such neurons are termed healdosterone-sensitive HSD2 neuronshf
(J. Comp. Neurol. 2006, 497: 223). Furthermore, a number of studies
have shown that the aldosterone-sensitive HSD2 neurons in the NTS are
involved in sodium appetite or in regulation of body fluid circulation as
they are selectively activated by sodium deficiency. In the present study,
we tested the hypothesis that the aldosterone-sensitive neurons in the
NTS regulate arterial baroreceptor reflex (baroreflex) function. To
address this hypothesis, we examined the sensitivity of baroreflex
induced by phenylephrine in high-sodium loaded rats. Experiments were
performed by our previous method (J. Pharmacol. Sci. 2007, 104, 402,).
(1) The baroreflex sensitivity was significantly reduced in the high
sodium-loaded rats. (2) The baroreflex sensitivity was reversed by microinjection of MR antagonist eplerenone into the nucleus tractus solitarius
(NTS). (3) HSD2 neurons and MRs were identified in the NTS. The
findings suggest that the aldosterone-sensitive neurons in the NTS may
play important roles of baroreflex functions. In addition, we examined
EDHF-mediated relaxation on mesenteric arteries in high sodium loaded
rats by using the type 2 diabetic model, Goto-Kakizaki rats (GK rat).
The endothelium-dependent relaxation to acetylcholine (ACh) ACh in
the high sodium-loaded GK rats significantly attenuated compared with
that in non-sodium loaded GK rats.
Paper No.: 2483
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
A NOVEL PROTOCOL FOR THE DISCOVERY OF
ANGIOGENESIS-MODULATORS
Chin-wen C Cho(1), J-C Yeh(1), T-P Fan(1)
University of Cambridge, Department of Pharmacology, Cambridge, UK
Current drug discovery regime employs cell models for screening synthetic compounds or natural products. However, the selection of experimental culture condition (e. g. foetal bovine serum and growth factors)
may mask potential therapeutic properties of the compounds tested. To
overcome this problem, we have developed a novel protocol for identify-
ing angiogenesis modulators, using two endothelial cell (EC) culture
media (EGM-2 and a modified medium, MM) in two in-vitro angiogenesis models (proliferation and tube formation assays). Our data show that
vascular-targetting agents (VTAs) taxol and combretastatin at nM range
produce similar anti-angiogenic effects under both conditions. Conversely,
phytosteriod ginsenosides Rh2 and PPD at lM range exhibit distinct
effects under these two conditions. At 20-40 lM, PPD causes cytotoxicity
in EGM-2, while promoting cellular survival in MM. Western blot analysis demonstrates differential expression of proteins involved in pro-apoptotic or pro-survival pathways in ECs treated with ginsenosides. However,
VTAs result in similar expression of proteins in both conditions. Taxol
and combretastatin are known to target microtubules whereas ginsenoside
Rh2 has been linked to inhibition of matrix metalloproteinases and
expression of junction adhesion molecules in tumour vasculature. Thus,
this novel protocol cannot only detect angiogenesis modulators but also
allow preliminary identification of their underlying mechanisms.
Acknowledgement: supported by Cambridge Overseas Trust.
Paper No.: 2149
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
ASSOCIATION OF THE ABCB1 3435C>T POLYMORPHISM
AND ADVERSE EVENTS OF CILOSTAZOLE IN HEALTHY
SUBJECTS
Sung Kweon Cho(1,2), LA Lim(1,2), SB Jang(1,2), YJ Lee(1,2),
DH Lee(1,2), HK Son(1,2), JY Chung(1,2), K Park(1,2)
(1) Yonsei University College of Medicine, Department of Pharmacology, Seoul, South Korea
(2) Yonsei University, Brain Korea 21 Project for Medical Science,
Seoul, South Korea
Cilostazole, a type III phosphodiesterase inhibitor, has been widely used
to control symptoms of lower extremity peripheral arterial disease and
intermittent claudication. Adverse events of Central Nervous System
(CNS) such as headache, nausea and dizziness are frequently reported
during the treament. This study was aimed to propose the role of ABCB1
polymorphism in the case of CNS adverse events of cilostazol. We
examined the the relationship between the genotypes and the adverse
events of cilostazol after single oral administration of Pletal¢ç(Otsuka) in
84 korean healthy volunteers who participated in clinical studies to
investigate safety and pharmacokinetics of newly developed cilostazol
formulations. Fifty-eight subjects were genotyped for ABCB1. Adverse
events were classified into 2 categories, headache or the other CNS
symptoms such as nausea, vomiting and dizziness. The relationship
between the adverse events and genotype was assessed by the chi-square
test by using the SPSS 17.0. The reported headache cases in 3435CC,
CT and TT group are 3, 12 and 4 out of 24, 24 and 10, respectively. A
significant difference was observed in the incidence of headache by a 2.5
fold increase (p = 0.02) in the subjects carrying 3435T allele compared
to those who do not carry that allele. All other symptoms were not significant. Our data suggest that ABCB1 3435T allele is associated with
the headache after taking cilostazole. Further study is required to establish and validate the role of ABCB1 polymorphism in the incidence of
headche during cilostazole treatment.
Paper No.: 2166
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
EFFECTS OF CYP2D6*10 ALLELE ON THE
PHARMACOKINETICS OF ATOMOXETINE IN HEALTHY
KOREANS
Chang-Ik Choi, J-W Bae, M-J Kim, C-G Jang, S-Y Lee
Sungkyunkwan University, College of Pharmacy, Laboratory of Pharmacology, Suwon, South Korea
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
235
Atomoxetine is a selective norepinephrine receptor inhibitor (SNRI), and
is usedfor the treatment of Atomoxetine is a selective norepinephrine
receptor inhibitor (SNRI), and is used for the treatment of attention-deficit hyperactivity disorder (ADHD) in children, adolescents and adults.
Atomoxetine is metabolized via aromatic ring-hydroxylation, benzylic
hydroxylation and N-demethylation in human. Among these, aromatic
ring-hydroxylation is the major metabolic pathway and is primarily mediated by CYP2D6, the highly polymorphic drug metabolizing enzyme.
CYP2D6*10 allele, causing the decreased CYP2D6 enzymatic activitiy,
is specifically and frequently distributed in Asians, including Korea. We
investigated effects of CYP2D6*10 allele on the pharmacokinetics of
atomoxetine. After genotyping for CYP2D6 with volunteers, 32 healthy
Korean subjects were selected for this study. They were grouped according to CYP2D6 genotype, group 1 (n = 9, CYP2D6*wt/*wt), group 2
(n = 13, CYP2D6*wt/*10), and group 3 (n = 10, CYP2D6*10/*10). A
single oral dose of 40 mg of atomoxetine was administered to each subject and plasma concentration of atomoxetine was determined by using
high-performance liquid chromatography-tandem mass spectrometry system. Mean AUC values of atomoxetine in group 2 (1903 ± 1289 ng•hr/
mL) and group 3 (2640 ± 742 ng•hr/mL) were significantly higher than
those in group 1 (768 ± 125 ng•hr/mL) (P < 0.05 and P < 0.001,
respectively). Elimination half-life (t1/2) of atomoxetine in group 3 was
significantly longer than in group 1 and group 2 (P < 0.001 and
P < 0.05, respectively). Oral clearance (CL/F) of atomoxetine in group 2
(26.8 ± 10.4 L/hr) and group 3 (16.2 ± 4.2 L/hr) was significantly lower
than in group 1 (53.2 ± 7.4 L/hr) (both P < 0.0001). In conclusion,
CYP2D6*10 allele is found to affect the pharmacokinetics of atomoxetine in Koreans.
Paper No.: 3314
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
DOSE ADJUSTMENT OF OMEPRAZOLE BASED ON CYP2C19
GENOTYPE
Chang-Ik Choi, Sung-Min Moon, Jung-Woo Bae, Mi-Jeong Kim,
Choon-Gon Jang, Seok-Yong Lee
Sungkyunkwan University, School of Pharmacy, Suwon, South Korea
Omeprazole is a proton pump inhibitors (PPIs) that suppresses gastric
acid secretion by specific inhibition of the H+/K+-ATPase in the gastric
parietal cell, and is used for the treatment of gastroesophageal reflux disease (GERD), the healing of erosive esophagitis, and Helicobacter pylori
(H. pylori) eradication. Omeprazole is extensively metabolized by cytochrome P450 enzymes, and CYP2C19 is the main metabolizing enzyme
which is known to be highly polymorphic. It is reported that CYP2C19
genetic polymorphism can affects the pharmacokinetic and pharmacodynamic changes of several PPIs, including omeprazole. So we evaluated
the optimal dose adjustment of omeprazole in relation to CYP2C19
genotype. Forty-two healthy Korean volunteers were selected and were
divided into three groups according to CYP2C19 genotype,
CYP2C19EM (CYP2C19*1/*1, n = 15), CYP2C19IM (CYP2C19*1/*2
and CYP2C19*1/*3, n = 15), and CYP2C19PM (CYP2C19*2/*2,
CYP2C19*2/*3, and CYP2C19*3/*3, n = 12). Each group was received
a single oral dose of 75 mg, 60 mg and 20 mg omeprazole, respectively,
and the plasma concentration of omeprazole was measured by using
LC-MS/MS system. AUC0-¥ of omeprazole in each genotype group
(CYP2C19EM, CYP2C19IM and CYP2C19PM group) was
3239 ± 1997 ng/mLÆhr, 4000 ± 1173 ng/mLÆhr and 3690 ± 1179 ng/
mLÆhr, respectively. These differences were not statistically significant,
indicating similar plasma concentration of omeprazole between three
genotype groups. Doses of omeprazole used in this study can be applied
to the CYP2C19 genotype-based omeprazole dose determination in
Koreans.
Paper No.: 3315
FOCUSED CONFERENCE GROUP: P02 - TRANSMEMBRANE
TRANSPORT: PERSPECTIVES FOR DISEASE AND DRUG
DISCOVERY
PHARMACOKINETIC INTERACTIONS OF CIMETIDINE AND
PRAMIPEXOLE IN RELATION TO OCT2
Chang-Ik Choi(1), S-M Moon(1), J-W Bae(1), M-J Kim(1), C-G Jang(1),
S-Y Lee(1)
Sungkyunkwan University, School of Pharmacy, Su-Won, South Korea
Pramipexole is in a class of non-ergoline dopamine agonist and is
approved for the treatment of Parkinson’s disease and restless legs syndrome (RLS). After administration, pramipexole is well absorbed and
undergoes little presystemic biotransformation. Approximately 90% was
excreted in the urine as unchanged form, with possible involvement with
organic cation transporters (OCTs) in the renal tubules, especially OCT2.
So we investigated the effects of cimetidine, known as an OCT2 inhibitor, on the pharmacokinetics of pramipexole. Eighteen healthy Korean
subjects with OCT2 808GG genotype enrolled an open-labeled, twophase parallel clinical study. In the first phase (control phase), each subject received a single oral dose of 0.25 mg pramipexole. In the second
phase (cimetidine phase), the subjects administered cimetidine 400 mg
twice daily for five days. On the morning of day 6, the subjects received
a single oral dose of 0.25 mg pramipexole 2 hours later the administration of a 400-mg single oral dose of cimetidine. Plasma concentration of
pramipexole was determined by using LC-MS/MS system. In cimetidine
phase, Cmax and AUC0-¥ of pramipexole was 21% and 43% higher than
those in control phase (P < 0.001 and P < 0.0001, respectively). Oral
clearance of pramipexole in cimetidine phase was 30% lower than that in
control phase (P < 0.0001). Other pharmacokinetic parameters were not
statistically significant. From these results, OCT2 is found to be associated with the renal excretion of pramipexole in vivo. Further studies on
the relationship between OCT2 genetic variants and pharmacokinetics of
pramipexole will be meaningful.
Paper No.: 2881
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
PHARMACOGENETIC STUDY ON METABOLIC SYNDROME
INDUCED BY ATYPICAL ANTIPSYCHOTICS AND MOOD
STABILIZERS
Eva Choong(1), M Etter(2), B Oneda(1), H Tayebi(3), G Bondolfi(2),
CB Eap(1,3)
(1) CHUV - Unit of Biochemistry and Clinical Psychopharmacology,
Centre for Psychiatric Neuroscience, Department of Psychiatry, Lausanne-Prilly, Switzerland
(2) University of Geneva, and University Hospital of Geneva, Department of Psychiatry, Switzerland
(3) University of Geneva and University of Lausanne, School of PharmaceuticalSciences, Geneva, Switzerland
Metabolic syndrome (weight gain, alteration of lipid and glycaemia profiles) induced by atypical antipsychotics and/or mood stabilizers are of
high clinical concern for the long term patient morbidity and mortality.
The major aim of this trial is to identify pharmacogenetic susceptibility
factors for metabolic syndrome. For this purpose, a transversal study was
performed in a Swiss out-patient psychiatric division, with the inclusion
of 196 psychiatric patients. 69% of patients were receiving atypical antipsychotics, and 31% lithium or valproate (two mood stabilizers also
known to induce weight gain). Genotyping of pharmacodynamic candidate genes was performed using rtPCR and allelic discrimination assays,
after validation of the method by direct sequencing. Uncoupling protein
2 (UCP2), a mitochondria membrane transporter involved in the release
of stored energy, leptin receptor (LEPR) and fat mass and obesity
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
236
associated gene (FTO), 2 genes playing a role in satiety, were chosen as
candidate genes. Result: UCP2 rs660339 polymorphism is associated
with differences in HDL-cholesterol levels (p = 0.002), and with obesity,
with CC and CT carriers presenting a 3.1-fold increased risk of obesity
(95%CI 1.2-9.9) as compared to TT carriers. Significant association
between BMI change and LEPR polymorphism were found in female
patients treated with all studied drugs (p = 0.039), and between BMI
change and FTO polymorphism in patients treated with risperidone or
olanzapine (p = 0.003). Predicting metabolic syndrome side effects
remains complex and requires further investigations. However, each relevant gene showing an association with this important side-effect could
assist in the choice of the appropriate treatment for each individual
patient.
Paper No.: 2262
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
QUANTITATIVE PHOSPHOPROTEOMICS DISSECTION OF
7TM RECEPTOR SIGNALLING USING FULL AND BIASED
AGONISTS
Gitte Lund Christensen(1), CD Kelstrup(2), C Lyngsøe(3), U Sarwar(1),
R Bøgebo(1), SP Sheikh(4), S Gammeltoft(1), JV Olsen(2),
JL Hansen(3),
(1) Glostrup Hospital, Department of Clinical Biochemistry, Glostrup,
Copenhagen, Denmark
(2) University of Copenhagen, Novo Nordisk Foundation Center for
Protein Research, Faculty of Health Sciences, Copenhagen, Denmark
(3) University of Copenhagen, Danish National Research Foundation
Centre for Cardiac Arrhythmia, Laboratory for Molecular Cardiology,
Department of Department ofBiomedical Sciences, Copenhagen,
Denmark
(4) University of Southern Denmark, Odense University Hospital,
Department for Biochemistry, Pharmacology & Genetics, Odense,
Denmark
Seven-transmembrane receptors (7TMRs), signal through heterotrimeric
G proteins, but can also activate G protein-independent signalling pathways of which the impact and complexity are less understood. The
Angiotensin II type 1 receptor (AT1R) is a prototypical 7TMR an important drug target in cardiovascular medicine. ‘‘Biased agonists’’ which
blocks Gaq protein activity, and simultaneously activitate G protein independent pathways, confer important perspectives in treatment of cardiovascular diseases. In this study we performed a global quantitative
phosphoproteomics analysis of the AT1R signalling network. Applying
high-resolution mass spectrometry (LTQ Orbitrap MS) we compared the
phosphoproteome of the AT1R agonist Angiotensin II with the biased
agonist SII Angiotensin II. We quantified more than elleven thousand
phosphorylation sites of which 879 were regulated by Angiotensin II.
45% of the Angiotensin II regulated phosphorylations were also regulated by SII Angiotensin II. Analysis of phosphorylation site patterns displays a striking distinction between protein kinases activated by Gaq
protein-dependent and –independent mechanisms. This study provides
substantial novel insight into Angiotensin II signal transduction and is
the first study dissecting the differences between a full and a biased agonist from a 7TMR on a systems-wide scale. Importantly, it reveals a previously unappreciated diversity and quantity of Gaq protein-independent
signalling and uncovers novel signalling pathways. We foresee that the
amount and diversity of G protein independent signalling may be more
pronounced than previously recognized for other 7TMRs as well. Quantitative mass spectrometry is a promising tool for evaluation of signalling
properties of biased agonists to other receptors in the future.
Paper No.: 2317
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
CHANGING DOCTORS PRESCRIPTION BEHAVIOUR. A
DRUG COMMITTEE PERSPECTIVE: TNF INHIBITORS
Hanne R Christensen, H Thomsen
Bispebjerg Hospital, Department of Clinical Pharmacology, Copenahgen,
Denmark
Introduction: The introduction of TNF inhibitors in rheumatoid arthritis
has resulted in improved treatment options but also a marked increase in
drug expenses. In 2009 adalimumab, infliximab and etanercept were
responsible for the highest drug expenditure in Denmark; 75 mil. Euro.
Of the five regions in Denmark, the Capital Region is the only one with
specific guidelines in this area. The TNF inhibitors adalimumab, etanercept and infliximab were considered to have equal clinical efficacy in the
treatment of rheumatoid arthritis, but use of infliximab was estimated as
the cheapest TNF inhibitor, and hence was chosen as 1. Line treatment
when a TNF inhibitor was needed. Methods: The drug committee did a
joint venture with the specialists in rheumatology in completing a guideline for the use of TNF inhibitors for treating patients with rheumatoid
arthritis. A goal was set, that 80% of all new patients diagnosed with
rheumatoid arthritis and approved for TNF inhibitor treatment, should
receive infliximab. Results: After implementation of the guideline all
new patients were monitored in the period January – October 2009, and
out of 136 patients, 111 received infliximab corresponding to 82%,
which was a marked change in prescription behaviour. In the four other
regions, no such change was observed. Conclusion: Even though it is
difficult to change doctors prescription behaviour, guidelines for the initial treatment of patients with rheumatoid arthritis can be complied with,
when implemented on rheumatology wards.
Paper No.: 1696
FOCUSED CONFERENCE GROUP: P02 - TRANSMEMBRANE
TRANSPORT: PERSPECTIVES FOR DISEASE AND DRUG
DISCOVERY
PHARMACOGENETICS OF METFORMIN
Mette Marie Hougaard Christensen(1), CB Andersen(2), P Damkier(2),
HB Nielsen(3), K Brøsen(1)
(1) University of Southern Denmark, Institute of Public Health & Odense
UniversityHospital, Department of Biochemistry and Pharmacology,
Odense, Denmark
(2) Odense University Hospital, Department of Biochemistry and Pharmacology, Odense, Denmark
(3) Odense University Hospital, Department of Endocrinology & University of Southern Denmark, Faculty of Health Sciences, Institute of Clinical Research, Odense, Denmark
Introduction: Genetic polymorphisms in metformin transporters and their
impact at the steady state metformin plasma concentrations is evaluated
in a large well characterized type 2 diabetes cohort given by South Danish Diabetes Study. SNPs in PMAT, OCT1-2 and MATE 1-2 have been
linked to reduced pharmacodynaminc response and large interindividual
pharmacokinetic variation. Materials/Patients: South Danish Diabetes
Study was designed as a 2 x 2 x 2 factorial prospective, randomized,
double-blinded, placebo-controlled, multi-centre study comprising 386 in
8 parallel groups. It was initialized to investigate the optimal pharmacological combination of metformin, rosiglitazon and insulin Asp/NPH in a
cohort of type 2 diabetics. One hundred and eighty-six patients were
allocated to metformin, and repeated measurements of steady state concentrations were collected. The final results will be adjusted for gender,
age, BMI, duration of disease, metformin dose, co-medication, smoking,
creatinine clearance, HbA1c-pretreatment and polymorphisms in PMAT,
OCT 1-2 and MATE 1-2. Results: The preliminary unadjusted result
for 456 repeated measurements of metformin confirm the large inter-
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
237
individual variation in steady state metformin concentration:
769 ± 649ng/ml (SD), range: 9.7 to 4133ng/ml. In the cohort 369 samples could be genotyped. In the renal transporter OCT2, the A270S genotype frequencies were determined: Heterozygous 19%, homozygous
0.5% and wild-type 80.5%, which corresponds well with the databases at
NCBI. Conclusion: The preliminary results strengthen the thesis that
Danish type 2 diabetics have the same allele frequencies in transporter
SNP as healthy Caucasian, and it indeed underscores the enormous interindividual variation in metformin steady state concentration.
Paper No.: 493
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
IMPROVEMENT OF MEMORY IN MICE AND INCREASE OF
HIPPOCAMPAL EXCITABILITY IN RATS BY GINSENOSIDE
RG1’S METABOLITES GINSENOSIDE RH1 AND
PROTOPANAXATRIOL
Shifeng Chu, J Chen, Y Wang, N Chen, J Zhang
Institute of Materia Medica, Chinese Academy of Medical Sciences &
PekingUniversity, Department of Pharmacology, Beijing, PR China
Abstract: The aim of present study is to observe the effect of ginsenoside
Rg1’s metabolites (primary metabolit ginsenoside Rh1 and end metabolite protopanaxatriol) on learning and memory function. We employed
the step through test and electrophysiological study to investigate the
effects of Rh1 and Ppton learning and memory as well as hippocampal
excitability. The behavioral study showed that both ginsenoside Rh1 and
Ppt significantly ameliorated memory-impaired model induced by scopolamine in mice. Consistently, electrophysiological work revealed that
Rh1 and Ppt as well as their precursor Rg1 all increased hippocampal
excitability in the dentate gyrus of anesthetized rats. The metabolism of
Rg1 in cerebrospinal fluid was detected by HPLC-MS, indicating that
Rg1 could not convert into Rh1 or Ppt. These results demonstrated that
both Rh1 and Ppt have similar effect on improving memory and hippocampal excitability, suggesting that the role of ginsenoside’s sugarmoeties in biological activities is not as necessary as traditionally
considered.
Paper No.: 1563
FOCUSED CONFERENCE GROUP: P08 - DEVELOPMENTS IN
THE TREATMENT OF SEXUAL DYSFUNCTION AND
DISEASES OF THE LOWER URINARY TRACT
FRACTIONATION OF SAW PALMETTO EXTRACTS AND
BIOASSAY FRACTIONS IN ISOLATED RAT PROSTATE
Thiam Chua, J Simpson, S Ventura
tude to the contractions produced by the crude extracts. These responses
were attenuated in the presence of cocaine (3 lM) (P < 0.001, n = 6,
repeated measures ANOVA) or prazosin (0.3 lM) (P = 0.001, n = 6,
repeated measures ANOVA). Analysis of NMR and mass spectra and
comparison with authentic material confirmed that this bioactivity was
due to tyramine in the active fraction. In conclusion, tyramine in saw palmetto extract causes indirect a-adrenoceptor mediated contractions via
the release of noradrenaline from sympathetic neurons.
Paper No.: 956
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
HPLC METHOD DEVELOPMENT & VALIDATION FOR THE
DETERMINATION OF WARFARIN IN HUMAN PLASMA
Yung An Chua, WZW Abdullah, SH Gan
University Sains Malaysia, Human Genome Center, Kelantan, Malaysia
Introduction: In many third world countries, not all laboratories have
sophisticated equipment such as Liquid chromatography Mass Spectrometry (LCMS) for fast analysis of serum drug levels. Objective: The aim
of our study is to develop a simple High Performance Liquid Chromatography (HPLC) method coupled with a UV detector for the determination of racemic serum concentrations of warfarin. Method: Warfarin and
phenylbutazone (internal standard) were spiked into 1 ml of human
serum before extraction using a liquid-liquid extraction (LLE) method
consisting of 3 ml ethyl-acetate. The samples were dried and reconstituted in 100 ml of acetonitrile before HPLC injection (10 ml). The
mobile phase comprised of 30% acetonitrile and 70% potassium dihydrogen orthophosphate buffer (0.01 M) with pH adjusted to 6.5. Separation
was performed on an endcapped C18 (250 X 4.6 mm I.D., 5 mm)
reversed phase column. The flow rate was set at 1.0ml/min with a total
run time of 14 min. The Photodiode Array (PDA) absorbance range for
warfarin was set as 210-350 nm. Result: The extraction recovery for warfarin was excellent (98%). Our limit of detection was 50 ng/ml. The
method was linear over the concentration range between 100 ng/ml and
6.0 mg/ml. The precision as well as accuracy meets the guideline laid by
the FDA.Conclusion: The developed HPLC method was found to be
suitable for the determination of serum warfarin concentrations for a clinical study.
Paper No.: 1997
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
PHARMAPOLY, GAME AS A TEACHING STRATEGY IN
PHARMACOLOGY CLASSES
Luis Cifuentes
Monash University, Monash Institute of Pharmaceutical Sciences, Parkville, VIC, Australia
University of the Andes, Faculty of Medicine, Bogota, Colombia
Extracts of saw palmetto berries (Serenoa repens) have been widely used
as a herbal remedy for benign prostatic hyperplasia (BPH). However, little is known about the beneficial physiological role of the bioactives and
which of the active phytochemicals are involved in improving the symptoms of BPH. The purpose of this study was to isolate and investigate
the bioactive compounds from saw palmetto extract that affect prostatic
smooth muscle. A commercially available saw palmetto extract (MediHerb Pty Ltd, Warwick, Queensland) was lyophilized and subjected to
fractionation using column chromatography. The composition of fractions was assessed by proton nuclear magnetic resonance spectroscopy
(1H NMR) and mass spectrometry (MS). Non-polar fractions were found
to contain fatty acids and their esters as the major constituents. In contrast, polar fractions revealed carbohydrate components (mono- and disaccharides) and alkaloids. In addition, the polar fractions were found to
produce contractions of isolated rat prostates that were similar in magni-
Introduction. Pharmacology teaching is an ongoing challenge for the teacher and students, in order to achieve that knowledge and motivation to
persist. The aim of this paper is the incorporation of games as a teaching
strategy (Ulrich D., et al. J of Nursing Education 2005;44:338-339) in
pharmacology classes, through the development of a material called
Pharmapoly. Materials. Using the principles of the game called Monopoly, I have developed a board with eighteen portraits of famous pharmacologists due to their discoveries. Cards and dices to play are related to
the area of pharmacology. Each pharmacologist has assigned several
questions related to the topics of research and development, pharmacokinetics and pharmacodynamics, which must be answered correctly by students to advance and finish with the highest number of pharmacologists
in their possession. Results. During the 1st half of 2009 it was made a
pilot with 43 students of the Faculty of Medicine, University of Los
Andes in Colombia, with the module of pharmacodynamics. The
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
238
comments at the end of the module by students were: 1. It was fun, 2.
We have learned a lot, and 3. We want to continue playing and learning.
Conclusion. The teaching of pharmacology can be facilitated by the incorporation of methodologies such as educational games, which complement
any teaching activity and generate continuous motivation in students.
Paper No.: 1998
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
SUMMARY OF PK AND PD IN A PRACTICAL WAY
Luis Cifuentes
University of the Andes, Faculty of Medicine, Bogota, Colombia
Introduction. One of the great barriers of medical students who are in
training of pharmacology and even the medical graduates is to understand and internalize the basic principles of pharmacokinetics (PK) and
pharmacodynamics (PD). The aim of this paper is to present an approach
of a summary of PK and PD in only one sheet for each concept. Materials: We have reviewed in 10 of pharmacology reference texts, the basic
principles of PK and PD, which has been supplemented with the teacher’s teaching experience in pharmacology. The result was an integration of all key concepts for PK and PD, in only one sheet, respectively.
Results. Each of the two sheets previously have been delivered to undergraduate medical students in teaching modules of PK and PD. Their conclusions after students have seen these sheets before a class of PK and
PD, were how easy the approach was, and after the module they mentioned that facilitates the recall the concepts, interaction with reference
texts and as continuous reference material. Conclusion. This tool has the
advantage of being able to access through the Internet via public domain
to help medical students and health professionals on the need to have
continued incorporation on PK and PD in the study and prescription of
drugs.
Paper No.: 1869
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
CROTOXIN: A TOXIN FROM RATTLESNAKE VENOM THAT
INDUCES A LONG-LASTING INHIBITORY EFFECT ON THE
PHAGOCYTIC ACTIVITY OF NEUTROPHILS
Maria Cristina Cirillo(1), TS Lima(1), SC Cataneo(1), SC Sampaio(1),
MC Della-Casa(2)
(1) Butantan Institute, Laboratory of Pathophysiology, São Paulo, Brazil
(2) Butantan Institute, Laboratory of Immunopathology, São Paulo,
Brazil
Crotalus durissus terrificus snake venom (CdtV) and crotoxin, its main
component, have long-lasting anti-inflammatory properties and inhibit
the phagocytic activity of macrophages. Recently, we demonstrated that
CdtV inhibits phagocytosis by neutrophils. Despite these findings, the
component of CdtV responsible for phagocytosis inhibition is unknow.
This study investigated the effect of crotoxin and other fractions (peak I
and III), obtained during crotoxin purification, on C3b-receptor-mediated
phagocytosis, by neutrophils. Neutrophils were obtained from peritoneal
cavity of Wistar rats, 4h after intraperitoneal administration of carrageenan (4.5 mg/kg) and phagocytosis of opsonizaded zymosan was evaluated after different treatments. In vitro, cells (1.2x106cells/mL) were
incubated with crotoxin (peak II), peak I or III, at different concentrations
(0.02, 0.04, 0.08, 0.16 or 0.32 lg/mL) or RPMI1640 (control). In vivo,
crotoxin (0.11 mg/kg) or saline (control) was administered subcutaneously to rats at different time periods: 2h, 1, 4 or 14 days before or 1h
after the administration of carrageenan. In vitro, crotoxin significantly
reduced the phagocytic activity of neutrophils (0.02 lg/mL: 29%,
0.04 lg/mL: 24%, 0.08 lg/mL: 28%, 0.16 lg/mL: 26% and 0.32 lg/
mL: 24%). A single dose of crotoxin also reduced the percentage of
phagocytosis (2h: 24%, 1 day: 31%, 4 days: 25%, 14 days: 18% and 1h
later: 35%). These results show that crotoxin is the component responsible for the inhibitory effect of CdtV on phagocytosis by neutrophils. Taking into account the role of neutrophils in inflammation, these data
reinforce that crotoxin is a potential natural product in controlling inflammatory diseases.
Support: FAPESP and CNPq/INCTTOX.
Paper No.: 1870
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
CROTOXIN: A TOXIN FROM RATTLESNAKE VENOM WITH
POTENTIAL ANTI-INFLAMMATORY PROPERTIES
Maria Cristina Cirillo(1), FPB Nunes(1), BC Zychar(1),
MS Della-Casa(2), SC Sampaio(1), LRC Gonçalves(1)
(1) Butantan Institute, Laboratory of Pathophysiology, São Paulo, Brazil
(2) Butantan Institute, Laboratory of Immunopathology, São Paulo,
Brazil
Crotalus durissus terrificus (CdtV) snake venom exhibit a long-lasting
anti-inflammatory effect, inhibiting paw edema and cell migration to the
peritoneal cavity induced by carrageenan. However, the contribution of
purified components of CdtV on this effect has not been evaluated yet;
thus the effect of crotoxin (CTX), the main toxin of CdtV, and other
components were investigated on paw edema, cellular migration and leukocyte-endothelium interaction induced by carrageenan. A single dose of
CTX (0.89lg/50lL s.c.), other components (1.0lg/50lL s.c.) or saline
(50lL) was injected in mice, 7 days before intraplantar injection of carrageenan (300lg/50lL) or saline (contralateral paw) and edema was evaluated. Total and differential cell counts were determined 4h after
intraperitoneal injection of carrageenan (300lg/200lL) in animals treated
with CTX, saline or other components 7, 14 or 21 days earlier. Leukocyte-endothelium interaction was induced 1h after injection of carrageenan or saline into the scrotum of animals, 7 or 14 days before the
injection of CTX, saline or other components. Adherent, or in rolling
leukocytes were counted by intravital microscopy 1h after carrageenan or
saline injection. The results showed that a single dose of CTX, but not
other components, reduced edema (30%), cell migration (30%) and
altered leukocyte-endothelium interaction, increasing the rolling (38%)
and reducing the adherent cells (63%) induced by carrageenan compared
with controls. Thus, CTX is the component involved in the long-lasting
anti-inflammatory effect of CdtV, indicating that it might be used as a
new approach for treating inflammatory diseases.
Supported by FAPESP, CAPES and CNPQ/INCTTOX.
Paper No.: 1871
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
INVOLVEMENT OF FORMYL PEPTIDE RECEPTORS ON THE
LONG-LASTING ANTI-INFLAMMATORY EFFECT OF
CROTOXIN, A TOXIN FROM RATTLESNAKE VENOM
Maria Cristina Cirillo(1), FPB Nunes(1), BC Zychar(1),
MS Della-Casa(2), SC Sampaio(1), LRC Gonçalves(1)
(1) Butantan Institute, Laboratory of Pathophysiology, São Paulo, Brazil
(2) Butantan Institute, Laboratory of Immunopathology, São Paulo,
Brazil
Crotalus durissus terrificus snake venom (CdtV) and crotoxin, the main
component of this venom, present long-lasting anti-inflammatory proprieties on paw edema and cell migration induced by carrageenan. Other
investigation showed that CdtV and crotoxin inhibit phagocytosis by neutrophil and macrophages. Moreover, crotoxin increases the production of
lipoxin A4, by macrophages. Lipoxins, anti-inflammatory mediators generated by lipoxygenase pathway exert their biological actions by binding
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
239
to specific receptors FPR2/ALX that belongs to formyl peptide receptors
family. In this study we investigated the involvement of formyl peptide
receptors on the long-lasting inhibitory effect of crotoxin on cell migration induced by carrageenan. The Boc-2 (10 lg/200 lL i.p.), a selective
antagonist of formyl peptide receptors was administered 30 minutes prior
to the subcutaneous injection of crotoxin (0.89 lg/50 lL) or saline. The
treatment with crotoxin (single dose) or saline was undertaken 1 hour, 7
or 14 days before intraperitoneal injection of carrageenan (300 lg/
200 lL). After 4 hours of carrageenan injection, cellular migration to the
peritoneal cavity was evaluated. Pretreatment of animals with crotoxin,
significantly diminished carrageenan-induced cell influx into the peritoneal cavity, when compared with the control group. These decreases
were 47% (1 hour), 44% (7 days) and 33% (14 days). Pretreating animals with Boc-2 abolished the decrease in cell migration induced by crotoxin in all time evaluated. In conclusion, results suggest that formyl
peptide receptors seem to play a role in the anti-inflammatory effect of
crotoxin and also indicate the involvement of the lipoxin in this effect.
Supported by FAPESP, CAPES, CNPQ/INCTTOX.
Paper No.: 3118
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
TYPE 2 DIABETES-ASSOCIATED NON-SYNONYMOUS
MUTATIONS IN THE GENE CODING FOR THE MELATONIN
MT2 RECEPTOR IMPAIR MELATONIN SIGNALLING
Nathalie Clement(1), A Bonnefond(2), K Fawcett(3), E Vaillant(2),
J-L Guillaume(1), B Balkau(4), M Marre(5), G Charpentier(6),
M Vaxillaire(2), I Barroso(3), R Jockers(1), P Froguel(2,7)
(on behalf of the MAGIC consortium)
(1) Institut Cochin, Inserm U567, CNRS-UMR8104, Paris, France
(2) CNRS-UMR8090, Pasteur Institute, Lille, France
(3) Wellcome Trust Sanger Institute, Metabolic Disease Group, Hinxton,
UK
(4) INSERM-U780, Villejuif, France
(5) INSERM-U695, Paris, France
(6) Corbeil-Essonnes Hospital, Endocrinology-Diabetology Unit,
Corbeil-Essonnes,France
(7) Imperial College, Department of Genomic Medecine, London, UK
Using genome-wide association study data, we have recently identified a
single nucleotide polymorphism (SNP) located within the MTNR1B gene,
encoding the MT2 melatonin receptor, associated with an increase of
both fasting plasma glucose (FPG) levels and risk of type 2 diabetes
(T2D). Melatonin is a neurohormone, which regulates the circadian
rhythm and may contribute to glucose homeostasis. Melatonin signalling
is mainly mediated by its G protein-coupled MT1 and MT2 receptors.
Though the association signal of the MTNR1B SNP rs10830963 is
strongly significant (P < 10-30), its effect on FPG and T2D risk is modest. We hypothesized that rare non-synonymous mutations in MTNR1B
may have a larger effect on T2D risk and may contribute to the haplotype association signal. We sequenced MTNR1B exons in 6,878 European samples including 2,271 individuals with T2D. All identified
mutations were generated and expressed as Myc-tagged fusion proteins.
Ligand binding properties, surface expression and signalling (cAMP,
ERK1/2) of mutant receptors was studied in HEK 293 cells. We identified a total of 40 non-synonymous variants including 35 rare mutations
(minor allele frequency < 0.001). Preliminary statistical analyses showed
that these mutations tend to be more prevalent in individuals with T2D,
compared to nondiabetic controls. So far we have found that three mutations are devoid of 125I-MLT binding, all present in individuals with elevated FPG. Subsequent analyses are in progress. Our present findings
show that MTNR1B coding regions are variable with putative loss of
function mutations being identified only in individuals with T2D or
impaired fasting glucose. Given that MTNR1B may mediate melatonininduced inhibition of insulin secretion, wouldn’t we expect loss-of-function mutations to decrease FPG/decrease risk of T2D.
Paper No.: 2665
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
CONTRIBUTION OF SMOOTH MUSCLE BKCA CURRENTS
TO OUTWARD CURRENTS RECORDED FROM
ENDOTHELIAL CELLS IN SITU
Belén Climent(1), P de la Villa(2), U Simonsen(3), A Garcı́a-Sacristán(1), L Rivera(1)
(1) University Complutense of Madrid Faculty of Pharmacy, Department
of Physiology, Madrid, Spain
(2) University of Alcala, Department of Physiology, Madrid, Spain
(3) University of Aarhus, Department of Pharmacology, Aarhus, Denmark
In rat mesenteric arteries large-conductance calcium-activated K channels
(BKCa) have been reported to be restricted to vascular smooth muscle
cells (VSMC) (Mistry & Garland, J Membr Biol 164:125 38, 1998). The
present study hypothesized that changes in K current in the vascular
smooth muscle layer will result in changes in endothel cell (EC) K currents. Simultaneous recordings of vascular tone and ionic currents by use
of in situ patch clamp were performed on rat mesenteric arterial segments
mounted in a microvascular myograph. Families of depolarizing voltage
pulses (from -60 to +140mV; holding potential -60mV) were applied
before and after: phenylephrine (Phe), acetylcholine (ACh)+Phe; and iberiotoxin (IbTx)+ACh+Phe or carbenoxolone+ACh+Phe. In mesenteric
arteries incubated with carbenoxolone (45-60 minutes), the same protocol was applied in the presence of Phe+ACh and after IbTx+Phe+ACh.
Outward potassium currents were recorded. ACh induced potassium-current amplitude was decreased both by IbTx (30 ± 3 to 11 ± 3 pA/pF;
n = 5; P < 0.01) and carbenoxolone (35 ± 5 to 19 ± 7 pA/pF; n = 5;
p < 0.05). Incubation with carbenoxolone abolished the effect of IbTx
on the potassium currents (n = 7, p < 0.05). IbTx-induced reversion of
the ACh induced arterial relaxation with or without carbenoxolone was
similar (87%, n = 7; 91%, n = 5, respectively, p > 0.05). Gap junctions
are active in EC in situ and the blockade with IbTx suggests that BKCa
channels contribute to these currents. These data suggest that the currents
recorded from EC in situ are modulated by smooth muscle cell BKCa
currents, an effect mediated most likely through gap junctions.
Supported by M.E.C., Spain, Grant No. BFU2007-67732/BFI.
Paper No.: 2666
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
INCREASED OUTWARD CURRENTS RECORDED IN SITU
FROM ENDOTHELIAL CELLS OF RABBIT CORONARY
ARTERIES AFTER ISCHEMIA REPERFUSION
Belén Climent(1), AL Garcı́a-Villalón(2), N Fernández(2), L Monge(2),
G Diéguez(2), A Garcı́a Sacristán(1), L Rivera(1)
(1) University Complutense of Madrid Faculty of Pharmacy, Department
of Physiology, Madrid, Spain
(2) University Autónoma of Madrid Faculty of Pharmacy, Department of
Physiology, Madrid, Spain
Ischemia–reperfusion is a clinical and experimental event that can produce dysfunction of the coronary vasculature, and evidences suggest that
endothelium dysfunction is involved. Rabbit hearts were perfused at constant flow and then exposed to 60 min global zero-flow ischemia followed by 30 min reperfusion. Segments of the descendent coronary
artery were mounted in a microvascular myograph. Simultaneous recordings of vascular tone and ionic currents by use of in situ patch clamp
were performed both on control and ischemic arteries. Families of depolarizing voltage pulses (from -60 to +140mV; Vh= -60mV) were applied
before and after endothelin 1(ET-1), and ET-1 + acetylcholine (ACh).
Outward potassium currents were recorded. The peak current value was
similar in ischemic and control segments (353 ± 59, n = 10, 281 ± 47
pA, n = 9; respectively; p = 0.167). However, the current density was
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
240
higher in ischemic vessels (48 ± 6, and 25 ± 5 pA/pF, respectively;
p < 0.05). In contrast, membrane capacitance values were lower in ischemic arteries (7 ± 1 vs 12 ± 1 pF; P < 0.01). ET-1 simultaneously
increased isometric tension and decreased outward currents in both arteries (14.9 ± 3 pA, n = 5, P < 0.05 and 7,3 ± 3, n = 4, P < 0.01, respectively). After ET-1, ACh induced relaxation and increased outward
currents in control but not in ischemic segments (23 ± 4, P < 0.01; and
10 ± 4 pA/pF p = 0.089, respectively). These preliminary data may suggest that increased current density along with capacitance reduction after
ischemia reperfusion injury might be due to an increase in K+ current
expression and to an impaired gap junctional communication. Further
experiments are needed to clarify this hypothesis.
Supported by M.E.C., Grants No. BFU2007-67732/BFI, and F.I.S., No
PS09/00394, Spain.
Paper No.: 2804
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
CERAMIDE IS INVOLVED IN THE HYPOXIC
VASOCONSTRICTION OF CHICKEN PULMONARY AND
CHORIALLANTOIC ARTERIES
Angel Cogolludo(1), E Villamor(2), J Moral-Sanz(1), R Scheepers(1), B
Barreira (1), C Menendez(1), L Escolano(1), L Moreno(1), F Perez-Vizcaino(1),
(1) University Complutense of Madrid School of Medicine, Department
of Pharmacology,Madrid, Spain
(2) Univertsity of Maastricht, The Netherlands
Ceramide is a sphingolipid-derived second messenger that plays an
important role in numerous physiological processes such as cell growth,
apoptosis and differentiation. We have recently reported that neutral
sphingomyelinase (nSMase)-derived ceramide is involved in hypoxic
pulmonary vasoconstriction in rats (Cogolludo et al, Cardiovasc Res.
2009; 82: 296-302). We hypothesized that ceramide may play a role in
other species and in other vessels exhibiting a hypoxic vasoconstrictor
response. Thus, in the present study we have studied if ceramide mediates the hypoxic responses in chicken pulmonary (PA) and choriallantoic
arteries (CA). Experiments were performed in CA and PA isolated from
chicken fetuses (15-day incubation) and newborns (1-3days), respectively, and in myocytes isolated from these tissues. Exposure to hypoxia
induced a contraction in both PA and CA. Similarly, hypoxia increased
the ceramide content measured by immunocytochemistry. The nSMase
inhibitor GW4869 (10 lM) reduced the contractile responses induced by
hypoxia but not those elicited by endothelin-1. Hypoxic responses were
also attenuated by an anticeramide antibody. Moreover, C6-ceramide
(10 lM) and bacillus cereus SMase (100 mU/ml) mimicked the effects
of hypoxia, i.e. they produced a sustained contractile response. Finally,
the mRNAs of nSMase2 was expressed in PA and CA as measured by
real-time RT-PCR. In conclusion, nSMase-derived ceramide plays a key
role in the hypoxic responses found in two specialized oxygen sensing
vessels from chickens.
Supported by ‘Fundación de Investigación Médica Mutua Madrileña’.
Paper No.: 1034
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
INHIBITION OF THE 26S PROTEASOME COMPLEX BY
CHEMICAL CONSTITUENTS OF SMILAX REGELII AND ITS
POSSIBLE CHEMOTHERAPEUTIC CAPABILITIES ON
MAMMARY TUMOURS IN SPRAGUE DAWLEY RATS
Damian Cohall(1), PDA Singh(2)
The folklore anti-neoplastic claim of Smilax regelii was investigated on
mammary tumour models in female Sprague Dawley rats by probing
the plant’s ability to inhibit the 26S Proteasome complex (Adams J,
Drug Discovery Today 2003; 8: 307-315). Bioactivity of the plant’s
extract was assayed in vitro with the 26S proteasome. Rats were treated
with the aqueous extract (35.7 mg plant material/Kg i.p.) in a spontaneous tumour model. A possible bioactive constituent of the extract and
MG-132, a known 26S proteasome inhibitor (Adams J, Drug Discovery
Today 2003; 8: 307-315), were administered (1.0 mg/Kg i.p.) to respective groups of animals in a 7, 12-Dimethylbenzanthracene induced
tumour model. Tumour sizes were estimated using the formula, 1/
6p[(d1 · d2)3/2] where d1 and d2 are the two perpendicular diameters
(Vigushin DM et al, Clinical Cancer Research 2001; 7: 971-976). Histopathology was done on tumour tissue samples. The extract inhibited the
26S Proteasome dose dependently. Chemical analysis of the extract
indicated that a possible bioactive constituent is Dioctyl Adipate.
Tumour growth within the groups treated with the aqueous extract
(p > 0.05, n = 12, Wilcoxon Signed Ranked test) and Dioctyl Adipate
(p > 0.05, n = 6, Student’s T test) was not significant compared to the
control groups. The tumours in both models were fibroadenomas and
remission of the neoplasia in the test groups was due to induced apoptosis. Dioctyl Adipate and its analogues are potential anti-neoplastic
agents. Further investigation of these agents as drug leads in the drug
discovery pipeline is recommended.
Paper No.: 3291
FOCUSED CONFERENCE GROUP: P18 - NUCLEAR
RECEPTOR TARGETS FOR TREATMENT OF DISEASES
ACTIVATION OF PEROXISOME PROLIFERATOR-ACTIVATED
RECEPTOR B/D (PPARB/D) BY GW501516 PREVENTS FATTY
ACID-INDUCED NF-JB ACTIVATION AND INSULIN
RESISTANCE IN SKELETAL MUSCLE CELLS
T Coll, AM Gomez-Foix, Manuel Vazquez-Carrera
University of Barcelona, Department of Pharmacology and Therapeutic
Chemistry, Barcelona, Spain
At present it is unknown whether Peroxisome Proliferator-Activated
Receptor (PPAR)b/d activation prevents fatty acid-induced inflammation and insulin resistance in skeletal muscle cells. In C2C12 skeletal
muscle cells, the PPARb/d agonist GW501516 prevented phosphorylation of IRS-1 at Ser307 and the inhibition of insulin-stimulated Akt
phosphorylation caused by exposure to the saturated fatty acid palmitate. This latter effect was reversed by the PPARb/d nantagonist
GSK0660. Treatment with the PPARb/d agonist enhanced the expression of two-well known PPARb/d-target genes involved in fatty acid
oxidation, carnitine palmitoyltransferase-1 (CPT-1) and pyruvate dehydrogenase kinase 4 and increased the phosphorylation of AMPK, preventing the reduction in fatty acid oxidation caused by palmitate
exposure. In agreement with these changes, GW501516 treatment
reversed the increase in DAG and PKC activation caused by palmitate. These effects were abolished in the presence of the CPT-1 inhibitor etomoxir, thereby indicating that increased fatty acid oxidation was
involved in the changes observed. Consistent with these findings,
PPARb/d activation by GW501516 blocked palmitate-induced NF-jB
DNA-binding activity. Likewise, drug treatment inhibited the increase
in interleukin 6 expression caused by palmitate in C2C12 and human
skeletal muscle cells as well as the protein secretion of this cytokine.
These findings indicate that PPARb/d attenuates fatty acid-induced
NF-jB activation and the subsequent development of insulin resistance
in skeletal muscle cells by reducing DAG accumulation. Our results
point to PPARb/d activation as a pharmacological target to prevent
insulin resistance.
(1) University of the West Indies, Faculty of Medical Sciences, Cave Hill
Campus, Black Rock, St Michael, Barbados
(2) University of the West Indies, Mona Campus, Jamaica
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
241
Paper No.: 729
FOCUSED CONFERENCE GROUP: P19 - GENERAL SESSION
EFFECT OF ENVIRONMENTAL TEMPERATURE ON
NOREPINEPHRINE-INDUCED CONTRACTION IN ISOLATED
CHICKEN CUBITAL VEIN: GENDER VARIATION
Simon Comerma-Steffensen(1), J Rojas(1), A Risso(1), M Rossini(1),
V De Basilio(2), I Oliveros(3), H Zerpa(1)
(1) Central University of Venezuela, Veterinary Faculty, Department of
Biomedical Sciences, Maracay, Venezuela
(2) Central University of Venezuela, Agronomy Faculty, Maracay, Venezuela
(3) National Institution of Agropecuary Research, Venezuela
The rise of global environmental temperature might impact thermoregulation of several organisms. To assess this issue in chickens (Ross linebroilers), segments of the wing cubital vein were harvested from 5 weeks
old individuals (n = 5-9) killed for human consumption. Isolated vessels
were obtained from male and female chickens, maintained at two breeding temperatures (30C ‘‘warm’’ and25C ‘‘cool’’) and prepared for isometric tension recording in organ baths. Endothelium integrity was
established by the relaxing response to acethylcolinein phenylephrinecontracted vessels. The response to a depolarising Krebs solution (DKS
118 mM KCl) was assessed followed by the construction of concentration response curves (CRC) to norepinephrine (10-9-10-4M). The Emax
and pD2 values were compared by unpaired Student’s tests. The DKS
peak height contraction was similar between males and females at both
breeding temperatures. Norepinephrine-induced Emax was also similar
between gender and temperatures; however, the norepinephrine-induced
Emax was higher (p<0.05) in vessels from females bred at ‘‘cool’’
(1.85 ± 0.4g/mg) vs. ‘‘warm’’ (0.88 ± 0.1g/mg) temperatures.These
results might suggest a reduced efficacy to norepinephrine in cutaneous
veins from females chickens under warming conditions, in contrast to the
response in males. The potential better physiological adaptability of
superficial veins from females chickens might contribute to overcome
higher environmental temperatures.
Paper No.: 1153
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
RECEPTOR MECHANISMS OF THE LONG-LASTING AND
PERSISTENT ARTERIAL CONTRACTILE EFFECTS OF
ENDOTHELIN-1
Matthijs Compeer(1), M Meens(1), C Höltke(2), W Neugebauer(3), J De
Mey(1)
(1) Maastricht University, Department of Pharmacology & Toxicology,
Maastricht, The Netherlands
(2) The University Hospital of Münster, Münster, Germany
(3) Sherbrooke University, Sherbrooke, QC, Canada
Introduction: Endothelin-1 (ET-1) acts as a high affinity, irreversible
polyvalent ligand on ETA-receptors. Relations between these characteristics have not been established. Materials: We used rat mesenteric resistance arteries, rhodamine-labeled ET-1 (to monitor ligand-binding by
2-photon laser scanning microscopy focusing on smooth muscle) and
native ET-1 (monitoring contractions by wire-myography in conditions
minimizing endothelial and sensory-nervous effects). Results: ET-1 (1 16nM) caused strong contractions, persisting in the absence of free agonist. The ETB-antagonist BQ788 (1uM) partly prevented binding, but
did not modify contractile effects of ET-1. The peptidergic ETA-antagonist BQ123 (1uM) prevented but not reversed binding persisting in presence of BQ788. The potent non-peptidergic ETA-antagonist PD156707
(1 - 100nM) and BQ123 (0.1 - 3uM) reduced sensitivity to ET-1-induced
contractions. They partly and reversibly reduced contractions in the pres-
ence and after exposure to ET-1. Preventive effects of Cy5.5-PD156707
and FITC-BQ123 were comparable to their unlabeled analogues. However, the bulky labeled antagonists were less effective in reducing contractions in presence and after exposure to ET-1. Conclusion: These
findings indicate sequential bitopic binding of ET-1 to ETA-receptors;
one agonist part interacts dynamically with an antagonist-sensitive binding-site (site H). Thereafter, another agonist part binds irreversibly to
another part of the receptor (site L). Signalling is triggered by agonistoccupied site L and can be enhanced by site H. Differences between
large versus small antagonists suggest ‘‘charnière’’ (hinge) behaviour of
the agonist molecule, irreversibly bound to one of its binding sites. This
study was performed within the framework of TI Pharma projects T2301 and T2-108.
Paper No.: 1013
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
JNK SIGNALLING PATHWAY INVOLVES IN AB1-40-INDUCED
NEURONAL DEATH AND PROTECTIVE EFFECT OF
EXTRACTS OF GINSENG AND GINKGO BILOBA
Weihong Cong(1), J Liu(1), L Sheng(1), C Lin(2)
(1) China Academy of Chinese Medical Sciences, Xiyuan Hospital,
Research Center, Beijing, PR China
(2) Shandong University of Traditional Chinese Medicine, Shandong, PR
China
b-amyloid peptide (Ab) has been implicated as a key molecule in the
neurodegenerative cascades of Alzheimer’s disease (AD). Meanwhile,
studies on the c-Jun N-terminal kinase (JNK) signalling pathway strongly
suggested that it plays an important role in mediating the neuronal death
induced by Ab. Therefore, we aimed to investigate the protective effects
of a combination of extracts of ginseng and Ginkgo biloba (NWK) on
neuronal death induced by Ab 1-40 in rats and the candidate molecular
mechanism. A 4-week NWK (150 mg/kg and 15mg/kg, respectively)
administration to rats was performed daily after bilateral injection of Ab
1-40 (4 lg/lL for each side) into hippocampus. Nissl staining, TUNEL
staining and phosphorylated JNK (p-JNK) immunohistochemical staining
were adopted to observe the changes of hippocampal neurons. Western
blot was used to detect the phosphorylated proteins of c-Jun and JNK.
The results showed that NWK (150 mg/kg) could prevent the loss of
Nissl staining neurons, reduce the TUNEL-positive and p-JNK-positive
neurons, and inhibit c-Jun and JNK phosphorylation in hippocampus.
Our study suggested that JNK signal transduction pathway involve in the
Ab 1-40-induced neuronal death and the combination of extracts of ginseng and Ginkgo biloba might attenuate the neuronal toxicity ofAb 1-40
by blocking JNK signalling cascades.
Paper No.: 1382
FOCUSED CONFERENCE GROUP: P08 - DEVELOPMENTS IN
THE TREATMENT OF SEXUAL DYSFUNCTION AND
DISEASES OF THE LOWER URINARY TRACT
VASCULAR INSULIN RESISTANCE IN PENILE ARTERIES
FROM A RAT MODEL OF METABOLIC SYNDROME
C Contreras, A Sánchez, B Climent, A Garcı́a-Sacristán, S Benedito,
Dolores Prieto
University Complutense of Madrid, Faculty of Pharmacy, Department of
Physiology, Madrid, Spain
Metabolic and cardiovascular abnormalities accompanying metabolic
syndrome, i.e. obesity, insulin resistance (IR) and hypertension, are all
associated to endothelial dysfunction and act as independent risk factors
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
242
for erectile dysfunction (ED). Our purpose was to investigate the vascular effects of insulin in penile arteries (PA) and whether these effects are
impaired in a rat model of IR/metabolic syndrome. PA from Obese
Zucker rats (OZR) and from Lean Zucker rats (LZR), were mounted on
microvascular myographs and the effects of insulin were assessed in the
absence and presence of endothelium and of specific inhibitors of nitric
oxide (NO) synthase (NOS), phosphatidil-inositol-3-kinase (PI3K), mitogen-activated protein kinase (MAPK), endothelin 1 (ET-1) receptors and
cyclooxygenase. The insulin-induced changes in the intracellular
Ca2 + concentration [Ca2 + ]i were also examined. OZR exhibited mild
hyperglycemia, hypercholesterolemia, hypertryglyceridemia and hyperinsulinemia. Insulin induced endothelium and NO-dependent relaxations in
LZR that were impaired in OZR. Inhibition of PI3K reduced the insulinand the b-adrenoceptor agonist isoproterenol-induced vasodilatation
mainly in arteries from LZR. Antagonism of ET-1 receptors did not alter
insulin-induced relaxation in either LZR or OZR, but MAPK blockade
increased this vasodilatation in OZR, which was inhibited by cyclooxygenase blockade. Insulin induced a decrease [Ca2 + ]i which was
impaired in OZR. In conclusion, insulin-induced vasodilatation is
impaired in PA of OZR due to altered NO release through the PI3K pathway and unmasking of a MAPK pathway-mediated vasoconstriction.
This vascular IR is likely to contribute to the endothelial dysfunction and
to the ED reported in IR states.
Supported by grant SAF2006-09191 from MICINN
Paper No.: 2077
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
HIGH LEVEL OF BLOOD ZOLPIDEM CONCENTRATIONS IN
THE MORNING AND IN THE EARLY AFTERNOON AFTER A
SINGLE DOSE OF A HYPNOTIC DRUG FOLLOWED BY A
NIGHT OF SLEEP IN HEALTHY ELDERLY SUBJECTS :
POTENTIAL LINK WITH IMPAIRED DRIVING
Antoine Coquerel(1,2), M Loilier(1,2), S Marie(1,2), F Bertran(3),
V Lelong-Boulouard(1,2), P Denise(2,3), M-L Bocca(2,4)
(1) University Hospital, Department of Pharmacology, Caen, France
(2) Caen University Research Team, ERI 27 Inserm - EA 3917, Caen,
France
(3) Caen University Hospital, Department of Pysiology, Caen, France
(4) Paris XI, University, Paris, France
Old people (OP) are drivers and may be hypnotic drug consumers.
Recently European community promoted research on psychotropic drugs
consumption to prevent traffic accidents. Usually the hypnotic tolerance
tests like driving ability are achieved with young healthy patients. We
tried to determine whether in OP - 55 to 65 - a single dose of a hypnotic
like Zolpidem (Zp), Zopiclone (Zc) or Flunitrazepam (FNZ) induce
residual effects versus placebo (PCB). Drugs were assayed with Gas and
Liquid Chromatograhy coupled to Mass Spectrometry to measure concentrations 10 and 16 h after drug intake. Methods: 16 OP. None consumed psychotropic medication. The night before a session, they took a
standard dose of either Zp (10 mg), Zc (7.5 mg), FLN (1 mg) or PCB at
23 h. The study was conducted double-blind, random order of treatments, each session being separated by >8 days. Results: 6/16 subjects
had residual effects and increased trough levels of Zc or Zp until midday.
Particularly 11/16 Zp had significant concentrations >15 mg/L as threshold. mean ± SEM at 9 h and 15 h were respectively 69 ± 17 and
41 ± 10 mg/L. The mean terminal half life was 4.0 h. Conclusion and
discussion: the incidence of high residual concentration of Zp is in contrast with our previous findings in young subjects (Bocca &al., Psychopharmacology 1999; 143: 373-9).The Zp residual effects were related to
pharmacokinetic profile. This suggests revisions of Zp dosage for people
over 55.
Paper No.: 1533
FOCUSED CONFERENCE GROUP: P10 - DRUGS FOR HALF
THE WORLD: PAEDIATRIC CLINICAL PHARMACOLOGY
DIAGNOSIS AND TREATMENT OF SEIZURES IN THE
NEONATAL INTENSIVE CARE UNIT
N Cosic-Cerovac(1), Milca Prostran-Veselinovic(2), N Jovic(1),
R Stojanovic(2), Z Stojanovic(2)
(1) Belgrade University School of Medicine, Department for Neurology
and Psychiatr yfor Children & Youth, Belgrade, Republic of Serbia
(2) Belgrade University School of Medicine, Department of Clinical
Pharmacology, Pharmacology and Toxicology, Belgrade, Republic of
Serbia
Neonatal seizures are heterogeneous in origin and are the most common
neurological emergency in newborn infants. We studied a group of 22
infants with clinically observed seizures. Gestation ranged from 34 to
41 weeks and time of first seizure ranged from 2 hours to 25 days of
age. Initial investigations included results of tests for blood sugar and
serum electrolytes, blood cultures, neurological examinations, and appropriate specialized studies (EEG and ultrasongraphy of infant brain). Subtle (70%) and multifocal clonic seizures (15%) were the most common
types of seizures. Status epilepticus was observed in 1 patient. Etiology
was determined in 92% of infants and the commonet was asphyxia. Interictal EEG discharges were noted in 25% of infants. AED treatment was
necessary in 15% of infants (phenobarbital, phenytoin and diazepam).
Continuous AED therapy was instituted in 10%. real time ultrasonography and EEG are very reliable methods of diagnosing these conditions
and should be part of the routine screening of all infants with seizures.
Paper No.: 2729
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
EFFECT OF CROTOXIN, THE MAIN TOXIN OF THE
RATTLESNAKE C.D.TERRIFICUS VENOM, ON SECRETORY
ACTIVITY OF PERITONEAL MACROPHAGES DURING
TUMOR PROGRESSION
ES Costa, AMSC Francisco, OJ Faiad, Y Cury, Sandra C Sampaio
Butantan Institute, Laboratory of Pathophysiology, São Paulo, Brazil
Crotoxin (CTX) inhibits tumor growth and modulates the function of
macrophages. Despite these evidence, the contributiuon of macrophage
inhibition to the decrease in tumor growth, caused by CTX, was not
determined yet. Macrophages provide a defense mechanism against
tumor cells and two distinct polarization states, M1 and M2, have been
described for these cells. In the beginning of tumor progression, M1
macrophages release reactive nitrogen/oxygen intermediates and the
cytokines TNF-a, IL-1b and IL-6. In contrast, during tumor development, the release of these mediators by tumor-associated macrophages
(M2 cells) is inhibited, contributing to tumor development. In the present
study, the effect of CTX on the activity (H2O2, nitric oxide-NO and cytokines release) of macrophages obtained from peritoneal cavity of Walker
256 tumor-bearing rats, was investigated. Male Wistar rats were inoculated (2x107, s.c.) with tumor cells and treated with CTX (18lg/300ll/
rat, s.c.) on day 1 (M1 macrophage) or 5 (M2 macrophage) after cell
injection. Macrophages were obtained on day 14 after cell inoculation.
CTX (day 1 and 5, respectively) stimulates H2O2 (149% and 162%) and
NO (77% and 94%) production and the release of the cytokines IL-1 b
(day 1: 71%) and TNF-a (day 5: 1.59 fold). These results indicate that
CTX modifies the secretory activity of M2 cells, which may contribute
to the inhibitory action of the toxin on tumor growth. These data reinforce the actions of CTX on defence mechanisms and bring new perspectives for the development of a new substance with therapeutic properties.
Supported by FAPESP(08/53840-8,09/52330-9), CNPq/PIBIC, INCTTOX.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
243
Paper No.: 3278
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
POSSIBLE ROLE OF THE ENDOCANNABINOID SYSTEM IN
THE ANALGESIC EFFECT OF PARACETAMOL-IBUPROFEN
ASSOCIATION - AN EXPERIMENTAL STUDY
Mihnea Costescu, H Paunescu, OA Coman, I Fulga
University Carol Davila School of Medicine and Pharmacy, Department
of Pharmacology and Pharmacotherapy, Bucharest, Romania
Aim: Evaluation of a potentiation of the analgesic effect between paracetamol and ibuprofen and implication of endocannabinod system. Materials and methods: 10 male albino mice groups were used. The
experiments were supervised by an institutional ethic board. The tested
substances were: paracetamol orally administered; ibuprofen and AM281
(CB1 receptor antagonist) intraperitoneally administered. Two analgesia
test were used: acetic acid 0,75%(v/v) writhing test, using as parameter
the antinociceptive percent; hot plate test at 55C, parameters used - time
until paw liking and until jump. Statistic analysis used ANOVA or nonparametric tests. Results: In writhing test a dose-analgesic effect relationship was obtained, with ED50 = 400 mg/kg bw for paracetamol and
75 mg/kg bw for ibuprofen. Using different proportion of these two substances the analgesic effect was additive. Association of 80 mg/kg bw
for paracetamol and 60 mg/kg bw for ibuprofen with AM281 decrease
the association effect. In hot plate test doses up to 1000 mg/kg bw of
paracetamol and up to 240 mg/kg bw ibuprofen had no analgesic effects,
the association of 600 mg/kg bw paracetamol and 30 mg/kg bw ibuprofen was analgesic and this effect was antagonized by AM281. Conclusions: Paracetamol and ibuprofen had an additive analgesic effect in
writhing test and reciprocal potentiation in hot plate test. Adding AM281
to the association paracetamol-ibuprofen decrease the analgesic effect in
the writhing test and eliminated the effect in the hot plate test. Influencing endocannabinoid system might be responsible for a part of analgesic
effect of paracetamol-ibuprofen associations. Key words: ibuprofen, paracetamol, endocannabinoids, analgesia.
Paper No.: 1989
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
OXALIS PES-CAPRAE INHIBIT NA
CONCENTRATION-RESPONSE CURVES IN HUMAN
INTERNAL MAMMARY ARTERY
Maria Cotrim(1), M Campos(1), M Gaspar(1), D Fonseca(1),
C Frigerio(1), N Gomes(1), M Antunes(2)
(1) University of Coimbra Faculty of Pharmacy, Center for Pharmaceutical Studies(CEF), Laboratory of Pharmacology, Coimbra, Portugal
(2) University Hospital of Coimbra (HUC), Department of Cardiothoracic Surgery, Coimbra, Portugal
Introduction: After a meta-analysis of the published papers in natural
sources for neurological bioactivity we verified that many are the natural
sources and the structures that can be utilised on this purpose (Gomes et
al., 2009). Duarte et al. (1993) found that Oxalis pes-caprae L. relax the
contractions induced by NA, KCl or phorbol 12-myristate-13-acetate in
rat aorta strips. Abdalla et al. (1994) reported a concentration-dependent
relaxation of the tone of ileum, epinephrine- precontracted pulmonary
artery and mild relaxation of acetylcholine- precontracted trachea for this
compound. We have study the effects of Oxalis pes-caprea leaves extract
on NA concentration-response curves in human internal mammary artery.
Material and methods: The flavonoid/phenolic acid profile was carried
out as been proposed by Campos (1997). The structural identifications
were made according to Campos & Markham (2007). Vascular rings of
the human internal mammary artery were cut into four segments. After
an equilibration period of 2 hours at a resting tension of 1.5 g, changes
in isometric tension were measured using the PowerLab data acquisition
package. A volume of 200 ll of the 0,2 mg/ml Oxalis pes-caprae L.
leaves extract, was used to test the effect on the NA concentrationresponse curves (10-7-10-4M) in human internal mammary artery.
Results: Oxalis pes-caprae L. inhibited NA concentration-response
curves on human internal mammary artery.
Results: Oxalis pes-caprae L. inhibited NA concentration-response
curves on human internal mammary artery.
Abdalla et al., 1994, Phytoth. Res., 8: 265-70; Campos M.G., Mitchel
K., Cunha A., Markham K., 1997, Phytochem. Anal., 8, 181-185; Campos M.G. and Markham K. R., Ed. Imprensa da Universidade de Coimbra, Portugal 2007; Duarte et al., 1993, Rev. Ciênc. Bioméd., São Paulo,
14:89-96 Gomes et al., 2009, Prog. Neuro-Psychopharmacol. & Biol.Psych, 33, 1372-89
Paper No.: 2404
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
BEHAVIORAL EFFECTS OF 5’-GNTI AND 6’-GNTI IN MICE
Alan Cowan(1), S Inan(1)
Temple University School of Medicine, Department of Pharmacology,
Philadelphia, PA, USA
We are comparing the in vivo pharmacology of 5’-GNTI (5’-guanidinonaltrindole), a selective j opioid receptor antagonist, and its 6’-regioisomer, commonly described as a selective agonist for the j-d receptor
heteromer. 5’-GNTI (0.03-3 mg/kg) precipitated frenzied scratching after
s.c. injection behind the neck of male, S.W. mice (25-30 g; n = 8-10), as
did 6’-GNTI (0.30-30 mg/kg). The A50 values were 0.17 and 2.28 mg/
kg, respectively. The nature of the spectacular behavioral arousal was
studied in the hexobarbital sleeping time test. Three groups of mice
(n = 9-10) received hexobarbital (100 mg/kg, i.p.), followed 3 min later
by s.c. saline, 5’-GNTI (0.30 mg/kg) or 6’-GNTI (4 mg/kg). The three
mean latencies (min) to recovery of the righting reflex (71 + /- 10,
s.e.m., 69 + /- 12 and 64 + /- 10, respectively) were not significantly different after Kruskal-Wallis nonparametric ranking analysis. We conclude
that, despite 6’-GNTI being heralded as a tissue-selective analgesic with
potentially reduced side effects, it provokes the same compulsive scratching as does 5’-GNTI, and this persistent behavior may be viewed either
positively as a useful preclinical model for drug discovery or negatively
as a portent of ultimate dermatological woe.
Paper No.: 3307
FOCUSED CONFERENCE GROUP: P02 - TRANSMEMBRANE
TRANSPORT: PERSPECTIVES FOR DISEASE AND DRUG
DISCOVERY
THE ROLE OF P-GLYCOPROTEIN IN BACTERIAL
ATTACHMENT TO INTESTINAL CELLS
Andrew Crowe(1,2)
(1) Curtin University, School of Pharmacy, Perth, WA, Australia
(2) Curtin Health Innovation Research Instutute, Perth, WA, Australia
This study investigated the effect of changes to P-glycoprotein function
and expression on bacterial attachment to Caco2 and RKO gastrointestinal cells using 6 species of bacteria (E. coli, Staph. aureus, Salmonella
typhimurium, Klebsiella aerogenes, Clostridium sporogenes or Pseudomonas aeruginosa). The RKO cell line was chosen to provide a cell line
with minimal P-gp expression. Following incubation of gastrointestinal
cells with the P-gp inhibitors, bacteria incubated with a stable fluorescent
dye (BacLight Green) were added and incubated at 37C for various
times between 30 and 240 min. Fluorescence intensity corrected for
background was used to compare bacterial attachment to these cell lines
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
244
with either the P-gp inhibited, induced (using rifampicin) or normal
expression. It was found that P-gp inhibition resulted in a significant
increase of all bacterial attachment to Caco2 cells. PSC-833 incubation
resulted in Pseudomonas attachment increasing by 65%, while E. coli
resulted in a doubling of attached bacteria. Salmonella, Klebsiella, Staphlococcus, and Clostridium had up to a 3 fold increased attachment in
Caco-2 cells. RKO cells did not alter their bacterial attachment with
PSC-833. Western blotting confirmed the presence of P-gp in Caco-2
cells, and absence in RKO cells. In addition, rifampicin, a P-gp inducer,
resulted in a reduction in Salmonella and Klebsiella attachment of up to
50%. These results suggest P-gp expression may contribute to the resistance of potential bacterial toxicity in the gut, by preventing them accumulating, which in turn reduces the risk of gastrointestinal disorders.
Paper No.: 1534
FOCUSED CONFERENCE GROUP: P03 - ION CHANNELS IN
ANALGESIA & ANAESTHESIA
POTENTIATING EFFECT OF DILTIAZEM ON PENTOBARBITAL HYPNOSIS MAY BE RELATED TO THE 5-HT1A AND
5-HT2A/2C RECEPTORS: THE DRN, TMN AND VLPO AS KEY
ELEMENTS IN THE PATHWAY
Su-Ying Cui, Y-H Zhang, X Zhao, X-Q Zhang
Peking University, School of Basic Medical Science, Department of
Pharmacology, Beijing, PR China
Our previous researches indicated that the augmentative effect of diltiazem on p entobarbital sleep may be mediated by the serotonergic system, and the VLPO-TMN neuronal circuit may play a key role. This
study was undertaken to investigate t he role of dorsal raphe nucleus
(DRN) in rats and two subtypes of serotonergic r eceptors (5-HT1A and
5-HT2A/2C) on the augmentative effect of diltiazem on pento barbital
sleep in mice and rats. The results showed that diltiazem obviously pro
longed the duration of pentobarbital-induced sleep with significant
increase in NREM sleep time including light sleep time and deep sleep
time. These effects we re significantly inhibited by the 5-HT1A agonist
8-OH-DPAT and 5-HT2A/2C agonist DOI and potentiated by atagonists
both of 5-HT1A receptor (p-MPPI) and 5-HT2A/ 2C receptor (ritanserin). Perfusion of diltiazem into DRN also dose-dependently increased
non-rapid eye movement (NREM) sleep including light sleep (LS) and
SWS. On the other hand, diltiazem significantly increased c-Fos expression which is suppressed by pentobarbital in rat DRN. From these
results, it should be presum ed that the augmentative effect of diltiazem
on pentobarbital-induced sleep may be influenced by 5-HT1A and 5HT2A/2C receptors and the DRN-VLPO-TMN neuronal ci rcuit may
play a key role.
Acknowledgement: This work was funded by grants from the National
Natural Science Foundation of China (no. 30640070 and 30772556) and
the 985 project from Peking University.
Paper No.: 3193
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
SELECTIVE MODULATION BY SINGLET OXYGEN OF
CCK1R AND OTHER G PROTEIN-COUPLED RECEPTORS IN
LIVE CELLS
Zong Jie Cui, Y Wang, TT Li, SF Zhang, B Nashun, HY Liang, ZQ Han
Beijing Normal University, Institute of Cell Biology, Beijing, PR China
respiratory burst. We have previously found that plasma membranelocalized singlet oxygen produced by photodynamic action permanently
activates CCK1 receptors in rat pancreatic acini. Materials: In the present work we used isolated rat pancreatic acinar cells, hepatocytes, cultured AR4-2J cells, to examine triggering by plasma membranelocalized singlet oxygen after brief photosensitization of sulphonated
aluminium phthalocyanine, of calcium oscillations in Fura-2-loaded
cells. Results: It was found that permanent calcium oscillations were
induced in pancreatic acinar cells, due to activation of CCK1 but not
M3 receptors. In hepatocytes singlet oxygen desensitized a1 adrenergic
and V1a vasopressin receptors. In AR4-2J cells, photodynamicallygenerated singlet oxygen activated CCK1 but desensitized NK1 receptors. Effects of membrane impermeant methionine / cysteine oxidant
chloramine T and cysteine oxidants PCMB, MTSEA, DTNB were
investigated. It was found that methionine / cysteine oxidation produced
reversible calcium oscillations via CCK1 receptor activation in pancreatic acinar cells. In AR4-2J cells, oxidation of extracellularly accessible
methionine residues induced permanent calcium oscillations due to
CCK1 receptor activation. In a mouse model of acute pancreatitis, pancreatic acini isolated during the initial but not later stages of pancreatitis
showed spontaneous but persistent calcium oscillations blockable by
CCK1 receptor antagonist FK480. Conclusion: Such data indicate that
oxidation of extracellular methionine residues in CCK1 receptor results
in its permanent activation, reversed only by methionine sulfoxide
reductase (MSR). This has important implications for GPCR modulation
in pancreatitis and other inflammatory diseases.
Paper No.: 2967
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
EPAC PROTEINS AS A DETERMINANT FOR
FORSKOLIN-INDUCED VASORELAXANT EFFECT AND
CAMP-INHIBITION OF STORE OPERATED CA2 + ENTRY IN
VASCULAR MYOCYTES
Andrea Cuı́ñas, J Elies, M Campos-Toimil
University of Santiago de Compostela, Department of Pharmacology,
Santiago de Compostela, Spain
The mechanism of the cAMP-induced vasorelaxation is poorly understood, although it may be due partially to interferences in the regulation
of cytosolic Ca2 + concentration ([Ca2 + ]i). The discovery of the cAMP
target Epac (exchange proteins directly activated by cAMP), have
explained previously unrecognized roles for cAMP that are independent
of its traditional effector cAMP-dependent protein kinase (PKA). We
have studied the effects of forskolin, an adenylylcyclase inhibitor, and
the role of the two main cAMP effectors in isolated endotheliumdenuded rat aorta. In rat aortic smooth muscle cells (RASMC), we have
measured [Ca2 + ]i with fura-2 and intracellular cAMP levels by fluorescence resonance energy transfer (FRET) using epac-cAMPs dyes. Phenylephrine-induced contractions were concentration-dependently inhibited
by forskolin, both in the presence or in the absence of extra cellular
Ca2 + . Forskolin also reduced the contractions elicited by reintroduction
of external Ca2 + after a phenylephrine-induced contraction in a Ca2 + free solution. Forskolin effects were partially reduced by PKA inhibitors (KT5720, Rp-cAMPs) and they were reproduced by the Epac activator 8-pCPT-2’-O-Me-cAMP. In RASMC, forskolin dose-dependently
inhibited increases in [Ca2 + ]i induced by arginine-vasopressin(AVP)
and the store-operated Ca2 + entry (SOCE) activated after depletion of
intracellular Ca2 + stores with AVP. FRET experiments shown that the
effects of forskolin on [Ca2 + ]i correlate well with the increases of
cAMP levels induced by this agent. In conclusion, forskolin-induced
increases of cAMP in RASMC inhibit intracellular Ca2 + release and
the subsequent transmembrane SOCE. The effects of cAMP are mediated by activation of Epac in these cells, although a role of PKA cannot be excluded.
Introduction: Singlet oxygen is produced by photodynamic action after
UVA irradiation of endogenous photosensitizers, or during neutrophil
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
245
Paper No.: 2849
FOCUSED CONFERENCE GROUP:
P04 - PHARMACOEPIDEMIOLOGY, CURRENT
CONTROVERSIES AND OPPORTUNITIES
ADHERENCE TO TREATMENT: FEASIBILITY OF A SPECIFIC
SELF-REPORTED QUESTIONNAIRE TO ASSESS
MEDICATION TAKING BEHAVIOUR
Josip Culig(1), M Leppee(1), J Boskovic(2)
(1) Andrija Stampar Institute of Public Health, Department of Pharmacoepidemiology, Zagreb, Croatia
(2) Pharmacy Simac, Zagreb, Croatia
Non-adherence to medication behaviour is observed in all situations
where self- administration is required. There is strong evidence that
many patients with chronic illnesses have difficulty adhering to their
treatment regimens. These results show poor management and control of
the illness, which reduces the patient’s quality of life and wastes health
care resources. Adherence is simultaneously influenced by various factors. The adherence assessment was done among the patients with
chronic diseases in the city of Zagreb. They were interviewed in different
community pharmacies by standardized self-reported questionnaires
offered by pharmacist. The questions concerned personal data, history,
health status, relationships with doctors and pharmacist, medication taking behaviour. According to 635 collected questionnaires there were
1357 diagnosed chronic diseases (roughly 2 per patient). The leading disease was essential hypertension (361, as first diagnosis 239), followed by
dislipidaemia (125, only 15 as first) and diabetes mellitus (120, as first
69).The non-adherent behaviour has been found in 58, 3% of patients.
As the main reason emerged forgetfulness (60%), followed by being
away from home (45, 4%), and running out of pills (44, 4%). The dosing
schedule was a problem for 40, 9% of the patients and polypharmacy for
39, 5%. Frequent changes of prescribed therapy were blamed by 26% of
patients for non-adherent behaviour (in hypertensives slightly higher 31,
9%) and avoiding side-effects were reason in 29, 6% (31% in hypertensives). The patient’s understanding and perception of the disease itself
might improve adherence behaviour significantly. The intervention strategies to improve the patient-health care provider relationship is crucial.
Paper No.: 2066
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD DISEASES
HYDROGEN SULFIDE IMPROVES NEUTROPHIL MIGRATION
AND SURIVAL IN SEPSIS VIA K+ATP CHANNEL ACTIVATION
Fernando Cunha, F Spiller, M Orrico, F Souto, J Alves-Filho,
S Ferreira, J Hothersall, A Freitas
University of São Paulo, School of Medicine Ribeirao Preto, Department
of Pharmacology, São Paulo, Brazil
The recovering of neutrophil migration to infectious focus improves the
survival in severe sepsis. Recently, we demonstrated that the cystathionine c-lyase (CSE)/hydrogen sulfide (H2S) pathway increases the neutrophil recruitment to inflammatory focus during acute localized
inflammation. The present study evaluates if H2S administration
increases the neutrophil migration to infection focus and survival of mice
with sepsis induced by cecal ligation and puncture (CLP). It was
observed that the pre-treatments of mice with H2S donors (NaHS or
Lawesson’s reagent) improved leukocyte rolling/adhesion in mesenteric
microcirculation as well as the neutrophil migration into the infection
focus. Consequently, bacteremia levels were reduced, hypotension was
prevented and the survival rate enhanced from ~13% to ~80%. Notably,
even when treatment was delayed (6 h post-CLP), a highly significant
reduction in mortality as compared to untreated mice was still observed.
Moreover, H2S-pretreatment prevented the down-regulation of CXCR2
and L-selectin and the up-regulation of CD11b and GRK2 in neutrophils
during sepsis. H2S also prevented the reduction of ICAM-1 expression
on endothelium of the mesenteric microcirculation in severe sepsis.
Confirming the critical role of H2S on sepsis outcome, pretreatment with
dl-propargylglycine (a CSE inhibitor) inhibited neutrophil migration to
infectious focus and induced high mortality in mice subjected to nonsevere sepsis (from 0% to ~80%). Finally, the beneficial effects of H2S
were blocked by glibenclamide (a K+ATP channel blocker). These results
showed that H2S restores the neutrophil migration to infectious focus
and improves survival outcome in severe sepsis by a K+ATP channel
dependent mechanism.
Paper No.: 2727
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
OPIOID RECEPTOR ACTIVATION AND EXPRESSION IS
REGULATED BY PERIPHERAL INJURY IN BOTH DORSAL
ROOT GANGLIA (DRG) AND NERVE PAW (NP) OF RATS
Yara Cury(1), V Zambelli(1), AC Fernandes(1), V Gutierrez(1),
C Parada(2)
(1) Butantan Institute, Laboratory of Pathophysiology, São Paulo, Brazil
(2) UNICAMP, Department of Physiology and Biophysics, Campinas,
São Paulo, Brazil
Previous data demonstrated that, in rats, prostaglandin E2- PGE2 and
chronic constriction injury-CCI of sciatic nerve increase peripheral analgesic efficacy of opioid drugs and also of crotalphine, a novel peptide
that induces analgesia mediated by activation of j- and d-opioid receptors. The aim of this study is to characterize the mechanisms involved in
the increase of the efficacy of CRP caused by tissue injury. For this purpose, the effect of PGE2 (intraplantar route) and CCI on opioid receptor
expression/activation in DRG and NP of rats was evaluated by immunoblotting and ELISA assays. In NP, PGE2 increases the expression of land j-opioid receptors (43 and 71%, respectively) and decreases (30%)
the expression of d-opioid receptors. l- and j-opioid receptor expression
is also increased in the DRG (79 and 168%, respectively). CCI up-regulates l-opioid receptors in NP (27%) and DRG (49%) and d-opioid
receptors in DRG (35%). In contrast, j-opioid receptors are down-regulated by CCI (51% in NP and 21% in DRG). Despite increasing the
expression of opioid receptors, PGE2, per se, did not alter the conformation of opioid receptors. CRP (0.0006lg/paw) or DAMGO (5lg/paw, lopioid agonist) activate, respectively, j- and l-opioid receptors, per se
(23 and 16%, respectively) or in the presence of PGE2 (15 and 50%,
respectively). These results indicate that peripheral opioid receptor
expression and activation are distinctly regulated by the presence of acute
or chronic injury and provide evidence regarding the effectiveness of
peripheral opioids in both acute and chronic pain.
Support: FAPESP (07/03404-4), INCTTOX Program, CNPq
Paper No.: 960
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
PD98059, A SPECIFIC MITOGEN-ACTIVATED PROTEIN
KINASE INHIBITOR, ATTENUATES THE DEVELOPMENT OF
BLEOMYCIN-INDUCED LUNG INJURY
Salvatore Cuzzocrea(1,2), M Galuppo(1), E Mazzon(1), E Esposito(1,2),
R Di Paola(1), T Genovese(1), P Bramanti(2)
(1) University of Messina School of Medicine, Department of Clinical &
Experimental Medicine & Pharmacology, Messina, Italy of Medicine,
University of Messina, Italy
(2) IRCCS Centro Neurolesi ‘Bonino-Pulejo’, Messina, Italy
Mitogen-activated protein kinase (MAPK) signalling pathways involve
two closely related MAPKs, known as extracellular signal-regulated
kinase (ERK)-1 and -2. The aim of the present study was to evaluate the
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
246
contribution of MAPK3/MAPK1 in the development of lung inflammation and fibrosis caused by bleomycin-administration (1 mg/kg) in mice.
To this purpose, we used 2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one (PD98059), which is an inhibitor of MAPK3/MAPK1. Mice
subjected to intra-tracheal administration of bleomycin developed significant lung injury characterized by marked neutrophil infiltration and tissue
oedema. An increase in immunoreactivity to nitrotyrosine, iNOS, TNF-a
and IL-1b was also observed in the lungs from bleomycin-treated mice.
PD98059 treatment (10 mg/kg, 10% DMSO, i.p.), shows therapeutic
effects on pulmonary damage, decreasing many inflammatory and apoptotic parameters: 1) cytokines production, 2) IkB-a degradation and NFjB nuclear traslocation 3) iNOS expression 4) nitrotyrosine and PARP
localization 5) the degree of apoptosis, evaluated as Bax and Bcl-2 balance, FAS ligand expression and TUNEL staining. Taken together, these
results clearly demonstrate treatment with the MEK1 inhibitor PD98059
reduces the development of lung injury and inflammation induced by
bleomycin in mice.
Paper No.: 2749
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
SEROTONIN IS INVOLVED IN AIRWAY HYPERREACTIVITY
AT BOVINE TRACHEAL SMOOTH MUSCLE
Darwin J Da Costa-Guevara(1), E Trejo(2), M Alfonzo(2)
(1) Central University of Venezuela Faculty of Pharmacy. Department of
Pharmacology, Caracas, Venezuela
(2) Central University of Venezuela Faculty of Medicine, Caracas,
Venezuel
Serotonin or 5-hydroxytryptamine (5-HT) has different effects in the
smooth muscle due to a great variety of receptors. The 5-HT is a common
regulator of the arterial pressure, gastrointestinal motility, etc. In addition,
serotonin role in the modulation of the smooth muscle tone of the upper
airways is a key factor of controversy in the pathophysiology of the bronchial hyperreactivity. In this sense, little is known about the receptors and
the mechanism that exactly are involved in bronchial hyperreactivity.
Some subtypes of 5-HT receptors are claimed to be associated with this
function are: 5-HT1, 5-HT3, 5-HT7 and more frequently 5-HT2 receptors.
Two pharmacological mechanisms have been proposed to explain the 5HT induced contraction of the upper airway smooth muscle: a) Acting
directly via binding to specific 5HT receptors localized on the smooth
muscle, or b) indirectly when 5HT facilitates the acetylcholine (ACh)
release from of neurons or other deposits by means of 5-HT receptors and
this release ACh contracts the airway smooth muscle. The aim of this
study was evaluated using isolated bovine trachea strips in organ bath the
effect of muscarinic inhibitors such as atropine (1lM) and 5-HT antagonists (1lM) on contraction of upper airway smooth muscle in the presence
of 5-HT. The atropine was capable of block the contractile effect of 5-HT
1lM in about 75% and the ritanserina almost completely. These result
suggest that the 5-HT2 receptors are important in this contractile effect of
the 5-HT and that the ACh participate in this action.
Paper No.: 2813
FOCUSED CONFERENCE GROUP: P11 - G PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR TO PHYSIOLOGICAL FUNCTION
LACK OF KININ RECEPTORS AFFECTED ACE EXPRESSION
IN MICE ABDOMINAL AORTA
Eliete da Silva Rodrigues, RP Martin, JB Pesquero, SI Shimuta
UNIFESP, Department of Biophysics, São Paulo, Brazil
Reduced responsiveness of abdominal aorta to bradykinin (BK), desArg9
BK and to angiotensin II (AngII) was described in mice deficient in kinin
B1 (B1KO) and B2 (B2KO) receptors. The expression levels of the kinin
receptors and of AngII AT1 receptor were low in these transgenic mice.
Then the role of angiotensin-converting enzyme (ACE), a link between
kallikrein-kinin and renin-angiotensin systems, was evaluated. Contractile responses to angiotensin I (AngI) and to BK were recorded in the
presence and absence of ACE inhibitor, lisinopril, in rings of aorta of the
transgenic animals. The efficacy of AngI induced responses was reduced
in B1KO and B2KO, suggesting that ACE activity was lower than in
WT mice. However BK-induced responses which were reduced in these
transgenic animals were potentiated in the presence of lisinopril, indicating that the enzyme activity was not inhibited. Noteworthy, AngI
induced contractions in B1KO was inhibited by lisinopril, whereas those
of B2KO were partially affected, suggesting that a role for ECA. The
finding that there was a significant reduction in the ACE expression level
in B1KO but not in B2KO mice is another fact to be considered. Our
present results suggest that the ACE inhibitor can potentiate BK-induced
effect by inducing cross talk between ACE and BK B2 receptor beyond
blocking BK inactivation. The finding that the efficacy of AngI was
decreased in both transgenic mice but lisinopril exerted partial inhibitory
effect on the B2KO reinforces our hypothesis that a regulation between
ACE, ACE inhibitor and kinin receptors may occur.
Paper No.: 1078
FOCUSED CONFERENCE GROUP: P19 - GENERAL
SESSION - ONCOLOGY
NOVEL IMIDAZO[1,2-A]PYRIDINES IN THE INDUCTION OF
COLONIC CELL DIFFERENTIATION AND APOPTOSIS
Nurit Dahan-Farkas(1), C De Koning(2), L Harmse(1), H Davids(1)
(1) University of the Witwatersrand, Department of Pharmacy and Pharmacology, Parktown, South Africa
(2) University of the Witwatersrand, Department of Chemistry, Parktown,
South Africa
Over the past few decades, a major research thrust has been directed
towards the development of anti-cancer agents through both empiric
screening and rational design of new compounds. These attempts are
made to reduce the severe adverse effects associated with existing cancer
chemotherapeutic agents, as well as to reduce the development of drug
resistance. The aim of the study was thus to screen novel imidazo[1,2a]pyridines for cytotoxic activity, as well as to determine the level of differentiation and the induction of apoptosis in the colonic carcinoma cell
lines, viz. HT-29 and Caco-2. All compounds were synthesised using
standard organic chemistry techniques. In vitro cytotoxicity of the synthetic compounds were tested against the HT-29 and Caco-2 cell lines. In
order to determine the cytotoxic effects of these compounds, the MTT
(3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) assay
was used. The measurement of cell differentiation was determined by the
alkaline phosphatase assay, and a colorimetric assay was used for the
determination of caspase activity. The percentages of apoptosis were
determined using the Annexin V-FITC apoptosis detection kit and cytochrome c levels were determined by Elisa. Compounds CL 3 and CL 6
showed significant cytotoxic activity (p < 0.05) against both cell lines,
with IC50 values ranging from 7.38-14.34 lM. The compounds did not
produce a significant reduction in white blood cell viability in comparison to camptothecin (p > 0.05). CL 3 induced the highest amount of
apoptosis. Maximal caspase 3 expression was observed between 412 hours after addition of the active compounds.
Paper No.: 941
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
A COMPARATIVE STUDY OF A-ADRENERGIC RECEPTOR
MEDIATED CA2 + SIGNALS AND CONTRACTION IN INTACT
HUMAN AND MOUSE VASCULAR SMOOTH MUSCLE
Jiazhen Minnie Dai(1,2), H Syyong(1), C van Breemen(1),
J Navarro-Dorado(2), S Redondo(2), M Alonso(3), T Tejerina(2)
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
247
(1) University of British Columbia, Child and Family Research Institute,
Department of Anesthesiology, Pharmacology and Therapeutics, Richmond. BC, Canada
(2) Universidad Complutense,Department of Pharmacology, School of
Medicine, Madrid, Spain
(3) Hospital Clinico San Carlos, Service of General Surgery. Madrid,
Spain
Paper No.: 1093
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
ACTIVATORS OF SMALL CONDUCTANCE
CALCIUM-ACTIVATED POTASSIUM CHANNELS
SELECTIVELY ENHANCE BRADYKININ NO-MEDIATED
VASODILATATION IN PORCINE RETINAL ARTERIOLES
In virtually all experimental animal studies on intact vascular smooth
muscle of a variety of blood vessels, physiological and pharmacological agonists initiate oscillatory fluctuations in intracellular Ca2 + to initiate and maintain vasoconstriction. In this study we compared
phenylephrine-mediated Ca2 + signals and contraction in native human
and mouse smooth muscle cells situated in the intact media of small
mesenteric arteries. In addition we employed electron microscopy to
examine the cytoplasmic distribution of the sarcoplasmic reticulum
(SR). Phenylephrine elicited tonic contractions in both vessel types,
asynchronous Ca2 + oscillations in the mouse mesenteric smooth muscle cells, but only single transient Ca2 + signals in the human mesenteric smooth muscle cells. While nifedipine inhibited 90% of the
phenylephrine-induced tonic contraction in mouse mesenteric arteries,
it only slightly attenuated those in human mesenteric arteries. The
remaining large nifedipine-resistant component in human mesenteric
arteries was abolished by the Rho-kinase blocker HA-1077. Significant
differences were also observed in the membrane ultrastructure of the
two types of arteries. Whereas the superficial SR was abundant in the
mouse vessels and many junctions were observed between the plasmamembrane (PM) and the SR, the smooth muscle of human mesenteric
arteries had far less peripheral SR and was almost devoid of PM-SR
junctions. As we had previously reported that the PM-SR junctions are
essential for maintenance of Ca2 + oscillations, we hypothesized that in
the relatively old human patients the change in Ca2 + signalling pattern
was due to impaired SR refilling via areas of close apposition between
the PM and SR.
Thomas Dalsgaard(1), C Kroigaard(1), M Misfeldt(2), T Bek(2),
U Simonsen(1)
Paper No.: 2207
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
COMPARATIVE STUDIES ON THE ANTIMUTAGENICITY
AND ANTIOXIDANT ACTIVITIES OF THE FRUITING BODY
AND MYCELIUM OF THAI GANODERMA LUCIDUM
Theera Dalodom, A Aramphongphan
Mahidol University, Faculty of Science, Department of Pharmacology,
Bangkok,Thailand
Ganoderma lucidum (GL, Lingzhi) has been suggested as a candidate
for prevention and treatment of different diseases, including cancer.
Thai Ganoderma lucidum (G2), which has been grown in Thailand as
part of the Royal Project since 1988 was studied for its safety and efficacy. The present study aimed at comparing the fruiting body (whole
fruiting) and mycelium of GL hot water extracts with regards to their
in vitro mutagenic and antimutagenic potential by bacterial reverse
mutation assay, antioxidant activity by free radical scavenging activity
(DPPH assay), and iron chelating activity. The results showed that both
GL extracts ranging from 0.15 to 3.0 mg/ml was neither mutagenic nor
antimutagenic in bacterial system. Higher antioxidant activity in term
of DPPH scavenging free radicals was found in fruiting body extract
when compared with the mycelium extract. Moreover, both GL extracts
showed a slight effect of chelating activity on Fe2 + . Significantly, the
antioxidant capacity correlated with the total phenolic content. While
no tested concentrations of GL extracts were toxic to TA 98 and
TA100 salmonella typhimurium, the highest non-mutagenic concentration was cytotoxic to lung carcinoma cell lines (A549) as determined
by Trypan blue assay. This is the first comparative study on the antimutagenic and antioxidant activities of Thai Ganoderma fruiting body
and mycelium extracts.
(1) Aarhus University, Department of Pharmacology, Aarhus, Denmark
(2) Aarhus University Hospital, Department of Ophthalmology, Aarhus,
Denmark
Introduction: The present study investigated whether the selective activator of small (SKCa) and intermediate (IKCa) conductance calcium-activated potassium channels, NS309 (6,7-dichloro-1H-indole-2,3-dione
3-oxime), or the selective activator of SKCa2 and SKCa3 channels, CyPPA (cyclohexyl-[2-(3,5-dimethyl-pyrazol-1-yl)-6-methyl-pyrimidin-4-yl]amine), enhanced endothelium-dependent vasodilatation in porcine retinal
arterioles. Methods: Localisation of SKCa3 and IKCa protein was examined by immunolabelling. Endothelial cell calcium was measured by fluorescence imaging with oregon green. For functional studies, arterioles
were mounted in microvascular myographs for isometric tension recordings and concentration-response experiments for bradykinin, NS309, and
CyPPA, were performed. Results: SKCa3 and IKCa protein was localised
in the endothelium. Bradykinin, but neither NS309 nor CyPPA increased
endothelial cell calcium. Pre-incubation with NS309 or CyPPA enhanced
bradykinin relaxation. This enhanced relaxation was abolished by blocking SKCa channels with apamin. In the presence of NS309 or CyPPA,
mainly inhibition of NO synthase with asymmetric dimethylarginine, but
also inhibition of cyclooxygenase with indomethacin, reduced bradykinin
relaxation. Bradykinin relaxation was completely abolished by inhibition
of NO synthase and cyclooxygenase together with a NO scavenger, oxyhaemoglobin. Conclusion: In porcine retinal arterioles, SKCa3 and IKCa
protein is localised to the vascular endothelium. Bradykinin increases
endothelial cell calcium followed by opening of SKCa and IKCa channels.
Without altering endothelial cell calcium, NS309 and CyPPA enhance the
NO-mediated bradykinin relaxation via activation of SKCa channels.
These results implicate that activating SKCa channels improves endothelium-dependent relaxation and make this channel a potential target for
treatments aimed at restoring retinal blood flow.
Paper No.: 1682
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
NITRITE AND NITROGLYCERIN PREFERENTIALLY DILATE
LARGE SYSTEMIC ARTERIES DURING HYPOXIA
Thomas Dalsgaard(1), C Kroigaard(1), A Fago(2), U Simonsen(1)
(1) Aarhus University, Department of Pharmacology, Aarhus, Denmark
(2) Aarhus University, Department of Biological Sciences, Aarhus, Denmark
Several mechanisms in the wall of large arteries have been proposed to
reduce nitrite to nitric oxide (NO) during hypoxia. This study investigated whether the vasodilating effect of nitrite and nitroglycerin (GTN)
is similar in large and small arteries. The results were compared to other
NO donors. Furthermore, the role of mitochondrial aldehyde dehydrogenase (ALDH2) in nitrite and GTN relaxation was investigated. Rat aorta,
small femoral, and mesenteric arteries (diameter (lm): 2044 8 n = 42, ±
8 n = 42, and ± 8 n = 24, respectively) were mounted in microvascular
myographs for isometric tension recordings and concentration-response
curves were performed for nitrite, GTN, NO solution, and S-nitrosogluthathione (GSNO), during normoxia (21% O2) and hypoxia (5% or 1%
O2). Nitrite, NO solution, GSNO, and GTN induced concentrationdependent relaxations in all arterial segments, but these relaxations were
more pronounced in aorta both during normoxia and hypoxia compared
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
248
to small systemic arteries. In aorta, nitrite and GTN relaxation was
increased during hypoxia compared to normoxia. During hypoxia,
ALDH2 inhibition with cyanamide, reduced nitrite and GTN relaxations
in aorta, whereas in small femoral arteries only GTN relaxation was
reduced. Guanylate cyclase inhibition with 1H-[1,2,4]oxadiazolo-[4,3a]quinoxalin-1-one (ODQ) reduced relaxations to nitrite, GTN and NO
in both aorta and small femoral arteries. These results suggest that nitrite
relaxation is more pronounced in large arteries compared with systemic
small arteries due to lower sensitivity to NO in small arteries. Moreover,
the effect of cyanamide in aorta during hypoxia suggests an involvement
of ALDH2 in the pronounced nitrite and GTN relaxation observed.
Paper No.: 2833
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
AN ENZYME-LINKED IMMUNOSORBENT ASSAY FOR
THERAPEUTIC DRUG MONITORING OF TRASTUZUMAB
François Darrouzain(1), I Teulon(2), T Ternant(3), D David(1),
D Degenne(4), H Watier(4), P Gilles(1)
(1) Tours University Hospital, Department of Pharmacology & Toxicology, Tours,France
(2) INSERM, Centre de Recherche en Cancérologie de Montpellier,
University ofMontpellier, Montpellier, France
(3) Tours University Hospital,Department ofObstetrics and Gynecology,
Tours, France
(4) Tours University Hospital, Department of Immunology, Tours,
France
Trastuzumab is a humanized recombinant monoclonal IgG1 antibody
directed against the extracellular domain of Human Epidermal Growth
Factor Receptor 2 (HER2) and used in the treatment of breast cancers
overexpressing HER2. Trastuzumab has a large inter-individual pharmacokinetic variability. An assay measuring trastuzumab serum concentrations in treated patients is therefore needed.Microtiter plates were
sensitised with three different proteins: a fusionprotein consisting in
recombinant HER2 extracellular domain linked to human immunoglobulin Fc (HER2-Fc) and two single chain variable fragments (SCFV) of an
anti trastuzumab antibody. After incubation with the sample, bound trastuzumab was detected using peroxidase-conjugated goat F(ab’)2 fragment
specific for human Fcc (HRP-anti hIgG). The analytical performances of
the three different methods were compared. The HER2-Fc method had
the best analytical performances and the larger standard calibrator range.
The intra-day precision for quality controls were (percent coefficients of
variation, CV%): 10.2%, 3.1% and 14.6% for 0.4 lg/mL, 3 lg/mL and
15 lg/mL, respectively. The corresponding bias measures (percent deviation) were +0.37%, -18.2% and -3.3%, respectively. The between-days
precision was 16.3%, 9.2% and 6.4% for for 0.4 lg/mL, 3 lg/mL and
15 lg/mL, respectively. The corresponding bias were 11.4%, -8.2% and
-2.6%, respectively. The limit of detection was 0.62 lg/mL. Lower and
upper limits of quantitation were 0.4 lg/mL and 15 lg/mL, respectively.
This method is rapid, accurate and reproducible, and can be used for
pharmacokinetic and therapeutic drug monitoring studies of trastuzumab.
Paper No.: 1193
FOCUSED CONFERENCE GROUP: P19 - GENERAL SESSION TOXICOLOGY
TOPOTECAN VITREOUS LEVELS AFTER PERIOCULAR OR
INTRAVITREAL DELIVERY IN HEALTHY ALBINO RABBITS:
AN ALTERNATIVE FOR ADVANCED RETINOBLASTOMA
CHEMOTHERAPY
Denisa Darsova(1), P Pochop(1), E Klapkova(2), D Kodetova(3), H Tesfaye(2), J Uhlik(4), L Vajner(4)
(1) Charles University 2nd Faculty of Medicine and Motol Hospital,
Department of Ophthalmology, Prague, Czech Republic
(2) Charles University 2nd Faculty of Medicine and Motol Hospital,
Department of Clinical Biochemistry and Pathobiochemistry, Prague,
Czech Republic
(3) Charles University 2nd Faculty of Medicine and Motol Hospital,
Department of Pathology and Molecular Medicine, Prague, Czech
Republic
(4) Charles University 2nd Faculty of Medicine and Motol Hospital,
Department of Histology and Embryology, Prague, Czech Republic
Background: Intravenous topotecan (TPT) was proved to be effective in
the treatment of advanced retinoblastoma. The local ocular TPT delivery
might result in higher intraocular chemotherapeutic levels eliminating
exposure to non-target tissues. Methods: Six albino rabbits received single
TPT dose of 2mg/1ml by periocular injection into right eyes (POI group).
Six albino rabbits (TII group) received a single transcorneal intravitreal
injection of TPT (1lg/0.1ml) into right eyes. Left eyes were treated with
the same volume of sterile water. Blood and vitreous samples were collected at 0,1,2,6,24,48,168 and 480 hours. Plasma and vitreous levels of
TPT were determined by HPLC. AUC was calculated using trapezoidal
rule. Clinical evidence of toxicity was classified into four grades according to the anatomical structure. Results: Mean AUC was higher for TII
group (780.365 lg/ml.h) than for POI group (mean: 640.3 lg/ml.h). Relatively high TPT concentration was measured in left eyes (mean: 1.86lg/
ml) and in plasma (mean: 0.79 lg/ml) in POI group. Two rabbits in POI
group developed grade 1 - periocular toxicity, two had grade 3 - ocular
toxicity and two grade 4 – systemic toxicity. In TII group, TPT vitreous
levels were observed until 480 hours. Five rabbits in TII group developed
grade 3 toxicity. Conclusion: Reached AUC in POI and TII groups was
higher than expected target AUC of 80-120ng/ml.h. TPT delivered periocularly developed grade 4, while TPT delivered intravitreally developed
grade 3 toxicity. Transcorneal intravitreal delivery of topotecan might be
promising in the treatment of advanced retinoblastoma.
Supported by the grant GAUK Nr. 63008.
Paper No.: 406
FOCUSED CONFERENCE GROUP: PW02 - SYMPOSIUM ON
ADVANCES IN GI PHARMACOLOGY
COULD VITAMIN K2 GUARD AGAINST EXPERIMENTALLY
INDUCED GASTRIC STRESS ULCERS IN RATS
Inas Darwish(1), A Mohamed(2), H Abdel Aziz(3)
(1) Alexandria University Faculty of Medicine, Department of Clinical
Pharmacology, Alexandria, Egypt
(2) Alexandria University Faculty of Medicine, Department of Physiology, Alexandria, Egypt
(3) Alexandria University Faculty of Medicine, Department of Medical
Biochemistry, Alexandria, Egypt
The present study was planned to evaluate the possible gastro protective
effect of vitamin K2 on water immersion plus restraint stress- induced
gastric ulcers in rats. It was carried out on 40 male albino rats that were
divided into 5 equal groups .Two of them served as control groups; the
first is normal and the second is subjected to stress ulcer induction but
not pretreated. Water immersion plus restraint stress (WIRS) was the
experimental model for gastric stress ulcer done in four groups. Pretreatment with vitamin K2 in a dose of 15mg/kg/day orally for 7days in the
third group was done. While in the fourth group with alendronate (ALD)
in an oral daily dose of 20mg/kg/day for 7 days, or combination of the
mentioned drugs in the same previous doses for 7 days in the fifth one.
Then they were subjected to WIRS-induced gastric ulcers. Ulcer index,
gastric tissue levels of myeloperoxidase(MPO)and serum levels of both
epidermal growth factor (EGF)and interleukin -10(IL-10) were assessed
in all groups. The present study revealed that Vitamin K2 induced significant reduction of the mean values of ulcer index and tissue levels of
MPO. Furthermore, there were significant increase in the levels of protective cytokines as IL-10 as well as to increase the levels of ulcer healing growth factor; EGF as compared to the control groups. Thus, vitamin
K2 could have a potential clinical benefit in preventing gastric ulceration
The Experimental protocol was approved by the Medical Ethical Committee of the Faculty of Medicine, Alexandria University August; 2009.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
249
Paper No.: 440
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
MENATETRONE AND NIGELLA SATIVA: A PROPHYLACTIC
ROLE IN CORTICOSTEROID INDUCED OSTEOPOROSIS IN
THE RAT
have proposed for the first time that endogenous hydrocortisone may
play a role in ischemic preconditioning phenomena.
Keywords: Hydrocortisone, Infarct size, Ischemia-reperfusion model,
Ischemic preconditioning, serum cortisole.
Suzan A Darwish, HM Khalifa, IE Darwish, EI Anwar
Paper No.: 892
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
EXPOSURE TO CONCENTRATED AMBIENT FINE
PARTICLES (PM2.5) OF URBAN SAO PAULO AIR POLLUTION
INDUCES ENDOTHELIAL DYSFUNCTION AND STRUCTURAL
CHANGES IN RAT PULMONARY ARTERIES
Alexandria University Faculty of Medicine, Department of Pharmacology, Alexandria, Egypt
The long term use of corticosteroids adversely causes osteoporosis and
consequently bone fracture. In the presented wok, osteoporosis was
induced by the SC injection of (methyl prednisolone acetate10 mg / kg
daily for 4 weeks) to male albino rats (CIOP) which served as our experimental model. The following drugs were tested: nigella sativa oil (NSO)
800 mg/ kg and menatetrenone 7.5 15, 30 mg / kg (vitamin K2). The
tested drugs were given with calcium (10mg/kg +vitamin D3(8IU/kg)
.The following parameters were assessed after 28days of treatment:
serum osteocalcin (OC), serum osteoprotegrin(OPG) serum total alkaline
phosphatase (ALP), 24 hr fasting urinary hydroxyproline (UHP). Histological examination of lumbar vertebrae was done at the end of the
experiment .Treatment with NSO, nearly demonstrated the classical
structure of the trabicular system. This was evidenced by a significant
increase in serum OC reduction of UHP increase in serum OPG and
reduction ALP The strong positive correlation between serum OC and
ALP reflected a perfect mineralization activity of NSO. Treatment with
menatetrenone (7.5mg / kg) demonstrated normalization of trabicular
architecture and bone markers. The combination of NSO (800 mg/kg)
and menatetrenone(7.5 mg/kg) caused a classical structure of the trabicular system. UHP and total serum ALP approached normal levels. Both
serum OPG and OC exceeded normal levels showing an excessive activity on bone formation. Our results suggest that nigella sativa oil and menatetrenone almost restore normal bone structure of CIOP in rats
especially if used in combination.
Paper No.: 490
FOCUSED CONFERENCE GROUP: P06 - THE HEART GONE
WRONG; STABILIZATION OF CARDIAC FUNCTION - BASIC
CARDIOPROTECTIVE EFFECTS OF EXOGENOUS AND
ENDOGENOUS HYDROCORTISONE IN THE RABBIT MODEL
OF ISCHEMIA-REPERFUSION
Ali Davarian, V Khori, O Shirinbeik Mohajer
Golestan Cardiovascular Research Centre, Faculty of Medicine, GolestanUniversity, Department of Pharmacology, Golestan, Iran
Reducing the infarct size in acute myocardial infarction is one of the
most important goals driving new drug research and development. During the last two decades, many clinical studies have found cardioprotective effects of corticosteroids, but their exact role in ischemic
preconditioning remains questionable. The aim of the present study was
to determine the protective effects of hydrocortisone sodium succinate on
myocardial preconditioning in rabbit hearts. Twenty-four male New Zealand rabbits were divided randomly & equally to four groups: 1) control,
2) Infarct, 3) Ischemic preconditioning (IP) and 4) Hydrocortisone
(HYD). The HYD group received 50mg/kg Hydrocortisone 45min before
major ischemia. Serum levels of cardiac troponin-T(cTNT) and cortisole
were measured before and after the protocols. Triphenyl-tetrazolium
chloride staining was used to determine the infarcted area. In the present
study, exogenous hydrocortisone decreased infarct size by 53% in comparison to the infarct group. Serum level of cortisole was increased in the
IP and HYD groups, and was significant in the HYD group (P < 0.01).
An increasing trend in cortisole level was associated with a decreasing
trend in infarct size and cTNT in the IP and HYD groups (p > 0.01). In
conclusion, we showed that hydrocortisone has cardioprotective effects
when injected before the onset of myocardial infarction. In addition, we
Ana Paula Davel, M Lemos, L Pastro, L Rossoni, P Saldiva
University of São Paulo, Department of Physiology and Biophysics, São
Paulo, Brazil
This study was design to investigate the effects of daily inhalation of
urban São Paulo PM2.5 in vascular reactivity and structure of pulmonary
arteries, the primary vascular target of inhaled air pollution. For this, rats
were daily exposed to 600lg/m3 concentrated PM2.5 from urban São
Paulo (n = 10) or filtered (n = 10) air, during 2 weeks, 5 hours/ day.
After exposure, rats were sacrificed by abdominal aorta exsanguination
and the lungs were removed. Extralobar pulmonary arteries were isolated
and cut into rings placed on organ bath containg Krebs-Henseleit solution (37C, pH=7.4). The maximal contraction of rings was tested with
KCl (75 mM). Concentration-response curves to acetylcholine, phenylephrine and sodium nitroprusside were obtained. Morphometric analysis
of pulmonary arterioles was performed, specifically targeting the bronchiole alveoli transition region. The results obtained demonstrated that pulmonary arterial rings of PM2.5 exposed animals showed significant
reduction of maximal relaxation induced by acetylcholine, whilst sodium
nitroprusside-induced relaxation was similar between exposed and control groups. Contractile responses of pulmonary arteries to KCl and phenylephrine did not modify after PM2.5 exposure. Morphometry of
intrapulmonary arteries showed that PM2.5 exposure promoted pulmonary arteriole narrowing, especially on the smaller arterioles. The present
results indicate that short exposure to urban levels of PM2.5 could
induce endothelial dysfunction and adverse structural alterations of pulmonary vascular territory.
Financial support: FAPESP.
Paper No.: 893
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
ROLE OF B2-ADRENOCEPTOR ON VASCULAR
INFLAMMATORY MEDIATORS AND OXIDATIVE STRESS
INDUCED BY ISOPROTERENOL TREATMENT
Ana Paula Davel, C Wenceslau, G Ceravolo, MH Carvalho, P Brum,
L Rossoni
University of São Paulo, Department of Physiology and Biophysics, São
Paulo,Brazil
Overactivation of b-adrenoceptors following chronic treatment with isoproterenol leads to hyperreactivity to phenylephrine, oxidative stress and
increases proinflammatory cytokines expression on rat aorta. The aim of
the present study was investigate the b-adrenoceptors subtype(s)
involved in these vascular effects of isoproterenol treatment. Male wildtype and b1- or b2-adrenoceptors knockout mice (b1-KO, b2-KO) were
daily treated with isoproterenol (15 lg/g, sc) or vehicle for 7 days. Vascular reactivity and biochemical experiments were conducted on thoracic
aortic rings. Isoproterenol treatment enhanced the vasoconstrictor
response to phenylephrine in aortas from wild-type and b1-KO mice.
L-NAME effect on phenylephrine-induced contraction was impairs and
superoxide dismutase improved vascular reactivity only in wild-type and
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
250
b1-KO mice treated with isoproterenol. Further, these vessels showed
enhanced fluorescence to oxidative products of dihydroethidine. Aforementioned effects of isoproterenol treatment were not observed in
b2-KO. Isoproterenol treatment increased aortic protein expression of interleukins-1b and -6, and p65 subunit of NF-jB in wild-type mice.
Moreover, inhibition of NF-jB activity with sodium salicylate normalised to control values the hyperreactivity to phenylephrine observed on
aortas from isoproterenol-treated wild-type mice. These proinflammatory
effects were abolished on aortas from b2-KO treated with isoproterenol.
Immunohistochemistry analysis reveals that isoproterenol treatment did
not modify aortic expression and distribution of b-adrenoceptors. The
results provides evidences suggesting a pivotal role of b2-adrenoceptor
subtype mediating oxidative stress and proinflammatory status associated
with the hyperreactivity to phenylephrine induced by persistent b-adrenergic stimulation.
Financial support: FAPESP and CNPq.
Paper No.: 1650
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
IMPACT OF AMBIENT EXTRACELLULAR CALCIUM ON
ANALYSIS AND INTERPRETATION OF AGONISM AND
ALLOSTERIC MODULATION OF THE EXTRACELLULAR
CALCIUM SENSING RECEPTOR (CASR)
(1) Federal University of Fluminense, Department of Physiology and
Pharmacology, Rio de Janeiro, Brazil
(2) Federal University of Rio de Janeiro, Rio de Janeiro, Brazil
Introduction: Leptin, an adipocyte-derived hormone, is an important regulator of energy metabolism with relevant vascular effects. Leptin may
increase blood pressure through enhanced sympathetic activity and it also
exerts direct depressor effects on vascular tone by stimulating endothelial
production of nitric oxide in a dose-dependent manner (Zanetti M et al,
Atherosclerosis 2004; 175: 253–259). The aim of the present work is to
study if neonatal leptin treatment develops vascular alterations. Methods:
Pups were divided into two groups. Leptin group, injected daily with leptin (8lg/100g sc) and Control group injected saline daily, for the first
10 days of lactation. The animal groups were studied at one, three and
five months of age. Rings (4mm long) from thoracic aorta artery were
used for assessing vascular reactivity. The maximal contraction of each
ring was determined by cumulative addition of phenylephrine (10-10 to
10-6 mol/l). Acetylcholine (10-9 to 10-5 mol/l) was added cumulatively
during a maximal contraction to phenylephrine. Results: No differences
were observed in phenylephrine dependent aorta contraction at the different ages studied. However, acetylcholine EC50 was statistically different
for Leptin group (1.36x10-7M) compared to Control group (6.53x107M) at 1 month of age. There was no difference at 3 and 5 months of
age. Discussion: The higher potency of acetylcholine in Leptin group at
1 month of age agrees with previous works that shows leptin decreasing
blood pressure via NO production. Possible adaptations can explain the
lost of this activity at the other ages studied.
Financial Support: FAPERJ, CNPq, CAPES, PROPPI/UFF.
Anna E Davey(1), K Leach(1), AD Conigrave(2), PM Sexton(1),
A Christopoulos(1)
(1) Monash University, Monash Institute of Pharmaceutical Sciences &
Department Pharmacology, Parkville, VIC, Australia
(2) University of Sydney, School of Molecular and Microbial
Biosciences, Sydney, Australia
The extracellular calcium sensing receptor (CaSR) is a family C G protein-coupled receptor (GPCR) that differs from most other GPCRs in that
physiological activation by its endogenous ligand, extracellular calcium,
occurs over a very narrow concentration range. Moreover, cell-based
assays used to study GPCRs usually need to be performed in the presence of different amounts of ambient calcium in the buffer media, introducing an additional layer of complexity into the analysis of CaSR
functional data. The current study compared different modes of analysis
of C/R data generated for the interaction between CaCl2 and the human
CaSR, stably expressed in a HEK293 TREx cell line. We find that a logarithmic parameterisation of the Hill equation that specifically incorporates the fixed concentration of ambient buffer calcium into the analysis
provides appropriate parameter estimates with minimal need for additional data transformations. Simulations also revealed that the effect of
increasing the concentration of ambient agonist is to decrease the apparent Hill slope and increase agonist potency. This effect on the Hill slope
was also noted experimentally (using intracellular calcium mobilisation
assay) in the presence of the positive allosteric modulator cinacalcet, suggesting that the mechanism underlying the phenomenon is potentiation
by the modulator of ambient as well as added calcium. Collectively,
these results provide new insights into some of the determinants of
experimentally derived CaSR C/R data, and suggest caution in the
(over)interpretation of mechanisms underlying allosteric modulator drug
action at this receptor.
Paper No.: 2698
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
CHANGES IN RATS VASCULAR REACTIVITY IN RESPONSE
TO NEONATAL INDUCED HYPERLEPTINAEMIA
Fernanda Carla Ferreira de Brito(1), NAV da Motta(1), EB Marques(1),
CBV Scaramello(1), LL da Silva(2)
Paper No.: 3387
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
VANCOMYCIN DOSING AND MONITORING IN CHILDREN:
COMPLIANCE WITH GUIDELINES IN A BELGIAN
TEACHING HOSPITAL
Pieter De Cock(1), E Haegeman(1), N Goerlandt(1),
P Vanhaesebrouck(2), V Stove(3), A Verstraete(3), P Schelstraete(4),
H Robays(1)
(1) Ghent University Hospital, Department of Pharmacy Department,
Ghent, Belgium
(2) Ghent University Hospital, Department,of Neonatology, Ghent, Belgium
(3) Ghent University Hospital, Department of Clinical Chemistry, Ghent,
Belgium
(4) Ghent University Hospital, Department of Pediatric Pulmonology and
Infectious Diseases, Ghent, Belgium
Introduction: the aim of this study was to evaluate the compliance of
vancomycin prescribing and therapeutic drug monitoring (TDM) in children, with updated hospital guidelines. Methods/patients: a retrospective,
observational study was conducted on 60 vancomycin therapies in 56
patients. Results: in 17 therapies (28.3%), a wrong dosing regimen was
initiated. From the 40 correctly initiated and evaluable dosing regimens,
37.5% resulted in a therapeutic, 32.5% in a subtherapeutic and 30% in a
potentially toxic trough level; 3 correctly initiated therapies were not
evaluable. Trough level monitoring was minimally needed a 138 times
from which 54.3% was monitored at the appropriate time, 18.1% too late
and 2.9% too soon; in 24.7% of cases no trough level was taken. Peak
level monitoring was inappropriate in 29 cases (100%). A dose adjustment was justified in 69 cases: 36.2% was done at the appropriate time,
29% too late (median of 3 doses, range : 1-22) and 34.8% was not performed at all. In 5 cases, the dose adjustment could not be justified from
the reported trough levels. 38 dose adjustments were evaluable: 50%
resulted in a therapeutic, 36.8% in a subtherapeutic and 13.2% in a
potentially toxic level; 7 dose adjustments were not evaluable. Conclusion: these results demonstrate the stringent need for thorough education
on guidelines to ensure efficacy and minimize toxicity of vancomycin
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
251
treatments. Moreover, this study highlights the value of TDM in therapy
optimisation, even when correct initiating dosing regimens are used.
Paper No.: 1962
FOCUSED CONFERENCE GROUP: PW02 - SYMPOSIUM ON
ADVANCES IN GI PHARMACOLOGY
CANNABIDIOL CONTROLS INTESTINAL INFLAMMATION
THROUGHT THE MODULATION OF ENTERIC GLIAL CELL
ACTIVATION
Daniele De Filippis(1), G Esposito(2), M Cipriano(1), C Scuderi(2),
J De Man(3), L Steardo(2), T Iuvone(1)
(1) University of Naples Federico II, Faculty of Pharmacy, Department
of Experimental Pharmacology, Naples, Italy
(2) University of Rome ‘La Sapienza’, ‘Vittorio Erspamer’ Faculty of
Pharmacy, Department of Human Physiology and Pharmacology, Rome,
Italy
(3) University of Antwerp, Faculty of Medicine, Division of Gastroenterology, Antwerp, Belgium
Introduction. Abnormalities of the enteric nervous system (ENS) are
essential elements in the pathogenesis of gastrointestinal disorders. During gut inflammation enteric glial cells (EGCs) proliferate and release
neurotrophines, growth factors and pro-inflammatory cytokines which, in
turn, amplify the immune response. The discovery of molecules able to
control ECG-activation by this way modulating inflammation is therapeutically interesting. Cannabidiol (CBD) represents a good candidate at
this attempt, since several evidence show that cannabinoids control neurogenic inflammation, both in CNS and ENS. Methods. Intestinal biopsies from patients suffering from ulcerative rectocolitis (URC) both in
active and remission phase, were stimulated with LPS plus INF-c for
24h in presence or absence of CBD. Moreover, mice with LPS-induced
acute intestinal inflammation were treated with CBD. Intestinal tissues
were processed for biochemical, histological and immunohistochemical
analysis. Results. Stimulation of URC biopsies with LPS+INF-c
increased glial cell activation and intestinal damage. CBD reduced
S100B and iNOS protein expression indicating an anti-inflammatory
effect of CBD. In in vivo experiments, a significant increase of S100B
protein in the intestine of mice treated with LPS was found. Moreover,
CBD pre-treatment reduced glial cell activation. Histological, biochemical and immunoistochemical analysis demonstrated that CBD decreased
mast cell and macrophage number in LPS-treated intestine. Finally, CBD
treatment of LPS-mice blunted the pro-inflammatory responses, as
revealed by the reduction of TNF-a expression. Conclusion. The presentr
results suggest that CBD, by modulating glial activation, was able to
control the inflammatory environment consequently protecting against
intestinal damage.
Paper No.: 1254
FOCUSED CONFERENCE GROUP:
P19 - GENERAL SESSION - ONCOLOGY
NEW HPLC-MASS SPECTROMETRY METHOD FOR THE
SIMULTANEOUS QUANTIFICATION IN HUMAN PLASMA OF
THE ANTILEUKEMIA DRUGS IMATINIB, DASATINIB, AND
NILOTINIB
Silvia De Francia(1), A D’Avolio(2), F De Martino(1), E Pirro(1),
F Piccione(1), S Racca(1), F Di Carlo(1)
(1) University of Turin, Department of Biological and Clinical Sciences,
Orbassano,Italy
(2) University of Turin, Department of Infectious Diseases, Turin, Italy
resistance may occur. Thus, two second-generation Bcr-Abl inhibitors,
dasatinib and nilotinib, active against most of imatinib resistant Bcr-Abl
mutants, have recently been tested in clinical trials. At present imatinib
pharmacokinetics has been well investigated, while only limited information on dasatinib and nilotinib are available. In support of current clinical
studies suggesting Bcr-Abl inhibitors use in sequential/simultaneous
administration, these knowledges should be achieved. Furthermore BcrAbl inhibitors plasma concentrations measurement has become an essential tool for CML patients management, useful to evaluate compliance,
potential drugs interactions, treatment efficacy, and drug-related adverse
events. Recently many laboratories reported the development of procedures for Bcr-Abl inhibitors quantification, but, no methods for their
simultaneous determination were available. Aim of our study has been
the development and validation of an original HPLC-mass spectrometry
method for simultaneous quantification of imatinib, dasatinib and nilotinib in human plasma. After a simple protein precipitation extraction,
drugs chromatographic separation was achieved on a C18 reverse-phase
column at 1ml/min flow rate. Validation results were all in concordance
with international guidelines (FDA, 2001): mean intra-day precision
4.3%; mean inter-day precision 11.4%; mean accuracy 1.5%; extraction
recovery 95-114%; mean regression coefficient 0.99. Method developed
then can be useful to achieve informations on dasatinib/nilotinib pharmacokinetics and due to its high extraction efficiency and to its good reproducibility it can be suitable to perform therapeutic drug monitoring in
clinical routine.
Paper No.: 2878
FOCUSED CONFERENCE GROUP: P10 - DRUGS FOR HALF
THE WORLD: PAEDIATRIC CLINICAL PHARMACOLOGY
THERAPEUTIC DRUG MONITORING: HYDROQUINIDINE
ADMINISTRATION IN A NEWBORN INFANT WITH SHORT
QT SYNDROME. A CASE REPORT
Silvia De Francia(1), E Pirro(1), E Banaudi(2), C Riggi(2),
F De Martino(1), F Piccione(1), F Di Carlo(1)
(1) University of Turin, Department of Biological and Clinical Sciences,
Orbassano,Turin, Italy
(2) O.I.R.M. - S. Anna Hospital, Department of Pediatric Cardiology,
Turin, Italy
The first-choice therapy in adult patients affected by short QT syndrome,
agenetic anomaly which may cause severe arrhythmias and death, is the
implantablecardioverter-defibrillator device (ICD). The most effective
pharmacologictherapy is hydroquinidine, indicated for patients who
refuse an ICD andchildren, even if there are very little data about pharmacological therapy inthis age range. On July 2005 cardiologists asked
the laboratory of clinicarmacology to measure hydroquinidine plasma
levels in a newborn with historyof short QT syndrome in her family and
recognized as being affected by amutation in the IKr encoding gene. The
ECG pattern at birth was consistent withthe diagnosis (QTc 310 msec).
Antiarrhythmic prophylaxis with oralhydroquinidine was started in the
first week of age. The plasma tests wereperformed by HPLC-UV. Separation was achieved with a C18 column. Mobile phasewas 0.1% triethylamine (pH 2.4) - acetonitrile (85:15) at constant flow rate of0.550 ml/
min and the eluate was monitored at 250 nm. Quinine was internalstandard. The starting hydroquinidine dose was 4 mg/kg TID, while some
monthslater dosage was increased up to 10 mg/kg TID in order to reach
a QTc interval >360 msec and basal hydroquinidine plasma levels > 0.6
ug/ml. It was observed astrong correlation between hydroquinidine
plasma levels and prolongation of QTinterval. An increase in drug biotransformation ability reached by liver duringthe first months of life is
probably responsible for the need of greater dosesof hydroquinidine for
maintaining therapeutic plasma levels.
Chronic myeloid leukemia (CML) is a myeloproliferative disorder. Current frontline therapy is imatinib, a Bcr-Abl kinase inhibitor. Although
most patients show excellent responses to imatinib treatment, clinical
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
252
Paper No. 2913
FOCUS GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
INFLUENCE OF PROLONGED-RELEASE ASPIRIN ON
PLATELET FUNCTION IN PATIENTS WITH CHRONIC
CORONARY DISEASE
Jose P de la Cortes(1), A Guerrero(1), E Rueda(2), J Muñoz-Marı́n(1),
JA Lopez-Villodres(1), JA Gonzalez-Correa(1)
(1) University of Malaga, School of Medicine, Department of Pharmacology, Malaga, Spain
(2) University Hospital ‘Virgen de la Victoria’, Service of Cardiology,
Malaga, Spain
Aim: to establish the influence on platelet function of prolonged-release
aspirin in patients with chronic coronary disease. Methods: clinical trial
parallel, double-blind, randomized study on the influence of two forms of
presentation of aspirin (immediate-release ASA, ASA-IR and prolongedrelease, ASA-PR), on the platelet function in patients with chronic coronary heart disease (Nordm; EudraCT: 2004-000398-76). Patients were
evaluated prior inclusion (V0) to 3 (V1), 6 (V2) and 12 (V3) months. We
collected clinical, analytical and biochemical parameters related to platelet function, endothelial and oxidative status. Statistical analysis was performed uni and bivariate (chi square and t-student). Results: included in
the study were 100 patients, 50 per treatment arm. Characteristics of the
population: 79% men, average age of 64.2 ± 10.1 years. Cardiovascular
disease: IAM (61%), coronary syndrome (57%), stable angina (45%).
CVRF: hypercholesterolemia (88%), hypertension (62%), diabetes
(28%), tobacco (13%), older than 70 years (30%), BMI> 30 (39%). Compliance was over 98%. The inhibition of platelet aggregation was similar
after taking ASA-IR and ASA-PR (1.41 vs. 0.28 ± . 1.99 ± 0.22 ohms V3-). The levels of thromboxane B2 were significantly higher in patients
who took ASA-PR (15.17 ± 2.03 vs. 30.90 ± 2.21 pg/mL), although adequate regarding the efficacy antiaggregant. The plasma levels prostacyclin
were significantly higher in patients who took ASA-PR (1.34 ± 0.08 vs.
0.63 ± 0.06 pg/mL). For none of the above parameters were observed
significant changes over time. Conclusions: The antiaggregant effect of
aspirin prolonged-release is similar to aspirin immediate-release, but
showed less activity on the synthesis of prostanoids.
Paper No.: 2061
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
HYDRO-ALCOHOLIC EXTRACT OF AÇAÍ (AN AMAZON
PLANT) PREVENTS ENDOTHELIAL DYSFUNCTION,
OXIDATIVE STRESS AND DEVELOPMENT OF
RENOVASCULAR HYPERTENSION
Miguel de Lemos Neto, CA Costa, RS de Moura, LCRM Carvalho,
PRB Oliveira, PJC Sousa, AC Resende
Rio de Janeiro State University, Department of Pharmacology. Rio de
Janeiro, Brazil
Hydro-alcoholic extract of açaı́ (an Amazon plant) prevents endothelial
dysfunction, oxidative stress and development of renovascular hypertension. Lemos M; Costa CA; Soares de Moura R; Carvalho LCRM; Oliveira PRB; PJC Sousa; Resende AC. UERJ. Polyphenols possess
antihypertensive, antioxidant and vasodilator activities. The aim of this
study was to investigate the protective effects of açaı́ stone extract (ASE,
200 mg/Kg/day) rich in polyphenols in 2-kidney, 1-clip (2K-1C) renovascular hypertension that is associated with an endothelial dysfunction
and oxidative stress. Young male Wistar rats used to obtain 2K-1C and
control rats (2K) received daily treatment with vehicle or ASE for
40 days, and systolic blood pressure (SBP, mm Hg) was measured by
plethysmography. The vasodilator effect of acetylcholine (ACh, 1-100
pmol) was studied in mesenteric arterial bed. Determination of oxidative
damage was estimated by formation of thiobarbituric acid reaction sub-
stances (TBARS) in mesenteric arteries and superoxide dismutase
(SOD), catalase (CAT) and glutathione peroxidase (GPx) activities (U/
mg protein) by spectrophotometry. SBP was increased in 2K-1C and
treatment with ASE (232 ± 12 vs 137 ± 8) prevented the development
of hypertension. The reduced vasodilator effect (% of relaxation) of ACh
in 2K-1C rats was recovered by the ASE (45 ± 2 vs 62 ± 2). ASE
decreased the levels of TBARS in 2K-1C rats (1.5 ± 0.2 vs 0.3 ± 0.1).
The activities of SOD, CAT and GPx were reduced in 2K-1C rats and
ASE increased these activities (14 ± 1.5 vs 24 ± 1.5; 0.8 ± 0.07 vs
2.1 ± 0.2; 0.06 ± 0.006 vs 0.12 ± 0.008, respectively). Chronic treatment
with ASE prevents the development of hypertension and improves endothelial dysfunction and oxidative stress in 2K-1C rats.
Paper No.: 2123
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
HEPARIN PREVENT THE CARDIOTOXIC EFFECT OF
BOTHROPSTOXIN I & II
Miguel de Lemos Neto(1), HD Ricardo(1), MM Machado(1),
VV Martins(1), BL Cons(1), A Cintra(2), PA Melo(1)
(1) Rio de Janeiro State University, Department of Pharmacology &
Medicinal Chemistry, Rio de Janeiro, Brazil
(2) University of São Paulo Faculty of Clinical Pharmaceutics of Riberão
Preto, Departament of Clinical Diagnostics, Roxicology and Barmatology, São Paulo,Brazil
We investigated the cardiotoxic effects of Bothropstoxin I and II (BthTX
I and II) which are toxins isolated from Bothrops jararacussu venom and
we also tested heparin (30-100mcg/mL) in these effects. We performed
the experiments on Langendorff preparation with isolated rat hearts.
These hearts were continuously perfused (2-5mL/min) with a Ringer’s
solution at 37C and recorded the electrocardiogram (EKG) and the cardiac tension. Both toxins at 10mcg/ml, induced a negative inotropic
effect over time leading to heart stoppage. BthTX I and II decreased the
EKG wave amplitude to 83.6% of the control. BthTX II increased the
perfusion pressure, enlarged the PR interval and decreased the QRS
amplitude on the EKG. Heparin 100mcg/ml abolished the BthTX I and
BthTX II cardiotoxic effects. Images obtained with heart incubated in
1% triphenyl tetrazolium chloride (TTC), showed the more damaged area
with BthTX II than BthTX I that were completely neutralized by heparin.
Heparin was able to antagonize completely the heart stoppage, changes
in the EKG and the damage induced by these toxins in the preparation of
isolated heart.
Support - CAPES, CNPq, PRONEX e FAPERJ.
Paper No.: 783
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
THE ANTI-INFLAMMATORY AND ANTIHYPERNOCICEPTIVE ACTIVITIES OF LASSBIO-1141: THE
ROLE OF ADENOSINE RECEPTORS
Cleverton KF de Lima, RB Lacerda, LL da Silva, CAM Fraga, E de
Jesus Barreiro, ALP de Miranda
University of Rio de Janeiro, Departament of Pharmacology, Rio de
Janeiro, Brazil
Introduction: Nowadays the therapies for chronic inflammatory diseases
are unsafe or inefficacious, so adenosine and its receptors arise as potential targets for the control of the inflammation. Materials: It was used
male and female Wistar rats and BALBc mice, weighing 120-180g and
25-35g, respectively. All the studies have been approved by the ethic
committee from UFRJ. Results: LASSBio-1141(100lmol/Kg, p.o.) was
able to inhibit the thermal hypernociception induced by carrageenan (La-
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
253
vich TR et al., Braz. J. Med. Biolog. Res. 2005, 38: 445-451), and by
capsaicin (Mizushima T et al. Pain 2005, 113:51-60) in 60% and 51%,
respectively. The compound inhibited the paw edema induced by carrageenan (Ferreira SHJ Pharmacy Pharmacol.1979, 31: 648) after 3h of
stimuli with Emax= 64% and ED50 = 14,3lmol/Kg, this effect, such as
antihypernociceptive effect, was totally blocked by the unspecific adenosine receptor antagonist, teophyline (10mg/Kg, i.p.), and by the adenosine deaminase (ADA, 2unit/mL, i.pl.), an enzyme which metabolize the
adenosine. In the end of the evaluation with LASSBio-1141 (100lmol/
Kg, p.o.) in the paw edema (6h after), the neutrophil recruitment was
indirectly measured through assay of myeloperoxidase (MPO) activity
(Posadas I et al. Br. J. Pharmacol. 2004 142: 331-338) and it has reduced
the infiltrated in 59%. LASSBio-1141 was able to inhibit the TNF-a production in LPS-stimulated peritoneal macrophage from mice, with Emax=
98% and IC50 = 292nM, this effect was blocked by ADA. Conclusion:
LASSBio-1141 arises as a potential candidate to control the inflammation
and its mechanism of action can involve the modulation of extracellular
levels of adenosine.
Paper No.: 1482
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
ROLE OF PROTEIN DISULFIDE ISOMERASE IN
ANGIOTENSIN II INDUCED CONTRACTION IN VASCULAR
SMOOTH MUSCLE CELLS FROM SPONTANEOUSLY
HYPERTENSIVE RATS
(1) University of Padova, Department of Pharmacology and Anesthesiology, Padova,I taly
(2) University of Padova, Department of Medical and Surgical Sciences,
Padova, Italy
Fluvoxamine, a selective serotonin reuptake inhibitor, is a first-choice
antidepressant agent for treatment of depression in elderly subjects and
patients with cardiovascular diseases. Since limited and conflicting data
are available regarding age-related modifications of fluvoxamine pharmacokinetics, this study aims to reassess the effect of age and ascertain the
possible impact of chronic heart failure (CHF) on the oral disposition
kinetics of fluvoxamine. To achieve this goal, a single 50 mg-dose of this
drug was administered orally to 10 healthy young adults, 10 healthy
elderly subjects and 10 elderly patients with CHF. Plasma fluvoxamine
concentration was measured for up to 96 hours. With the exception of
apparent distribution volume, all main pharmacokinetic parameters of the
drug were significantly modified in elderly with respect to young subjects: peak concentration was about doubled (31 ± 19 vs 15 ± 9 ng/ml;
P < 0.05); the area under the concentration-time curve was almost three
times higher (885 ± 560 vs 304 ± 84 ng•h/ml; P < 0.05); half-life was
prolonged by 63% (21.1 ± 6.2 vs 12.9 ± 6.4 h; P < 0.01), and oral clearance was halved (1.12 ± 0.77 vs 2.25 ± 0.66 l/h•kg; P < 0.001). CHF
had no significant effect on any pharmacokinetic parameters. Thus, aging
causes considerable impairment of fluvoxamine disposition, whereas no
significant pharmacokinetic alterations are produced by CHF. Therefore,
adjustment of initial dose and subsequent dose titrations may be required
in elderly subjects, whereas no further dose reduction is necessary in
patients with CHF.
Livia de Lucca Camargo, ACD Androwiki, GS Ceravolo, AZ Chignalia,
Alexandre Denadai-Souza, MN Muscará, MHC de Carvalho, ZB Fortes,
M Janizewski, LR Lopes
University of São Paulo, Institute of Clinical Biomedics, Department of
Pharmacology, São Paulo, Brazil
Enzymatic ROS generation is an inherent component of redox signalling.
We demonstrated that the redox chaperone protein disulfide isomerase
(PDI) modulates NADPH oxidase dependent ROS generation in VSMCs.
However, the role of PDI in oxidative stress and in vascular alterations
observed in hypertension is unknown. Inhibition of PDI was evaluated in
vascular reactivity to angiotensin II (Ang II) in isolated mesenteric arteriolar bed from SHR and Wistar rats. To investigate the mechanisms
involved, VSMCs from mesenteric arteries were cultured. The expression
of PDI in VSMCs was analysed by WB. Ang II-induced ROS production, c-Src activation and intracellular free calcium concentration ([Ca2 +
]i) were measured by dihydroethidium derived fluorescence, WB and
fluo-3AM fluorescence, respectively. Experiments were done in the presence of the PDI inhibitor, bacitracin (0.5 mM), and the NADPH oxidase
inhibitor, apocynin (30lM). Vasoconstrictor responses to Ang II were
substantially suppressed with bacitracin in Wistar (P < 0.05, n = 6) and
SHR (P < 0.05, n = 5). Additionally, PDI expression was higher in SHR
VSMCs as compared to Wistar and this was concomitant to an increase
in ROS generation (P < 0.05, n = 3). PDI inhibition significantly
reduced Ang II dependent ROS generation, c-Src phosphorylation and
[Ca2 + ]i responses in both Wistar (P < 0.01, n = 4) and SHR cells
(P < 0.01, n = 3). Therefore, the reduction of Ang II mesenteric vasoconstriction after PDI inhibition could be related to a decrease in ROS
production, c-Src phosphorylation and [Ca2 + ]i in VSMC from Wistar
and SHR. In summary, our data suggest that PDI could be a new player
in the oxidative stress and vascular dysfunction observed in
hypertension.
Paper No.: 1888
FOCUSED CONFERENCE GROUP: P17 - NEW APPROACHES
AND TARGETS IN PSYCHIATRY
FLUVOXAMINE PHARMACOKINETICS IN ELDERLY
SUBJECTS AND PATIENTS WITH CHRONIC HEART FAILURE
Paper No.: 2062
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
PROTECTIVE ACTION OF HYDRO-ALCOHOLIC EXTRACT
OF AÇAÍ (AN AMAZON PLANT) ON EXPERIMENTAL
METABOLIC SYNDROME IN C57BL/6 MICE
RS de Moura, PRB Oliveira, CA Costa, LCRM Carvalho, Miguel de Lemos Neto, PJC Sousa, AC Resende
Rio de Janeiro State University, Department of Pharmacology, Rio de
Janeiro, Brazil
Protective action of hydro-alcoholic extract of açaı́ (an Amazon plant)
on experimental metabolic syndrome in C57BL/6 mice. Soares de Moura R, Oliveira PRB, Costa CA, Carvalho LRM, Lemos M, Sousa
PJC, Resende AC. Rio de Janeiro State University.This study was
designed to determine the protective effects of açaı́ stone extract (ASE,
300 mg/Kg/day) in C57BL/6J mice fed a high-fat diet (HFD) that
delineate components of metabolic syndrome. C57BL/6 mice at
6 weeks of age were fed a control diet and received water (CD; 10%
fat) or ASE (CD+ASE) and a HFD (60% fat) that received water
(HFD) or ASE (HFD+ASE) for 12 weeks. The vasodilator effect of
acetylcholine (ACh, 0.1-1000 nmol) was studied in pre-contracted and
perfused mesenteric arterial bed of the mice. Body weight, plasma total
cholesterol, triglyceride, glucose and insulin levels were determined
and the insulin resistance measured by HOMA index. Oral glucose tolerance test (OGTT) was also assessed by oral administration of glucose (2g/Kg) at time 0 and measurement of glucose at different times.
Vasodilator response to ACh was reduced in HFD and ASE restored
the response. Increased body weight, plasma triglyceride, total cholesterol, glucose levels and insulin resistance were observed in HFD and
reduced by ASE. Treatment with ASE also reduced glucose intolerance
observed by OGTT in HFD. The results demonstrate a protective
effect of an extract obtained from açaı́ stone in metabolic syndrome,
since the endothelial dysfunction, obesity, hypercholesterolemia, hyperglicemia and insulin resistance induced by HFD were reduced by oral
treatment with the extract.
Sara De Martin(1), R Orlando(2), M Floreani(1), P Palatini(1)
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
254
Paper No.: 2124
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
PREVENTIVE ACTION OF AÇAI (AN AMAZON PLANT)
HYDROALCOHOLIC EXTRACT ON DELETERIOUS EFFECT
OF CIGARETTE SMOKE IN MOUSE LUNGS
Roberto Soares de Moura, AC Resende, M de Lemos Neto, MAS Silva,
ETL Trajano, JN Alves, RT Nesi, PJC Sousa, LC Porto, SS Valença
Rio de Janeiro State University, Department of Pharmacology, Rio de
Janeiro, Brazil
Cigarette smoke-induced lung damage is frequently associated either
with unbalanced protease, antiprotease production or increased oxidative
status. The present study was undertake in order to find out if an hydroalcoholic extract obtained from açai fruits can reduce the lung damage
observed in mice that inhales smoke from regular cigarette. C57BL/6
male mice were exposed to 12 commercial full-flavor filtered Virginia
cigarettes per day, for 7 days/week during 60 days (CS) or smoke from
cigarette containing açai extract (CS+A). Control group was shamsmoked. Lung histological analysis of CS group showed a typical pattern
of murine emphysema, with alveolar enlargement and areas of initial
fibrosis while CS+A presented a reduction in the alveolar enlargement
and no areas of initial fibrosis. The number of leukocytes (macrophages
and neutrophils) in the bronchoalveolar lavage was significantly higher
in the CS group than in the CS+A and control. GPx activity was reduced
in CS but fully restored in CS+A. The nitrite was reduced in the CS
compared to control group and CS+A group. Myeloperoxidase activity,
lipid peroxidation (4-HNE), metalloelastase (MMP-12) and neutrophil
elastase were elevated in CS but not in control and CS+A. The results
suggest that harm induced by cigarette smoke may be significantly
reduced by the presence of açai extract inside the cigarette.
Paper No.: 2430
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
GASTROPROTECTIVE EFFECT OF OLEA EUROPAEA LEAF
EXTRACT IN EXPERIMENTALY INDUCED GASTRIC
LESIONS IN RATS
Dragana Dekanski(1), S Ristic(1), V Piperski(1), N Radonjic(2),
N Petronijevic(2), D Mitrovic(3)
(1) Galenika a.d. R&D Institute, Biomedical Research, Belgrade,
Republic of Serbia
(2) Institute for Medical Biochemistry, School of Medicine, Belgrade
University, Belgrade, Republic of Serbia
(3) Institute for Medical Physiology, School of Medicine, Belgrade
University, Belgrade, Republic of Serbia
In this study we investigated the gastroprotective effect of dry olive leaf
extract (DOLE) in two established preclinical models of gastric damage:
gastric lesions induced by absolute ethanol and by cold restraint stress
(CRS) in Wistar rats. Different doses of standardized olive leaf extract
were applied intragastrically 30 minutes prior to absolute ethanol administration or before CRS induction. Gastric lesions were evaluated and
ulcer index was calculated. Malondialdehyde (MDA) concentration as an
index of lipid peroxidation, was determined in gastric mucosa since lipid
peroxidation has important role in pathogenesis of these conditions. The
effects of applied DOLE were compared with effect of pretreatment of
quercetin as positive control. Absolute ethanol and CRS caused severe
gastric lesions in all non-pretreated animals. Pretreatment with DOLE
(40, 80 and 120 mg/kg), as well as with quercetin (100 mg/kg), significantly (p < 0,001) attenuated induced gastric lesions. MDA concentration significantly increased in gastric mucosa after ethanol administration
(593 ± 3.86) and in CRS exposed rats (445.4 ± 43.6) vs. 287.1 ±
15.6 nmol/mg prot. in control, non-pretreated group. Pretreatment with
reference drug, quercetin, restricted rise in MDA concentration, and this
effect was statistically significant. All doses of DOLE decreased MDA
level in ethanol experimental model significantly while MDA was
reduced significantly by pretreatment with two higher doses of DOLE in
CRS. The results obtained indicate that DOLE possesses strong gastroprotective potential in experimentally induced gastric lesions, possibly
related to its antilipid peroxidative activity.
Paper No. 619
FOCUS GROUP: FC16 - NATURAL PRODUCTS: PAST AND
FUTURE?
PHARMACOLOGICAL PROFILE AND THERAPEUTIC
POTENTIALITY OF NATURAL PRODUCT OBTAINED FROM
STEM BARK OF MANGIFERA INDICA L (VIMANG) AND ITS
GLUCOSILXANTONA, MANGIFERIN
Rene Delgado-Hernandez(1), B Garrido Suarez(1), D Garcia-Rivera(2),
GL Pardo-Andreu(3), P Hernandez Casaña(4), I Rodeiro-Guerra(5),
I Hernandez-Balmaseda(5), N Merino -Garcia(2), MM GuevaraGarcia(1), A Alvarez-Leon(2), Y Maragiño(3), AJ Nuñez-Selles(6)
(1) National Center Coordinating of Clinical Trials, Havana, Cuba
(2) Center of Biomolecular Chemistry, Havana, Cuba
(3) Pharmacy Institute, Havana University, Havana, cuba
(4) Center of Molecular Immunology, Havana, Cuba
(5) Center of Marine Bioactive Compound, Havana, Cuba
(6) Department of Science. Ministry of Health, Havana, Cuba
The aqueous extract from stem bark of Mangifera indica L (VIMANG)
has been used in Cuba during several years in ethnomedical practices for
the improvement of quality of life of patients with different pathologies.
A phytochemical characterization of the extract has led to the isolation of
nine phenolic constituents, with the glucosylxanthone mangiferin as a
major component, and different microelements as zinc, copper, and selenium. The extract has demonstrated as the main pharmacological property its antioxidant activity. Others studies have shown that the extract
also possesses others pharmacological activities, such as: anti-inflammatory, antiallergic, analgesic and inmunomodulador, with very complex
and multifactorial mechanisms of action involved. These properties are
related to its scavenger capacity of different reactive oxygen species. The
interaction of mangiferin and others component of the extract with iron
represent an important antioxidant mechanism recently characterized in
our studies. On the other hand, mangiferin and Vimang have the property
of modulating different mediators involved into immune response. In
general, the total extract and its xanthone, mangiferin are involved in
several inmunomodulatory processes, properties that confer an important
therapeutic potentiality as adjuvant in the preparation of phytopharmaceuticals products for the treatment of pathologies where oxidative stress
and immunomodulator disorders are related with their etiology. Different
clinical studies are conducted at this moment in order to get new knowledge about its therapeutic potentiality.
Key words: Mangifera indica L, Vimang, mangiferin, antioxidant, antiinflammatory, immunomodulation
Paper No.: 2191
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
MECHANISMS OF THE DILATOR ACTION OF A DANSHEN
AND GEGEN FORMULATION ON RAT BASILAR ARTERY
Shirlly Y Deng(1), FFY Lam(1), ESK Ng(1), JHK Yeung(1),
YW Kwan(1), CBS Lau(2), JCM Koon(2), PC Leung(2), KP Fung(1,2)
(1) The Chinese University of Hong Kong, School of Biomedical
Sciences, Shatin, Hong Kong, China
(2) The Chinese University of Hong Kong, Institute of Chinese
Medicine, Shatin, Hong Kong, China
Danshen (root of Salvia miltiorrhiza) and Gegen (root of Pueraria lobata) are two herbs used in traditional Chinese medicine, most commonly
for their putative cardioprotective and anti-atherosclerotic effects. In this
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
255
study, the actions of a Danshen and Gegen formulation (DG; ratio 7:3)
were investigated on rat isolated basilar artery rings precontracted with
100nM U46619. DG produced concentration-dependent relaxation of the
artery rings with an IC50 of 0.90 ± 0.12mg/ml. Mechanical removal of
the endothelium or pre-treatment with 10mM of a potassium channel
inhibitor tetraethylammonium (TEA) had no effect on the DG-induced
vasodilator response, but pre-treatment with 100mM TEA suppressed
the maximum response to DG by 18% (P < 0.05). Involvement of
Ca2 + channels was investigated in artery rings incubated with Ca2 + -free
buffer and primed with 100nM U46619 for 5 minutes prior to adding
CaCl2 to elicit contraction. Pre-incubation of the artery rings with DG
for 10 minutes produced concentration-dependent (1, 3 and 7mg/ml) and
total inhibition on the CaCl2-induced vasoconstriction. These findings
suggest DG relaxes the basilar artery primarily by inhibiting Ca2 + influx
in the vascular smooth muscle cells, and a minor component is mediated
by opening TEA-sensitive K+ channels. DG could be a useful cerebroprotective agent in some patients with occlusive cerebrovascular disease.
Paper No.: 813
FOCUSED CONFERENCE GROUP: PW02 - SYMPOSIUM ON
ADVANCES IN GI PHARMACOLOGY
PROTECTIVE EFFECT OF VEGF ON ETHANOL-INDUCED
HEMORRHAGIC GASTRIC EROSIONS IN RATS
Xiaoming Deng(1,2), X Xiong(1), Z Sandor(1,2), S Szabo(1,2)
(1) VA Long Beach Healthcare System, Department of Diagnostic &
Molecular Medicine HCG, Long Beach, CA, USA
(2) University of California, Irvine, CA, USA
Treatment with the peptide or gene of vascular endothelial growth factor/
vascular permeability factor (VEGF/VPF) accelerated healing of experimental gastric and duodenal ulcers (Szabo et al., DDS 1998;43:40S-45S;
Jones et al., Gastroenterology 2001;121:1040-7; Deng et al., J Pharmacol
Exp Ther 2004;311:982-988.). Since VEGF/VPF increases vascular permeability, we tested the hypothesis that it might create a perivascular
‘‘histodilutional barrier’’, hence delay the absorption of damaging chemicals that are known to induce vascular injury and hemorrhagic erosions.
Fasted S-D rats were given rat VEGF164 (ProSpec) 0.01, 0.04, or 0.2
ug/100g by gavage at 30 min before 75% ethanol (1 ml/kg) intragastrically. Control rats were pretreated with equal amount saline. All the rats
were euthanized 1 hr after ethanol administration. Hemorrhagic gastric
mucosal lesions were measured by an Imaging Analysis System (MetaMorph) and calculated as % of glandular stomach. The results showed
that VEGF/VPF pretreatment with 0.01, 0.04, or 0.2 ug/100g prevented
gastric mucosal lesions from 10.6 ± 1.7% (controls) to 8.9 ± 1.8,
0.9 ± 0.2, or 2.8 ± 0.7%, respectively. Statistical analysis indicated that
the doses (0.04 and 0.2 ug/100g) of VEGF/VPF significantly reduced the
gastric mucosal lesions (p < 0.01 and 0.05, respectively) when compared
with controls. Conclusions: 1) This is the first demonstration that pretreatment of VEGF/VPF offered dose-dependent gastroprotection in rats
exposed to ethanol. 2) The likely mechanisms might be related to a histodilutional barrier induced by VEGF/VPF. 3) Further studies are needed
to elucidate the molecular mechanisms on this new form of gastroprotection by VEGF/VPF.
Paper No.: 2853
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
THE RELAXANT EFFECT OF HYDROGEN SULFIDE IS
POTENTIATED WHILE THAT OF ACETYLCHOLINE IS NOT
ALTERED IN PULMONARY AND MESENTERIC ARTERIES
ISOLATED FROM STREPTOZOTOCIN-INDUCED DIABETIC
RATS
Merve Denizalti(1), TE Bozkurt(1), I Sahin-Erdemli(1), N Abacioglu(2)
(1) Hacettepe University, Faculty of Pharmacy, Department of Pharmacology, Ankara,Turkey
(2) Gazi University, Faculty of Pharmacy, Ankara, Turkey
Hydrogen sulfide (H2S) is an endogenous gas which has potent relaxant
effect in vascular and nonvascular smooth muscles. The involvement of
H2S in several pathological states including diabetes has also been
reported. In the present study, we have investigated the relaxant effect of
H2S in mesenteric and pulmonary arteries isolated from 12 weeks-diabetic rats. Diabetes was induced by tail vein injection of streptozotocin
(STZ) (35 mg/kg) dissolved in 0.1 M citrate buffer. The control group
received citrate buffer alone. Insulin-treatment was applied by using insulin implants. At the end of 12 weeks the mesenteric and pulmonary arteries were isolated and the relaxation response to H2S donor sodium
hydrogen sulfide (NaHS) (0.1 mM-3 mM) and acetylcholine (10 nM-30
ı̀M) was elicited. The relaxation responses of the arteries isolated from
12 weeks-diabetic rats were found to be increased when compared to that
obtained from non-diabetic controls. ATP-sensitive potassium channel
(KATP) blocker glibenclamide inhibited the relaxation response to NaHS
in the arteries isolated from either diabetic or non-diabetic group of rats.
Concurrent insulin treatment of STZ-injected rats prevented the potentiation of the relaxant effect of NaHS in the mesenteric and pulmonary
arteries. However, acetylcholine-induced relaxation responses were not
altered in diabetic group of rats. In conclusion, STZ-induced experimental diabetes resulted in enhanced relaxation response to H2S in mesenteric and pulmonary arteries of rats. The potentiation of the relaxation
response to H2S in diabetic arteries may indicate a role against the vascular complications of diabetes.
Paper No.: 1686
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
SIGNALLING VIA IRAG REGULATES NO- AND
ANP-MEDIATED SMOOTH MUSCLE RELAXATION
Matthias Desch(1), B Hieke(1), D Bernhard(2), K Hoecherl(3),
F Hofmann(2), J Schlossmann(1)
(1) University of Regensburg, Department of Pharmacology and
Toxicology, Regensburg, Germany
(2) Technical University of Munich, FOR 923, Munich, Germany
(3) University of Regensburg, Department of Physiology, Regensburg,
Germany
NO / ANP / cGMP signalling is important for many physiological functions e.g. vascular tone regulation, vascular smooth muscle proliferation,
gastrointestinal motility or neurotransmission. IRAG (IP3 Receptor Associated cGKI substrate) is highly expressed e.g. in smooth muscle cells
and might play an important role for cGKI downstream signalling. To
elucidate the role of IRAG for NO- and ANP-mediated smooth muscle
tone regulation, cGKI localization and blood pressure adjustment we
generated IRAG knockout mice by targeted deletion of exon 3. Hormone
precontracted aortic tissues of IRAG deficient mice showed an impaired
relaxative response to exogenous and endogenous cGKI activating compounds like NO, ANP, Acetylcholine and cGMP. The cGKI dependent
reduction of tonus by high potassium depolarization was not affected in
IRAG deficient tissue in comparison to wild type tissue. Reduction of
hormone-induced calcium rise by NO, ANP and cGMP was abolished in
IRAG deficient vascular smooth muscle cells (VSMC) in contrast to wild
type VSMC. The localization of cGKIa- and cGKIb-isozymes in cultured aortic smooth muscle cells was not altered in absence of IRAG.
Basal mean arterial blood pressure, heart rate and activity were not changed in IRAG-/- mice. Interestingly, under pathophysiological conditions
like sepsis (induced by E.coli lipopolysaccharide treatment) IRAG deficient mice were resistant to blood pressure reduction. These findings
indicate that blood pressure regulation via cGKI / IRAG under physiological terms might be counterbalanced whereas this signalling pathway
might play an important role for blood pressure modulation under pathophysiological conditions.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
256
Paper No.: 1282
FOCUSED CONFERENCE GROUP: P08 - DEVELOPMENTS IN
THE TREATMENT OF SEXUAL DYSFUNCTION AND
DISEASES OF THE LOWER URINARY TRACT
ROLE OF CX43 IN THE MAINTENANCE OF SPONTANEOUS
ACTIVITY IN THE GUINEA PIG PROSTATE
Anupa Dey, S Kusljic, RJ Lang, B Exintaris
Monash University, Department of Medicinal Chemistry and Drug
Action, Melbourne, VIC, Australia
Background: Investigate the role of CX43 in the maintenance of spontaneous activity in young and aging guinea-pig prostates. Methods: Conventional intracellular microelectrode and tension recording techniques
were used. Results: Experiments were performed using gap junctional
uncouplers 18b GA (40lM), CBX (50lM) and octanol (0.5mM), on
cells displaying slow wave activity and on spontaneously contracting tissue in young and aging guinea-pig prostates. 40lM 18b GA and 50lM
CBX both abolished slow wave activity in young and aging prostates
and significantly depolarised the membrane potential by approximately
5mV (n = 5, student paired t-test p < 0.05). 50lM CBX and 40lM 18b
GA abolished all spontaneous contractions in young and aging guineapig prostate tissue (2-way ANOVA, Bonferroni post test, P < 0.01)
(n = 5). Although, lower concentrations of CBX (10lM) and 18b GA
(10lM) significantly reduced the spontaneous contractions within aging
guinea pig prostates while the young tissue remained unaffected from
control (students paired t-test p > 0.05). Octanol abolished slow wave
activity in both age groups of guinea-pig prostates (paired students-t test,
p < 0.05). Octanol also significantly depolarised the membrane potential
from the control values in both age groups (paired students-t test,
p < 0.05). Within both age groups of animals octanol inhibited all contractile activity at two different concentrations (0.5mM and 1mM) (2-way
ANOVA, Bonferroni post test, P < 0.01) (n = 5). These effects were
reversed upon washout. Conclusion: Results demonstrated that when gap
junctional communication via CX43 was impaired spontaneous activity
was abolished at a cellular and whole tissue level; CX43 is therefore
essential for the maintenance of activity in the guinea-pig prostate gland.
Paper No.: 2401
FOCUSED CONFERENCE GROUP: P19 - GENERAL
SESSION - TOXICOLOGY
OXALIPLATIN-DEPENDENT NEUROPATHY IN THE RAT:
PAIN BEHAVIOUR AND PERIPHERAL NERVOUS SYSTEM
ALTERATIONS
Lorenzo Di Cesare Mannelli(1), L Bonaccini(2), A Pacini(2),
C Ghelardini(1), A Bartolini(1)
(1) University of Florence, Department of Pharmacology, Florence, Italy
(2) University of Florence, Department of Anatomy, Florence, Italy
Oxaliplatin, a platinum-based chemotherapeutic agent, has become a
standard treatment for advanced colorectal cancer and a valid option for
patients in the adjuvant setting. The dose-limiting toxicity of this
compound is the development of a peripheral neuropathy with gloveand-stocking distribution sensory loss, combined with paresthesia, dysesthesia, and pain. In order to delucidate the morphological and molecular
alterations that occur in the peripheral nervous system during neuropathy,
we daily injected rats with 2,4 mgkg-1 oxaliplatin intraperitoneally.
Twenty-one days later, the paw-pressure test evidenced a significantly
reduced pain threshold consisting in mechanical hyperalgesia. The histochemical study of the peripheral nerve did not highlight variations in
myelin thickness or axonal diameter. On the contrary, the treatment
increased the incidence of eccentric nucleoli and multinucleolated neurons in the lumbar dorsal root ganglia (DRG). Moreover, a significant
reduction in the somatic area of small and medium neurons in the oxaliplatin groups was observed. The immunohistochemical analysis showed
an oxaliplatin-induced alteration of gene expression profile. The expression of the cellular injury marker ATF3 was enhanced both in axons and
in myelinating Schwann cells of the sciatic nerve, as well as in neurons
and satellite cells of DRG. The expression of the structural protein
NF200 was unaltered in the nerve, whereas the majority of DRG neurons
displayed a dramatic decrease. An understanding of the factors involved
in the development and maintenance of neuropathy and the study of the
neuron vs glia signalling may lead to mechanism based therapies that
prevent/treat neuropathic pain and improve neurorestoration.
Paper No.: 2000
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
GENETIC TARGETING OF COX-1 AND COX-2 IN MURINE
AORTIC ENDOTHELIAL CELLS REVEALS THE
ANTIOXIDANT EFFECT OF ENDOTHELIAL COX-2
Andrea Di Francesco(1), L Di Francesco(1), M Dovizio(1), F Seta(2),
C Funk(2), P Patrignani(1)
(1) ‘‘G. d’Annunzio’’ University, Department of Medicine and Center of
Excellence on Ageing, Chieti, Italy
(2) Queen’s University, Departments of Physiology and Biochemistry,
Kingston, Canada
Prostacyclin restrains inflammation in endothelial cells through the induction of heme oxygenase(HO)-1, an antioxidant enzyme. Using genetic
strategies of selective inhibition of cyclooxygenase (COX)-1 and COX-2
in mice, we explored the contribution of COX-isozymes to prostanoid
generation and HO-1 expression in murine aortic endothelial cells (MAECs) in response to interleukin(IL)-1b(10ng/ml). MAECs were isolated
from wild-type (WT), COX-1 and COX-2 knock-down(KD) mice, and
cultured in EBM-2 supplemented with 10% FBS for 6-24 hours. COX-1,
COX-2, HO-1 and terminal prostaglandin synthases(tPGS) levels were
assessed by Western blot analysis, and released prostanoids were evaluated by specific immunoassays techniques. COX-1, but not COX-2, was
detectable in unstimulated MAEC. IL-1b caused COX-2 induction by 4and 8-fold in WT and COX-1 KD cells, respectively. COX-1 and tPGS
levels were not significantly modulated. The induction of COX-2 was
associated with significant (P < 0.05) increase of 6-keto-PGF1a>PGE2 >
PGF2a in a time dependent fashion, both in WT and COX-1 KD cells.
PGD2 was not detected. In contrast, COX-2 KD cells were associated
with complete inhibition of prostanoid generation. COX-1 KD cells
exhibited substantially enhanced (2 fold) expression of COX-2 wich translated into significant (P = 0.007) higher levels of prostacyclin versus WT
(11611 ± 1678 vs 4342 ± 674.2 pg/ml, respectively). HO-1 expression
was strongly suppressed in COX-2 KD MAEC but unaffected by COX-1
manipulation. In conclusion, COX-2 may counteract cytokine-dependent
activation of endothelial cells through the induction of HO-1. Interestingly
COX-2 expression seems to be under the negative control of COX-1.
Whether this phenomenon is mimed by pharmacological inhibition of
COX-1 remains to be elucidated.
Paper No.: 1774
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
ANALYSIS OF THE IMMUNOGENICITY OF AN INTERFERON
B1A BIOSIMILAR
Roberto Diez(1), M Kauffman(1), A Sterin-Prync(1), M Papouchado(1),
E Gonzalez(2), A Grossberg(3), S Chuppa(3), B Odoriz(4), C Vrech(5),
H Ferro(1)
(1) Bio Sidus S.A., Buenos Aires, Argentina
(2) CDM, Buenos Aires, Argentina
(3) Medical College of Wisconsin, WI, USA
(4) Hospital Central, UNC, Mendoza, Argentina
(5) Sanatorio Allende, Córdoba, Argentina
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
257
Introduction: Interferon (IFN) b, like other therapeutic proteins derived
from human genes by recombinant DNA technology, can be immunogenic, potentially influencing its efficacy and safety. Blastoferon is a
pharmaceutical product of human IFN-b1a currently marketed in Argentina and Latin America as a biosimilar to the innovator IFNb1a product
for the treatment of multiple sclerosis (MS). The aim of this paper is to
present the assessment of Blastoferon immunogenicity. Materials and
patients: Immunological recognition by monoclonal and polyclonal antibodies (with either neutralizing and non-neutralizing activity) was followed by IFN biological activity testing, comparing Blastoferon to other
IFN-b products, including WHO standard. After marketing approval, a
pharmaceutical program was established, including active surveillance at
two sentinel sites in Buenos Aires and Córdoba cities. Serum of their
patients (n = 37) were tested for anti-IFNb antibodies by ELISA and in
positive cases, also for neutralizing activity against the antiviral effect of
IFN-b on Wish cells challenged with Vesicular Stomatitis Virus. Results:
recognition by the panel of antibodies showed that Blastoferon shares
immunological determinants with other human interferon-b products,
especially IFNb1a. At two years of follow-up, IFNb -binding antibodies
have been detected by enzyme immunoassay (EIA).in 72.9% of the
patients, whereas neutralizing antibodies were found in only 8.1% of the
MS patients registered in the ongoing Blastoferon pharmacovigilance
program. Conclusion: Blastoferon appears to generate antibodies at a
rate similar to the innovator IFNb1a product administered by subcutaneous route.
Paper No.: 1775
FOCUSED CONFERENCE GROUP: P04 - PHARMACOEPIDEMIOLOGY, CURRENT CONTROVERSIES AND OPPORTUNITIES
EMERGENCE OF H274Y MUTATION AND OSELTAMIVIR
RESISTANCE DURING TREATMENT IN TWO PATIENTS IN
THE 2009 EPIDEMICS OF SWINE FLU IN ARGENTINA
Paper No.: 2114
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
BISPHOSPHONATE-RELATED OSTEONECROSIS OF THE
JAW (BONJ)
G Dimosiari, Konstantinos Imprialos, Xxxxx Mironidou-Tzouveleki
Aristotle University of Thessaloniki, Medical School, A’ Laboratory ofPharmacology, Thessaloniki, Greece
Background: Bisphosphonates are primary agents in the current pharmacological arsenal against osteoclast-mediated bone loss due to osteoporosis, Paget disease and hypercalcaemia associated with certain malignant
tumors. They are also frequently prescribed for prevention and treatment
of low bone density and osteogenesis imperfecta. They have been shown
to reduce skeletal morbidity in multiple myeloma as well as a wide range
of solid tumors affecting bone. Despite these uses, the current recognition
that bisphosphonate use can be associated with pathologic complications
such as osteonecrosis of the jaw has brought skepticism and has planted a
new field for research regarding the widespread prescription of these
drugs. Currently the American Dental Association defines bisphosphonate-associated osteonecrosis as an exposed area of necrotic jaw bone
present for at least eight weeks in patients using bisphosphonates. Recent
studies have documented that osteonecrosis, associated with the use of
bisphosphonates, primarily refers to intravenously administered drugs,
nitrogen-containing, on a long-term basis, and few cases have been
reported after short-term or oral delivery. Purprose and Methods : Since
2003, the number of publications on the subject has dramatically
increased. The purpose of this paper is to report on characteristics, mechanisms of action, risk factors and incidence of BONJ by comprehensively
reviewing the literature via pub med research. Conclusion: Though evidence supports clinical decisions in favor of the patient taking oral bisphosphonates, it is ultimately the prescribing physician’s duty to weigh
the benefits and risks of oral bisphosphonate use individually before determining the call for temporary or permanent termination of medication.
Roberto Diez(1), ML Valinotto(2), P Barrero(2), M Viegas(2), MC Alvarez López(3), E López(3), AS Mistchenko(2)
(1) University of Buenos Aires, School of Medicine, Department of
Pharmacology, Buenos Aires, Argentina
(2) Hospital de Niños R Gutiérrez, Virology Lab, Argentina
(3) Hospital de Niños R Gutiérrez, Department of Medicine, Argentina
Introduction: Oseltamivir was the most commoly used antiviral in the
2009 epidemics of swine-origin influenza A H1N1 in Argentina and elsewhere. The emergence of resistance is an expected result of exposure to
and use of any anti-infective agent. The most frequent event resulting in
oseltamivir resistance is the substitution of histidine by tyrosine at position 275 of neuraminidase, encoded by the H274Y mutation The aim of
this report is to present genotypic and phenotypic evidence of oseltamivir
resistance emergence, mediated by the H274Y mutation in two cases during the 2009 epidemics in Argentina. Materials and patients: Complementary DNAs including the 274 codon were obtained by RT-PCR using
viral RNAs extracted from nasopharyngeal aspirates. Sequencing and pyrosequencing was performed following the WHO Influenza A (H1N1)
NA-H274 protocol. Neuraminidase activity in the presence or absence of
oseltamivir was performed by chemiluminescence with comercial
reagents (NA-Star Influenza Neuraminidase Inhibitor Resistance Detection Kit). Results: Sequential samples of two pediatric patients under
oseltamivir treatment were analyzed. Pre-treatment samples were composed of 100% oseltamivir-sensitive variants. In Case 1, the oseltamivirresistant variant was found eight days after the beginning of treatment. In
Case 2 the viral population became resistant on the second day of
treatment, with 83% of the viral population bearing the mutation and
reaching 100% on the seventh day. Conclusion: We describe the intratreatment emergence of oseltamivir resistance in two pediatric patients.
Pyrosequencing allowed us to detect variant mixtures, showing the
precise moment when the viral population changed from sensitive to
resistant.
Paper No.: 1921
FOCUSED CONFERENCE GROUP: P04 - PHARMACOEPIDEMIOLOGY, CURRENT CONTROVERSIES AND OPPORTUNITIES
A COMMUNITY PHARMACY MULTINATIONAL PROJECT TO
INVESTIGATE THE PRESCRIPTION DRUG ABUSE
Paola D’Incau(1), M Lapeyre-Mestre(2), A Carvajal(3), U Bergman(4),
ER Heerdink(5), RV Stichele(6), D Macias(3), L Pourcel(2),
A Conforti(1),
(1) University of Verona, Department of Medicine and Public Health,
Section of Pharmacology, Verona, Italy
(2) Université Paul Sabatier, CEIP-A, Unit of Pharmacoepidemiology,
Toulouse, France
(3) University of Valladolid, Institute of Pharmacoepidemiology,
Valladolid, Spain
(4) Karolinska Institute, Department of Physiology & Pharmacology,
Stockholm, Sweden
(5) University of Utrecht, Department of Pharmacoepidemiology,
Utrecht, TheNetherlands
(6) Ghent University Hospital, Department of Pharmacology, Ghent,
Belgium
Prescription drug abuse and diversion have been reported as a critical
concern in terms of patient care and public health, receiving conspicuously targeted consideration from health authorities. The aim of this article was to analyse the feasibility of carrying out the OSIAP system to
detect the potential abuse of marketed drugs in a multinational community pharmacy setting. An average of 2105 community pharmacies took
part in the OSIAP project during 2006 and 2007. They reported a total
of 862 suspect prescriptions concerning 1220 different drugs. The mean
age of the total sample of subjects presenting suspect prescriptions was
45 years (SD, 16.7) and the majority of them were women. The most
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
258
frequently reported criteria of suspicion was ‘‘modification of the prescription’’ and most suspect prescriptions regarded the ATC N class. Of
these drugs, 54% were psycholeptics (54% anxiolytics, 40% hypnotics
and sedatives and 6% antipsychotics), 23% analgesics (72% opioids,
23% other analgesics and antipyretics and 5% antimigraine preparations)
and 11% psychoanaleptics (66% antidepressants, 29% psychostimulants,
5% anti-dementia drugs and 1% psycholeptics and psychoanaleptics).
The OSIAP system provided useful information resulting from the
patients’ everyday life, thus confirming potential role of a pharmacy network in limiting drug diversion. Further projects should be developed
taking into consideration a variety of intervention strategies, from psychiatric intervention to practical law enforcement strategies. They should
entail the collaboration of multidisciplinary efforts involving the abusers
themselves in frontline educational activities.
Boeuf O & Lapeyre-Mestre M. Drug Safety 2007; 30: 265-276.
Paper No.: 1922
FOCUSED CONFERENCE GROUP:
P04 - PHARMACOEPIDEMIOLOGY, CURRENT
CONTROVERSIES AND OPPORTUNITIES
GENDER DIFFERENCES OF ADRS RELATED TO
PSYCHOTROPIC DRUG USE: A SURVEY FROM FRANCE,
ITALY AND SPAIN
Paola D’Incau(1), M Lapeyre-Mestre(2), M Sáinz(3), M Donati(1),
A Carvajal(3)
(1) University of Verona, Department of Medicine and Public Health,
Section of Pharmacology, Verona, Italy
(2) Université Paul Sabatier, CEIP-A, Unit of Pharmacoepidemiology,
Toulouse, France
(3) University of Valladolid, Institute of Pharmacoepidemiology, Valladolid, Sain
It is well recognized that being female appears to be a risk factor for
developing ADRs. A number of studies clearly suggest that ADRs are
50 to 75% more likely in women than men. At the same time, nervous
system agents represents one of the most frequent drug classes reported
(20%) to elicit adverse events. A female propensity to experience or
report drug-related adverse effects may result from gender-related differences in drug exposure as well as in the number of drugs prescribed, in
the drug pharmacokinetics and pharmacodynamics. Nonetheless, the reasons for this increased risk in female patients are not entirely clear, notably whether adverse drug reactions among women reflect an
inappropriate use of psychotropic medicines. The results from the European Study of the Epidemiology of Mental Disorders (ESEMED) and
from the Ordonnances Suspectes Indicateur d’Abus Possible (OSIAP)
project suggest that among European countries, France, Italy and Spain
reordered one of the highest percentages of psychotropic drug use. On
the basis of these assumptions, the aim of this study is to compare in
males and females incidence, type, seriousness and drugs involved in
ADRs reported in a regional pharmacovigilance centre of each of these
countries respectively, Midi-Pyrénées, Veneto and Castilla-Leon, using
spontaneously reported cases between 2007 and 2009. This analysis will
also compare the incidence, type, seriousness and psychotropic drugs
involved in ADRs in males and females, as spontaneously reported to
these three regional pharmacovigilance centres.
Van der Heyden JHA et al. Pharmacoepidemiology and Drug Safety
2009;18: 1101–1110
Paper No.: 3353
FOCUSED CONFERENCE GROUP:
P19 - GENERAL SESSION - TOXICOLOGY
ROLE OF SISTER CHROMATID EXCHANGE ANALYSIS IN
ASSESSMENT OF PROGUANIL GENOTOXICITY IN
CULTURED HUMAN LYMPHOCYTES
Domagoj Dinter(1), G Gajski(2), V Garaj-Vrhovac(2)
(1) Pliva Croatia Ltd., Oral Solid Forms, Zagreb, Croatia
(2) Institute for Medical Research and Occupational Health, Mutagenesis
Unit, Zagreb, Croatia
Proguanil hydrochloride is a prodrug of cycloguanil, a dihydrofolate
reductase inhibitor, which in malaria parasites Plasmodium falciparum
blocks DNA synthesis. In this study, evaluation of DNA damage in
human lymphocytes after proguanil treatment in vitro was made by virtue
of measuring frequency of sister chromatid exchanges (SCE) as a sensitive biomarker in assessment of DNA integrity since SCE are known to
increase as a result of the exposure to various genotoxic agents. Different
concentrations of proguanil obtained from the plasma concentrations;
130 ng/ml for prophylactic treatment and 520 ng/ml for the treatment of
malaria were used with and without metabolic activation (S9) in different
lenghts of time. SCE technique allows metaphases in first, second and
third division to be recognized after culturing. For this reason, the SCE
analysis was also employed in estimation of proliferation kinetics after
the proguanil treatment. Results of the frequencies of SCE in human lymphocytes did not show statistically significant deviation from the control
samples even though mean SCE per cell in all the exposed samples was
slightly higher than in corresponding controls and was more prominent
with metabolic activation (S9). In addition, the percentage of high frequency cells (HFC) for each sample was estimated using the pooled distribution of all SCE measurements. This parameter also did not show
significant deviation compared to the control samples. Results revealed
that proguanil did not inhibit lymphocyte proliferation in significant manner, therefore proguanil can be considered relatively safe from the aspect
of genotoxicity, especially if used for prophylactic treatment.
Paper No.: 2951
FOCUSED CONFERENCE GROUP: P04 - PHARMACOEPIDEMIOLOGY, CURRENT CONTROVERSIES AND OPPORTUNITIES
ALUMINIUM PHOSPHIDE POISONING - A RETROSPECTIVE
STUDY
Nandlal Disania, L Sharma, S Janani, BM Gupta
SMS Medical College, Department of Forensic Medicine, Jaipur, India
Aluminium Phosphide Pesticide poisoning is an important cause of morbidity and mortality in India, there has been an explosion of reports from
parts of India predominantly being northwest and central India. The
study was aimed to generate a baseline data on the epidemiological factors contributing to the incidence and mortality due to Aluminium Phosphide poisoning, so as to highlight the problem which requires planned
and concentrated efforts in dealing with it on a broader horizon. Since
prevention is the only logical approach there is an urgent need to take
appropriate steps to prevent loss of lives. The analysis of the data
revealed that the cases of Aluminium Phosphide poisoning brought to
the mortuary of S M S Medical College, Jaipur(Rajasthan) for medico –
legal autopsy, during five years period i.e. January 2005 to December
2009, with highest incidence in 2008. The age group ranged between 14
to 65 years, with maximum incidence between 22 to 29 years and males
outnumbering females. The main mode of poisoning was suicidal. The
use of poisoning Aluminium phosphide for suicidal purpose has markedly increased during the last decade contributing 6.2% of the total mortality .The mechanism of action of Aluminium Phosphide is not clearly
known. More basic research is required to find exact mechanism of
action of phosphine so that a specific antidote can be developed.
Paper No.: 1891
FOCUSED CONFERENCE GROUP: P06 - THE HEART GONE
WRONG; STABILIZATION OF CARDIAC FUNCTION - BASIC
ENGAGEMENT OF IMIDAZOLINE RECEPTORS IN EFFECTS
OF IMIDAZOLINE DRUGS ON ISOLATED RAT HEART ATRIA
Justyna Dlugokecka, A Radwanska, R Kaliszan, A Nasal
Medical University of Gdañsk, Department of Biopharmaceutics and
Pharmacodynamics, Gdansk, Poland
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
259
The main sites of action of imidazoline drugs are located in the brain
and in peripheral vasculature. Recently, attention has been payed to the
role of imidazolines on physiology of the heart. However, no systematic
comparative studies were reported regarding the activity of imidazoline
drugs towards imidazoline receptors in the heart preparations. In this project the effect of I1- (clonidine, rilmenidine, moxonidine and
AGN192403) and I2-imidazoline receptor ligands (2-BFI, BU239) were
studied on isolated rat heart atria. The spontaneously beating right and
electrically driven left atria were treated with cumulative concentrations
of the agents studied. The inotropic and chronotropic responses of imidazolines were measured at the presence of fixed concentrations of aadrenergic (phentolamine or yohimbine) or I1/I2-imidazoline (idazoxan)
receptor blockers. These effects were calculated as per cent of changes of
the control value of atria rate or amplitude preceding the administration
of each agent studied. Log EC50 parameters were also calculated. The
positive inotropic effect were evoked with the rank order of potency: clonidine > BU239 > rilmenidine > moxonidine. These effects were diminished by idazoxan. AGN192403 did not change the amplitude of left
atria. 2-BFI weakly diminished the rate ofbeating of atria; moxonidine
and rilmenidine had no effect. In conclusion, imidazoline receptors of I1
subtype may be involved in inotropic reaction of the agents studied, but
this effect depends mainly on the a2/a1 adrenergic receptors. Engagement
of I2 imidazoline receptors, along with the a2 adrenergic ones, in chronotropic activity of isolated right atria of rat has been demonstrated.
Paper No.: 2990
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
THALIDOMIDE PRODUCES SMOOTH MUSCLE
RELAXATION
James R Docherty, SW Seto
Royal College of Surgeons in Ireland, Department of Physiology, Dublin,
Ireland
We have found that thalidomide produces hypotension in conscious and
anaesthetised rats, and have investigated the mode of action. Male Wistar
rats (250g) were killed by CO2 overdose, and the tail artery, aorta and
vas deferens was removed and rings were set up in small vessel myographs. Thalidomide (10-100 uM) produced significant relaxations of
phenylephrine contracted tail artery and inhibited contractions to calcium
restoration in the presence of phenylephrine but not KCl. Hence, thalidomide did not act as an L-type calcium channel blocker. Thalidomide also
inhibited nerve-evoked contractions of epididymal, but not prostatic, portions of rat vas deferens, an action shared with the T-type calcium channel blocker NNC 55-0396. Thalidomide did not reduce the maximum
contraction to noradrenaline in aorta. Relaxant actions in vitro do not
seem to involve a1-adrenoceptors, caffeine sensitive calcium stores, glibenclamide sensitive potassium channels, PKC or P38 MAP kinase. Like
thalidomide, the T-type calcium channel blocker NNC 55-0396 (100
uM) inhibited contractions to calcium restoration in the presence of phenylephrine but not KCl. combination with NNC 55-0396, thalidomide
failed to produce any further inhibition of the contraction to calcium restoration in the presence of phenylephrine. It is concluded that thalidomide produces vascular and non-vascular relaxations by a mechanism
distinct from L-type calcium channel block, but its actions resemble
those of T-type calcium channel blockers.
Paper No.: 2300
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
CYP2D6 AND CYP2C19: A COMPARISON OF DIRECT VERSUS
PREDICTED PHENOTYPING IN A SOUTH AFRICAN
POPULATION
Tyren Dodgen(1), D Cromarty(1), H Fickl(2), M Pepper(3,4)
(1) University of Pretoria, Department of Pharmacology, Pretoria, South
Africa
(2) University of Pretoria, Unit for Inflammation and Immunity and Academic Division NHLS, Department of Immunology, Pretoria, South
Africa
(3) University of Pretoria, Department of Immunology, Pretoria, South
Africa
(4) Geneva University Medical Centre, Department of Genetic Medicine
and Development, Geneva, Switzerland
Knowledge of cytochrome P450 (CYP) isoenzyme activity is used to
predict inter-individual variability in drug metabolism. Several genetic
screening tests are available, including the FDA-approved Roche AmpliChip CYP450 test, which predicts for poor, intermediate, extensive and
ultra-rapid metabolisers (PM, IM, EM and UM respectively). However,
several non-genetic factors influence drug metabolism and contribute to
the final phenotype, thereby confounding predicted phenotyping based
on genetic screening alone. Samples collected from subjects treated with
specific probe drugs (dextromethorphan for CYP2D6 and omeprazole for
CYP2C19) at 0, 2, 3 and 4 hours post administration were genotyped
using the AmpliChip CYP450 test. A semi-automated on-line solid phase
extraction liquid chromatograph tandem mass spectrometer (LC-MS/MS)
method was developed and validated in-house for simultaneous phenotyping using plasma concentrations of the drugs and metabolites to determine a metabolic ratio. For CYP2D6, 55% of the sampled population
were EM, 42% were IM, 1% were PM and 2% could not be classified
using the AmpliChip. For CYP2C19, 93% were EM and 7% PM. Phenotyping 3 hours post administration reveals that the sampled population
was 48.1% EM, 44.2% IM and 7.7% PM for CYP2D6 and 45.5% EM,
42.4% IM and 12.1% PM for CYP2C19. A significant discordance was
thus observed between genotype and phenotype, and the reasons for this
discrepancy are being addressed. Direct rather than predicted phenotyping may prove to be a simple and efficient means of determining interindividual variability in drug metabolism.
Paper No.: 1081
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
THYROXINE MEDICATION AND CYP 3A4 ACTIVITY AS
MEASURED BY THE URINARY 6B-HYDROXYCORTISOL/
CORTISOL RATIO: A PILOT STUDY
Josef Donnerer(1), J-P Siest(2), E Beubler(1)
(1) Medical University of Graz, Institute of Experimental & Clinical
Pharmacology, Graz, Austria
(2) Stabiligen, Nancy, France
Introduction: The aim of the study was to investigate a possible CYP
3A4- inhibiting activity of long-term oral thyroxine hormone treatment.
An inhibition of the CYP 3A4 enzyme has so far only been demonstrated in vitro for triiodothyronine in cultured hepatocytes. Methods: We
performed a pilot study by using urinary 6b-hydroxycortisol and urinary
6b-hydroxycortisol/cortisol ratio as a non-invasive marker of microsomal
CYP 3A4 activity. Ten individuals, male and female, age 20 – 64 years
and taking a daily dose of thyroxine between 75 and 160 lg for at least
1 year were compared with 10 age-matched control persons. Morning
spot urine samples collected from 8:00 a.m. to 12:00 p.m. were studied
using ELISA kits from Stabiligen. Results: The mean ± SEM of the 6bhydroxycortisol/cortisol ratio was 11.19 ± 1.38 in the control group and
16.43 ± 3.52 in the thyroxine group, with no significant difference
between the 2 groups. The individual values differed by a factor of 4 in
the control group and by a factor of 9 in the thyroxine group. The ratio
of 6ß-hydroxycortisol [nmol/l] versus creatinine [mmol/l] was
163.5 ± 17.1 in the control group and 179.1 ± 21.9 in the thyroxine
group. Conclusion: The data from our pilot study confirm the marked
inter-individual variation in the morning spot urine 6b-hydroxycortisol/
cortisol ratio. They further show that it is most unlikely of thyroxine
medication being a microsomal CYP 3A4 inhibitor in vivo.
Acknowledgement: This study was supported by the Franz-Lanyar-Stiftung (project 336).
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
260
Paper No.: 2657
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
PROTEIN DISULFIDE ISOMERASE REGULATES REACTIVE
OXYGEN SPECIES GENERATION AND ANGIOTENSIN II
INDUCED CONTRACTION IN AORTA FROM WISTAR RATS
Ana Alice Dos Santos Dias(1), LL Camargo(1), GS Ceravold(1),
ACD Androwiki(1), MHC Carvsalho(1), FRM Laurindo(2),
M Janiszewski(1), LR Lopes(1)
(1) University of São Paulo, Department of Pharmacology, São Paulo,
Brazil
(2) University of São Paulo, Heart Institute (InCor), São Paulo, Brazil
Angiotensin II (Ang II) reactive oxygen species (ROS) generation is
involved in the modulation of vascular resistance. We have shown previously that the generation of ROS by Ang II can be regulated by a thiol
oxidoreductase, protein disulfide isomerase (PDI). Thus, in the present
study we investigated the role of PDI in ROS generation and Ang IIinduced aorta contraction in Wistar rats. PDI was found in all layers of
the aorta. ROS generation (dihydroethidium fluorescence) increased after
stimulation with Ang II (30 min), an effect diminished by the PDI inhibitor, bacitracin (BAC; 1mM, 30 min). The role of PDI and ROS in vasoconstriction was evaluated in concentration-effect curves to Ang II (1nM
to 1 lM) in aortic rings with (E+) and without endothelium (E-) in the
presence and absence of BAC (1mM, 15 min) and antioxidant enzymes
superoxide dismutase (SOD, 150U/ml, 30 min) and catalase (CAT;
300l/ml, 30 min). Ang II increased contraction in both E+ and E- rings.
This effect was reduced in the presence of BAC, (P < 0.01; n = 9),
SOD (p < 0.05; n = 7) and CAT (p < 0.05; n = 7). These results suggest
that PDI modulates ROS production and the contractile response to Ang
II in aortic rings. Therefore, PDI may be a new player in the control of
vascular tone.
Paper No.: 2463
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
RESULTS ABOUT THE INFLUENCE OF METFORMIN ON
THE CONCENTRATIONS OF MAGNESIUM,ZINC AND
COOPER IN NIDDM PATIENTS
Monica Daniela Dosa(1), LC Petcu(2), C Gales(3), M Nechifor(4)
(1) Constanta University Faculty of Medicine, Department of Pharmacology, Constanta, Romania
(2) Constanta University Faculty of Dentistry, Department of Biophysics
and Medical Informatics, Constanta, Romania
(3) UMF Gr.T.Popa, Department of Histology, Iasi, Romania
(4) UMF Gr.T.Popa, Department of Pharmacology, Iasi, Romania
Introduction: The aim of this study was to investigate the influence of
the treatment with metformin in NIDDM patients, who didn’t received
medication, on the concentrations of magnesium, cooper and zinc. Materials: We studied 30 newly diagnosed adults with NIDDM, in diabetes
clinic of the Faculty of Medicine,Constanta, who received metformin,
1000 mg/day, and 17 control healthy subjects. Methods.We determined
the concentrations for: glicemia, cholesterol, tryglicerides, HDL, LDL,
magnesium, copper and zinc, HbA1c, urine magnesium, copper and zinc
and intraerythrocyte magnesium, before and after 3 months of therapy.
The results were statistically interpreted. Results: Our data showed in
diabetic patients compared with healthy group: lower plasma levels in
magnesium (1.95 ± 0.19, vs 2.2 ± 0.18 mg/dl; p < 0.05) and in zinc
(67.56 ± 6.12 vs 98.41 ± 20.47 lg/dl; p < 0.05), increased levels of cooper (111.91 ± 20.98 vs 96.33 ± 8,56 lg/dl; p < 0..05) increased urinary
magnesium (237.28 ± 34.51 vs 126.25 ± 38.82 mg/24h; p < 0.05)
increased urinary zinc (1347.54 ± 158.24 vs 851.65 ± 209.75 lg/24;
p < 0.05), lower intraerhythrocyte magnesium (5.09 ± 0.63. vs 6.38 ±
0.75 mg/dl; p < 0.05). The treatment with metformin reduced significant
urinary concentration in magnesium (t = 5,35, df= 23,93; P < 0.001),
increased significant the intraerythrocyte magnesium (t= - 3,09, df= 45,
p = 0.002), and did not modify significant the urinary zinc and cooper
concentrations.There was a positive correlation between cooper and glycated haemoglobin (r = 0.517;p < 0.003) and a negative correlation
between zinc and glicemia (r= -0.399;p < 0.029). Conclusions: In diabetic patients plasma magnesium and zinc was decreased, after metformin treatment the ratio of plasma between cooper and zinc, increased in
a positive correlation with the increase of glycated haemoglobin, and the
rate of intraerythrocyte magnesium increased in a positive correlation
with the decrease of glycated haemoglobin.
Paper No.: 2311
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
NCX4040, A NITRIC OXIDE-DONATING ASPIRIN, EXERTS
ANTI-INFLAMMATORY EFFECTS THROUGH INHIBITION OF
IKB-A DEGRADATION IN HUMAN MONOCYTES
Melania Dovizio(1), E Ricciotti(2), L Di Francesco(1), P Anzellotti(1),
T Salvatore(1), A Di Francesco(1), MG Sciulli(1), G Pistritto(3),
A Monopoli(4), P Patrignani(1)
(1) ‘‘G. d’Annunzio’’ University and ‘‘G. d’Annunzio’’ University Foundation, Ce.S.I, Chieti, Italy
(2) University of Pennsylvania, Philadelphia, PA, USA
(3) University of Rome Tor Vergata Medical School, Rome, Italy
(4) NicOx Research Institute, Bresso, Milan, Italy
NO-donating aspirins consist of aspirin to which a NO-donating group is
covalently linked via a spacer molecule. NCX4040 and NCX4016 are
positional isomers with respect to the-CH2ONO2 group (para and meta,
respectively) on the benzene ring of the spacer. Because positional isomerism is critical for antitumor properties of NO-donating aspirins (Kashfi
et al., JPET, 2005, 312:978-988), we aimed to compare their anti-inflammatory effects versus aspirin in vitro. Thus, we assessed their impacts on
cyclooxygenase(COX)-2 activity(by measuring PGE2 levels), protein
expression, and cytokine generation(IL-1b, IL-18, TNF-a, and IL-10) in
human whole blood and isolated human monocytes stimulated with LPS.
Interestingly, we found that micromolar concentrations of NCX4040, but
not NCX4016 or aspirin, affected COX-2 expression and cytokine generation. We compared the effects of NCX4040 versus NCX4016 or aspirin
on IkB-a stabilization and proteasome activity in the LPS-stimulated
human monocytic cell line(THP1). At micromolar range, NCX4040
inhibited IkB-a degradation, as opposed to aspirin and NCX4016. In
fact, NCX4040 caused concentration-dependent accumulation of IkBa
and phospho-IkB-a. This effect was not reversed by 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one, an inhibitor of guanylyl cyclase, thus excluding the contribution of NO-dependent cGMP generation. In contrast,
IkB-a accumulation by NCX4040 may involve an inhibitory effect on
proteasome functions. Indeed, NCX4040 inhibited 20S proteasome activity when incubated with intact cells but not in the presence of cell lysate
supernatants, thus suggesting an indirect inhibitory effect. In conclusion,
NCX4040 is an inhibitor of IkB-a degradation and proteasome function
and it should be taken into consideration for the development of novel
anti-inflammatory and chemopreventive agents.
Paper No.: 1904
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
STATINS CONSUMPTION IN THE REPUBLIC OF CROATIA IN
2005, 2006, 2007 AND 2008
Pero Draganic, S Zezelic, V Macolic-Sairinic, S Tomic
Agency for Medicinal Products and Medical Devices (ALMP), Zagreb,
Croatia
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
261
Objectives: Drug usage evaluation is a system of continuous, systematic,
criteria-based drug evaluation that ensures the appropriate use of drugs.
Statins are drugs used to prevent cardiovascular diseases. The importance
of collecting data on statins consumption is in correlation with the
improvement on cardiovascular diseases therapy. Methods: ALMP Croatia collected and processed data on consumption of the medicines, and
the usage between 2005-2008 was processed by the DDD/1000inh/day
and ATC classification, and analyzed according to the financial indicators. Results: The percentage of statins (C10AA group) within total
financial resources indicates: in 2005 5.20%, in 2006 6.58%, in 2007
8.17%, and in 2008 8.22%. By processing consumption data as DDD,
between 2005-2008 within C10AA group shows 35.62, 45.77, 67.89 and
68.37 DDD/1000inh/day, respectively. The individual expenditure of statins indicates: simvastatin 20.23, 20.87, 23.5, and 23.77 DDD/1000inh/
day in 2005-2008 periods; atorvastatin 13.03, 20.73, 39.86 and 36.89
DDD/1000inh/day in 2005-2008 periods. Conclusion: Between 20052008 all statins obtained in the Croatian market showed a continuous
increase in DDD/1000inh/day as well as in financial costs. For the entire
period atorvastatin and simvastatin were the most prescribed drugs. The
overall consumption of statins in Croatia (35-67x DDD/1000inh/day) is
lower than in EU and other Western countries, where a consumption of
statins is about 80-200 DDD/1000inh/day.
Paper No.: 2769
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
INTERACTION BETWEEN CICLETANINE AND
PRECONDITIONING IN CONCIOUS RABBITS WITH INSULIN
RESISTANCE
László Drimba, B Peitl, T Patonay, Z Szilvássy
University of Debrecen, Department of Pharmacology and
Pharmacotherapy and Pharmapolis Debrecen Ltd, Debrecen, Hungary
Cicletanine, a cyclic GMP phosphodiesterase inhibitor has been shown
to potentiate the anti-ischaemic effect of preconditioning. It has also been
shown that hypercholesterolemia blocks preconditioning in conscious
rabbits. The aim was therefore to asses if cicletanine restores the rapid
pacing (RP)-induced preconditioning phenomenon in insulin resistant
rabbits. Right ventricular electrode and left ventricular polyethylene catheter-instrumented conscious rabbits were subjected to two consecutive 5min periods of RP with an interpacing period of 5 minutes. The 1st RP
served as an ischemic challenge and to induce preconditioning, the 2nd
one was to test the preconditioning effect. The resulting intracavital STsegment elevation and increase in left ventricular end-diastolic pressure
(LVEDP) were measured after pacing periods. Both variables were significantly reduced after the 2nd RP compared to those measured after the
1st one. Cicletanine (20 mg/kg) applied intravenously 20 min prior to
the 1st preconditioning attenuated both postpacing ST-segment elevation
and LVEDP-increase and the protection was further amplified by preconditioning. The same animals were then made insulin resistant by partial
hepatic sensory denervation by perineurial treatment with 2% capsaicin
solution around the anterior hepatic plexus. This decreased whole body
insulin sensitivity 2 weeks later. The development of insulin resistance
resulted in the loss of the RP-induced preconditioning effect on both
parameters, nevertheless, cicletanine was found to significantly reduce
both ST elevation and LVEDP-increase produced by the 1st preconditioning. This protection also was amplified by preconditioning. We conclude that CIC produces an anti-ischemic effect and preserves PC in
conscious rabbits with neurogenic insulin resistance.
Paper No.: 2885
FOCUSED CONFERENCE GROUP: P04 - PHARMACOEPIDEMIOLOGY, CURRENT CONTROVERSIES AND OPPORTUNITIES
MANAGEMENT OF ACUTE MYOCARDIAL INFARCTION IN
FRANCE: RESULTS FROM INCLUSION OF A NATIONAL
COHORT
C Droz-Perroteau(1), C Dureau-Pournin(1), D Thomas(2),
N Danchin(3), J Tricoire(4), J Bénichou(5), F Paillard(6),
P Ducimetière(7), S Hercberg(8), H Maı̈zi(1), E Guiard(1),
M-A Bernard(1), P Blin(1), Nicholas Moore(1)
(1) Université de Bordeaux, INSERM CIC 0005, Bordeaux, France
(2) Hôpital Pitié-Salpétrière, Paris, France
(3) Hôpital Européen Georges Pompidou, Paris, France
(4) Cardiologist, Toulouse, France
(5) CHU de Rouen-INSEM U657, Rouen, France
(6) CHU de Pontchaillou, Rennes, France
(7) INSERM, Villejuif, France
(8) INSERM U557-INRA-CNAM-CRNH, Bobigny, France
This study was performed at the request of the Health Authorities to
assess the impact of recommended cardiovascular treatments in secondary prevention after acute myocardial infarction (AMI) in real-life practice. A national cohort study was designed to include 5000 recent AMI
(<3 months) recruited by hospital and non-hospital cardiologists. At
inclusion, a standardized questionnaire was used to collect socio-demographic data, AMI characteristics, cardiovascular risk factors, history and
associated cardiovascular diseases, the last biological balance sheet, cardiovascular drugs used, prescription of cardiovascular rehabilitation program. Between May 2006 to June 2009, 5540 patients were included
(3306 by 346 hospital cardiologists and 2234 by 389 non-hospitals);
77.6% were male, mean age was 62.1 years. It was the first AMI for
86.7%, 70.5% had the 3 AMI criteria (symptomatic, electrical, enzymatic), 8.2% a LVEF <40%. Concerning the cardiovascular risk factors,
9.6% were current smokers, 16.7% had diabetics, 44.6% had history of
hypercholesterolemia, 43.6% history of arterial high blood pressure. The
4 recommended cardiovascular drugs in secondary prevention were prescribed to 70.5% (73.7% of hospital cardiologists and 65.8% of non-hospitals): aspirin or other anti-platelet agents to 99.3% (99.4% of hospital
cardiologists and 99.2% of non-hospitals), statins or others hypolipidemic drugs to 96.1% (96.6%, 95.3% respectively), b-blockers to 88.9%
(90.5%, 86.5% respectively) and IEC or ARA II to 80.7% (82.9%,
77.5% respectively). A cardiovascular rehabilitation program at inclusion
was prescribed for 40.5% (46.6%, 31.5% respectively). This study shows
that the 4 recommended cardiovascular drugs in secondary AMI prevention are now widely used in France.
Paper No.: 2080
FOCUSED CONFERENCE GROUP: P19 - GENERAL SESSION HEPATOLOGY
EFFECT OF SIRNA-MEDIATED KNOCKDOWN OF ALDH2 ON
GTN-INDUCED CGMP FORMATION IN PK1 CELLS
Yohan D’Souza, BM Bennett
Queen’s University, Department of Pharmacology & Toxicology, Kingston, Ontario, Canada
Recent studies suggest a primary role for aldehyde dehydrogenase 2
(ALDH2) in mediating the biotransformation of organic nitrates, such as
glyceryl trinitrate (GTN), to the proximal activator of soluble guanylyl
cyclase (sGC), resulting in increased cGMP accumulation and vasodilation. In studies using stably transfected porcine renal epithelial cells
(LLC-PK1 cells) overexpressing ALDH2, we found that overexpression
of ALDH2 did not significantly alter GTN-induced cGMP accumulation
compared to wild type cells. However, it was possible that GTN bioactivation mediated by endogenous ALDH2 was maximal in wild type cells,
and that overexpression of ALDH2 had no further impact on GTN bioac-
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
262
tivation. Accordingly, we used siRNA-mediated knockdown of ALDH2
to examine the effect of decreased ALDH2 expression on GTN-induced
cGMP formation. PK1 cells treated for 72 hours with 50 nM of two siRNAs directed against ALDH2 resulted in a greater that 95% reduction in
ALDH2 protein. Exposure of cells to 1 uM GTN for 3 minutes resulted
in a 5-6 fold increase in cGMP accumulation, with no differences in the
GTN-induced cGMP response in wild type cells compared to cells treated with the two siRNAs or an siRNA negative control. Preincubation of
cells with 10 lM GTN for 2 hours resulted in a similar inhibition of the
cGMP response on subsequent exposure to GTN (‘GTN tolerance’) in all
treatment groups. Together, these results indicate that ALDH2 does not
mediate the mechanism-based biotransformation of GTN to activators of
sGC, and that GTN tolerance can occur in the absence of ALDH2.
Paper No.: 1675
FOCUSED CONFERENCE GROUP: PW12 - EXPERIMENTAL
PAIN METHODS IN DEVELOPMENT AND VALIDATION OF
ANALGESICS
COMPARISON OF THE PD/PK OF ROTUNDINE CRYSTAL
FORM A AND C IN MICE
Lida Du(1), X Li(1), J Ying(2), X Bai(2), S Yang(2), H Zhang(2),
G Wang(1), Y Lu(2)
(1) China Pharmaceutical University, School of Pharmacy, Nanjing, PR
China
(2) Institute of Materia Medica, Chinese Academy of Medical Science,
Nanjing, PR China
Rotundine is isolated from Chinese traditional herb, and have been used
as analgesics clinically for decades. But the absorption of rotundine in
gastrointestinal tract is rare and unstable. We researched the crystal forms
of rotundine and investigated their relationship of PD/PK in mice. The
crystal forms of rotundine were prepared by the physical methods and
tested by X-ray diffraction, DSC and infraredspectrum. The analgesic
effects of rotundine were tested by the methods of hot plate and writhing
induced by acetic acid in mice. The concentrations of rotundine in blood
were measured by the method of HPLC. The results show that the crystal
form C is a new one and has high absorption ratio in mice. The number
of writhing induced by acetic acid in mice from 36.67 + /-10.19
decreased to 27.00 + /- 13.39 and 21.17 + /- 13.73 for form A and C
respectively when administrated 200 mg/kg orally. The Cmax for the
form A, C are 4.8872 (mg/L) and 5.5410 (mg/L), the Tmax(min) for two
different forms are 45.0 min(form A) and 75.0 min (form C). Compared
the results of PD and PK of rotundine in mice, it can be concluded that
the new crystal form C of rotundine is a optimal form for the drug preparation with a good absorption and exert better analgesic effects than other
forms.
Acknowledgment: this work was supported by the National Major project No:2009ZX09302-003 and basic science and techniques research
project: No: 2008IM022200
Paper No.: 1730
FOCUSED CONFERENCE GROUP: P06 - THE HEART GONE
WRONG; STABILIZATION OF CARDIAC FUNCTION - BASIC
EFFECTS OF ULTRA-LOW DOSES OF PERORAL ANTIBODIES
TO ANGIOTENSIN II RECEPTOR IN NON-INVASIVE AND
INVASIVE RAT MODELS OF HEART FAILURE
Julia Dugina, IA Kheyfets, SA Sergeeva, OI Epstein
Materia Medica Holding Company, Department of Research & Development, Moscow,Russian Federation
failure:isoproterenol-induced (ISO) HF and HF induced by permanent
coronary occlusion(CO). In ISO HF male and female Wistar rats
(220-250 g) were administered per osdaily with cardosten (2.5 ml/kg),
losartan (20 mg/kg) for 28 days starting onday 7 after twice repeated sc
injection of isoproterenol (80 mg/kg). In CO HFmodel male Wistar rats
(240-340 g) were treated per os daily with vehicle,cardosten (7.5 ml/kg),
losartan (20 mg/kg) for 30 days starting one day beforecoronary occlusion. In ISO HF cardosten was considerably better than placebo andalmost as effective as losartan in improving exercise performance
(a tendencyin males, and significant changes by 27.8% and 34.5% vs
baseline respectively infemales), cardiac output, LV function. In CO HF
model cardiac output measuredboth in vivo and in vitro is tended to
increase by cardosten (32.9% and 47.4 vsvehicle respectively), while losartan had no any noticeable effect. Totalperipheral resistance was
decreased by both of losartan and cardosten (p < 0.05).Increase in stroke
volume in cardosten (42% vs vehicle) and aortic flow (60.3% 66.3% vs
vehicle) was registered. Heart rate was not changed in the all groupsas
compared to control. In summary, cardosten had comparable beneficial
effectsto that of losartan in two models of CHF.
Paper No.: 758
FOCUSED CONFERENCE GROUP: P06 - THE HEART GONE
WRONG; STABILIZATION OF CARDIAC FUNCTION
EFFECT OF ATORVASTATIN ADDITION TO CARDIOPLEGIC
SOLUTION ON MYOCARDIAL PROTECTION IN ISOLATED
RAT HEART
Deniz Kaleli Durman(1), M Agirgol(1), M Kucur(2), F Basinoglu(3),
O Ozdemir(1), BS Uydes-Dogan(1)
(1) Istanbul University, Faculty of Pharmacy, Department of
Pharmacology, Istanbul, Turkey
(2) Istanbul University, Cerrahpasa Medical Faculty, Fikret Biyal Central
Research Laboratory, Istanbul, Turkey
(3) Haseki Training and Research Hospital, Biochemistry Laboratory,
Istanbul, Turkey
Cardioplegia is a selective cardiac arrest technique, which aims to prevent myocardial damage and provide adequate heart hemodynamics in
cardiac surgery. For minimising myocardial damage, various agents
accepted to be cardioprotective are added to cardioplegic solution. Statins
are reported to have protective effects in myocardial ischemia-reperfusion
(I/R) injury independently from their lipid-lowering effects. Herein, we
aimed to investigate the effect of atorvastatin addition to cardioplegic
solution on myocardial protection. Isolated rat hearts perfused in Langendorff system at constant pressure (80mmHg) were subjected to 40min.
ischemia/60min. reperfusion. Cardioplegia was administered 3min.
before ischemia by St.Thomas2 cardioplegic solution (+4C) at a constant flow (10ml/min). Atorvastatin was added to cardioplegic solution at
increasing concentrations (5x10-6M, 10-5M, 5x10-5M). Cardiac function
(left ventricular developed pressure, heart rate, double-product), coronary
hemodynamics (coronary flow, coronary vascular resistance) and myocardial necrosis (creatine kinase, troponin-T, necrotic area) parameters
were evaluated and compared with cardioplegia. For investigating the
mechanism of action, atorvastatin (10-5M) was studied in the presence
of PI3 kinase inhibitor, wortmannin (10-7M). When compared to cardioplegia, addition of atorvastatin to cardioplegic solution significantly
enhanced postischemic cardiac recovery, whereas no change was
observed in coronary hemodynamics. Furthermore, cardiac enzyme levels and infarct area were also significantly reduced. Wortmannin inhibited
beneficial effects of atorvastatin on cardioplegia. These findings demonstrated that, atorvastatin strengthens protective effect of cardioplegia via
PI3 kinase related mechanism which may provide an additive clinical
benefit in minimising myocardial damage during cardiac surgery.
The present work was supported by the Research Fund of Istanbul University. Project No. T-1892.
Ultra-low doses of antibodies to C-terminal of AT1 receptor to angiotensin II(cardosten) is a novel antibody-based therapeutic for the treatment
of chronicheart failure (CHF). The drug was tested in rat models of heart
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
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Paper No.: 2776
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
CROSSTALK BETWEEN GABAB AND GROUP III MGLU
RECEPTORS FOLLOWING INTRANIGRAL DRUG
ADMINISTRATION IN THE RESERPINE-TREATED RAT
Susan Duty, M Broadstock
King’s College London, Wolfson Centre for Age-Related Diseases,
London, UK
Cross talk between Class C GPCRs has received much attention at the
molecular level, though little is known about this phenomenon in vivo.
Here, we examined whether crosstalk between group III mGlu and
GABAB receptors was seen in vivo using reversal of reserpine-induced
akinesia as our measure. Male rats, cannulated above the substantia nigra
pars reticulata (SNpr) were rendered akinetic with reserpine (5 mg kg-1
s.c.). 18h later, rats received a unilateral injection of either L-SOP (group
III agonist; 750 nmol in 2.5 ll PBS), baclofen (GABAB agonist; 800ng
in 0.5 ll PBS) or vehicle into the SNpr. 4h later rats received a second
injection of either L-SOP or baclofen (doses as above). Contraversive
360o rotations were quantified for up to 60 min following each injection.
Data are expressed as mean ± s.e.m (n = 6). The rotational responses to
L-SOP and baclofen were reduced following second injection of the
same agonist (P < 0.05; paired t-test between injections). The response
to L-SOP was also significantly reduced following initial injection with
baclofen (7 ± 3 rotations 30 min-1 versus 30 ± 6 after initial vehicle
injection; P < 0.05; unpaired t-test). In contrast, the response to baclofen
was significantly enhanced following initial injection with L-SOP
(329 ± 29 rotations 60 min-1 versus 106 ± 4 rotations following initial
vehicle injection; P < 0.05; unpaired t-test). These data confirm that crosstalk occurs in vivo between group III mGlu receptors and GABAB
receptors but that the consequences of this can produce opposite effects,
depending on the order of receptor activation.
Paper No.: 2492
FOCUSED CONFERENCE GROUP: P14 - ADDICTION AND
DOPING: NEUROBIOLOGICAL AND CLINICAL
RUSSIAN BIOSIMILAR EPOETINS AND THEIR POSSIBLE
USE IN SPORT
Yuliya I Dykhal, GI Krotov, PV Postnikov, GM Rodchenkov
FGUP Antidoping Centre, Department of Peptide Doping Analysis, Moscow, Russian Federation
Biopharmaceuticals have revolutionized the treatment of many diseases.
The advent of recombinant erythropoietins has greatly benefited patients
with anemia related to chronic kidney disease and cancer. In parallel they
became potential agents for use in sports as doping due to their similarity
to endogenous EPOs. Recent expire of the patent restriction leads to
development of new versions of these products that may stimulate
demand and reduce prices. For now several biosimilar epoetins are available on Russian biopharmaceuticals market: Epocrine, Erythrostime,
Vero-epoetine, Eralphone and some illegal forms. These epoetins were
studed for possible differences in erythropoietin content, potency, possible
immunogenesity and isoform distribution. For those purposes different
methods were used. For isoform distribution isoelectric focusing with
dobble-blotting were performed, and differences in molecular mass were
shown in SDS-electrophoresis study. Total protein was determined by
Bradford assay meantime concentration of EPO was determined by ELISAs and LIA. Some urine samples which contain mentioned epoetines
were analysed and they shown that results sometimes are very complicated and almost impossible to interpret as positive case following identification criteria set in doping control analysis even if it’s very likely that
athlet abused epoetine. The study shows that epoetin products from different manufacturers differ in composition. Some of the products failed to
meet their own specifications, indicating that some of the manufacturers
do not have adequate quality control over their production process. This
could result in undesired clinical effects and even lead to lethal outcome.
Paper No.: 1747
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
TAURINE AND ITS ANALOGUE TAUROPYRONE:
COMPARATIVE PHARMACOLOGICAL ACTIVITIES
Zane Dzirkale(1), J Pupure(1), J Rumaks(1), M Vanina(1), S Svirskis(1),
R Mezhapuke(2), MA Fernandes(3), G Duburs(2), R Muceniece(1),
V Klusa(1)
(1) University of Latvia, Department of Pharmacology, Riga, Latvia
(2) Latvian Institute of Organic Synthesis, Riga, Latvia
(3) University of Coimbra, Faculty of Sciences, Department of Zoology,
IMAR-CMA, Coimbra, Portugal
Taurine, a sulphur-containing amino acid, is characterized by its hydrophilicity and poor absorption. Tauropyrone, a taurine-containing 1,4-dihydropyridine derivative, is suggested to have a higher activity than that
of taurine due to the lipophylic cyclic 1,4-dihydropyridine (DHP) moiety
that might serve also as a carrier element capable of facilitating the compound’s delivery to cell targets. In the present study, we compared the
effects of taurine and tauropyrone in in vitro experiments: binding to
GABA-A receptor, and influence on mitochondrial processes; and in in
vivo tests: influence on behavioral effects caused by GABA-A receptor
ligands bicuculline, diazepam and ethanol. Our results demonstrated: 1)
unlike taurine, tauropyrone lacked GABA-A receptor binding affinity,
and did not influence bicuculline convulsant activity; 2) neither taurine
nor tauropyrone influenced mitochondrial processes, and did not alter
diazepam myorelaxing action; 3) tauropyrone showed more pronounced
‘‘anti-ethanol’’ effect (shortening of ethanol-sleeping time). One may suggest that tauropyrone molecule due to its DHP moiety has a hindered
access to GABA-A receptor GABA site, but the enhanced modulatory
influence on ethanol-mediated cellular events.
Acknowledgements: ESF Nr.2009/0217/1DP/1.1.1.2.0/09/APIA/VIAA/
031, Latvian Council of Science Grant Nr.05-1418; ESF 2004/3; COST
D34.
Paper No.: 2867
FOCUSED CONFERENCE GROUP: P07 - SIMULATION AND
DATA MODELLING IN DRUG DEVELOPMENT.
MODELLING OUTCOMES OF ANTIBIOTIC THERAPY BY
DISCRETE EVENT SIMULATION (DES) OR
MULTI-AGENT-BASED SIMULATION (MABS) OF
NOSOCOMIAL PNEUMONIA
Mario Eandi(1), P De Luca(1), L Pradelli(2), S Iannazzo(2)
(1) University of Turin, Department of Anatomy, Pharmacology and
Forensic Sciences, Torino, Italy
(2) Adres srl - Turin, Torino, Italy
Discrete-event-simulation (DES) and multi-agent-based-simulation
(MABS) are two approaches to Modelling complex processes. DES
micro-simulates therapeutic pathways of individuals and is increasingly
used in cost-effectiveness studies. MABS is a new scientific instrument
well-suited to study how interactions of autonomous agents (e.g. microorganisms-host-antibiotic) driven by a simple local rule can generate the
emergence of global outcomes from the bottom up. We developed a
DES and a MABS model to analyze outcomes of nosocomial pneumonia
(NP) treated with carbapenems. Both models predict individual outcomes
by simulating in vivo microbiological efficacy according to PK-PD relationships. To evaluate the clinical results, the DES model integrates the
surrogate parameter ‘‘time-supra-MIC’’ (%T>MIC) with the baseline risk
of death. The MABS model evaluates outcomes by replicating the
dynamics of pathogen (replication and death rates) as influenced by host
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
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and antibiotic, and of patients (healing or death rate, resistance) as influenced by pathogen and antibiotic. Outcomes predicted by both models
compare well to those registered in comparative trials in response assessment at end of treatment (EOT), test-of-cure (TOC), and late-follow-up
(LFU). Despite non-inferiority results in cure and death rates, results
obtained from DES model showed a superior effect of doripenem over
comparators in a number of areas including seizures, emerging resistance, length of stay (LOS) in general ward and in ICU on mechanical
ventilator. The analysis performed by MABS model lead to similar conclusions. Yet, MABS Modelling is even more flexible and is suitable to
find the optimal solution to critical drug management by computer-experimenting virtual reality.
Paper No. 3393
FOCUSED CONFERENCE GROUP: P11 - G
PROTEIN-COUPLED 7TM RECEPTORS: FROM MOLECULAR
TO PHYSIOLOGICAL FUNCTION
FUNCTIONAL EXPRESSION OF CB1 RECEPTORS IN RAT
BRAIN NUCLEAR FRACTIONS
Leire Echeazarrra(1), S Barrondo(1), GG del Caño(2), M Montaña(1),
ML de Jesús(1), J Sallés(1)
(1) Departamento de Farmacologı́a. Facultad de Farmacia (UPV/EHU);
Centro de Investigación Biomédica en Red de Salud Mental. CIBERSAM, Vitoria-Gasteiz, Spain
(2) Departamento de Neurociencias, Facultad de Farmacia (UPV/EHU).
Vitoria-Gasteiz, Spain
The aim of the present work was to study the presence of CB1 receptors
in rat brain nuclear fractions and its functional coupling to Gi/o proteins.
To this aim, inmunohistochemistry, double inmunofluorescence, immunoblotting, and [3H]SR141716A and GTPc[35S] binding assays were
performed. Inmunohistochemistry revealed the classical presynaptic profile of CB1 distribution together with a nuclear inmunolabelling. Double
inmunofluorescence of CB1 receptors and lamin B1 (a marker of the
nuclear envelope) confirmed the nuclear localization of CB1. Employing
enriched nuclear preparations, the nuclear location of CB1 receptors and
their downstream signalling partners Gi/o was also demonstrated by western blot analysis. The selective CB1 antagonist radioligand,
[3H]SR141716A, recognized a specific population of binding sites in
nuclear membranes, with a density (Bmax of 1,04 ± 0,24 pmol/mg) and
affinity (KD of 1,6 ± 0,5 nM) similar to those obtained in cytoplasmatic
membranes. In order to evaluate the functional coupling of CB1 receptors GTPc[35S] binding assays were performed in nuclear membranes.
The cannabinoid agonist WIN55,212-2 was able to stimulate the
GTPc35S] binding with and efficacy (Emax of 89,5 ± 7.3% over basal)
and a potency (EC50 of 0,63 ± 0,06 lM) in the same order as those
obtained in cytpolasmatic membranes.Our results demonstrate the functional expression of CB1 receptors in nuclear fractions. Due to the role
of these receptors in the regulation of cell proliferation, survival and
apoptosis, our results open and interesting subject for future studies about
the functional responses specifically mediated by nuclear CB1 receptors.
Funding: SAIOTEK S-PEO8UN29 and SE-PE07UN21, UPV/EHU
NUPV08/05 and CIBERSAM.
Paper No.: 642
FOCUSED CONFERENCE GROUP: P02 - TRANSMEMBRANE
TRANSPORT: PERSPECTIVES FOR DISEASE AND DRUG
DISCOVERY
EXTRACELLULAR LOOPS 2 AND 4 OF GLYT2 ARE
REQUIRED FOR N-ARACHIDONYL-GLYCINE INHIBITION
OF GLYCINE TRANSPORT
Concentrations of glycine are regulated via the Na+/Cl--dependent glycine transporters, GLYT1 and GLYT2. N-Arachidonyl glycine (NAGly)
is an endogenous inhibitor of GLYT2 with no effect on GLYT1. Understanding the molecular basis of NAGly interactions with GLYT2 may
allow for the development of novel analgesics. We investigated whether
extracellular loops 2 and 4 (EL2/4) are important for NAGly sensitivity
between GLYT1 and GLYT2 and also a series of related N-arachidonylamino acids. Chimeras were constructed between GLYT1 and GLYT2
with their EL2 and/or EL4 regions switched. Point mutations of all
GLYT2 EL4 residues which differed from GLYT1 were mutated to the
corresponding residue in GLYT1. Transporters were tested for their
sensitivity to GLYT2 inhibitors: NAGly, N-arachidonyl-L-alanine (NALAla), N-arachidonyl-D-alanine (NADAla) and N-arachidonyl-c-aminobutyric acid (NAGABA) using electrophysiology (n ‡ 5/transporter).
GLYT2 is inhibited (and not GLYT1) by NAGly, NADAla and NAGABA whereas GLYT2(GLYT1EL2) and GLYT2(GLYT1EL4) had
reduced sensitivities. Interestingly, GLYT2 and GLYT2(GLYT1EL2) are
inhibited by NALAla whereas GLYT2(GLYT1EL4) is not. GLYT2R531L
and GLYT2K532G had reduced sensitivity to NAGly (IC30; 13 ± 2lM;
9 ± 1lM, respectively) compared to GLYT2 (IC30: 3.4 ± 0.6lM) while
GLYT2I545L had markedly reduced sensitivity to NAGly (IC30:
>30lM). GLYT2R531L, GLYT2K532G and GLYT2I545L also had
reduced sensitivities to NALAla (IC30: 14 ± 1lM; 12 ± 1lM; and
21 ± 1lM, respectively) compared to GLYT2 (IC30: 5.9 ± 0.7lM). In
conclusion, EL2 and EL4 of GLYT2 are important in the inhibition of
GLYT2 by NAGly, NADAla and NAGABA while only EL4 of GLYT2
is required for NALAla inhibition of transport. Key residues in GLYT2
EL4 required for NAGly and NALAla sensitivity are R531, K532 and
I545.
Paper No.: 1806
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
EVIDENCE FOR FUNCTIONAL TWO-PORE DOMAIN
POTASSIUM CHANNELS (K2P) IN RAT MESENTERIC
ARTERIES
Gillian Edwards, E Porter, N Ashton, A Gurney, A Weston, I Johnson
University of Manchester, Faculty of Life Sciences, Manchester, UK
The K2P channels TREK and TRAAK are activated by the polyunsaturated fatty-acid (PUFA) arachidonic acid (AA) and are present in rat mesenteric arteries (Gardener et al., Br J. Pharmacol. 2004; 142:192-202).
Although Blondeau et al., (Circ Res. 2007; 101:176-84) showed PUFAs
activate K2P channels in murine arteries, their functional role in rat arteries is unknown. In our study, isolated Wistar rat (225-250g) mesenteric
arteries were pre-contracted with U46619 and then exposed to AA (1100microM) in the absence and presence of chlorpromazine (TREK/
TRAAK inhibitor; 1microM). No differences were detected between
AA-induced relaxation log EC50 values in the absence and presence of
chlorpromazine (-5.4 ± 0.1 vs -4.9 ± 0.2, n = 4, respectively). However,
in the continuing presence of the nitric oxide synthase inhibitor
L-nitroarginine (L-NOARG, 100microM) and the cyclo-oxygenase
inhibitor indomethacin (INDO, 10microM), the AA log EC50 value was
shifted significantly by chlorpromazine at 1microM (-5.1 ± 0.05, AA
with L-NOARG/INDO vs. -4.7 ± 0.05, AA + chlorpromazine with
L-NOARG/INDO, p < 0.0001; n = 4) and 10microM (-5.2 ± 0.07, AA
with L-NOARG/INDO vs. -4.6 ± 0.2, AA + chlorpromazine with
L-NOARG/INDO, p = 0.0053; n = 3). These data suggest that in the
absence of relaxing factors such as nitric oxide and endogenous cyclooxygenase products, activation of K2P channels by PUFAs generates
measurable mechano-inhibitory effects in rat mesenteric arteries.
Work funded by the British Heart Foundation.
Amelia Edington, R Ryan, R Vandenberg
University of Sydney, School of Medical Sciences, Discipline of Pharmacology, NSW, Australia
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
265
Paper No.: 2976
FOCUSED CONFERENCE GROUP: P06 - THE HEART GONE
WRONG; STABILIZATION OF CARDIAC FUNCTION
DISPROPORTIONALLY IMPAIRED MICROVASCULAR
STRUCTURE IN PATIENTS WITH VERY MILD ESSENTIAL
HYPERTENSION
Ashkan Eftekhari(1,2), ON Mathiassen(1,2), NH Buus(2,3), O Gotzsche(2), MJ Mulvany(2), KL Christensen(1,2)
(1) Aarhus University, Department of Pharmacology, Aarhus, Denmark
(2) Aarhus University Hospital, Department of Medicine and Cardiology
A, Aarhus, Denmark
(3) Aarhus University Hospital, Department of Renal Medicine, Aarhus,Denmark
Essential hypertension (EH) is characterized by reduced vasodilatory
capacity of the resistance vasculature. The purpose of our study was to
investigate if coronary flow reserve (CFR) and minimum forearm vascular resistance (Rmin) are affected proportionally to the increased blood
pressure (BP). 75 previously untreated EH patients (age 48 yr, 60%
males) without previous cardiovascular disease and with 24-h systolic
BP (SBP) l 130 mmHg or diastolic BP (DBP) ¡Ý 80 mmHg were
assigned into two groups: either Very Mild EH (125/79 mmHg) or MildModerate EH (145/89 mmHg). Twenty five healthy normotensive
controls were used . CFR was measured with trans-thoracic echocardiography, forearm Rmin was measured using plethysmography. Systemic
vascular resistance index (SVRI) was calculated from cardiac output
measurements using a gas rebreathing technique. Compared to control,
24-h mean BP was raised 15% in Very Mild EH and 29% in Mild-Moderate EH. CFR was decreased by 23% in Very Mild EH and 29% in
Mild-Moderate EH, respectively. Rmin was elevated by 49% in Very
Mild EH and 75% in Mild-Moderate EH, as compared to control persons. Thus both CFR (P < 0.01) and Rmin (P < 0.01) were altered more
than could be expected from the raised BP level in Very Mild EH. In
contrast, SVRI increased proportionally to the increased BP (19%, Very
Mild EH; 33%, Mild-Moderate EH). In patients with very mild EH,
structural remodelling of the myocardial and the forearm resistance
vasculature seems disproportionally impaired compared to that expected
from the modest rise in BP.
Paper No.: 915
FOCUSED CONFERENCE GROUP: PW12 - EXPERIMENTAL
PAIN METHODS IN DEVELOPMENT AND VALIDATION OF
ANALGESICS
NEUROTROPIN RELIEVES THE OXALIPLATIN-INDUCED
PERIPHERAL NEUROPATHY BY INHIBITING
NEURODEGENERATION
Nobuaki Egashira, T Kawashiri, H Watanabe, Y Ikegami, T Yano,
H Ikesue, R Oishi
Kyushu University Hospital, Department of Pharmacy, Fukuoka, Japan
Oxaliplatin is a key drug for colorectal cancer, but it causes acute peripheral neuropathy (triggered by cold) and chronic neuropathy (sensory and
motor dysfunction). Oxaliplatin is metabolized to oxalate and dichloro(1,2-diaminocyclohexane)platinum [Pt(dach)Cl2]. Recently, we demonstrated the involvement of oxalate in oxaliplatin-induced cold
hyperalgesia but not mechanical allodynia, and preventive effect of preadministration of Ca2 + or Mg2 + on the cold hyperalgesia in rats (Sakurai
et al, Pain 2009; 147: 165-174). Neurotropin, a non-protein extract from
the inflamed rabbit skin inoculated with vaccinia virus, has been used to
treat various chronic pains. We previously reported that repeated administration of neurotropin reverses the paclitaxel-induced neuropathy without affecting anticancer activity in rats (Kawashiri et al, Eur J Cancer
2009; 45: 154-163). In the present study, we investigated the effect of
neurotropin on the oxaliplatin-induced neuropathy in male Sprague-Dawley rats. Repeated administration of oxaliplatin (4 mg/kg, i.p., twice a
week) caused cold hyperalgesia (acetone test) in the acute phase and
mechanical allodynia (von Frey test) in the chronic phase. Repeated
administration of neurotropin relieved the oxaliplatin-induced mechanical
allodynia but not cold hyperalgesia, and inhibited the oxaliplatin-induced
axonal degeneration in rat sciatic nerve. Neurotropin also inhibited the
oxaliplatin-induced neurite degeneration in cultured PC12 and rat dorsal
root ganglion (DRG) cells. On the other hand, neurotropin did not affect
the oxaliplatin-induced cell injury in rat DRG cells. These results suggest
that repeated administration of neurotropin relieves the oxaliplatininduced mechanical allodynia by inhibiting the axonal degeneration and
it is useful for the treatment of oxaliplatin-induced neuropathy clinically.
Paper No.: 2807
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
USAGE OF AND ATTITUDES TOWARDS THE DRUG-DRUG
INTERACTION DATABASE SFINX – A QUESTIONNAIRE
STUDY WITHIN SWEDEN
Birgit Eiermann(1), M Andersson(2), M-L Ovesjö(2), P Bastholm(1),
A Veg(1), M Edlert(1), Y Böttiger(2)
(1) Stockholm County Council, Department of Drug Management and
Informatics, Stockholm, Sweden
(2) Karolinska Institutet, Division of Clinical Pharmacology, Department
of Laboratory Medicine, Stockholm, Sweden
Introduction: Drug-drug interactions (DDI:s) are a common cause for
adverse drug events or reduced drug effect. Therefore avoidance of
DDI:s through usage of a decision support system for DDI:s can improve
quality of care. The DDI database SFINX, developed and used in Sweden and Finland is available through a web application, presenting colour
coded alerts when entering a drug or substance list. Clicking on the alert
button will lead to a short description of the medical consequence of a
DDI followed by useful clinical recommendations, mechanism and background information. Method: In a questionnaire study 11 763 registered
users of SFINX were asked about their use of SFINX, its influence on
decision making and the perceived benefits. Results: 2707 (23%) answers
from physicians (43%), pharmacists (18%) and other health care personnel (20%) were received. The majority of physicians and pharmacists
used the database during patient contact (60 resp. 50%). 75% of the users
always read the alert messages instead of just reacting to a warning signal. Over 56% of the physicians take actions receiving a DDI alert such
as change of drug therapy and informing the patients. The most important benefits using SFINX are that physicians learn about DDIs (77%)
and that they receive confirmation of the patients drug lists regarding
interactions (73%). Conclusion: DDI databases with focus on concise
and short recommendation texts are seen as useful tools to support physicians‘ decision making process during drug prescribing and to increase
their knowledge about DDI:s.
Paper No.: 1878
FOCUSED CONFERENCE GROUP: P14 - ADDICTION AND
DOPING: NEUROBIOLOGICAL AND CLINICAL
TESTOSTERONE INHIBITS ENDOTHELIAL NITRIC OXIDE
SYNTHASE MRNA EXPRESSION AND NITRIC OXIDE
PRODUCTION
Lena Ekström, C Skogastierna, A Rane
Karolinska Institute, Department of Laboatory Medicine, Stockholm,
Sweden
Background: Accumulating evidence indicates that abuse of anabolic
androgenic steroids (AAS) cause cardio-vascular adverse effects and disease. One early event in cardiovascular diseases is endothelial dysfunction. Several vasoactive mediators are released from the endothelium,
including the vasodilator nitric oxide (NO) which is synthesized by
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
266
endothelial nitric oxide synthase (eNOS). Surprisingly, there are no studies available on how AAS affect the endothelial metabolism of NO.
Aims: To investigate the effects of testosterone on the endothelial production of NO in vitro and in vivo. Methods: Testosterone (500 mg) was
administrated as a single dose to healthy volunteers and the morning
urine was collected prior to and 2 days after injection. The urinary excretion level of NO was assessed by a colorimetric assay. Moreover, testosterone was added to the medium of a human endothelial cell-line and
eNOS mRNA was quantified by real-time PCR. Results: The in vivo
results show that the urinary NO level significantly decreased by 25%
two days after testosterone administration. The in vitro studies show that
testosterone inhibits the eNOS mRNA expression after 48 hours. When
the antioxidant selenomethionine was included, the eNOS mRNA
expression was not inhibited by testosterone, indicating that the downregulation of eNOS may partly be induced by oxidative stress. Conclusion: These results show that a supra-physiologic dose of testosterone
decreases the expression of eNOS and consequently the formation of
NO, indicating that testosterone may induce endothelial dysfunction. It is
possible that this effect contributes to the cardiovascular adverse effects
observed in AAS abusers.
Paper No.: 1867
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
CANNABINOIDS AND CALCIUM MOBILISATION IN
MICROGLIA CELLS
Khalil El Deeb(1), SPH Alexander(2), D Pritchard(3), DA Kendall(2)
(1) University of Nottingham Medical School, School of Biomedical
Sciences, Queen’sMedical Centre, Nottingham, UK & Al-Azhar University, Faculty of Medicine, Department of Pharmacology, Damietta, Egypt
(2) University of Nottingham Medical School, School of Biomedical
Sciences, Queen’s Medical Centre, Nottingham, UK
(3) University of Nottingham Medical School, Queen’s Medical Centre,
Nottingham, UK
(4) University of Nottingham Medical School, School of Pharmacy,
Queen’s Medical Centre, Nottingham, UK
Introduction Activation of microglial cell is a key feature in a wide range
of neuroinflammatory CNS disorders such as stroke and multiple sclerosis. Cannabinoid compounds are known to have an immunomodulatory
effects through their actions on CB2 receptors and, potentially GPR55, a
cannabinoid-related receptor (Kreitzer & Stella, 2009). In the present
study, we investigated the effects of cannabinoid receptor ligands against
LPI and ADP on Ca2 + levels in BV-2 mouse microglial cells. Materials:
Intracellular calcium levels ([Ca2 + ]i) were monitored using Fluo-4
using a FlexStation-96 (Kasorn et al., 2006). Results expression of
GPR55 mRNA in BV-2 cells was indicated by Taqman qRT-PCR analysis. While AEA and 2AG, as well as HU210, CP55940, WIN55212-2
and rimonabant did not alter [Ca2 + ]i (9 ± 1, 8.3 ± 3, 10 ± 1, 9.8 ±
1.3, 8.8 ± 1.3 and 8.4 ± 1.9% ATP response respectively n = 5) cannabidiol and THC produced modest increases in [Ca2 + ]i (15 ± 2 and
16 ± 1.9%, respectively n = 5). Pre-treatment with cannabidiol or THC
(10 lM) produced significant attenuations of LPI- (9 ± 1, 2 ± 0.5%
respectively n = 3) and ADP-evoked (31 ± 8, 6.3 ± 1.6% respectively
n = 3) [Ca2 + ]i responses. Conclusion these finding are consistent with
a non-selective inhibition of evoked Ca2 + release by cannabinoid
ligands in BV-2 microglial cells. The functional implications of these
effects on microglial calcium mobilization remain to be determined.
(Kasorn A et al. (2006). pA2 online 3: 24P.Kreitzer FR, Stella N (2009).
Pharmacol Ther 122: 83-96).
This work was made possible by a scholarship to Khalil Eldeeb from the
Egyptian government.
Paper No.: 2119
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
TUMOR NECROSIS FACTOR ALPHA INHIBITION BY
PENTOXIFYLLINE PROTECTS FROM THE IMPAIRMENT IN
VASCULAR REACTIVITY ASSOCIATED WITH INSULIN
RESISTANCE
Hany El-Bassossy(1), M El-Moselhy(2), M Fouad(1)
(1) Zagazig University Faculty of Pharmacy, Department of Pharmacology, Zagazig, Egypt
(2) University of Minia Faculty of Pharmacy, Department of Pharmacology, Minia, Egypt
Alterations of vascular reactivity due to insulin resistance play roles in
diabetic vascular complications. Tumor necrosis factor a (TNF-a) is an
important pro-inflammatory cytokine. Here we have investigated the
potentially protective effect of TNF-a inhibition against the changes in
vascular reactivity associated with insulin resistance. Insulin resistance
was induced by including fructose (10%) in drinking water. while TNF-a
was inhibited by pentoxifylline (50mg.kg-1) for 8 weeks. Serum levels
of glucose, insulin, TNF-a was determined. Rings of rat isolated thoracic
aorta were used for histopathological examination and to measure the
responses to receptor (phenylephrine, PE) and depolarization (KCl) vasoconstrictors, and the endothelial dependent (ACh) and independent
(sodium nitroprusside, SNP) relaxants. Fructose drinking induced a significant increase in insulin level and insulin resistance index. The insulin
resistance was accompanied with a significant elevation in TNF-a level.
While pentoxifylline administration prevented the elevation in TNF-a
level and inhibited the developed insulin resistance. Insulin resistance
significantly increased contraction of aortic rings to PE and KCl and
decreased relaxation to ACh and SNP. TNF-a inhibition by pentoxifylline protected against insulin resistance-induced hyperresponsiveness to
PE without affecting responses to KCl while it prevented the hyporesponsiveness to ACh and SNP. Insulin resistance was associated with
marked infiltration of leukocytes in the adventitia and endothelial cells
pyknosis. TNF-a inhibition by pentoxifylline prevented the leukocyte
infiltration and significantly inhibited the observed pyknosis. In conclusion, inhibition of TNF-a by pentoxifylline protects from the impairment
in aortic vascular reactivity associated with insulin resistance, by a mechanism involving an inhibition in leukocyte infiltration.
Paper No.: 3309
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
PROTEIN KINASE C INHIBITION PROTECTS FROM THE
IMPAIRMENT IN VASCULAR RESPONSIVENESS
ASSOCIATED WITH INSULIN DEFICIENCY BUT NOT
INSULIN RESISTANCE
Hany El-Bassossy, A Fahmy, W Barakat, N Dsokey
Zagazig University, Faculty of Pharmacy, Department of Pharmacology,
Zagazig, Egypt
Alterations in vascular reactivity play important roles in diabetic vascular
complications. In addition, stimulation of protein kinase C (PKC) has
been reported in diabetes. Here we have investigated the potential protective effect of PKC inhibition against the changes in vascular reactivity
associated with diabetes. Two animal models of diabetes were used.
Insulin deficiency was induced by injecting rats with streptozotocin
(50 mg.kg-1) while, insulin resistance was induced by fructose (10%) in
drinking water. Rings of isolated thoracic aorta were used to measure the
responses to receptor (phenylephrine, PE) and depolarization (KCl) vasoconstrictors, and the endothelial dependent (ACh) and independent
(sodium nitroprusside, SNP) relaxants. Chelerythrine, a selective PKC
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
267
inhibitor, was added to organ bath in a final concentration of 10 lM.
Streptozotocin injection significantly decreased serum insulin level while
increasing glucose level. The produced insulin deficiency significantly
increased contraction of aorta to PE and KCl without affecting relaxation
to SNP. PKC inhibition by chelerythrine completely protected against
insulin deficiency-induced hyperresponsiveness to PE and KCl. Fructose
drinking induced a significant increase in insulin level and insulin resistance index. The developed insulin resistance significantly increased contraction of aorta to PE and KCl and decreased relaxation to ACh without
affecting relaxation to SNP. Despite completely restoring responsiveness
in insulin deficiency model, PKC inhibition by chelerythrine had no
effect on the impaired responsiveness to PE and KCl in insulin resistance
model. In conclusion, inhibition of PKC by chelerythrine protects from
the impairment in aortic vascular reactivity associated with insulin defeciency but not insulin resistance.
Paper No.: 2796
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
PERIPHERAL EFFECTS OF THE MARIJUANA SMOKE IN
THE LUNG OF MICE
Krisztian Elekes(1), K Sandor(1), E Szoke(1), D Toth(3), A Markovics(1),
I Szitter(1), L Jakab(2), B Szakacs(1), L Kereskai(4), J Szolcsanyi(1),
T Molnar(2), Z Helyes(1)
(1) University of Pécs Faculty of Medicine, Department of Pharmacology
and Pharmacotherapy, Pécs, Hungary
(2) University of Pécs Faculty of Medicine, Surgery Clinic, Pécs,
Hungary
(3) University of Pécs Faculty of Medicine, Analgesic Research
Laboratory, Pécs and Gedeon Richter Plc, Budapest, Hungary
(4) University of Pécs Faculty of Medicine, Department of Pathology,
Pécs, Hungary
Although the central nervous system effects of marijuana smoke has
been extensively studied, there are less and contradictory data on its
peripheral actions. Since human case reports described emphysema and
lung inflammation in marijuana smokers, the aim of the present study
was to examine these alterations in a predictive mouse model. Male CD1
mice were exposed to smoke of cigarettes containing chopped dried marijuana (0.4 + 0.02w/w%. THC; 2 cigarettes twice a day throughout 1-3months). Experiments with research cigarettes were paralelly performed
for comparison. Airway reactivity to inhaled carbachol was measured
with unrestrained whole body plethysmography and the total cannabinoid
content of the urine was determined every week. Histological examination was performed, myeloperoxidase activity as a marker of granulocyte
accumulation and IL-1b concentration were measurement from the lung
samples. Prominent bronchial hyperreactivity was observed from the second week in the marijuana-exposed group and only from the middle of
the third month in cigarette smoke-exposed mice, the extent was significantly smaller in the latter group. The histological pictures showed definitive inflammatory changes. At 2 and 3 months the number of
inflammatory cells and the empysema markedly increased, goblet cell
hyperplasia, perivascular granulocyte accumulation and large number of
eosinophilic giant cells were also observed. In the cigarette smokeexposed group inflammation and emphysema appeared later and the histopathological changes were markedly less severe. Myeloperoxidase content and IL-1b concentrations were markedly elevated in both groups.
These data clearly show that marihuana smoke induces more severe
inflammation, emphysema and hyperresponsiveness in the mouse airways than the conventional cigarettes.
Paper No.: 1175
FOCUSED CONFERENCE GROUP: P17 - NEW APPROACHES
AND TARGETS IN PSYCHIATRY
EFFECT OF GABA MIMETICS ON THE DURATION OF
IMMOBILITY IN BEHAVIORAL DESPAIR SWIM TEST
Abdalla Elhwuegi, N Zahaf
Alfateh University of Medical Sciences Faculty of Pharmacy, Department of Pharmacology and Clinical Pharmacy, Tripoli, Libya
Introduction: Studies regarding the role of GABA in depression are conflicting. Therefore, it was decided to study the antidepressant effect of
different drugs that enhance the GABA system by different mechanisms
against forced swim test (FST). Materials and methods: Adult Albino
mice divided to several groups of 6 animals, each received an intraperitoneal injection of either imipramine (10, 20, or 30 mg/kg), diazepam (0.5,
1 or 2 mg/kg) vigabatrin (100,200, or 300 mg/kg), zolpidem (2.5, 5, or
10 mg/kg), or alprazolam (1, 2.5 or 5 mg/kg). Control groups received
the appropriate vehicle. One hour after injection the duration of immobility for 5 minutes was calculated in the FST. The percentage change in
the duration of immobility from control group was calculated. Statistical
significance (p < 0.05) between treated and control group was calculated
using unpaired t-test. Results: Imipramine produced a significant dose
dependent decrease in the duration of immobility (78%, 74% and 56%
respectively). Diazepam, vigabatrin and zolpidem produced a significant
dose dependent increase in the duration of immobility (119%, 126% and
128%), (116%, 124% and 128%) and (108%, 109% and 119%) respectively. The 2 low doses of alprazolam produced a significant increase
(115% and 120%), while the high dose produced a significant decrease
in the duration of immobility (74%) Discussion: Except for the high dose
of alprazolam, all of the selected GABAmimetic drugs had a depressant
like action in FST. This can be the result of a direct and/or an indirect
effect of these drugs on other neurotransmitter systems including GABA.
Paper No.: 2920
FOCUSED CONFERENCE GROUP: P10 - DRUGS FOR HALF
THE WORLD: PAEDIATRIC CLINICAL PHARMACOLOGY
NEPHROTIC SYNDROME IN CHILDREN: GENETIC
VARIABILITY OF THE CLINICAL PRESENTATION AND
THERAPEUTIC RESPONSE
Valery Elie(1), M Fakhoury(1), Y Medard(1), A Rousseau(3),
G Deschenes(2), E Jacqz-Aigrain(1)
(1) Hospital Robert Debre, Department of Paediatric Pharmacology and
Pharmacogenetics, Paris, France
(2) Hospital Robert Debre, Department of Paediatric Nephrology, Paris,
France
(3) Clinical Research Unit (URC-EST), Paris, France
Introduction: Nephrotic syndrome is the most common kidney disease in
childhood, of unknown origin. In its steroid sensitive form, remission is
usually achieved within 4 weeks of prednisone but almost 70% of
patients will experience relapses. This ongoing multicentric prospective
case-control study aims to explore the impact of genetic polymorphisms
affecting glucocorticoid disposition and response on the outbreak, outcomes and interindividual variability of response to glucocorticoids in
pediatric nephrotic patients. Materials: Enrolled patients were treated
according to recommendations of the Society of Paediatric Nephrology
and paired unaffected controls selected according to sex and age. Patients
were genotyped for 4 single additional single nucleotide polymorphisms
(SNPs): glucocorticoid receptor (GR) rs6196, rs6198, rs33388 and
FK-binding protein 5 rs1360780. GR individual haplotypes were inferred
using HapMap database, PHASE and Haploview softwares. All patients
were also genotyped using Illumina Infinium Whole-Genome Genotyping beadarray for 17826 SNPs (1395 genes). Statistical analyses are performed with Plink 1.07, Thesias and SPSSE Statistics 17. Results: 132
patients with a first outbreak of NS were included: 78 boys and 54 girls
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
268
(sex ratio: 1.44/1), aged 5.25 ± 3.26 [0.10-15.10] years. Minor allele frequencies were: rs6196 (13.40%), rs6198 (15.17%), rs33388 (38.99%)
and rs1360780 (31.30%). Preliminary results using a logistic regression
model of relapse predicted 80.6% of overall outcomes (83.3% of relapsers and 76.9% of non relapsers). Conclusion: Further statistical analyses
will be performed to evaluate the impact of genetic background on the
clinical outcome and variability of steroid treatment response in nephrotic syndrome.
Paper No.: 1180
FOCUSED CONFERENCE GROUP: P02 - TRANSMEMBRANE
TRANSPORT: PERSPECTIVES FOR DISEASE AND DRUG
DISCOVERY
THE EFFECT OF LOPERAMIDE AND FENTANYL ON THE
DISTRIBUTION KINETICS OF VERAPAMIL IN THE LUNG
AND BRAIN IN SPRAGUE DAWLEY RATS
Iman El-Kiweri(1), MT van Patot(2), YL Zhang(2), U Christians(2),
T Henthorn(2)
(1) King Saud Bin Abdulazziz University for Health Science, College of
Nursing, Department of Basic Science, Jeddah, Saudi Arabia
(2) University of Colorado, Denver Health Sciences Center, Denver, CO,
USA
Verapamil has been shown to inhibit fentanyl uptake in vitro and is a
potent P-glycoprotein inhibitor. Tissue partitioning of loperamide, a commercially available opioid, is closely controlled by the P-gp efflux transporter. The following studies were designed to evaluate the effect of
opioids on verapamil partitioning in the lung and brain, in vivo. Opioid
(fentanyl or loperamide) was administered by intravenous infusion to
Sprague Dawley rats alone or in combination with verapamil and plasma,
with lung and brain tissues were collected at 1, 5, 6, 8, 10 and 60 minutes. Drug dispositions were modeled by recirculatory pharmacokinetic
models. Fentanyl slightly increased the verapamil lung (PL) partition
coefficient yet decreased the brain (PB) partition coefficient. Furthermore, loperamide significantly increased PL and PB. Fentanyl reduced
the verapamil volume of distribution (V1) and verapamil elimination
clearance (ClE). Fentanyl decreased verapamil brain partitioning, yet
increased verapamil lung partitioning. Also, loperamide increased lung
and brain partitioning in vivo. These results suggest that verapamil and
fentanyl may be substrates of an unidentified inward transporter in brain
tissue and confirm that verapamil and loperamide are substrates of the
efflux transporter P-gp.
Paper No.: 972
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
COMPARATIVE EFFICACY AND BIOAVAILABILITY OF
DIFFERENT PRAZIQUANTEL BRANDS
Naglaa El-Lakkany(1), S Seif el-Din(1), AN Sabra(1), M Ibrahim(2),
S Botros(1)
(1) Theodor Bilharz Research Institute (TBRI), Department of
Pharmacology, Giza, Egypt
(2) Egyptian International Pharmaceutical Industries Company (EPICO),
El-Asher Men, Ramadan City, Egypt
The efficacy, bioavailability and drug metabolizing enzymes mainly
involved in the metabolism of the commercial brands of praziquantel
(PZQ) in Egypt were examined in comparison with the original pure
powder. Mice infected with PZQ-susceptible (CD) or insusceptible (EE2)
S. mansoni isolates were divided each into 7 groups, 6 of them received
PZQ brands (Distocide, Epiquantel, Biltricide, Bilharzid, Praziquantel,
and pure PZQ), while the seventh one was left as infected untreated.
Seven weeks PI, worms were quantified and hepatic CYP450 and cyt b5
were examined. To study PZQ pharmacokinetics, groups of normal mice
were given different PZQ brands and subdivided into subgroups, killed
at 2, 5, 15, 30, 60, 90, 120,150, 180, 240 and 360 minutes post dosing.
Significant decrease in worm burden was recorded in all PZQ brands
treated mice infected with CD isolate and it was to a less extent in mice
infected with the EE2 isolate. Biltricide and Distocide showed higher
worm reduction with least inhibition of CYP450 and cyt b5 as that of
pure powder. Pharmacokinetic data revealed higher Cmax and AUC0-6h
for both formulations versus that for PZQ in pure powder. PZQ T3A,
Bilharzid and Epiquantel showed in addition to less efficacy, higher Kel
and lower t½, Cmax and AUC0-6h. The 32-46% reduction of their bioavailability was shown to reflect on their antischistosomal efficacy and
recovery of drug metabolizing enzymes. Conclusion; the quality of generic PZQ should include in addition to examining the physicochemical
characteristics of the brands, biological testing including efficacy and
bioavailability studies.
Paper No.: 973
FOCUSED CONFERENCE GROUP:
P19 - GENERAL SESSION - HEPATOLOGY
POTENTIAL ANTIFIBROTIC EFFECTS OF LOSARTAN OR
SILYMARIN COADMINISTRATION WITH PRAZIQUANTEL
AGAINST EXPERIMENTAL SCHISTOSOMA
MANSONI-INDUCED LIVER FIBROSIS
Naglaa El-Lakkany(1), W El-Maadawy(1), A Ain-Shoka(2),
A Badawy(3), F Ebeid(1)
(1) Theodor Bilharz Research Institute (TBRI), Department of Pharmacology, Giza, Egypt
(2) Cairo University Faculty of Pharmacy, Department of Pharmacology,
Cairo, Egypt
(3) Theodor Bilharz Research Institute (TBRI), Department of Pathology,
Giza, Egypt
The main cause of mortality and morbidity in human schistosomiasis is
hepatic fibrosis, which characterized by excessive accumulation of extracellular matrix (ECM). This study investigates the possible antifibrotic
effects of losartan and silymarin at different stages of schistosomal hepatic fibrosis. Parasitological, biochemical and histological parameters
which reflect disease severity and morbidity were examined. Moreover,
the effects of both drugs on ECM turnover were assessed by determination of hydroxyproline (Hyp), serum MMP-2 and TGF-b1. S. mansoniinfected mice were divided into 7 groups: Control untreated (i), treated
with either losartan (ii), or silymarin (iii) alone in doses of 50 or 750 mg/
kg/day for 5 days/wk for 6 wks respectively starting from the 4th, 8th,
12th wks PI, treated with praziquantel (PZQ), 500 mg/kg/day for 2 days
7 wks PI (iv), treated with losartan or silymarin in addition to PZQ (v &
vi) and treated with both losartan and silymarin in addition to PZQ (vii).
Administration of losartan or silymarin alone caused remarkable decrease
in worm burden, hepatic and intestinal tissue egg load with an increase
in dead eggs. Best results were obtained when losartan or silymarin were
co-administered with PZQ in the acute stage of infection as revealed by
normalization of ALAT, Hyp, MMP-2, TGF-b1 and of GSH versus PZQ
treated group. Taken together, these data suggested diminution of liver
fibrosis with a decrease in the severity of murine schistosomiais and
point to losartan and silymarin as a convenient and promising coadjuvant
therapeutic tool in the therapy of early schistosomal hepatic fibrosis.
Paper No. 1087
FOCUS GROUP: P04 - PHARMACOEPIDEMIOLOGY,
CURRENT CONTROVERSIES AND OPPORTUNITIES
ANALYSIS OF PHARMACOTHERAPY FOR PREGNANT
WOMEN IN PRIMORYE REGION
Ekaterina Eliseeva, Y Feoktistova, L Lozinskaya
Vladivostok State Medical University, Department of Clinical Pharmacology,Vladivostok, Russian Federation
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
269
Materials: 703 medical charts of pregnant women were analyzed by continuous sampling method (2004-2007). All the women consulted clinical
pharmacologist or geneticist regarding possible fetotoxic effects in pregnancy age from 3 to 32 weeks. Results: taking medication was carried
mostly in 1-3 and 6-7 weeks of pregnancy age. Average number of drugs
taken by a pregnant woman was 2.95 + 0.22. Total number of medicines
was 2078. Polypharmacy was noted in 85 (12.1%). Indications for pharmacotherapy were pelvic inflammatory disease, respiratory infections,
UTI, GI disorders, connective tissue diseases, arterial hypertension and
epilepsy. Prescriptions included antibiotics 423 (60.17%), NSAID 98
(13.9%), CNS-affecting drugs 69 (9.8%), female sex hormones 68
(9.67%), corticosteroids 44 (6.25%), vaccines 37 (5.26%), mucolytics and
antitussives 50, bronchodilators (21). Also were used anticonvulsants
(21), opiate receptor agonists and amphetamines (9), PKKN drugs (32),
nootropics (35). ACE inhibitors were used in 2 patients. Two women took
veroshpiron in I trimester. According to the FDA classification 41.81% of
prescribed medicines is potentially toxic to the fetus (category C, D and
X of the FDA). Only 3,75% of drugs could be considered safe (Category
A). 26,27% prescriptions were relatively safe (category B). In our study
28,15% of the prescribed medications are not included in the FDA classification. Their risk for pregnancy is not known (NR). Conclusions: our
results suggest no sufficient drug safety for the fetus in pregnancy.
Paper No. 1088
FOCUS GROUP: P04 - PHARMACOEPIDEMIOLOGY,
CURRENT CONTROVERSIES AND OPPORTUNITIES
MEDICAL PROGNOSIS OF PERIOPERATIVE PROPHYLAXIS
EFFICIENCY IN CHOLECYSTECTOMY
Ekaterina Eliseeva, V Shevtsov
Vladivostok State Medical University, Department of Clinical Pharmacology, Vladivostok, Russian Federation
Materials: 487 medical records of patients in three hospitals of the Primorsky Territory (2007-2008) were analyzed by sample comparing method
according to kea words like: surgical treatment of cholelithiasis, the
absence of infectious diseases, rare complications. Exclusion criteria
were: intolerance to antibacterial drugs in medical history; incomplete
record in medical charts, death during surgery or within 30 days after.
Results: Antibiotic perioperative prophylaxis (APP) has been performed
15.22% of patients, subjected to lap cholecysyectomy (cefazolin, cefuroxime and cefoperazone). APP was used in 48% of open cholecystectomy cases. Cefazolin, cefuroxime combined with metronidazole and
cefoperazone were used in 48% of the open colecystectomies. In laparoscopic cholecystectomy cases the frequency of postoperative infections
(PI) was 4,89% versus open surgical method - 32,77%. Atypical location
of the gall bladder was the very high risk for the development of PI. Relatively possible risks in laparoscopic cholecystectomy were: concrements
in the common bile duct (OR 0,23), recent exacerbations of cholecystitis
(OR 0,16). The relative PI risks after open cholecystectomy were: age
(OR 0,98), of obstruction of common bile duct (OR 0,78), recent episodes
of acute cholecystitis OR (0,36), presence of jaundice before surgery (OR
0 43), duration of surgery (OR 0,99). Conclusion: APP does not reduce
the incidence of PI in laparoscopic cholecystectomies. In open cholecystectomy APP reduces incidence of postoperative infection (OR 9,24). Further study is needed to reduce the routine use of cefoperazone for APP.
Paper No.: 2102
FOCUSED CONFERENCE GROUP: P18 - NUCLEAR
RECEPTOR TARGETS FOR TREATMENT OF DISEASES
GASTROPROTECTIVE EFFECT OF PEROXISOME
PROLIFERATOR-ACTIVATED RECEPTOR-C (PPARC) AGONIST, ROSIGLITAZONE AGAINST STRESS-INDUCED
GASTRIC MUCOSAL INJURY IN RATS
Mohamed A EL-Moselhy(1), WH Nazmy(2), HM Ibrahim(2)
(1) University of Minia, Department of Pharmacology, Minia, Egypt
(2) University of Minia, Department of Physiology, Minia, Egypt
This study was designed to investigate the protective effects of rosiglitazone against acute gastric mucosal lesions induced by cold-restraint stress
ulcer (CRS) in rats and the mechanisms underlying these effects. Rats
were pylorically ligated and divided randomly into following groups:
control, CRS & CRS pretreated with rosiglitazone (3 mg/kg/day, orally
for7days) groups. Blood samples were collected from the heart; left to
clot and the sera were separated for determination of tumor necrosis factor-a (TNF-a) level. Animals were sacrificed and gastric juice was collected for determination of volume, free and total acid outputs, mucin
concentration and pepsin activity. The gastric mucosal lesions were
assessed and the stomachs were scraped and the gastric mucosa was analyzed for determination of lipid peroxides, nitric oxide (NO), and prostaglandin E2 (PGE2) levels. Our results revealed that CRS induced gastric
mucosal ulceration mainly through an increase in gastric acidity, pepsin
activity, lipid peroxides and TNF-a levels with concomitant reduction in
NO, mucin and PGE2 levels compared to the control non-stressed rats.
Rosiglitazone pre-treatment significantly attenuated the gastric mucosal
lesions induced by CRS, which was accompanied by significant reduction in lipid peroxides and TNF-a levels and concomitant increase in
mucin concentration. Additionally, rosiglitazone caused a significant rise
in gastric mucosal nitrite and PGE2 levels. In conclusion: Rosiglitazone
exerts a protective effect against CRS induced gastric ulcer possibly via
NO, PGE2 and mucus production as well as anti-inflammatory and antioxidant mechanisms.
Paper No.: 2971
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
RHO KINASE-DEPENDENT CALCIUM SENSITIZATION IN
THE INTERPLAY BETWEEN PRESSURE- AND
AGONIST-INDUCED CONSTRICTION OF CEREBRAL
ARTERY
Ahmed El-Yazbi(1), RP Johnson(1), K Takeya(2), EJ Wlash(1),
MP Walsh(2), WC Cole(1)
(1) University of Calgary, Department of Pharmacology and Physiology,
Calgary, ALB, Canada
(2) University of Calgary, Department of Biochemistry, Calgary, ALB,
Canada
Calcium sensitization is the mechanism by which smooth muscle cells
attain an elevated level of myosin regulatory subunit phosphorylation
and contraction with no change in cytosolic calcium concentration. This
process has been implicated in the myogenic response of resistance
arteries and in agonist-induced constriction of conduit arteries. Only
limited information is available concerning the interplay of myogenicand agonist-induced calcium sensitization in resistance arteries due to
technical difficulties associated with quantification of phosphoprotein
markers of this mechanism; i.e. myosin light chain phosphatase targeting subunit (MYPT1) and CPI-17. Here, we used a high sensitivity
western blotting technique to detect a role for Rho kinase-mediated calcium sensitization in the myogenic response of cerebral arteries, to
assess the role of this mechanism in serotonin (5-HT)-evoked constriction, and whether there is an interplay between these constrictor stimuli
in the control of calcium sensitization. Pressure in the range of 10100 mmHg increased phospho-MYPT1-T855, but not -T697. 5-HT
evoked calcium sensitization only in the presence of myogenic constriction at 60 and not at 10 mmHg, and increased phosphorylation was
detected at MYPT1-T855 and -T697. Dual phosphorylation may be
required to permit increased force, a view that is supported by phosphorylation of both sites at an extreme pressure of 140 mmHg. No role
for CPI-17 was identified. These findings provide the first biochemical
evidence of an interplay myogenic and agonist dependent mechanisms
for control of calcium sensitization.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
270
Paper No.: 923
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
PHARMACOKINETICS OF FLORFENICOL (WATER SOLUBLE
FORMULATION) IN HEALTH AND PASTEURELLA INFECTED
BROILER CHICKENS
Paper No.: 1798
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
ANTIVIRAL ACTIVITY OF ANAFERON FOR CHILDREN IN
PANDEMIC A/H1N1 IN MICE
Hesham Elzorba, H El-Banna
Oleg Epstein, JL Dugina, SA Tarasov, SA Sergeeva
Cairo University, Department of Pharmacology, Cairo, Egypt
Materia Medica Holding Company, Department of Research & Development, Moscow, Russian Federation
Florfenicol have been approved in the European Union for use in cattle
and pigs as injectable solution for treatment of respiratory diseases in
cattle but now it introduced in some countries as oral solution for the
treatment of several poultry diseases. The aim of the present study is
to describe the Pharmacokinetics of florfenicol (water soluble formulation) in broiler chickens after either a single intravenous and/or oral
administration at a dose of 30 mg/kg body weight. Meanwhile, its disposition in control healthy and Pasteurella-infected broilers was compared. Following the IV administration of the drug in healthy and
diseased birds, the drug plasma concentration declined in a biphasic
pattern. The maximum plasma concentration of florfenicol in control
healthy and diseased was reached one hour after its oral administration,
but the peak level detected in control broilers was higher than that
detected in infected birds. Data of the present study showed that volume of distribution (Vdss, 1.98 L/kg), total body clearance (ClB,
0.55 L/kg/h) in infected birds were higher than that determined in control birds compared to values determined in healthy ones (Vdss, 1.3 L/
kg, ClB, 0.38 L/kg/h). On the other hands, systemic bioavailability
were significantly lower (F %,55.6%) in diseased broiler compared to
values determined in healthy ones (F %,71.5).
Paper No.: 491
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
ANTIBIOTIC ACTIVITY AGAINST URINARY PATHOGENS IN
HOSPITALIZED MEDICAL PATIENTS IN EASTERN NIGERIA
Oguamanam Enwere, K Duruewuru, V Okwara
Imo State University Teaching Hospital, Department of Internal Medicine, Orlu, Imo State, Nigeria
Urinary tract infection causes high morbidity and increases cost of care
in clinical practice. Quinolones have been recommended as first line
drugs for uncomplicated urinary tract infection. Changing antibiotic
sensitivity patterns are becoming a concern in Nigeria due to the high
rate of self medication and substandard drugs in the Nigerian market.
This study, scheduled to last a one year period, assessed sensitivity of
commonly available antibiotics to isolated urinary pathogens in admitted patients who had urine analysis and culture as part of their
workup. 29 positive cultures (20 males and 9 females) were recorded
so far. 9 patients had self medicated with antibiotics prior to admission
(all men). 12 samples grew escherichia coli (41.4%), 15 for staphylococcus aureus (51.7%) and one each for proteus mirabilis and enterococcus spp (3.4% each). There was a high resistance rate against the
quinolones (33%-50%), nitrofurantoin and nalidixic acid (85.7% &
68.8% respectively). Resistance against gentamycin was 67.9%, streptomycin (17.4%) and coamixiclav (68.8%). E.coli did not have any
resistance against levofloxacin. Only 82.8% of patients had pyuria
while 79.3% had proteinuria, 44.8% had hematuria and 24.1% had
glycosuria. The rising resistance to antibiotics is alarming; discouraging
self medication is very important at preventing rising resistance to antibiotics and physicians must be notified on the changing antibiotic sensitivity patterns in clinical practice.
Antiviral activity of anaferon for children, ultra-low doses of oral antibodies to interferon c, in the mice models of non-lethal and lethal A/California/07/09 (H1N1) infection was studied. First study included 96
female Balb/c mice (15-17 g) infected intranasally with 2.3 lg EID50/
mouse. Anaferon for children and distilled water (control) were administered intragastrically BID 0.2 mL/mouse starting 5 days prior to and
continuing for 8 days after inoculation. Oseltamivir was given 20 mg/kg/
day BID for 5 days with first treatment 1hr after the challenge. Virus
titers in lungs on days 2, 4, 6, 8 after inoculation were assessed. Both
anaferon for children and oseltamivir significantly reduced A/H1N1
titers: by 2.7 and 2.4 times respectively. Second study included 105
female Balb/c mice (16-20 g) infected intranasally with 3xLD50. Anaferon for children and distilled water were administered intragastrically BID
0.2 mL/mouse in combination with free access to drinking water (in
active treatment group anaferon for children serve as drinking water)
starting 5 days prior to and continuing for 13 days after challenge. Oseltamivir was given 20 mg/kg/day BID for 3 days with first treatment 4hr
before challenge. Virus titers in lung tissue on days 3 and 6 after inoculation were assessed. Anaferon for children significantly reduced mortality
of mice: in control group survival was 12.5%, in oseltamivir group –
10%. The titet measurements in lungs confirmed the survival data. This
study provides evidence of antiviral activity of anaferon for children
against pandemic A/California/07/2009(H1N1) influenza virus infection
in mice.
Paper No.: 2257
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
RESISTANCE OF ESCHERICHIA COLI AND KLEBSIELLA
PNEUMONIAE ISOLATES IN THREE TERTIARY INTENSIVE
CARE UNITS: A RESULT OF INSUFFICIENT EMPLOYMENT
OF INFECTION CONTROL MEASURES OR INADEQUATE
ANTIBIOTIC CONSUMPTION?
Viktorija Erdeljic, I Francetic, Z Bosnjak, A Budimir, S Kalenic, L Bielen,
R Likic, K Makar-Ausperger
University Hospital Zagreb, Department of Medicine, Zagreb, Croatia
Introduction: Escherichia coli (E.coli) and Klebsiella pneumoniae
(K.pneumoniae) are among the most important causes of serious nosocomial bacterial infections, and their resistance to antimicrobial agents has
become an increasingly relevant problem. We evaluated reasons (insufficient employment of infection control measures vs. inadequate antibiotic
consumption) for high resistance levels of E.coli and K.pneumoniae isolates to fluroquinolones, aminoglycosides and beta-lactams in three tertiary intensive care units. Methods: Data on monthly fluroquinolones,
aminoglycosides and beta-lactams consumption and resistance rates of
K.pneumoniae and E.coli isolates to these antibiotics were analyzed for
the period April 2006-July 2007 using distributed lags time series analysis. In addition, pulsed-field gel electrophoresis (PFGE) was done on a
representative sample of 78 E.coli and 66 K.pneumoniae isolates (17%
and 23% of all isolates, respectively) to evaluate the type of resistance:
clonal or polyclonal. Results: A significant temporal relationship between
fluoroquinolones, aminoglycosides and beta-lactams consumption and
the resistance rates of E.coli and K.pneumoniae isolates to these antibiotics was identified. PFGE typing showed a wide diversity of strains: 57%
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
271
of E.coli and 61% of K.pneumoniae isolates were considered distinct as
they demonstrated <90% similarity with any other isolate. The remaining
isolates clustered into 16 and 10 clonal groups, respectively. For 10/16
and 8/10 clonal groups, respectively, the isolates belonging to each group
came from the same ICU. Conclusion: We were able to demonstrate antibiotic use as the main reason for the emergence of resistant E.coli and
K.pneumoniae strains, thus confirming the usefulness of antimicrobial
use restrictions as a method of resistance control.
Paper No.: 2460
FOCUSED CONFERENCE GROUP:
P04 - PHARMACOEPIDEMIOLOGY, CURRENT
CONTROVERSIES AND OPPORTUNITIES
CLINICAL PHARMACOLOGY OUTPATIENT CLINIC:
OUTCOMES AND MATERNAL CHARACTERISTICS
ASSOCIATED WITH PREGNANCY EXPOSURE TO FDA
CATEGORY X DRUGS
Viktorija Erdeljic(1), I Francetic(1), K Makar-Ausperger(1), R Likic(1),
M Radacic-Aumiler(1), D Juricic(1), L Bielen(1)
University Hospital Zagreb, Department of Medicine, Zagreb, Croatia
Introduction: Risk classification systems have been set up to guide safe
drug use during pregnancy. The most frequently used is the FDA pregnancy risk categorisation which places the drug in one of five categories
(A, B, C, D, X) according to the level of risk to the infant, FDA X category being the category of contraindicated drugs in pregnancy. However,
drug safety classifications give a very crude estimation of risk and should
only be used as general guidelines. Methods/Patients: We retrospectively
evaluated medical records on 130 consultations on the use of FDA X
drugs in pregnancy (2001-2007). Results: Out of 1003 pregnant women
consulted, 130 were taking an FDA X drug during the 1st trimester of
pregnancy. Most women were reffered during the first trimester of pregnancy (88%). Maternal characteristics were as follows: 25% women had
an university degree, 36% had comorbidity, 26% were cigarette smokers,
for 39% this was the 1st pregnancy, 20% women were prescribed >2
drugs during pregnancy. The most frequently FDA X prescribed drugs
were oral contraceptives (75%), followed by oral anticoagulants, retinoids, methotrexate, etc. Follow-up was available for 50% women: Normal fetal outcomes were reported for 76% pregnancies, spontaneous
abortions for 9%, and artificial abortions for 12% pregnancies. One baby
was born with coarctation of the aorta. In more than 2/3 of cases the clinical pharmacologist’s risk estimation was lower than X. Conclusion: Our
study shows high prevalence of consultations for drug exposures to FDA
X drugs in pregnancy in the clinical pharmacology outpatient clinic.
Paper No.: 2411
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD DISEASES
SUBSTANCE P INTERACTING PROTEINS: POTENTIAL
MECHANISMS FOR NON-RECEPTOR MEDIATED EFFECTS
Nuray Erin(1), V Bui(1), G Clawson(2)
(1) Akdeniz University, Department of Pharmacology, Antalya, Turkey
(2) Penn State University, Department of Pharmacology, Hershey, USA
Substance P (SP) belongs to the tachykinin family of peptides which is
involved in chronic inflammation, cancer, and neurodegenerative disorders. Inflammatory and carcinogenic effects of SP are believed to be
mediated by neurokinin 1 receptors (NK1Rs). SP also has anti-inflammatory as well as anti-carcinogenic properties, although the mechanisms
underlying these activities are not known. We hypothesized that direct
interaction of SP with key binding partners mediate certain effects of
SP.Biotinylated SP was incubated with skin extracts and binding proteins
were precipitated using streptavidin beads. After PAGE, novel bands that
differed from controls were excised and analyzed using nanocapillary
reverse phase HPLC coupled with mass spectrometry. We found eight
high confidence proteins, each of which contained three or more peptide
sequences with high homology scores and good spectra. Three of the
proteins (collagen type 1a1, procollagen type 1a2, and keratan sulfate
proteoglycan lumican) were extracellular matrix components. Another
two (myosin and actin) are part of a specialized membrane structure
involved in cell migration.The final protein was identified as Histone 3.
Interaction of SP with three of the high confidence proteins was verified
in skin extracts by immunoblot analyses. Interaction of SP with keratan
sulfate proteoglycan was also demonstrated in primary murine tumors.
Paper No.: 2751
FOCUSED CONFERENCE GROUP: P17 - NEW APPROACHES
AND TARGETS IN PSYCHIATRY
THE EFFECTS OF SERTRALINE ON BEHAVIORAL
ALTERATIONS CAUSED BY ENVIRONMENTAL
ENRICHMENT
Kevser Erol(1), E Yildirim(1), E Ulupinar(2)
(1) Eskisehir Osmangazi University, School of Medicine, Department of
Pharmacology, Eskisehir, Turkey
(2) Eskisehir Osmangazi University, School of Medicine, Department of
Anatomy, Eskisehir, Turkey
Introduction: Depression, anxiety, motor skills and learning ability could
be affected by rearing environment. Neurotransmitters such as norepinephrine, dopamine and 5-HT may participate in the plasticity of environmental enrichment. In the present study, the effects of enriched
environment and social isolation on motor activity, anxiety, depression,
and also the effects of sertraline on these behaviors were evaluated.
Materials and Methods: After weaning at postnatal day 21, Wistar rats
were divided in different rearing conditions (Standard:SC, Enriched:EC,
Isolated:IC). Animals in SC were housed as group of 4 rats in regular
size cages. Animals in EC were housed as group of 12 rats in three large
cages connected via tunnels, with different toys. Animals in IC were
housed individually in metal cages. Six weeks later activitymeter, elevated plus maze, rota rod, Grip, forced swimming and sucrose preference
tests were applied to all animals. All these tests were repeated after i.p.
injection of sertraline (10 mg/kg/day) for 7 days. Results: Environmental
enrichment reduced stereotypic behavior, improved motor coordination
and facilitated learning skills. There were no significant changes in anxiety and depression in EC and IC. Sertraline reversed the effects of
enriched environment on the stereotypic behavior; It reduced depression
in SC and EC but did not change it in IC. Sertraline was anxiolytic in
EC. The animals in IC prefered more sucrose than the others, and sertraline increased sucrose consumption. Conclusion: Rearing conditions
affect the behavioral and motor functions. The effects of sertraline
change depend on rearing conditions of animals.
Paper No.: 1174
FOCUSED CONFERENCE GROUP: P08 - DEVELOPMENTS IN
THE TREATMENT OF SEXUAL DYSFUNCTION AND
DISEASES OF THE LOWER URINARY TRACT
DEVELOPMENTS IN THE TREATMENT OF SEXUAL
DYSFUNCTION AND DISEASES OF THE LOWER URINARY
TRACT
Saintmoses Eromosele(1), V Eruanga(2), PA Francis(3)
(1) The Pedagogues Educonsult, Benin City, Nigeria
(2) University of Benin, Department of Pharmacology, Benin City, Nigeria
(3) Ostee Trust Services Limited, Nigeria
Objective: To evaluate the relationship between lower urinary tract symptoms and erectile dysfunction in different male populations. Methods:
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
272
Data sources: PubMed (Medline), clinical evidence, Embase, Cochrane
reviews, and articles from reference lists. Selection criteria: Selection criteria in search databases were lower urinary tract symptoms, LUTS, comorbidity (MeSH), impotence (MeSH), sexual dysfunction, and male.
Studies on these subjects, and concerning men aged 30 years or older,
were eligible for inclusion in this review. Both community-based and
clinical-based studies were included. Results: 30 studies were eligible for
inclusion, representing 72 400 men. These studies showed a significant
positive relationship between lower urinary tract symptoms and erectile
dysfunction.
Paper No.: 763
FOCUSED CONFERENCE GROUP: P04 - PHARMACOEPIDEMIOLOGY, CURRENT CONTROVERSIES AND OPPORTUNITIES
PRESCRIPTION OF POTENTIALLY INAPPROPRIATE
MEDICATIONS: COMPARISON BETWEEN INTERNAL MEDICINE SERVICE AND GERIATRIC UNIT
obstructive pulmonary disease (COPD) develop a disproportionate
response leading to pulmonary hypertension. The aim of this study was
to analyze whether polymorphisms in nSMase2 (SMPD3 gene) can make
patients more prone to suffer from pulmonary hypertension. After
informed consent, genomic DNA was isolated from blood samples of 14
patients suffering from COPD or interstitial lung disease, 5 of them without and 9 with associated pulmonary hypertension (group III Who classification), from 3 patients with pulmonary arterial hypertension (PAH,
group I) and from 7 controls. Nine fragments covering the whole gene
sequence were amplified by PCR and sequenced. A missense mutation
(C617Y) in heterozygosis was found in one of the patients with class I
and in one the patients with class III pulmonary hypertension but in none
of the COPD patients without pulmonary hypertension or in controls. In
conclusion, this pilot study suggests that mutations in the cysteine 617 of
the nSMase2 which belongs to the catalytic center of the enzyme might
be associated with the development of pulmonary hypertension. The
sample size does not allow to draw definitive conclusions but the results
guarantee further research.
Leslie Escobar(1), P Jara(1), M Jirón(1), M Rain(1), G Martinez(2),
C Dechent(2)
(1) Universidad de Chile, Facultad de Ciencias Quı́micas y Farmacéuticas, Santiago,Chile
(2) Hospital Clinico de la Universidad de Chile, Santiago, Chile
With the exception of specialized geriatric services, in Chile there is a
scarcely knowledge about potentially inappropriate medications (PIM)
prescription to elderly. This study was conducted for comparing frequency of PIM to elderly patients attended in an internal medicine (IM)
service and a geriatric unit(GU), and to assess their outcomes over functionality. A prospective follow-up study was performed in GU and IM
services of a teaching hospital. Patients’ inclusion criteria were: 65 years
old, both sexes, without terminal disease, >3 days of hospitalization.
PIM was evaluated using 2003 Beers’ criteria and functionality by Barthel index. Data were analyzed using STATA 10.1. Eighty-one IM
patients and 70 GU patients were enrolled. In total 86(57%) were
females. GU patients were significantly older than IM patients
(80.5 ± 7.7 versus 76.5 ± 7.3 years) and IM patients had lower mean
number of co-morbidities than GU patients (3.9 ± 1.9 versus 5.3 ± 2.6,
respectively). During hospitalization, 1029 medications were prescribed
in IM versus 684 in GU. 4.5% and 4.7% of them corresponded to PIM,
respectively. These results show that a 44.4% of IM patients received at
least 1 PIM compared with 37.1% in GU patients, mainly NSAIDs concomitantly with anticoagulation therapy and amiodarone. Mean number
of medication/patients was also significantly higher in IM (12.7 ± 5.3)
compared with GU (9.8 ± 4.5). GU patients lost more than 14 points in
Barthel score during hospitalization, but there were no statistically differences between both services. More data analysis is needed for establishing associations between functionality and PIM. Also, additional studies
with an extended sample are necessary.
Paper No.: 2037
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
DOES MISSENSE MUTATION IN THE SMPD3 GENE
INCREASE THE RISK OF PULMONARY HYPERTENSION?
Lucı́a Escolano(1), MA Nieto(2), L Moreno(1), A Cogolludo(1), A
Roman(1), Jose Luis Alvarez-Sala(2), F Perez-Vizcaino(1)
Paper No.: 1364
FOCUSED CONFERENCE GROUP: P14 - ADDICTION AND
DOPING: NEUROBIOLOGICAL AND CLINICAL
IN VITRO NEUROCHEMICAL PROFILE OF MDMA AND ITS
METABOLITE ALPHA-METHYLDOPAMINE
Elena Escubedo, S Abad, I Torres, J Camarasa, D Pubill,
University of Barcelona, Department of Pharmacology and Therapeutic
Chemistry, Barcelona, Spain
Several studies suggest that MDMA-induced neurotoxicity is dependent
on its metabolic disposition. In rats, the N-demethylation of MDMA
leads to the formation of a-methyldopamine (MeDA). Both compounds
have different affinity profile for the serotonin transporter (SERT) (Ki =
3.10 ± 1.48 lM for MDMA and 0.12 ± 0.15 mM for MeDA) but similar affinity for the dopamine transporter (DAT) (assessed by [3H]WIN
35428 binding)(Ki = 14.78 ± 5.58 lM and 16.94 ± 8.73 lM respectively). They inhibited [3H]DA uptake with IC50 values: 3.90 ± 2.30 and
0.11 ± 0.02 lM respectively. The DA uptake inhibition by MDMA but
not by MeDA is dependent on NOS, PKC and nicotinic receptor. Conversely, MeDA preincubation followed by washout induces a reduction
in [3H]WIN 35428 binding, but not MDMA, pointing to a direct effect
of the metabolite on DAT. A hypothesis about MDMA neurotoxicity
states that DA enters in serotonergic terminals where it is deaminated by
MAO-B, resulting in free-radical formation and the selective degeneration of the serotonergic axons. The effect of both compounds on MAOB was very different (IC50 values of 1.00 ± 0.23 mM MeDA and
70.10 ± 13.20 lM MDMA). MeDA but not MDMA inhibited DA
uptake via SERT (IC50 about 20 nM at DA 5 nM) in hippocampal synaptosomes. Both MeDA effects, MAO-B and DA uptake inhibition via
SERT, point to the absence of neurotoxic effect of this metabolite.
Paper No.: 2091
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
PULMONARY PRESSURE REDUCTION ATTENUATES
EXPRESSION OF PROTEINS IDENTIFIED BY LUNG
PROTEOMIC PROFILING IN PULMONARY HYPERTENSIVE
RATS
(1) University of Complutense, Department of Pharmacology, Madrid,
Spain
(2) Hospital Clinico San Carlos, Madrid, Spain
Yvonne Eskildsen-Helmond(1), L Østergaard(1), B Honore(2), L Bech
Thorsen(1), J Baandrup(1), B Elmedal Laursen(1), H Vorum(2),
M Mulvany(1), U Simonsen(1),
Previous results show that neutral sphingomyelinase type 2 (nSMase2) is
involved in the signalling cascade of hypoxic pulmonary vasoconstriction
both in vitro and in vivo. In humans the magnitude of hypoxic vasoconstriction is variable: some patients suffering hypoxia due to chronic
(1) Aarhus University, Department of Pharmacology, Aarhus, Denmark
(2) Aarhus University, Department of Medical Biochemistry, Aarhus,
Denmark
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
273
The present study was designed to analyze protein regulation in the lungs
of chronic hypoxic rats in order to identify novel signalling pathways,
which could provide new targets for the treatment of pulmonary hypertension. This was achieved by proteomic studies in which proteins from
lung homogenates from five hypoxic rats were compared to those from
five normoxic rats. In a second study the expression of these proteins
and correlation to alterations in vascular muscularization was also investigated in lungs from hypoxic rats that had received treatment with either
an activator of soluble guanylyl cyclase, BAY 412272, or an inhibitor of
phosphodiesterase type 5, sildenafil. The proteomic study revealed upregulation of guanine nucleotide-binding protein beta (Gb-beta), glutathione
S transferase omega 1, cathepsin D (CatD), chloride intracellular channel
subunit 5, Annexin A4, F-actin capping protein CapZ (CapZalpha), and
the translation factor elongation factor 1 delta in lungs from chronic hypoxic rats with pulmonary hypertension. Immunohistochemistry showed
that many of these proteins were expressed in the pulmonary vascular
wall (e.g. CapZalpha, CatD, and annexin A4) and immunoblotting
showed these proteins correlated to alterations in muscularization. Both
treatments inhibited hypoxia-induced increase in right ventricular systolic
pressure and pulmonary arterial muscularization and prevented most of
the protein regulations observed after hypoxia. These findings suggest
that pulmonary pressure is an important factor for initiating signalling
pathways leading to protein expression and muscularization in the pulmonary vasculature.
Paper No.: 3117
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
UNFOLDED PROTEIN RESPONSE INVOLVED IN OXIDIZED
LDL-EVOKED ENDOTHELIAL DYSFUNCTION
Yvonne Eskildsen-Helmond(1), M Jørgensen(1), E Stankevicius(1),
N Christophersen(1), E Falk(2), U Simonsen(1)
(1) Aarhus University, Department of Pharmacology, Aarhus, Denmark
(2) Aarhus University Hospital, Department of Cardiological Medicine,
Aarhus, Denmark
Accumulation of cholesterol in the endoplasmatic reticulum (ER) contributes toapoptosis of macrophages and smooth muscle cells in atherosclerotic lesions. Thepresent study was designed to investigate whether
unfolded protein response(UPR) is activated in endothelial cells in atherosclerotic aortae and cellsexposed to oxidized low density lipoprotein
(ox-LDL). Immunohistochemistry wasperformed for glucose-regulated
protein (Grp78) and measurements of nitric oxide(NO) and calcium in
human umbilical vein endothelial cells (HUVEC). Grp78, asmarker for
UPR activation, was extensively expressed in atherosclerotic plaques,in
endothelium and adventitia localised in the ascending part of aorta
(ApoE-/-mice (n = 8)). GRP78 expression was also increased in HUVEC
exposed to acomponent of ox-LDL, lysophosphatidyl choline (LPC), and
a chemical inducer ofER stress tunicamycin (TM). In HUVEC, endothelial NO synthase (eNOS) expressionwas unaltered, while eNOS phosphorylation at serine1177 was unaltered afterincubation with LPC and
increased with tunicamycin. However, histamine (1ı̀M)-induced increases
in NO concentration (dNO, 22.9 ± 5.9 nM) were decreasedafter incubation with LPC (7.8 ± 1.7) and TM (7.9 ± 2.1), and inhibited in thepresence of asymmetric dimetylarginine. Our results indicate activation of
theUPR system associated with reduced NO release, suggesting involvement inendothelial dysfunction caused by Ox-LDL.
Paper No.: 2972
FOCUSED CONFERENCE GROUP: PW04 - ANTIOXIDANTS AS
THERAPEUTIC TARGETS
MELATONIN MODULATES OXIDATIVE STRESS AND
PREVENTS CELL DEATH IN HUMAN LEUKOCYTES. EFFECT
OF INTRACELLULAR CALCIUM OVERLOAD AND AGE
Javier Espino(1), I Bejarano(1), SD Paredes(1), D González(1),
RJ Reiter(2), JA Pariente(1), AB Rodrı́guez(1)
(1) University of Extremadura, Faculty of Science, Department of Physiology,Badajoz, Spain
(2) University of Texas Health Science Center, Department of Cellular &
Structural Biology, San Antonio, USA
Ageing is associated with an increased production of free radicals and
alterations in the mechanisms of adaptation to oxidative stress. In fact,
the free radical theory of ageing proposes that deleterious actions of free
radicals are responsible for the functional deterioration associated with
ageing. Moreover, a close relationship exists between calcium homeostasis and oxidative stress. The current work was aimed at proving the
effect of melatonin on the levels of reactive oxygen species (ROS) and
cell viability in human leucocytes collected from young (20-30-year-old)
and elderly (65-75-year-old) individuals under both basal and oxidative
stress-induced conditions. Treatments with 10 nM FMLP and/or 1 uM
thapsigargin induced a transient increase in cytosolic free-calcium concentration ([Ca2 + ]c) in human leucocytes due to calcium release from
internal stores, and led in turn to oxidative stress, as assessed by intracellular ROS measurement. Non-treated leucocytes from aged individuals
exhibited higher ROS levels and lower rates of cell survival when compared to leucocytes from young individuals. Similar results were
obtained in FMLP and/or thapsigargin-treated leucocytes from elderly
individuals when compared to those from the young individuals. Melatonin treatment significantly reduced both hydrogen peroxide (H2O2) and
superoxide anion levels, likely due to its free-radical scavenging properties, and enhanced leucocyte viability in both age groups. Therefore, melatonin may be a useful tool for the treatment of disease states and
processes where an excessive production of oxidative damage occurs.
Funded by BFU2007-60091 (MEC-DGI) and PRI07-A024 (Junta de Extremadura)
Paper No.: 961
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
THE ABSENCE OF A FUNCTIONAL PEROXISOME
PROLIFERATOR-ACTIVATED RECEPTOR-ALPHA GENE IN
MICE ENHANCES DOPAMINERGIC NEURON SENSITIVITY
TO 1-METHYL-4-PHENYL-1,2,3,6-TETRAHYDROPYRIDINE
Emanuela Esposito(1,2), E Mazzon(1), I Paterniti(1), T Genovese(1),
S Cuzzocrea(1,2)
(1) University of Messina School of Medicine, Department of Clinical &
Experimental Medicine & Pharmacology, Messina, Italy
(2) IRCCS Centro Neurolesi ‘‘Bonino-Pulejo’’, Messina, Italy
The nuclear receptor peroxisome proliferator-activated receptor (PPAR)a plays a prominent role in several physiological processes including the
inflammatory response, and its activation mediates a reduced production
of proinflammatory factors. Using PPAR-a knockout (KO) mice, we
report a detailed investigation of the role of PPAR-a in cerebral damage,
associated inflammatory and antioxidant processes in a subacute model
of Parkinson’s disease (PD). N-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), is used in rodent and cell models to elicit neurochemical
alterations associated with PD. Immunohistochemistry and Western blotting analysis were performed to detect the expression of neuronal and
inducible nitric oxide synthase (NOS) and cyclooxygenase (COX)-2,
matrix metalloproteinase (MMP)-2 and -9 activity, 3-nitrotyrosine formation, MAPK and NF-jB activation, apoptosis, comparing mice lacking
PPAR-a KO with wild-type (WT) mice. Brain oxidative stress was dramatically enhanced in KO mice, as documented by an increased content
of malondialdehyde, decreased levels of glutathione, a marked increase
of oxidative products, and no induction of uncoupling protein 2. PPAR-a
deficiency appears to aggravate the severity of brain injury through an
increase in extracellular matrix formation and inflammatory parameters.
These data indicate that PPAR-a plays an important role in integrating
and regulating central inflammation, antioxidant mechanisms in the
process of MPTP-induced toxicity, and suggest that the use of PPAR-a
agonists may be of interest for the prevention of PD.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
274
Paper No.: 1210
FOCUSED CONFERENCE GROUP: P08 - DEVELOPMENTS IN
THE TREATMENT OF SEXUAL DYSFUNCTION AND
DISEASES OF THE LOWER URINARY TRACT
PACEMAKER CELLS AND CONTRACTILITY IN THE
PROSTATE GLAND
Betty Exintaris, A Dey, S Kusljic, M Lam, D-T Nguyen, R Lang
Monash University, Department of Medicinal Chemistry and Drug
Action, Parkville, VIC, Australia
Introduction: Morphologically-distinct, c-Kit immunoreactive prostaticinterstitial cells (PIC) are likely to generate the pacemaker signal thatmanifests as slow wave activity and resultant contractility in the smooth
musclecells of the guinea-pig prostate. Methods: In this study, we have
compared thespontaneous activities and cell types in younger (300-500g)
and older animals(>900g), using tension-recording techniques, intracellular micro-electrodes andimmunohistochemistry. Results: Slow waves
recorded within the guinea pigprostatic stroma had similar frequencies to
that of the spontaneous contractionswithin both age groups of animals.
In addition, older prostates (n = 78) had ahigher basal tension of
6.0 ± 0.3mN than the younger prostates (n = 51) 4.7 ± 0.66mN.Pacemaker activity was recorded in approximately 10% of all electricalrecordings in the younger guinea-pig prostate (n=>100). In the older guineapigprostate, pacemaker activity was not recorded (n = 84), however cells
exhibitinglarge-amplitude spontaneous transient depolarisations (STD)
were recorded in 9%of all electrical recordings. Microscopic examination
revealed a distinct layerof c-Kit and connexin 43-immunoreactive cells
in the region between the smoothand glandular layers in younger and
older guinea-pigs; c Kit positive cells werealso sparsely distributed
within the glandular layer and among smooth musclecells (n = 5). Conclusion: These results suggest that that older prostates have anincreased
level of smooth muscle tone. In addition, with age, there is achange in
the proportion of cells exhibiting pacemaker activity; however c-Kitimmunoreactive cells are present, indicating that the characteristics of the
PICmay change with age.Supported by the NH&MRC
Paper No.: 2454
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
CAN STEADY STATE PLASMA CONCENTRATION OF
EFAVIRENZ PREDICT VIROLOGIC OUTCOMES IN
TREATMENT NAÏVE HIV ETHIOPIAN PATIENTS?
Abiy H Eyakem(1,2), W Amogne(3), G Yimer(1), E Makonnen(1),
G Aderaye(3), A Worku(4), L Lindquist(6), L Bertilsson(2),
J Burhenne(5), E Aklillu(2)
(1) Addis Abeba University, Faculty of Medicine, Department of Pharmacology, Addis Ababa, Ethiopia
(2) Karolinska Institute, Department of Laboratory Medicine, Stockholm,
Sweden
(3) Department of Internal Medicine, Faculty of Medicine, Addis Abeba
University, Addis Abeba, Ethiopia
(4) Addis Abeba University, Faculty of Medicine, School of Public
Health, Addis Abeba, Ethiopia
(5) University of Heidelberg, Department of Clinical Pharmacology and
Pharmacoepidemiology, Heidelberg, Germany
(6) Karolinska Institute, Department of Internal Medicine, Stockholm,
Sweden
Previously, the therapeutic plasma concentration of efavirenz has been
suggested to range between 1 – 4 lg/mL. Efavirenz plasma concentration < 1 lg/mL is associated with therapeutic failure while > 4lg/mL is
associated with CNS toxicity. This study was designed to investigate the
steady-state plasma concentration of efavirenz and compare its relationship with the treatment outcome measured by virologic success among
treatment naı̈ve HIV infected Ethiopian patients. A cohort of HIV
infected adult participants (n = 118), naı̈ve for HAART with CD4 counts
less than 200 cells per mm3 were recruited. Participants received 600mg
daily dose of efavirenz. 16 h ± 1 h post-dose, blood samples were collected on the 4th week after initation of efavirenz based treatment. Steady
state plasma concentrations of efavirenz were determined using LC/MS/
MS and viral loads (VLs) were determined on week 24 using Abbott M2000 RT. The mean plasma efavirenz concentration at week 4 was
1.5 ± 0.12 lg/ mL while the median was 1.1 lg /mL. Proportion of participants in sub-therapeutic (<1 lg/mL), therapeutic (1- 4 lg/mL) and
toxic (< 4 lg/mL) ranges were 41.5%, 55.1% and 3.4% respectively.
The proportion of participants with virologic success among participants
with plasma efavirenz concentration <1, 1-4 and < 4lg/mL were 100%,
91.4% and 100% respectively. Our result indicates no association
between the described therapeutic range (1-4 lg/mL) categorization
based on plasma concentrations of efavirenz and therapeutic success
measured by VL among Ethiopian HIV infected individuals.
Paper No.: 1495
FOCUSED CONFERENCE GROUP: P12 - ION
CHANNELOPATHIES: NEW WINDOWS ON COMPLEX
DISEASE AND THERAPY
VARIABLE EXPRESSION OF ABCC8 AND ABCC9
TRANSPORTERS IN PREGNANT RAT MYOMETRIUM:
INFLUENCE OF GESTATION AGE
George Falkay, N Lovasz, E Ducza, R Gaspar
University of Szeged Faculty of Pharmacy, Department of Pharmacodynamics and Biopharmacy, Szeged, Hungary
Potassium channel activation has been shown to decrease uterine tone
which is considered as a target for inhibition of uterine activity in the
treatmet of preterm labour. This study was designed to examine the
expression and role of SUR1 (ABCC8) and SUR2 (ABCC9) subunits of
KATP channels in pregnant rat myometrium specially with regard to the
contractility. RT-PCR was performed to detect the presence of sulphonyurea binding regulatory subunit 1 and 2. The effect of the KATP channel opener pinacidil on the contraction provoked by electric field
stimulation (EFS) (50Hz) and by oxytocin were investigated on the isolated pregnant uterus. The SUR 1 and SUR 2 were markedly increased
at the early stage of the pregnancy. The highest level was detected at day
6 of the pregnancy while at late stage the levels of the transporters were
significantly decreased. Pinacidil inhibited the oxytocin and EFS induced
contraction. Glibenclamide as a selective blocker of KATP channel antagonised the pinacidil induced inhibition of contraction. The pharmacological reactivity of KATP channels might be attributed to the levels of SUR1
and SUR2 subunits. This is the first direct study to define the potencial
role of SUR subunits in the pregnant rat myometrium. These findings
may have a potencial therapeutic implication for managing preterm
labour in the future.
Acknowledgement:This work was supported by a Hungarian Otka
Research Grant (K62707).
Paper No.: 561
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
TANSHINONE IIA INHIBITS ER-A ACTIVITY IN MCF-7
BREAST CANCER CELL
Guanwei W Fan(1), XM Gao(1), H Wang(1), Y Zhu(1), J Zhang(2),
LM Hu(1)
(1) Tianjin University of Traditional Chinese Medicine, Tianjin, PR
China
(2) Nankai University, Tianjin, PR China
Phytoestrogens are a diverse group of plant-derived compounds that
structurally or functionally mimic mammalian estrogens and show potential benefits for human health. An increase in phytoestrogen research
over the past few decades has demonstrated the biological complexity of
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
275
phytoestrogens. Phytoestrogens can bind to estrogen receptors (ERs) and
mimic some actions of human estrogen through the activation or inactivation of certain genes.Tanshinone IIA (Tan IIA), a major component
extracted from a traditional herbal medicine Salvia miltiorrhiza BUNGE,
has been implicated as a chemopreventive agent against breast cancer.
However, the mechanisms behind it cancer-protective effects are not
clear. The goal of this study was to determine whether the antiproliferative action of this compound on breast cancer cell (MCF-7) is mediated
by estrogen receptors (ERs). The percentage of cell viability was evaluated by MTT assay. It was found that Tan IIA exhibited antiproliferative
effects in MCF-7 cells in the range of 10-6M–10-5M. In RT-PCR assay,
Tan IIA(10-5M) was found to inhibit pS2 expression at 6h (0.35 ± 0.27fold), at 24h (0.80 ± 0.32-fold) when compared to untreated control. In
transfection assay, Hela cells were co-transfected with pERE-luc, pTKRen, ER-a or ER-b, Tan IIA(10-6M–10-5M) decreased the ER-a transactivation. Our data unveil, that the anticancer action of Tan IIA in ER-a
positive breast cancer cells mediated, in part, by inhibit the activity of
ER-a and the expression of pS2.
Paper No.: 3218
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
VASORELAXANT EFFECTS OF COPTISINE IN RAT AORTIC
RINGS
Lian-Hua Fang, L-L Gong, J-J Hu, H-L Qin, Y Lv, G-H Du,
Chinese Academy of Medical Science & Peking Union Medical College,
Institute of Materia Medica, National Center for Pharmaceutical Screening, Beijing, PR China
Coptisine is an isoquinoline alkaloid isolated from Coptidis Rhizoma.
Several biological actions of Coptidis Rhizoma have been reported. The
present study was performed to investigate the vasorelaxant effects of
coptisine and its mechanism in isolated rat aorta. Coptisine(1~200?M)
inhibited norepinephrine(NE) and KCl induced sustained contraction
with pEC50 values of 4.49 ± 0.48 and 4.85 ± 0.57, respectively. Pretreatment of coptisine also suppressed the concentration-response curves
to NE and KCl. The vasorelaxant effect of coptisine was endothelium
dependent. In addition, the vasorelaxant effect of coptisine was partially
reduced by pretreatment with NjØ-nitro-L-arginine methyl ester, methylene blue and indomethacin, respectively. In endothelium-denuded aortic
rings, the vasorelaxant effect of coptisine was reduced significantly by 4aminopyridine. However, glibenclamide and tetraethylammonium failed
to do so. Moreover, coptisine inhibited not only the NE-induced transient
contraction caused by cytosolic Ca2 + release from endoplasmic reticulum in Ca2 + -free medium, but also the contractions induced by 60 mM
KCl buffer with different Ca2 + concentrations. This study showed that
coptisine inhibited both endothelium-dependent and -independent contraction in rat aortic rings. These results suggested that NO-cGMP pathway may be involved in the endothelium-dependent relaxation of
coptisine, and activation of voltage-dependent K+ channel contributes in
part to the endothelium-independent relaxation of coptosine. Furthermore, coptisine may inhibit the extracellular Ca2 + influx by blocking
both voltage- and receptor-operated Ca2 + channel.
Acknowledgements: This study was supported by special foundation on
scientific & technological basic work(2007FY130100) and national scientific & technological major special project(2009ZX09302-003,
2009ZX09501-021) provided by China Ministry of Science and Technology. Corresponding authors: dugh@imm.ac.cn (G.-H. Du).
Paper No.: 2956
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
DISTRIBUTION OF METHYLPHENIDATE AND RITALINIC
ACID IN ORAL FLUID
Magi Farre(1), R Pardo(1), E Marchei(2), O Garcia-Algar(3), M Pellegrini(2), R Pacifici(2), S Pichini(2)
(1) IMIM-Hospital del Mar, UAB, Department of Human Pharmacology
and Neurosciences, Barcelona, Spain
(2) Department of Therapeutic Research and Medicines Evaluation, Istituto Superiore di Sanita, Rome, Italy
(3) IMIM-Hospital del Mar, UAB, Department of Pediatrics, Barcelona,
Spain
Introduction: Oral fluid as an alternative matrix for monitoring drug use
due to its ease-of-collection and non-invasiveness. The aim of this study
was to evaluate the pharmacokinetic profile of methylphenidate (MPH)
and its metabolite ritalinic acid (RA) in oral fluid and in plasma after
controlled MPH administration. Materials/Patients: Eight male healthy
volunteers participated in the study. Subjects received a single dose of
20 mg short-release (n = 5) or extended-release MPH (n = 3) by oral
route. Samples of oral fluid and plasma were collected along 24 hours.
Concentrations of MPH and RA in oral fluid and plasma were measured
by liquid chromatography–electrospray ionization– mass spectrometry
using a validated method. Apparent pharmacokinetic parameters of MPH
in plasma and oral fluid were estimated. Results: MPH and RA were
detected in oral fluid. Whereas parent drug concentrations in oral fluid
were one order of magnitude higher than those in plasma, the opposite
was observed for RA, which resulted 10 times more concentrated in
plasma. Oral fluid concentrations of MPH ranged between 0.5-466.7
mcg/L and peaked at 0.5 h after short release administration and ranged
between 0.7-89.5 mcg/L and peaked at 2 h after extended release administration. Conclusions: The results obtained support the measurement of
MPH in oral fluid as a valuable alternative to drug monitoring in plasma
both in clinical and toxicological studies. Acknowledgements.
Supported in part by grants from ISS-Rome, FIS-RTA (RD06/0001/
1009), 2009 SGR 718 and FIS (R. Pardo is the recipient of a fellowship
Rio Hortega, no. CM08/00051).
Paper No.: 2957
FOCUSED CONFERENCE GROUP: P04 - PHARMACOEPIDEMIOLOGY, CURRENT CONTROVERSIES AND OPPORTUNITIES
MINI-CASES AS A METHOD TO DYNAMIZE LECTURES OF
AUTONOMIC NERVOUS SYSTEM PHARMACOLOGY IN A
MEDICAL SCHOOL
Magi Farre(1), J-E Baños(2)
(1) IMIM-Hospital del Mar, UAB, Dept. Human Pharmacology and Neurosciences, Barcelona, Spain
(2) Universitat Pompeu Fabra, Department of.Experimental and Health
Sciences, Barcelona, Spain
Introduction: We report here a preliminary study using a series of minicases or very short problems in conjunction with traditional lectures for
teaching medical students. Materials: We used ten mini-cases in our lectures on autonomic nervous system pharmacology. Mini-cases consist of
a five-line text with two questions on the subject of the lecture. They
were delivered just before the lecture, and students were given five minutes to answer the questions at the end. Teachers evaluated their correctness and were able to award extra points to the overall mark in the
regular exam. At the end of term the experience was evaluated by asking
students to answer ten questions anonymously. Results: Attendance to
lectures increased when mini-cases where used in comparison to other
lectures of different parts of the Pharmacology program. Students handed
in a mean of 7-8 mini-cases and gave a mean score of 7.5 for their usefulness in understanding the subject better and 7 for their interest. When
asked about their reasons for taking part, they scored 8.5 for the possibility of obtaining an extra mark, 7.5 for the idea that they would understand the subjects better. Around 70% reported that mini-cases increased
attendance at lectures and 95% said they would participate even if extra
marks were not available. All participants reported that they would participate in similar experiences in the future. Conclusions: We conclude
that mini-cases may be a cost-effective method for improving students’
attendance at lectures, as well as their understanding of the autonomic
nervous system pharmacology.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
276
Paper No.: 1026
FOCUSED CONFERENCE GROUP: P17 - NEW APPROACHES
AND TARGETS IN PSYCHIATRY
ANXIOLYTIC-LIKE EFFECT OF SKF-38393 IN COMBINATION
WITH LOW DOSE OF 17B-ESTRADIOL IN
OVARIECTOMIZED RATS
Paper No.: 1048
FOCUSED CONFERENCE GROUP: P02 - TRANSMEMBRANE
TRANSPORT: PERSPECTIVES FOR DISEASE AND DRUG
DISCOVERY
CANNABIDIOL INHIBITS BCRP AND P-GP IN HUMAN
PLACENTAL TROPHOBLAST CELL LINES BEWO AND JAR
Yulia Fedotova
Valeria Feinshtein(1), G Holcberg(2), Z Ben-Zvi(1)
Institute for Experimental Medicine, Department of Neuropharmacology,
St Petersburg, Russian Federation
(1) Ben-Gurion University, Department of Pharmacology, Beer-Sheva,
Israel
(2) Soroka University Medical Center, Beer-Sheva, Israel
Dopamine systems are activated by stress and involved in stress-related
behaviors such as anxiety. On the other hands, estrogens have been
implicated in anxiety behavior. Recent evidence has shown that estrogen
receptors can be activated by a variety of neurochemical compounds,
including the dopaminergic ligands acting on D1-receptor. The aim of
the present study was to explore the antianxiety-like effects of D1-receptor agonist SKF-38393 (0.1 mg/kg, i.p.) or D1-receptor antagonist SCH23390 (0.1 mg/kg, i.p.) alone or in a combination with low dose (0.5
mkg/rat, s.c.). The anxiety-like behavior was assessed in the white model
(BWM) and the open field test (OFT). In the BWM, SKF-38393 significantly reduced anxiety behavior in OVX females as compared with control group. SKF-38393 in a combination with low dose of 17b-E2
induced more profound anxiolytic effect in OVX rats as compared to
OVX rats treated with SKF-38393 alone. In the OFT, SKF-38393 alone
or in combination with 17b-E2 increased grooming and rearing without
significant changes in locomotor activity as compared to OVX and control groups. SKF-38393 simultaneously restored mRNA expression of
D1- and 17b-E2-receptors in the hypothalamus. In conclusion, SKF38393 alone or in combination with low dose of 17b-E2 may exert antianxiety-like effect on OVX females through D1- and 17b-E2-receptors,
via altering the expression of these receptors in the brain. Thus, the
results demonstrate that 17b-E2 and SKF-38393 behaviorally and neurochemically interact to enhance anxiolytic action and can potentiate effects
of each other.
Paper No.: 2274
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
ROPREN IS A POLYPRENOL PREPARATION FROM
CONIFEROUS PLANTS THAT AMELIORATES COGNITIVE
DEFICIENCY IN A RAT MODEL OF B-AMYLOID
PEPTIDE-(25-35)-INDUCED AMNESIA
Yulia Fedotova(1), V Soultanov(2), V Roschin(3), T Nikitina(4)
(1) IP Pavlov Institute of Physiology RASCi, Department of Neuroendocrinology, St.Petersburg, Russian Federation
(2) Solagran Limited, Melbourne, VIC, Australia
(3) State Forest Technical Academy, Department of Wood Chemistry, St
Petersburg,Russian federation
(4) IM Sechenov Institute of Evolutionary Physiology and Biochemistry,
RussianAcademy of Science, Laboratory of Functional Biochemistry of
Invertebrates, StPetersburg, Russian Federation
This study assesses the efficacy of a fixed dose of a novel preparation
Ropren (a plant preparation isolated from the neutral fraction of an
extract of spruce and pine needles) on cognitive impairments in rats with
b-amyloid peptide-(25-35)-induced amnesia. Ropren was administered
at a dose of 8.6 mg/kg for 28 days, per os, to rats with b-amyloid peptide-(25-35)-induced amnesia. Cognitive performance was assessed using
the passive avoidance paradigm and the Morris water maze. The behavior of animals was assessed in the open field test. After four weeks,
Ropren treatment significantly improved non-spatial and spatial learning in rats with b-amyloid peptide-(25-35)-induced amnesia. The results
of the present study suggest that Ropren, a novel plant preparation,
ameliorates cognitive deficiencies in an animal model relevant to Alzheimer’s disease.
Cannabis is the most commonly used illegal drug, when approximately
4% of women in the USA report using illicit drugs during pregnancy,
with marijuana being the most popular one (75%). BCRP and P-gp are
efflux transporters expressed at the apical membrane of the syncytiotrophoblast and are putatively important protective proteins for the fetus.
To day, very little is known about the influence of cannabinoids on
BCRP and P-gp in the placental barrier. Our study focused on cannabidiol (CBD) effect on the functionality of BCRP and P-gp in BeWo and Jar
cells, while MCF-7 cells were used as control for BCRP function properties. CBD cytotoxycity was tested first. Non toxic concentrations of
CBD were then used in Mitoxantrone (MX) (substrate of P-gp and
BCRP) uptake assay. Cells were preincubated with/without CBD for
30min, followed by MX addition and further incubation for 30min.
Nicardipine was used as P-gp inhibitor. CBD 25 and 10 lM significantly
inhibited BCRP dependant MX efflux in MCF-7, BeWo and Jar cells
(for 10uM – 40%, 25% and 22%, respectively). Nicardipine inhibition of
MX efflux in Jar and BeWo cells was significantly lower compared to
CBD inhibition of MX efflux, indicating that CBD inhibited both BCRP
and P-gp dependant MX efflux. We showed that BCRP and P-gp efflux
function can be inhibited after only 1h exposure to CBD. These findings
indicate that marijuana consumption during pregnancy (even at the very
early stages of gestation) could jeopardize fetal wellbeing by inhibiting
active efflux placental barrier transporters.
Paper No.: 2701
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
EFFECT OF ULVA LACTUCA IN LIVER CONSERVATION IN
WISTAR RAT MODEL
H Ferchichi, I Salouage, S Bacha, L Ouanes, M Lakhal, Anis Klouz
(National Center of Pharmacovigilance, Laboratory of Clinical Pharmacology, Tunis, Tunisia
The purpose of our work is to evaluate the extracts stemming from the
green seaweed Ulva lactuca to improve the organs conservation solution
against cold hepatic-ischemia effects in Wistar rats. Rats were randomized between 3 groups: - Sham group (shortly preserved): livers were
washed with 0.9% NaCl, and then placed in a preservation solution
(Krebs–Henseleit) for 30 min at 4 C. - For the other two groups (no
treated and treated group), livers were preserved under the same conditions for 24 h. For treated group, Ulva lactuca solution was added to the
preservation solution at a concentration of 40 ı̀g/ml. After liver conservation, we realized an extraction of mitochondria for the dosage of malondialdehyde (MDA) and to evaluate the mitochondrial respiration.
Transaminase levels (AST and ALT) were assessed in the preservation
liquid. A fragment of liver used for histological analysis. The results of
AST and ALT levels decrease significantly in the treated group (201 and
163 U/I) compared with no treated group (716 and 466 U/I) respectively.
The amelioration percentage of lipoperoxidation inhibition was 15,21%
and the amelioration percentage of mitochondrial activity was 13%,
between treated and no treated groups. The necrotic areas were irreversible in the no treated group. Extensive vacuolar degeneration (reversible
lesion) was found in the treated group. The addition of the Ulva extract
in standard conservation solution permitted us to avoid deleterious effects
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
277
of the cold ischemia in Wistar rats. The use of this extract will improve
the graft quality before its transplantation.
Paper No.: 1522
FOCUSED CONFERENCE GROUP: P14 - ADDICTION AND
DOPING: NEUROBIOLOGICAL AND CLINICAL
ACETALDEHYDE STIMULATES IL-6 MRNA EXPRESSION
AND IL-6 PROTEIN SECRETION IN CULTURED RAT
ASTROCYTES
Polonca Ferk(1), L Sarc(2), B Wraber(3), M Lipnik-Stangelj(3)
(1) University of Maribor Faculty of Medicine, Department of Pharmacology and Toxicology, Maribor, Slovenia
(2) University Medical Centre, Poison Control Centre, Ljubljana,
Slovenia
(3) University of Ljubljana, Faculty of Medicine, Ljubljana, Slovenia
Several disorders in the CNS, related to excessive ethanol consumption,
seem to be associated with altered cytokine production. Astrocytes are
important source of several cytokines, including a pro-inflammatory
cytokine IL-6, and significantly contribute to the immune response in the
brain. Recent studies have shown that some of the ethanol effects are
mediated through acetaldehyde, the primary metabolite of ethanol. We
hypothesized that acetaldehyde could be, at least partly, responsible also
for ethanol immunomodulatory effects in the brain. Specifically, we
investigated a possible influence of acetaldehyde on the IL-6 mRNA
expression and IL-6 protein secretion in cultured rat astrocytes. Primary
cultures of cortical astrocytes were prepared from the brain of neonatal
Wistar rats. The cells were incubated with 1mM acetaldehyde, for
4 hours, 24 hours (acute exposure) or for 7 days (chronic exposure).
Total RNA was isolated and IL-6 mRNA expression assessed by quantitative real-time PCR. IL-6 protein secretion was determined using appropriate ELISA protocol. Acetaldehyde increased the IL-6 mRNA
expression after 4 hours of incubation and IL-6 protein secretion after
acute and chronic exposure. The IL-6 protein secretion was dose-dependent and time-dependent. Short-term exposure to acetaldehyde showed
higher stimulation of IL-6 protein secretion compared to long-term exposure. We confirmed that ethanol immune effects in the brain could be at
least partly due to its major metabolite, acetaldehyde. However, further
studies are needed to quantify the contribution of acetaldehyde to the
overall brain damage after ethanol exposure.
Paper No.: 2837
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
TRPA1 PLAYS AN IMPORTANT ROLE IN TNFA-INDUCED
MECHANICAL HYPERALGESIA
carried out using mixed ANOVA followed by the Sidak test. P < 0.05
was considered significant. Animals received TNFa (10 pmol/paw, ipsilateral paw) and Tyrode (50 ll/paw, contralateral paw). TRPA1 involvement was analyzed in WT and TRPA1 knockout (TRPA1KO) mice and
CD1 mice treated with the selective TRPA1 receptor antagonist AP-18
(i.pl., 25 nmol/paw, co-injected with TNFa or intrathecal, prior to
TNFa). TNFa evoked a significant bilateral hyperalgesia in WT mice
that lasted up to 7 days. In contrast, TRPA1KO mice exhibited significant less hyperalgesia after TNFa injection. Additionally, both i.pl. and
intrathecal treatment with AP-18 significantly prevented TNFa-induced
hyperalgesia in CD1 mice. Herein, these results show that TRPA1 is
important to the development of TNFa-induced mechanical hyperalgesia.
Thus, we present novel evidence of TRPA1 participation in pain processing. We suggest TRPA1 plays a role in maintaining inflammatory hyperalgesia associated with TNFa. Keeble, J. et al. (2005) Arthritis Rheum.
52: 3248-3256. Russell FA, et al. (2009) Pain. 142: 264-274. This work
was supported by ARC and the BBSRC/Pfizer.
Paper No.: 791
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
MEROPENEM CONTROLLED-RELEASE SYSTEM
Gonzalo Fernández Govantes, JJT Durán, ST Durán
Universidad Complutense de Madrid, Farmacia y Tecnologı́a
Farmacéutica, Madrid, Spain
Surgical infections are one of the main problems in hospitals, not only
because they pose a risk for the patients, but also due to their economic
implications. The aim of this project is to obtain controlled-release systems of meropenem for implantation and use them as a local treatment in
surgery, as it is a broad spectrum antibiotic and it can prevent the colonisation of bacteria. The materials used are meropenem and polylacticcoglycolic acid, which is a biodegradable polymer. Initially, meropenem
was validated following the USP method, based on HPLC. Afterwards,
the polymer was atomized by spray drying, obtaining microspheres of
PLGA. Finally, tablets baring a small diameter were obtained by direct
compression (Murakami H et al, J. Controlled Release. 2000, 67:29-36).
Both microspheres and tablets have been assayed. The microsphere
assays included size, morphology and residual solvents by GC; and
weight, porosity, morphology, delivery assay, and other properties for the
tablets. It resulted that microspheres had a diameter of 10 lm, were
roughly spherical and had an insignificant proportion of dichloromethane. On the other hand, the tablets produced had diameter of 7 mm,
weight about 95 mg and provided an optimal delivery of meropenem. In
the future, these tablets will be assayed in laboratory animals. In conclusion, it is possible to obtain local delivery systems of meropenem with
favourable properties, which could provide higher efficiency and lower
adverse effects than current therapies.
Elizabeth S Fernandes(1), FA Russell(1), D Spina(2), R Graepel(1),
C Gentry(3), J Keeble(4), S Bevan(3), SD Brain(1)
(1) King’s College London, Cardiovascular Division, London, UK
(2) Sackler Institute of Pulmonary Pharmacology, London, UK
(3) Wolfson Centre for Age-related Diseases, London, UK
(4) King’s College London, Pharmaceutical Sciences Division, London,
UK
The mechanisms linking TNFa and inflammatory pain are unclear. Previously, we showed that TRPV1 is involved in TNFa-induced thermal hyperalgesia (Russell et al., 2009) and that TNFa levels are raised in a
model of CFA-induced joint inflammation (Keeble et al., 2005). Here,
we investigated the contribution of TRPA1 receptors in inflammatory hyperalgesia induced by intraplantar (i.pl.) TNFa injection. Mechanical hyperalgesia was assessed in both hind paws by using an automated Von
Frey system before (baseline) and after treatment. Statistical analysis was
Paper No.: 2714
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
ASPARAGUS OFFICINALIS REGULATES CHOLESTEROL
METABOLISM AND IMPROVES ANTIOXIDANT STATUS IN
HYPERCHOLESTEROLEMIC RATS
Aangeles Fernández-Arche, D Garcı́a -Gonzalez, R Saenz-Rodriguez,
R Puerta-Vazquez
University of Sevilla, Department of Pharmacology, Sevilla, Spain
Asparagus officinalis L. has a high antioxidant capacity, associated to its
total phenolic content. There are large population studies showing that
vegetables rich in antioxidants are associated with a low incidence of
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
278
cardiovascular diseases. Objetive: To evaluate the effect of Asparagus officinalis on plasma lipids levels in rats fed a cholesterol-rich diet. Methods: The rats in the control group were fed a standard laboratory diet; the
rats in the hypercholesterolemic group were fed a cholesterol-rich diet .
The rats in others groups were treated with freeze-dried asparagus at a
dose of 500, 250, and 125 mg/Kg of body weight /day, while being fed
the cholesterol-rich diet. At the end of treatment animals were sacrificed
and the livers were collected and weighed. According to the standard
methods, total cholesterol (TC), triglicerides (TG), and high density lipoprotein cholesterol (HDLc) of the plasma, were analyzed. Low density
lipoprotein cholesterol (LDLc) and atherogenic index was calculated by
using Friedenwald’s formula and the equation: TC/HDL-c, respectively.
Results: Asparagus, significantly decreased the plasma level of TC
(P < 0.01) against the hypercholesterolemic group. LDLc values in the
asparagus groups were lower than the hypercholesterolemic group
(p < 0.05), HDL-c levels, a beneficial effect was observed although the
increase was not significative. TG was slightly reduced but the effect
was not significant against the reference group. The antioxidant status
(catalase and superoxide dismutase activities) and protection against lipid
peroxidation was also improved.
Paper No.: 1157
FOCUSED CONFERENCE GROUP: P19 - GENERAL SESSION HEPATOLOGY
CONGENITAL-NEONATAL HYPOTHYROIDISM IMPACTS
HEPATIC LIPID METABOLISM IN ADULTHOOD
Leandro Fernández-Pérez(1), R Santana-Farré(1), L Henrı́quez-Hernández(1), C Bocos(2), N Kahlon(3), E Herrera(2), N Gunnar(3), P Parini(4), A Flores-Morales(3),
(1) University of Las Palmas de Gran Canaria, Department of Clinical
Sciences, Molecular and Translational Endocrinology Group, Cancer
Research Institute of The Canary Islands (ICIC), Spain
(2) University of San Pablo-CEU, Department of Biochemistry and
Molecular and Cellular Biology, Faculties of Pharmacy and Medicine,
Spain
(3) Karolinska Institute, Department of Molecular Medicine and Surgery,
Sweden
(4) Karolinska Institute Huddinge, Division of Clinical Chemistry,
Department ofLaboratory Medicine, and Molecular Nutrition Unit,
Center for Nutrition andToxicology, Sweden
Thyroid hormones (TH) are essential for development, growth, and
metabolism. If not treated immediately, congenital hypothyroidism (CH)
has a profound impact on physiology and can imprint neurological and
endocrine systems, which, in turn, led to mental retardation, growth
arrest, and metabolic disturbances. To test if CH may influence liver gene
expression program in adulthood, pregnant rats were given the antithyroid drug methimazole between gestational day 12 and post-natal day
(PND) 30 to induce CH in male offsprings. Upon weaning at PND30,
exposure to MMI was ceased. On PND80, we analyzed changes in
serum and hepatic lipids and liver transcriptome to explore whether CH
influences liver physiology in adulthood in comparison with age-matched
rats without CH (INTACT). Body weight of PND80 rats with CH was
reduced; however, serum levels of T3, cholesterol, and triglycerides
showed no significant differences in relation to INTACT. In contrast,
hepatic content of fatty acids, triglycerides, cholesteryl esters, and phospholipids were reduced in the CH group. Liver transcriptome was significantly modified by CH. The mRNA levels several genes associated with
metabolism and transport of lipids were significantly modified and differently-expressed genes were mostly mapped into lipid metabolism GO
categories as well as PPARalfa and complement-coagulation cascades
signalling pathways. In summary, we provide in vivo evidence that CH
causes long-lasting effects on hepatic functions in part through changes
in transcriptome. This underscores the critical role of T3 during neonatal
life for maintaining a normal developmental program of growth and lipid
metabolism.
Paper No.: 1105
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
MODULATION OF TH2-CYTOKINES PRODUCTION AND
ADHESION MOLECULE EXPRESSION ON BONE MARROW
AND PERIPHERAL BLOOD EOSINOPHILS IN ALLERGIC
MICE BY INDUCIBLE NITRIC OXIDE SYNTHASE
Heloisa HA Ferreira(1), EH Pellaquini(1), LR Benetti(1), LGR Fernandes(2), WMSC Tamashiro(2)
(1) San Francisco University, Laboratory of Inflammation Research, São
Paulo, Brazil
(2) UNICAMP, Department of Microbiology and Immunology, Campinas, SP, Brazil
We investigated the role of the inducible nitric oxide isoform (iNOS) in
the migration of eosinophils to the lungs and the mechanisms by which
NO exerts these effects. Results demonstrate that the peak of eosinophil
infiltration into the lungs of ovalbumin-sensitised control (non-treated)
mice, seen at 48h after antigen challenge, was significantly reduced by
treatment of allergic mice with iNOS inhibitor (1400W). Flow cytometric
analysis showed that treatment resulted in a significant increase in Mac-1
expression (85% increases) on bone marrow (BM) eosinophils at 24h, as
compared to control mice. However, at 24h, 1400W decreased Mac-1
and VLA-4 expressions on eosinophils by about 40% and 76%, respectively, in the peripheral blood (PB). In contrast, increases in both Mac-1
and VLA-4 expressions (42% and 64%, respectively) were observed at
48h on PB eosinophils. These results show a temporal effect of NO upon
the expression and function of Mac-1, which is the chief adhesion molecule involved in eosinophil efflux from the BM at 24h. In contrast, Mac1 and VLA-4 were involved in eosinophil mobilization from PB to lungs
at 48h after antigen challenge. Furthermore, 1400W significantly reduced
lung tissue levels of IL-4 at 24 and 48h and IL-5, IL-13 and eotaxin at
48 and 72h after OVA-challenge. In conclusion, results suggest the
involvement of NO in the modulation of Th2-derived cytokines and
eotaxin secretion in the lungs and in the expression of adhesion molecules on BM and PB eosinophils that may influence cell migration to the
lungs of allergic mice.
Support: FAPESP
Paper No.: 786
FOCUSED CONFERENCE GROUP: PW12 - EXPERIMENTAL
PAIN METHODS IN DEVELOPMENT AND VALIDATION OF
ANALGESICS
CHARACTERIZATION OF THE ANTINOCICEPTIVE AND
ANTI-INFLAMMATORY ACTIVITIES OF NICOTINAMIDE
AND ITS ISOMERS IN DIFFERENT EXPERIMENTAL MODELS
Wallace Ferreira, A Godin, L Rocha, R Vieira, E Nascimento, J Seniuk,
M Coelho
Federal University of Minas Gerais, Department of Pharmacology &
Pharmaceutic Products, Belo Horizonte, Brazil
Nicotinamide, member of the vitamin B3 family, presents antinociceptive
activity and there are also evidences of its anti-inflammatory property
(CUZZOCREA, S et al., Life Sci., 1999; v. 65: p. 1297,). However, there
is not any information about effects induced by picolinamide or isonicotinamide, isomers of nicotinamide, on the nociceptive or inflammatory
responses. To establish a structure-activity relationship, we have investigated the effects induced by the three isomers in models of nociceptive
and inflammatory pain and edema. Female Swiss mice were used. Nicotinamide, isonicotinamide (500 or 1000 mg/kg) or picolinamide (62.5 or
125 mg/kg) were administered per os. The nociceptive response was
evaluated in the formalin model. In the model of carrageenan-induced
edema, each isomer was administered 1 h before and 2 h after the
inflammatory stimulus. The motor activity was investigated using a
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
279
rota-rod apparatus. The results were analyzed by one way Anova followed by Newman-Keuls test (p < 0.05). Nicotinamide (1000 mg/kg)
inhibited the first and second phases of the formalin model. Only the second phase of this response was inhibited by isonicotinamide (500 or
1000 mg/kg) or picolinamide (125 mg/kg). Nicotinamide (1000 mg/kg),
picolinamide (125 mg/kg) and isonicotinamide (500 or 1000 mg/kg)
inhibited the paw edema. None of the substances impaired the motor
activity of mice. This study shows that the antinociceptive and antiinflammatory activities of nicotinamide extend to its isomers. Despite
some evidences of potential molecular targets, the mechanisms that contribute to these activities are still uncertain. The substances may prove to
be useful as anti-inflammatory and analgesic drugs.
Paper No.: 787
FOCUSED CONFERENCE GROUP: PW12 - EXPERIMENTAL
PAIN METHODS IN DEVELOPMENT AND VALIDATION OF
ANALGESICS
CHARACTERIZATION OF THE ANTINOCICEPTIVE AND
ANTI-INFLAMMATORY ACTIVITIES OF NICOTINIC ACID
AND ITS ISOMERS IN DIFFERENT EXPERIMENTAL MODELS
Wallace Ferreira, A Godin, L Rocha, R Vieira, E Nascimento, J Seniuk,
M Coelho
Federal University of Minas Gerais, Department of Pharmacology &
Pharmaceutic Products, Belo Horizonte, Brazil
Nicotinic acid is one of the two forms of the B3 vitamin. In addition to
being a nutrient, it is a clinically applied drug. In high doses, it has been
used as a lipid-lowering agent (BODOR, ET et al., Br. J. Pharmacol.,
2008; v. 153: p. 568,). Nicotinamide, the amide derivative of vitamin B3,
exerts a number of anti-inflammatory properties (CUZZOCREA, S et al.,
Life Sci., 1999; v. 65: p. 1297,). Thus, we investigated the effects
induced by nicotinic acid and its isomers, picolinic and isonicotinic
acids, in models of nociceptive and inflammatory pain and also edema,
as these effects have not been investigated. In the formalin model in
female Swiss mice, nicotinic (250 or 500 mg/kg), picolinic (62.5 or
125 mg/kg) or isonicotinic (250 or 500 mg/kg) acids were administered
per os, 1 h before the injection of the noxious stimulus. The evaluation
of motor activity was evaluated in the rota-rod. When paw edema was
induced by carrageenan, each isomer was administered 1 h before and
2 h after the inflammatory stimulus. Results were analyzed by one way
Anova followed by the Newman-Keuls test. The results show the antinociceptive activity of nicotinic and picolinic acids and also the anti-inflammatory activity of nicotinic acid. The study shows the structure-activity
relationship of the molecules. Despite the lack of precise information of
its mode of action, nicotinic acid is safe, approved for clinical use, and
represents a potential analgesic and anti-inflammatory agent. More
research is needed to elucidate its mechanisms of action.
Paper No.: 1790
FOCUSED CONFERENCE GROUP:
P19 - GENERAL SESSION - ONCOLOGY
BEER POLYPHENOLS AGAINST HUMAN GLIOBLASTOMA:
EFFECTS OF XANTHOHUMOL ON T98G CELL APOPTOSIS
Michela Festa, A Capasso, L Parente, CW D’Acunto, C Pizza, S Piacente
(1) University of Salerno, Department of Pharmaceutical Science, Fisciano, Salerno, Italy
Glioblastoma multiforme (GBM) is the most common primary brain
tumor in humans and has a median survival of 6 months from the time
of diagnosis. The lack of effective current therapy, including surgery,
radiation and chemotherapy, has not significantly improved the prognosis
of patients with GBM (Terzis et al, Opin Biol Ther. 2006; 6:739-749).
Thus, new therapeutic strategies need to be explored. Xanthohumol
(XH), a prenylated chalcone of the hop plant (Humulus lupulus L.), has
shown cancer chemopreventive, antiproliferative and anticancer activities
in several human malignancies (Zanoli & Zavatti, J Ethnopharmacol.
2008; 116:383-96). The aim of this study was to investigate the effect of
XH on human glioblastoma T98G and U87MG cells. Xanthohumol
induced apoptosis in concentration-dependent manner in T98G but not in
U87MG cells. XH induced apoptosis in T98G cells by elevating intracellular oxidative stress generating reactive oxygen species (ROS). XH
treatment resulted in activation of the mitogen-activated protein kinase
(MAPK) ERK1/2, and pretreatment with an ERK inhibitor PD98059 partially reduced XH-induced apoptosis. We found that XH-mediated production of a large amount of ROS caused activation of ERK1/2 and
changes in mitochondrial membrane potential. Pretreatment with the antioxidant N-acetylcysteine (NAC) strongly inhibited the XH-induced generation of ROS, phosphorylation of ERK1/2, and apoptotic cell death.
XH-induced apoptotic cell death was mediated by activation of the effectors caspases and cleavage of PARP-1 and was blocked by pretreatment
with Z-VAD-FMK (a pan-caspase inhibitor). These results suggest the
potential application of xanthohumol as a novel chemiotherapic agent for
the treatment of glioblastoma.
Paper No.: 2979
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
CENTRAL NERVOUS SYSTEM PATHOLOGY DURING
PREGNANCY. STUDY OF 25 CLINICAL CASES USING AN
EVIDENCE-BASED APPROACH
Isabel Vitória Figueiredo(1,2), AC Rama(1,2,3), O Isabel(3),
F Tavares(4), T Santos(4), F Fernández-Llimós(5),
MM Castel-Branco(1,2), A Cabral(1), M Caramona(1,2)
(1) University of Coimbra, Faculty of Pharmacy, Department of Pharmacology, Coimbra, Portugal
(2) University of Coimbra, Center for Pharmaceutical Studies, Coimbra,
Portugal
(3) Coimbra University Hospitals, Pharmacy Department, Coimbra,
Portugal
(4) Coimbra University Hospitals, Department of Maternal Foetal
Medicine, Coimbra, Portugal
(5) University of Lisbon, Faculty of Pharmacy, Department of Social
Pharmacy, Lisbon, Portugal
Central nervous system (CNS) pathology during pregnancy is becoming
more frequent among our pregnant women. Confronted with the complexity in managing foetal risk, efficient literature searching in addition
to the type and accuracy of clinical and medication data collection are
becoming crucial skills for evidence based practice as a support to clinical decision. Study of 25 cases of women with CNS diseases that were
pregnant or planned to be and were exposed to drugs. Assessment of
quality/utility of information provided to support clinical decision. Medical records analysis for pregnant, foetus and newborn follows-up. Evaluation of the parameters related to the likely effects described. Clinical
queries (2005-2009) of 25 cases were examined. 23 pregnants: 6 with 512/weeks and 17 with 13-24/weeks of gestation. Ages ranged from 1639 years. Pathologies occurring more frequently: depression, anxiety and
epilepsy. Associated conditions: hypertension, hypercholesterolaemia,
asthma. They were exposed to 33 CNS drugs, alone or in association
(with higher prevalence of diazepam, paroxetine and alprazolam) and to
22 other drugs. Information provided was considered reliable, accessible,
complete and applicable and used for clinical decision in all cases. There
was the possibility of establishing a causal relationship in 5 cases, where
newborns had abstinence and/or floppy child syndromes. One quarter of
pregnant women came to us in the first trimester. There were 5 newborns
in 16 presenting withdrawal syndrome and floppy child syndrome. We
propose to create a structured formulary for collecting information oriented to medication, disease and clinical situation of the pregnant, foetus
and newborn.
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
280
Paper No.: 1633
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
ADVERSE DRUG REACTIONS IN INFANTS AND CHILDREN
Houda Filali, A Elgharbi, F Hakkou
Ibn Rochd Hospital, Department of Pharmacology, Casablanca, Morocco
The cause of iatrogenic accidents drug preferentially affects extreme
ages. Objective: To analyze the case reports concerning children and
infants (18 years or younger) in Ibn Rochd Hospital Materials and methods: Cases report of adverse drug accidents (ADRs) in patients aged 18
and under are analyzed in a retrospective study of 1 year (Sept.2008Sept.2009). Finding: Eighty-two ADRs reported, representing 23.4% of
all ADRs collected. The main events are 24 toxidermas, 24 digestive disorders, 10 haematological and neurological disorders, 6 disturbances of
general condition, 5 events nephro-urological. We also identified 14
overdose caused by therapeutic errors. The major drug makers are antiinfectives (37.8%), antineoplastics (23.1%), anti-inflammatories (14.6%),
drugs of neuro-psychiatric (10%) with antiepileptic (6%). The high number of daily medications promotes the risk of drug interactions responsible for iatrogenic injuries in 14.6% of patients in this study. Off label
prescriptions were identified in 10.9% of cases. Conclusions: Adverse
drug reactions are not common in paediatric patients, and most are mild.
However, due to limitations of clinical trials in children, pharmacoepidemiological studies may be the only source of information on the benefitrisk profile of drugs received by these patients, and as such require special attention.
Paper No.: 1634
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
CAUSES OF THERAPEUTIC ERRORS IN OLDER ADULTS
Houda Filali, H Elkarimi, D Mzah, A Miftah, F Hakkou
Ibn Rochd Hospital, Department of Pharmacology, Casablanca, Morocco
Introduction: The Sweet syndrome or acute febrile neutrophilic dermatosis is a table whose toxidermie stereotype is that of fever, leukocytosis with neutrophils, a rash made with painful plaques and dense
dermal infiltration with neutrophils histologically. The diagnosis is
based on a series of arguments. The forms of the drugs heal after
stopping the drug responsible. Observations: We report two cases of
Sweet’s Syndrome Drug: the first is a woman of 52 years treated for
10 days amoxicillin-clavulanate for furunculosis foot. 2 days after
completion of treatment, she presented an eruption febrile generalized
erythemato-papular. Laboratory studies showed a major inflammatory
syndrome with leukocytosis with neutrophils. Histological examination
of a skin biopsy confirmed the diagnosis of Sweet’s syndrome. Systemic corticosteroids led to the disappearance of the clinical and biological symptoms. The second observation concerns a woman aged
60, also treated with amoxicillin-clavulanate for angina. At days 5 of
the treatment, she developed papulopustular rash erythematous confluent bunched on the hands and feet. Laboratory tests and histology
were similar to the first case. The evolution was marked regression of
lesions after steroid therapy. Analysis and discussion: In two cases,
Sweet’s syndrome is well defined criteria of Sweet’s syndrome drug
proposed by Walker and Cohen. In addition, the study of accountability inherent in accordance with the WHO method has proved plausible
for the two cases, based on chronological criteria C2 and semiotic S2,
while the extrinsic accountability found a score B1 not known for the
amoxicillin-clavulanate.
Paper No.: 1350
FOCUSED CONFERENCE GROUP:
P11 - G PROTEIN-COUPLED 7TM RECEPTORS: FROM
MOLECULAR TO PHYSIOLOGICAL FUNCTION
THE SCAFFOLD PROTEIN NHERF2 DETERMINES
NEURONAL ION CHANNEL COUPLING OF GLUTAMATE
MGLUR5 AND NUCLEOTIDE P2Y1 RECEPTORS
Alexander K Filippov(1), J Simon(2), EA Barnard(2), DA Brown(1)
Ibn Rochd Hospital, Department of Pharmacology, Casablanca, Morocco
Objectives: To evaluate the reasons of unintentional therapeutic errors in
older people, the types of drugs most frequently involved, and medical
outcomes associated with these adverse drug events. Materials and methods: Retrospective analysis of cases reported between 2005 and 2009 at
pharmacovigilance center of hospital Ibn Rochd, involving adults aged
65 and over with a potentially toxic exposure due to inadvertent errors
therapy. Results: Of the 65 notifications in older adults who reported
treatment errors, including 27 cases have been followed by a known
medical outcome. A major effect or death occurred in 16 cases. The most
common reasons for therapeutic errors were made inadvertently or medication administered twice, wrong drug given or taken, and the other
wrong dose. The reasons for the higher rate of major impact or deaths
were drug interactions, health professional or iatrogenic errors, and more
than one product containing the same ingredient. Certain classes of drugs
such as analgesics, anticoagulants, psychotherapy and some cardiovascular drugs were associated with high risk factors. Conclusion: interest to
evaluate the therapeutic errors in older adults to determine the causes of
errors related to the most frequently reported, and the reasons and drugs
involved with errors that result in serious consequences. Knowing why
they occur can help Causes of Therapeutic Errors in Older Adults.
Paper No.: 1635
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
THE SWEET SYNDROME : TWO CASES INDUCED BY
AMOXICILLIN- CLAVULANIC ACID
(1) University College London, Department of Neuroscience, Physiology
and Pharmacology, London, UK
(2) Cambridge University, Department of Pharmacology, Cambridge, UK
Expressed mGluR5 and P2Y1 receptors show promiscuous ion channel
coupling in sympathetic neurons: their stimulation inhibits M-type (Kv7,
K(M)) potassium currents and N-type (CaV2.2) calcium currents
(Kammermeier & Ikeda, Neuron 1999; 22: 819; Brown et al., J.Aut.Nerv.Syst. 2000; 81: 31). These effects are mediated by Gq and Gi/o
G proteins respectively. In contrast, activation of endogenous mGluR5
and P2Y1Rs in hippocampal pyramidal cells inhibited K(M)-current
(Filippov et al., J. Neurosci. 2006; 26: 9340), but not CaV (unpublished).
We ask why the difference? One possibility is that it relates to the differential distribution of the mGluR5/P2Y1R-binding scaffold protein
NHERF2 (Fam et al, PNAS 2005; 102: 8042; Paquet et al., JBC 2006;
281: 29949). Thus, we found that hippocampal neurons showed strong
NHERF2 immunoreactivity whereas sympathetic neurons showed none.
We therefore tested the effect of expressing NHERF2 (by intranuclear
cDNA injection) on the effects of mGluR5 and P2Y1R activation on
CaV2.2 and K(M) currents in sympathetic neurons. NHERF2 co-expression did not affect the amount of M-current inhibition produced by either
receptor but strongly reduced CaV inhibition by mGluR5 activation
(from 18 ± 2.8%, n = 11 to 4 ± 1.2%, n = 10), and by P2Y1R activation
(from 38 ± 4.3%, n = 10 to 10 ± 3.0%, n = 10). NHERF2 expression
had no significant effect on CaV inhibition by norepinephrine (via a2adrenoceptors, which do not bind NHERF2), or on CaV inhibition produced by an expressed P2Y1 receptor lacking the NHERF2-binding
DTSL motif. Thus, NHERF2 selectively restricts downstream coupling
of mGluR5 and P2Y1Rs in neurons to Gq-mediated responses such as
K(M) inhibition.
Support: Wellcome Trust grant 090370.
Houda Filali, I Rahmoun, A Elgharbi, F Hakkou
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
281
Paper No.: 687
FOCUSED CONFERENCE GROUP: P14 - ADDICTION AND
DOPING: NEUROBIOLOGICAL AND CLINICAL
THE EFFECT OF CENTRALLY INJECTED GLY-L-GLN
(GLYCYL-GLUTAMINE) ON NUCLEUS ACCUMBENS GABA
OUTPUT AFTER ACUTE MORPHINE ADMINISTRATION
Nesrin Filiz Basaran(1), S Cavun(1), RL Buyukuysal(1),
WR Millington(2)
(1) Uludag University, Faculty of Medicine, Department of Medical
Pharmacology, Bursa,Turkey
(2) Albany Collage of Pharmacy, NY, USA
Substance abuse and dependence is growing health problem all around
the world. At present still there is not effective and accurate treatment for
substance addiction. The most important site of action of all addictive
drugs is mesolimbic dopaminergic pathway. We previously showed that
Gly-Gln, a dipeptide synthesized by proteolytic clevage from endogenous b-endorphin, blocked the positive reinforcing and rewarding effects
of morphine, also delayed tolerance to morphine, inhibited morphine
addiction and suppressed withdrawal syndrome symptoms. After this
results we have focused on the possible mechanism of action of Gly-Gln.
We first studied on dopamine and its metabolite DOPAC. After swowing
inhibitor activity on morphine evoked dopamine release in nucleus
accumbens we tested Gly-Gln’s effect on GABA release. In this study
we used 200-300g Sprague Dawley male rats. Microdialysis probes were
placed in to Nacc (bregma 0 point, AP:+1.8; L:-1.0; V:-6.8) and intracerebroventricular (ICV) canulla to left lateral ventricul (bregma 0 point,
AP:-1.6; L:+0.9; V:-4.0) for injection Gly-L-Gln. Gly-L-Gln was administered 2 minutes before morphine injection. Morphine injection was performed via intraperitoneal (IP) canulla. Our results showed that after
morphine (2.5 mg/kg, IP) injection, Nacc GABA output start to increase
in 20. minute about 426% of baseline values and goes significantly high
during first hour. Gly-L-Gln (100 nmol/5ll ICV) blocked this increased
GABA release significantly. The effect of Gly-L-Gln on addiction and
rewarding impact of morphine, nicotine and ethanol is known. Gly-LGln is thought to show this effect also by altering GABAergic tonus
besides dopamine in Nacc.
Key-words: Gly-Gln, Morphine, GABA, Nucleus accumbens, Microdialysis,
_
Supported by TÜBITAK
and Uludag University (BAP)
Paper No.: 1530
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
CONTRIBUTION OF CYP2C8 TO THE METABOLISM OF
MONTELUKAST IN VITRO
Anne Filppula, J Laitila, PJ Neuvonen, JT Backman
University of Helsinki and Helsinki University Central Hospital, Department of Clinical Pharmacology, Helsinki, Finland
The leukotriene receptor antagonist montelukast is extensively metabolised. The majority of its oral dose is excreted in bile, mainly as a secondary metabolite formed by oxidation of metabolite 6 (M6). In vitro
studies with high montelukast concentrations have shown that cytochrome P450 (CYP) 3A4 catalyses montelukast sulfoxidation (M2
formation) and 21-hydroxylation (M5), and CYP2C9 catalyses 36hydroxylation (M6) (Chiba et al. Drug Metab Dispos 1997;25:10221031). We have studied the contributions of different CYP enzymes,
particularly the contribution of CYP2C8, to the hepatic microsomal
metabolism of clinically relevant montelukast concentrations (0.021 lM) in vitro. The effects of CYP isoform inhibitors on the metabolism
of montelukast were examined using pooled human liver microsomes
(HLM). In addition, montelukast metabolism by a range of human
recombinant CYP isoforms was investigated. Both inhibition and recombinant enzyme studies verified the central role of CYP3A4 in the formation of M2 and M5b (the more abundant diastereomer). Recombinant
CYP3A5 also mediated M2 and M5b formation. However, the CYP2C8
inhibitors trimethoprim and gemfibrozil 1-0-b glucuronide inhibited the
depletion of 0.02 lM montelukast and formation of M6 more potently
than did the CYP2C9 inhibitor sulfaphenazole. Similarly, M6 formation
was more effectively catalysed by recombinant CYP2C8 than by
CYP2C9 at low montelukast concentrations. At <0.1 lM montelukast,
the contribution of CYP2C9 was minimal, less than 10% of that of
CYP2C8. Moreover, recombinant CYP2C8 also mediated the further
metabolism of M6. In conclusion, CYP2C8 plays a major role in the
main metabolic pathway of montelukast at clinically relevant montelukast concentrations in vitro.
Paper No.: 2265
FOCUSED CONFERENCE GROUP:
P11 - G PROTEIN-COUPLED 7TM RECEPTORS: FROM
MOLECULAR TO PHYSIOLOGICAL FUNCTION
THE CONSERVED PROLINE OF HELIX 6 PLAYS A ROLE IN
THE FOLDING AND EXPRESSION OF A1B AND
B2 –ADRENERGIC RECEPTORS
Angela M Finch, SJ Humphrey, AP Campbell
University of New South Wales, School of Medical Sciences, Department of Pharmacology, Sydney, NSW, Australia
It has been proposed that prolines within transmembrane a-helices play a
role in the regulation of protein folding or alternatively have dynamic
structural roles as hinges, pivots or switches. To investigate the role of
proline in the transmembrane a-helices of G-protein coupled receptors
(GPCRs), we focused on the conserved proline of helix 6 (designated
P6.50 according to the Ballesteros-Weinstein numbering scheme) of the
Family A GPCRs. We mutated P6.50 of the a1B and b2 -adrenergic
receptors (AR). Mutation of P6.50 to alanine or glycine in the a1B-AR
resulted in no change in agonist binding or receptor activation; however,
receptor expression was halved. In contrast in the b2-AR these mutations
resulted in a 75-85% loss in receptor binding sites (Bmax wildtype:
25.1 ± 4.5; P6.50A: 3.8 ± 0.7, P6.50G: 6.3 ± 1.2 pmol/mg, n = 4-6).
The b2-AR mutants had unaltered [3H]-DHA antagonist binding affinities
(KD wildtype: 1.78 ± 0.66; P6.50G: 1.03 ± 0.37; P6.50A: 2.35 ± 0.68,
nM, n = 4-6) however they had significantly increased agonist (epinephrine) affinity (Ki wildtype: 16.72 ± 2.0; P6.50G: 6.90 ± 0.79; P6.50A:
2.80 ± 0.76 lM, n = 4, p < 0.05). These data suggest that the mutant
receptors may favour an active conformation. The use of a pharmacological chaperone (propanolol 100nM) lead to a substantial recovery of surface expression of both b2-AR mutants. The chaperoned b2-AR mutants
displayed wildtype agonist and antagonist binding properties. These findings support the notion that P6.50 is critically involved in folding and
expression of the b2-AR, however once the receptors are inserted into
the plasma membrane P6.50 is not required for maintaining the structural
integrity of the receptor.
Paper No.: 2266
FOCUSED CONFERENCE GROUP: PW01 - PHARMACOLOGY
OF ADRENOCEPTORS: EIGHTH SATELLITE MEETING
VIRTUAL SCREENING USING A1–ADRENERGIC RECEPTOR
PHARMACOPHORES IDENTIFIES A NOVEL STRUCTURAL
FRAMEWORK FOR A1A/D-ADRENERGIC RECEPTOR
SELECTIVE LIGANDS
Angela M Finch(1), ES Stoddart(1), IJA MacDougall(2), R Griffith(1,2)
(1) University of New South Wales, School of Medical Sciences, Department of Pharmacology, Sydney, NSW, Australia
(2) The University of Newcastle, School of Environmental and Life
Sciences, Australia
Pharmacophores are in silico tools for drug discovery that represent an
exciting opportunity to increase the efficiency and reduce the cost of
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
282
identification of ‘hit’ compounds. In this study a1- adrenergic receptor
(AR) pharmacophores were used to identify novel a1-AR selective compounds. Using five-feature pharmacophores for selective antagonists at
each of the a1-AR subtypes the National Cancer Institute (NCI), Tripos
LeadQuest (TDD) and Maybridge 2004 databases were mined. 12 compounds from these databases were selected, based on diversity of structure, predicted high affinity and selectivity at the a1D- subtype compared
to a1A- and a1B-ARs. The selected compounds were subsequently tested
in vitro for affinity and efficacy. 9 out of 12 of the tested compounds displayed affinity at the a1A and a1D -AR subtypes and 6 displayed affinity
at all three a1-AR subtypes, no a1B-AR selective compounds were identified. These results represent a high ‘hit’ rate, indicating our pharmacophores can successfully screen for ligands with a1-AR affinity. The
efficacy of the compounds that displayed affinity for the a1-AR subtypes,
measured as total inositol phosphate production, indicate 8 of the 9 compounds with a1-AR affinity displayed antagonist efficacy, with one compound (NCI-2) displaying partial agonist characteristics (pEC50
5.26 ± 0.15 M, n = 3). This virtual screen has successfully identified an
a1A/D-AR selective antagonist (NCI-5), with low lM affinity (pKi, a1A
5.75 ± 0.14, a1B 4.96 ± 0.12, a1D 5.82 ± 0.1, n = 3) with a novel structural scaffold of a an isoquinoline ring system with a rigid four ring
structure and good lead-like qualities ideal for further drug development.
Paper No.: 2909
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
PROSTAGLANDIN E2 PROMOTES ANGIOGENESIS BY
SYNERGIZING WITH FIBROBLAST GROWTH FACTOR
RECEPTOR-1/FIBROBLAST GROWTH FACTOR-2 SYSTEM
Federica Finetti, S Donnini, L Morbidelli, A Giachetti, M Ziche,
University of Siena, Department of Molecular Biology, Siena, Italy
Prostaglandin E2 (PGE2), the major product of cyclooxygenase, has been
shown to promote angiogenesis, however the functional and molecular
mechanism by which PGE2 induces this process is not completely understood. We investigated the PGE2 effects on the pro-angiogenic fibroblast
growth factor receptor-1 (FGFR-1)/fibroblast growth factor-2 (FGF-2)
signalling in cultured microvascular endothelial cells and in ex vivo and
in vivo assays. We demonstrated that PGE2 synergizes with FGF-2
and activates FGF-2 pathway. In particular PGE2 promotes FGFR1 and
FRS2á (FGFR substrate 2a) phosphorylation, endothelial nitric oxide synthase, the MAPK ERK 1/2 and the transcription factor STAT-3 activation
and FGF-2 protein up-regulation. In cells KO for FGF-2, PGE2 loses the
pro-angiogenic activity supporting the key role played by FGFR1/FGF-2
signalling cascade in PGE2 effects. FGF-2 over-expression, in turn, activates an autocrine/paracrine cycle in endothelial cells, which controls cell
survival and proliferation. In vivo, in the rabbit cornea and Matrigel plug
assay, the synergism between PGE2 and FGF-2 promotes a robust angiogenesis. In conclusion we demonstrated that the synergistic interaction of
PGE2 and FGF-2 promotes the angiogenic switch through activation of
the autocrine/paracrine FGF-2/FGFR1 signalling cascade. The synergism
between the PGE2 and FGF-2 signalling pathways here described may
substantiate the use of antitumor drug combinations, the most notable
being cyclooxygenase inhibitors with growth factors or growth factor
receptor inhibitors.
Paper No.: 1017
FOCUSED CONFERENCE GROUP: PW19 - INFLUENCE OF
DEGENERATION AND REPAIR IN THE CNS AND PERIPHERY
COMPARISON OF GABA-A IN VIVO OCCUPANCY IN RAT
BRAIN AND SPINAL CORD
GABA-A receptor occupancy has been validated clinically using [11C]
flumazenil PET imaging in human brain (Van Laere, Biol Psychiatry
2008; 64: pp153-161) and preclinically using [3H]flumazenil binding in
rodents (Atack et al, Neuropsychopharmaology 1999; 20: 255-262).
Technically PET imaging in the spinal cord can be more challenging than
using brain. This aim of this study was to compare GABA-A in vivo
receptor occupancy in rat whole brain vs spinal cord in order to determine if brain occupancy can be used as a surrogate for spinal cord occupancy when the spinal cord is the target tissue. Male CD rats (200-250g,
Charles River) were dosed orally with vehicle, and a range of GABA-A
selective ligands including SL651498 (3-100mg/kg), TPA023B (0.110mg/kg), or TP-13 (0.1-10mg/kg). At 3 min prior to sacrifice all rats
received, an iv dose of [3H]flumazenil (10lCi/kg). At 60 min rats were
sacrificed, brain/spinal cord removed, homogenised and filtered over GF/
B filters by washing with 2x5mls ice cold buffer. Non specific binding
was determined in a separate group of rats pre-treated with bretazenil
(5mg/kg, i.p. for 30 min). Dose - occupancy curves have shown similar
Occ50 values (a dose that produces 50% receptor occupancy) in rat
whole brain versus spinal cord for SL651498 (194 and 165mg/kg respectively), TPA023B (0.09 and 0.1mg/kg respectively), TP-13 (3.7 and
1.9mg/kg respectively). This data from a range of GABA-A ligands provides confidence that whole brain PET may be used as a surrogate for
spinal cord occupancy in translatable human studies.
Paper No.: 2281
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
MULTI-TARGET DRUGS FOR COGNITIVE IMPAIRMENT
THERAPY: IN VITRO INHIBITION OF HISTAMINE H3
RECEPTORS AND CHOLINESTERASE BY ORIGINAL
DONEPEZIL HYBRIDS
L Flammini(1), S Bertoni(1), C Giorgio(1), M Tognolini(1),
Vigilio Ballabeni(1), G Morini(2), M Incerti(2), E Barocelli(1)
(1) University of Parma, Department of Pharmacological, Biological
Sciences and Applied Chemistries, Parma, Italy
(2) University of Parma, Pharmacy Department, Parma, Italy
Introduction: A new paradigm called ‘‘multi-target–directed ligand’’
design is emerging for the development of new successful drugs for the
treatment of neurodegenerative diseases such as Alzheimer’s disease. A
current treatment of this multifactorial syndrome is based on acetylcholinesterase inhibitors endowed with a poor ability to alter disease progression and coupled with severe adverse side effects. The selective
blockade of histamine H3 receptors, auto- and hetero–receptors negatively modulating the synthesis and release of histamine and other neurotransmitters (such as acetylcholine) in different brain areas, has been
proven to ameliorate learning and memory in experimental models.
Therefore, some hybrid molecules have been developed combining the
structure of Donepezil with that of new H3 antagonists, to meet the goal
for development of original neuroprotective agents with improved tolerability. Materials: Donepezil and five new hybrids were evaluated in vitro
for their affinity (pKi), antagonistic potency (pKB) at rodent and human
histamine H3 receptors and for their ability to inhibit rat brain cholinesterase activity (pIC50). Results: Donepezil that inhibits cholinesterase
(pIC50 = 7.45) shows a two order of magnitude lower H3 binding affinity
(pKi = 5.30) than hybrids. All the new hybrids were potent H3 antagonists (pKB from 7.47 to 8.10; pKi from 7.34 to 8.18) exhibiting lower
anti-cholinesterase activity (pIC50 from 5.83 to 6.09) than Donepezil. No
species-specific differences were detected between human and rat histamine H3 receptor affinity. Conclusion: From this study the compound
M122, possessing a quite balanced H3 antagonistic/anti-cholinesterase
activity, can be selected for further in vivo studies.
Rebecca Fish, S Able, C Powell, L Booth
Pfizer Global Research and Development, Department of Discovery
Biology, Sandwich, UK
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
283
Paper No.: 750
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
ACUTE ORAL AND ACUTE DERMAL TOXICITY STUDY OF
DIOSCOREA HISPIDA (NAMI) AQUEOUS PLANT EXTRACT IN
SPRAGUE-DAWLEY RATS
Mona Liza Flores(1,2)
(1) University of the Philippines College of Medicine, Department of
Pharmacology and Toxicology, Manila, The Philippines
(2) Philippine Council for Health Research and Development - Department of Science and Technology, Manila, The Philippines
Dioscorea hispida, popularly known as nami in the Tagalog regions,
belongs to the family Dioscorecae. It has a wide variety of species found
growing in different countries and is widely distributed in the Philippines. Nami is believed to possess therapeutic potentials and is used for
various purposes such as medication to pains, bilious colic, nausea of
pregnancy, for spasmodic conditions, prevention of miscarriage, anodyne
and maturative in cases of tumors and buboes, to reduce swelling due to
insect bites or jelly fish sting, and as foodstuff and was found to have
sedative effects. Tubers of nami contain the alkaloid dioscorine responsible for its toxicity that can result to death. The activity of the Dioscorea
species has been attributed to the presence of and the actions of various
steroidal saponins (diosgenin – an aglycone), alkaloids (dioscorine, dioscine), and other alkaloids which are derived from nicotinic acid. In this
study, nami was prepared by maceration and the aqueous extract was
lyophilized. Administration and application of the plant material on sprague-dawley rats follows the OECD guidelines for testing of chemicals Up-and-Down-Procedure Guideline 425 for acute oral test and FixedDose-Procedure Guideline 434 for acute dermal test. Preliminary acute
toxicity test showed depression as the major toxidrome observed for the
live test subjects dosed between 175 mg/kg and 375 mg/kg. Mortality
was seen at 470 mg/kg. Acute dermal toxicity test showed no evident
dermal toxicity at the dose of 200 mg/kg.
Paper No.: 3147
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
POPULATION PHARMACOKINETICS OF OSELTAMIVIR
CARBOXYLATE IN MEXICAN PATIENTS WITH INFLUENZA
A(H1N1)
Francisco Flores-Murrieta(1,2), M Barranco-Garduño(1),
M Carrasco-Portugal(1), H León-Molina(1), A Cervantes-Nevárez(1),
G Castañeda-Hernández(3)
(1) Instituto Nacional de Enfermedade Respiratorias, Mexico City,
Mexico
(2) Escuela Superior de Medicina del Instituto Politécnico Nacional,
Mexico City, Mexico
(3) CINVESTAV-IPN, Mexico City, Mexico
The orally active neuraminidase inhibitor oseltamivir has shown to be
effective against the 2009 (H1N1) virus, being widely used for both,
treatment and prophylaxis. Oseltamivir is readily absorbed from the gastrointestinal tract and then extensively biotransformed to its active metabolite, oseltamivir carboxylate (OC). OC is minimally bound to plasma
proteins exhibiting a volume of distribution (Vd) equivalent to the extracellular volume of body water. OC is eliminated by renal clearance (CL)
by both, glomerular filtration and tubular secretion. Almost all this information has been generated in healthy volunteers or patients with mild
influenza, being data on severe ill patients very scarce. The purpose of
this study was to evaluate OC pharmacokinetics in adult hospitalized
patients diagnosed with influenza A(H1N1) using a population approach.
Plasma samples from 15 healthy volunteers and from 77 patients receiving 75 or 150 mg b.i.d. were obtained and analyzed by HPLC. Vd and
CL of OC were respectively increased and reduced, compared with
healthy volunteers. Notwithstanding, therapeutic plasma concentrations
were achieved with 75 mg bid. This was observed regardless of BMI or
if oseltamivir was given via nasogastric tube. Hence, higher doses are
not required for obese of severely ill patients. Moreover, oseltamivir
combinations with probenecid are not justified, as patients already exhibit
a reduced CL. Our data support current recommendations for oseltamivir
dosing in adult patients with influenza A(H1N1), and point that higher
doses are not rational from a pharmacokinetic point of view.
Paper No.: 2844
FOCUSED CONFERENCE GROUP: P18 - NUCLEAR
RECEPTOR TARGETS FOR TREATMENT OF DISEASES
SYNERGISTIC INTERACTION BETWEEN PPAR AGONISTS
AND ALBUTEROL ON HUMAN BRONCHIAL SMOOTH
MUSCLE CELL PROLIFERATION
Stefano Fogli, L Picchianti, F Stefanelli, MC Breschi
University of Pisa, Department of Psychiatry, Neurobiology,
Pharmacology & Biotechnology, Pisa, Italy
An important aim in asthma therapy is to prevent the accelerated growth
of bronchial smooth muscle cells (BMSC) which leads to hyperplasia
and bronchial hyperreactivity. In this study, we investigated the antiproliferative effect of the combination of albuterol with different PPAR-c agonists on human BSMC maintained in an optimized medium containing
epidermal growth factor and basic fibroblastic growth factor. RT-PCR
and Electrophoretic Mobility Shift Assay were used to assess PPAR-c
gene and protein expression, respectively. Apoptosis and necrosis were
assessed by ELISA. Synergism or antagonism was quantified by the
combination index (CI) method (Chou TC et al, J Natl Cancer Inst 1994;
86: 1517-1524). PPAR-c gene was highly expressed in BSMC and the
protein was identified in cell nuclei. Stimulation of BSMC with growth
factors for 48 h induced a 4-fold increase in cell proliferation which was
significantly prevented by single-agent albuterol, rosiglitazone or 15deoxy-d 12, 14-prostaglandin J2 with IC50 mean values of 7.37 ± 1.75,
8.52 ± 1.26, and 10.1 ± 1.37 mM, respectively. Selective b2-adrenoceptor (b2-AR) and PPAR-g antagonists dose-dependently block the antiproliferative effect of drugs. Combination of albuterol with PPAR-g agonists
at nanomolar concentrations, allows obtaining the 50% growth inhibition
at levels from 2 to 10-fold lower than those required for each drug alone
(CI <1), without induction of apoptosis or necrosis. These data indicate
the presence of a synergistic interaction between PPAR-g agonists and
bronchodilators on airway smooth muscle cell proliferation, thus supporting the therapeutic potential of this combination in chronic airway
diseases.
Paper No.: 3181
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
MONTELUKAST REVERSES ALBUTEROL-INDUCED
B2-ADRENOCEPTOR TOLERANCE IN AIRWAY SMOOTH
MUSCLE
Stefano Fogli, F Stefanelli, L Picchianti, MC Breschi
University of Pisa, Department of Psychiatry, Neurobiology, Pharmacology & Biotechnology, Pisa, Italy
The regular use of b2-adrenoceptor (b2-AR) agonists has been associated
with proasthmatic-like changes that limit their efficacy and potentially
increase the risk of asthma exacerbation (Spitzer WO et al, N Engl J
Med 1992; 326: 501-506). We sought to determine whether montelukast
prevents homologous b2-AR desensitization in tracheal muscular tissues
derived from guinea-pigs and in human bronchial smooth muscle cells
(BSMC). The guinea-pig model of albuterol-induced b2-AR tolerance
described in the present study closely resembles the clinical setting since
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
284
it was characterised by b2-AR agonist hyporesponsiveness (-25%) and
hyperreactivity to cholinergic stimuli (+53%). Montelukast, given intraperitoneally at 5 mg/kg/d for 6 consecutive days, completely reversed
albuterol-induced b2-AR tolerance in homologously desensitised tracheal
smooth muscle, without significantly affecting the cholinergic constrictor
response. In vitro findings demonstrated that albuterol-induced cAMP
stimulation was significantly reduced in homologously desensitised
BSMC (P < 0.001), in the absence of changes in b2-AR mRNA levels.
In this experimental setting, montelukast 30 lM for 24 h was able to
reverse albuterol subsensitivity induced by homologous desensitization
(P < 0.01). No substantial change in b2-AR transcription was observed
in desensitised cells, as compared with those treated with montelukast.
Overall, our data suggest the presence of a positive pharmacodynamic
interaction between montelukast and b2-AR agonists, thus supporting
the use of this combination to improve the therapeutic index of
bronchodilators.
Paper No.: 1406
FOCUSED CONFERENCE GROUP: P14 - ADDICTION AND
DOPING: NEUROBIOLOGICAL AND CLINICAL
EXERCISE INFLUENCE ON THE
AMPHETAMINE-DEPENDENT BEHAVIOUR AND DOPAMINE
IN STRIATUM, FRONTAL CORTEX AND HIPPOCAMPUS
Carlos Alberto Fontes-Ribeiro, E Marques, F Pereira, AP Silva,
T Macedo
University of Coimbra Faculty of Medicine, Institute of Pharmacology
and Experimental Therapeutics, Coimbra, Portugal
The dopaminergic mesocorticolimbic and nigrostriatal systems are considered to be involved in drug addiction. However, there is little information about the effect of repeated exercise on the synthesis, metabolism
and action of dopamine (DA) in the central nervous system, in the presence of amphetamines. Thus, our aims were to verify the influence of
exercise on addiction using an amphetamine (AMPH)-induced conditioned place-preference (CPP) in rats, and to determine DA and metabolites in striatum, frontal cortex and hippocampus. Adult male SpragueDawley rats were trained under a 8 week treadmill running program.
The CPP test was performed in trained and untrained groups. Conditioning was performed for 8 days with i.p. injection of 2 mg.Kg-1 AMPH or
saline. Animals were sacrificed 24 h after the last AMPH or saline, and
striatum, hippocampus and frontal cortex were dissected for DA and
metabolites measurement by HPLC-EC. Rats without exercise showed
preference for the compartment associated with AMPH, an effect which
did not occur with the animals with training. Amphetamine decreased
striatal DA content and turnover from both trained and untrained rats.
Training reduced and increased the DA content in frontal cortex and hippocampus, respectively. According to these results, exercise may prevent
the AMPH-seeking behaviour but not changes the impact of AMPH on
striatal DA synthesis. Data also suggest that exercise may correlate with
changes in DA dynamics in the frontal cortex and hippocampus.
Paper No.: 2321
FOCUSED CONFERENCE GROUP:
P04 - PHARMACOEPIDEMIOLOGY, CURRENT
CONTROVERSIES AND OPPORTUNITIES
MACRO-STUDY ON-LINE OF SEARCHING IN INTERNET
FOR HEALTH AND DRUG PRODUCTS: A SOCIAL
PHARMACOLOGY APPROACH
Juan Antonio Formigos(1), M Palmero(1), JL Alloza(2)
(1) University of Alicante, Department of Optic, Pharmacology and
Anatomy, San Vicente del Raspeig, Alicante, Spain
(2) University of Alcalá, Department of Pharmacology - Social Pharmacology Group, Alcalá de Henares, Madrid, Spain
Internet is a new stage of our civilization that provides any kind of information. In recent years searching on health and medical treatments is one
of the most widely used on the Internet. We have developed an epidemiological transversal and observational study, using a self-administrated
questionnaire on the Web, with free access at any time. The target was
the community of The University of Alcalá de Henares, Madrid (Students, Lecturers and Researchers, and the administrative staff services),
over 30.000. 45.2% of Internet users have sought information about
medications on occasion. The searchers were characterized by size
of internet use (OR=2.43, 95%CI=1.97-2.99) related with a greater
confidence in the contents of the network (OR=1.55, 95%CI=1.30-1.85).
It was more evident for those health related careers (OR=2.07,
95%CI=1.69-2.53), the chronically ill people (OR=1.41, 95%CI=1.091.82). The latter were more concerned about the medicines’ safety
(OR=1.33, 95%CI=1.09-1.62), and also read the prospectus over the rest
(OR=1.66, 95%CI=1.26-2.18). The knowledge provided by internet was
used positively by 21.3% in a doctor’s consultancy. Our results on the
internet and health care are applicable to the current European society.
This study supports that new social trends run in a similar way that has
been assessed in the US in recent years. Internet plays a definite role in
health care.
Paper No. 3423
FOCUSED CONFERENCE GROUP: PW19 - INFLUENCE OF
DEGENERATION AND REPAIR IN THE CNS AND PERIPHERY
SELF-ASSESSMENT WITH MOODLE FROM ACTIVITIES
PROPOSED BY STUDENTS
Juan Formigós-Bolea, M Palmero, V Maneu, C Garcı́a-Cabanes
University of Alicante, Department of Optics, Pharmacology & Anatomy, Alicante, Spain FARMAGITE
Preliminary results of an experiment in the area of pharmacology. Background: The B-learning improves academic performance [1]. Blended
learning is very popular and well received among students [2]. Students
can learn through taking the role of a teacher [3]. Methods: The experience was performed in the subject of ‘Pharmacology, Nutrition and Dietetics’ of the second year of the Nursing Degree during the 2009-2010
academic years at the University of Alicante (Spain). A total of 180 students enrolled were invited to perform self-assessment tests (J-click and
hot potatoes) into a Moodle platform. The activities and test questions
were proposed by students themselves. The role of teachers has been
limited to monitor the consistency of the questions asked, respecting the
characteristic form of expression and the students’ vocabulary. Results:
The interim results show strong trends in favour of this procedure. These
findings indicate that students who participated by providing questions
and solving quizzes obtain better academic performance than those who
were limited to answering the tests and these, in turn, stand on that have
prevented their participation in the test. Conclusions: The approach of
self-assessment exercises collaboratively by students, under supervision
of faculty, is shown as a useful tool and could be useful in the transition
to new teaching strategies suggested by the European Higher Education
Area [1]. [1]. Ábalos C et al . RCCV. 2009; 3:198-207. [2]. Shah IM et
al. Emerg Med J. 2008; 25:354-357. [3]. Yeung MA et al. Australasian
Journal of Peer Learning. 2008; 1: 26-39.
Paper No.: 887
FOCUSED CONFERENCE GROUP: SOCIETY WORKSHOP SW14
A RAPID AND SENSITIVE HIGH-PERFORMANCE LIQUID
CHROMATOGRAPHIC METHOD TO QUANTIFY CARBAMAZEPINE, OXCARBAZEPINE, ESLICARBAZEPINE ACETATE
AND THEIR MAIN METABOLITES IN HUMAN PLASMA
Ana Fortuna(1,2), G Alves(2,3), L Santos(1,2), A Falcão(1,2), P Soaresda-Silva(4)
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
285
(1) Faculty of Pharmacy, University of Coimbra, Coimbra, Portugal
(2) Centre for Neurosciences and Cell Biology, University of Coimbra,
Coimbra, Portugal
(3) Health Sciences Research Centre, University of Beira Interior,
Covilhã, Portugal
(4) Department of Research and Development, BIAL, S. Mamede do
Coronado, Portugal
A simple and fast HPLC-UV method was developed and validated for
simultaneous quantification in human plasma of three structurally related
antiepileptic drugs, carbamazepine, oxcarbazepine and eslicarbazepine
acetate, and their main metabolites carbamazepine-10,11-epoxide, racemic licarbazepine and trans-diol. Aliquots (500ll) of blank plasma from
healthy subjects were spiked with known amounts of all compounds and
internal standard (10,11-dihydrocarbamazepine). The analytes were
extracted from plasma by solid-phase extraction (SPE) and, thereafter,
separated at 40C on the reversed-phase column LiChroCART Purospher Star (C18, 3lm, 55mm x 4mm) using an isocratic elution with a
mixture of water, methanol and acetonitrile (64:30:6, v/v/v) as mobile
phase and detected at 235 nm. Under these chromatographic conditions,
all studied drugs and metabolites were baseline separated in less than
9 minutes and no peaks due to the plasma matrix interfered at their retention times. The assay was linear (r2 > 0.997) in the ranges of 0.05-30,
0.05-20, 0.15-4, 0.1-30, 0.1-60 and 0.1-10lg/ml for carbamazepine,
oxcarbazepine, eslicarbazepine acetate, carbamazepine-10,11-epoxide,
licarbazepine and trans-diol, respectively. The overall intra and interday
precision did not exceed 15% and the accuracy was within ±15% considering all compounds. Precision and accuracy at the lower limit of quantification was also demonstrated. In addition, the assay afforded high
recoveries for all analytes. Thus, the selective SPE procedure combined
with the simple and fast chromatographic run make this assay a useful
tool to support the therapeutic drug monitoring of such antiepileptic
drugs and future pharmacokinetic clinical studies.
Paper No.: 888
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
LACK OF INTERFERENCE OF CARBAMAZEPINE AND ITS
METABOLITES IN THE ENANTIOSELECTIVE THERAPEUTIC
DRUG MONITORING OF ESLICARBAZEPINE ACETATE
Ana Fortuna(1,2), G Alves(2,3), J Sousa(1,2), R Direito(2,3),
M Rocha(2,4), A Falcão(1,2), P Soares-da-Silva(5)
(1) Faculty of Pharmacy, University of Coimbra, Coimbra, Portugal
(2) Center for Neurosciences and Cell Biology, University of Coimbra,
Coimbra, Portugal
(3) Pharmacology Deapratment, Faculty of Pharmacy, University of
Coimbra, Coimbra, Portugal
(4) Pharmaceutical Services, Coimbra University Hospital, Coimbra,
Portugal
(5) Department of Research and Development, BIAL, S. Mamede do
Coronado, Portugal
As add-on therapy, eslicarbazepine acetate (ESL) phase III clinical trials
conducted in patients with refractory partial-onset seizures have evidenced good efficacy, safety and tolerability, even though ~60% of the
enrolled patients were taking carbamazepine (CBZ). Considering the
pharmacological disadvantages of CBZ and the similar spectrum of efficacy of CBZ and ESL, the switching to ESL may be successful in many
patients. As ESL is extensively converted to S-licarbazepine (S-Lic;
~95%), the enantioselective assessment of S-Lic is expected to be useful.
This study investigated the plasma levels of S-Lic and R-licarbazepine
(R-Lic) in patients under CBZ long-term treatment to assess the potential
interference of CBZ or its metabolites in the enantioselective therapeutic
drug monitoring (TDM) of ESL. For that a chiral HPLC-UV assay developed and validated within our research group was used. Twenty-four
patients admitted to the Coimbra University Hospital and receiving CBZ
long-term treatment were identified. Blood samples were collected and
serum CBZ levels were measured by the usual TDM protocol. Aliquots
of plasma were also submitted to a chiral HPLC-UV analysis. The bioan-
alytical data indicated that S-Lic and R-Lic were not present at detectable
levels in plasma samples of the CBZ-treated patients. The chromatograms generated by the analysis of patient plasma samples, when compared with those obtained from blank plasma samples spiked with S-Lic
and R-Lic, showed the absence of interferences. These data support the
usefulness of the chiral HPLC-UV method validated by our group for enantioselective TDM of ESL (mainly S-Lic) during the switching from
CBZ to ESL.
Paper No.: 1759
FOCUSED CONFERENCE GROUP:
P19 - GENERAL SESSION - ONCOLOGY
MORPHOLOGICAL EVALUATION OF RAT LIVER TISSUE
AFTER THE SUBCUTANEOUS INJECTION OF
1,2-DIMETHYLHYDRAZINE
Ana Fortuna(1,2), R Direito(3), G Alves(2,4), J Sousa(1,2),
A Falcão(1,2), A Cabrita(5)
(1) University of Coimbra, Faculty of Pharmacy, Coimbra, Portugal
(2) University of Coimbra, Center for Neurosciences and Cell Biology,
Coimbra, Portugal
(3) University of Coimbra, Faculty of Sciences and Technology,
Biochemical Department, Coimbra, Portugal
(4) University of Beira Interior, Health Sciences Research Centre,
Covilhã, Portugal
(5) University of Coimbra, Faculty of Medicine, Institute of Experimental Pathology, Coimbra, Portugal
The exposure to 1,2-dimethylhydrazine (DMH) is usually associated to
colorectal cancer induction. However, one can predict that the liver may
be also an important target for DMH toxicity, since it is the primary
organ of metabolism, playing a major role in the bioactivation and detoxication of xenobiotics. Therefore, the present study was carried out to
evaluate the morphological and histopathological modifications in liver
tissue following the administration of DMH to rats. Groups of adult male
Wistar rats were treated twice-weekly with subcutaneous injections of
DMH at a dose of 15 mg/kg body weight for 8 weeks. Hematoxylin and
eosin histological analysis using optic microscopy revealed the existence
of hepatic necrosis following the 8-week treatment period with DMH.
Aiming to evaluate the regenerative capacity of liver in the sequence of
the administration of DMH, groups of rats were sacrificed at different
time points after the last dose: immediately after and, 4 and 24 weeks
later. The histological examination showed that, in all exposed animals,
the observed stasis, cytoplasmic changes, steatosis, inflammatory infiltration and necrosis, were not reversible six months after the last administration of DMH. It was also noticeable that at the end of six months no
evidences compatible with development of hepatic neoplasia were found.
Accordingly, these results give enough support for the experimental evidence of DMH-induced hepatotoxicity in rats. In addition, this wellestablished experimental model will allow us to study, in the future, the
signalling pathways involved in such histopathological alterations
induced by DMH in the liver of rat.
Paper No.: 1841
FOCUSED CONFERENCE GROUP: P02 - TRANSMEMBRANE
TRANSPORT: PERSPECTIVES FOR DISEASE AND DRUG
DISCOVERY
GENE EXPRESSION AND COMPARATIVE GENOMIC
HYBRIDIZATION PROFILES OF 6-MERCAPTOPURINE- AND
6-THIOGUANINE-RESISTANT HUMAN LEUKAEMIA CELLS
Alan Fotoohi, J Hashemi, A Moshfegh, C Larsson, F Albertioni
Karolinska Institute, Department of Medicine & Clinical Pharmacology,
Stockholm, Sweden
Treatment of the MOLT4 human T-cell leukemia cells with 6-mercaptopurine (6-MP)and 6-thioguanine (6-TG), resulted in acquired resistance as
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
286
a result of low expression of concentrative nucleoside transporter 3
(CNT3) and equilibrative nucleoside transporter 2 (ENT2) genes. The
activity of thiopurine methyl transferase enzyme and other enzymes in
metabolism pathway of 6-MP and 6-TG was unchanged. The exception
was >40% reduction in expression of guanine monphosphate synthetase
(GMPS) in both resistant sub-lines. In order to identify other modifications at RNA and DNA levels associated with 6-MP and 6-TG resistance, we analyzed the patterns of gene expression and comparative
genomic hybridization profiles by resistant sublines and compared with
the wild-type MOLT4 cells. Expression of several nucleoside transporter
genes at mRNA level were down-regulated in the both thiopurine-resistant sub-lines. Besides mRNA expression of genes encoding the purine
de novo synthesis enzymes was reduced equally in both resistant cell
lines; 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase
by 20%, and glycinamide ribonucleotide transformylase by 40%, respectively, which along with a significant reduction in expression of GMPS
indicate defected purine metabolism in these resistant sublines. Interestingly, reductions observed in some apoptotic genes: caspase 1 and 4
encoding genes were down-regulated in 6-MP resistant cells by 75 and
80%, respectively. While the expression of mRNA of Fas receptor was
reduced by >50% in 6-TG resistant cells, the apoptosis-inducing serinethreonine kinase 17b gene was down-regulated in both 6-MP- and 6-TGresistant sub-lines by 50 and 40%, respectively. Our results suggest contribution of several genes in development of resistance to thiopurines.
Paper No.: 1706
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
HIGH SALT INTAKE ABOLISHES AT2-MEDIATED
VASODILATION OF PIAL ARTERIOLES IN RATS
Sebastien Foulquier(1), C Perrin-Sarrado(1), F Dupuis(1), F Soussi(1),
P Liminana(1), Y-W Kwan(2), C Capdeville-Atkinson(1), I Lartaud(1),
J Atkinson(1),
(1) Nancy University Faculty of Pharmacy, EA3452 ‘Drug Targets, Formulation & Preclinical Drug Evaluation’, Cardiovascular Pharmacology
Laboratory, Nancy, France
(2) Chinese University of Hong Kong Faculty of Medicine, Basic Medical Sciences, Hong Kong, PR China
Objectives: Angiotensin II (Ang II) has a dual effect on the cerebral circulation, inducing constriction via AT1 and dilatation via AT2 receptors
(VINCENT et al., Stroke 2005, 36, 2691-2695). We hypothesized that
HS specifically changes functional Ang II-responses of cerebral arterioles. Methods: Three m.o. male Wistar rats received (drinking water)
Na+ 172 mM or 4 lM (control). Internal diameter of pial arterioles (ID)
was measured (cranial window) following Ang II 10-6M (AT1-mediated
vasoconstriction) and Ang II 10-8M + losartan 10-5M (AT2-mediated
vasodilation) suffusion at day 4 or 30. Vasoactivity was evaluated following serotonin (5-HT) 10-6 M and sodium nitroprusside (SNP) 10-5M.
Results (m+/)sem; n = 5-10; * p < 0.05 vs control same day, 2 way
ANOVA + Bonferroni): HS did not modify mean arterial pressure (control d4; d30 106 + /-5; 109 + /-4; HS 103 + /-3; 110 + /-6 mmHg).
Vasoconstrictor responses to Ang II (d4; d30, control -16 + /-1; -14 + /1; HS -18 + /-2; -14 + /-1% baseline ID) did not change. AT2-mediated
vasodilation (control d4; d30 8 + /-2; 5 + /-2%) was abolished by HS at
d4 (-2 + /-2* %) and even reversed to vasoconstriction at d30 (-7 + /-2*
%). Vasoconstriction to 5-HT (control -15 + /-3; -8 + /-1; HS -12 + /-2;
-10 + /-1%) did not change, nor did vasodilation to SNP (control 84 + /12; 87 + /-8; HS 86 + /-11; 82 + /-8%). Conclusion: HS does not change
arterial pressure in Wistar rats, but specifically abolishes AT2-mediated
vasodilation. This effect is immediate (d4) and maintained in time (d30).
Were AT2-mediated vasodilation be protective, its abolition would be
deleterious in case of stroke.
Paper No.: 2854
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
INHIBITION OF NF-JB ACTIVATION BY CYMBOPOGON
CITRATUS LEAVES AND THEIR POLYPHENOLS IN HUMAN
MACROPHAGES: A POTENTIAL NATURAL SOURCE OF
ANTI-INFLAMMATORY DRUGS
Vera Francisco(1,2,3), A Figueirinha(1,4), BM Neves(2,3),
C Garcia-Rodriguez(5), MC Lopes(2,3), MT Cruz(2,3), MT Batista(1,3)
(1) Centro de Estudos Farmacêuticos, Faculdade de Farmácia-Universidade de Coimbra, Coimbra, Portugal
(2) Centro de Neurociências e Biologia Celular, Universidade de
Coimbra, Coimbra,Portugal
(3) Faculdade de Farmácia, Universidade de Coimbra, Coimbra, Portugal
(4) Departamento de Ambiente, Instituto Politécnico de Viseu, Viseu,
Portugal
(5) Instituto de Biologia y Genética Molecular, Universidad de Valladolid
- CSIC,Valladolid, Spain
Introduction: Most of pharmaceutical drugs presently available are
derived from natural products because the bioactivity of their phytoconstituents. Phenolic compounds have important biological properties, being
a potential source of new therapeutic agents for the treatment of inflammation and related diseases like cancer and cardiovascular pathologies.
Since nuclear transcription factor (NF)-jB regulates the expression of
many pro-inflammatory genes we evaluated the effect of an infusion of
Cymbopogon citratus (Cc) leaves and its polyphenols on lipopolysaccharide (LPS)-induced NF-jB activation, in human macrophages. Materials:
Cc dry leaves were extracted by infusion (5g/150ml H2O). Subsequently,
lipids and essential oils were removed with n-hexane and the extract was
freeze-dried (Cc extract). Different Cc polyphenolic fractions (PFs),
namely phenolic acids, flavonoids and tannins, were obtained for Cc
extract fractionation on a reverse phase semipreparative column Lichroprep RP-18 and in a gel chromatography on a Sephadex LH-20 column. As an in vitro model of inflammation, human monocyte-derived
macrophages stimulated with LPS from E. coli were used. The effect of
Cc extract and PFs on NF-jB activation was evaluated by Western blot
using anti-inhibitory-jB (IkB) and anti-phospho-IjB antibodies. Results:
We observed that Cc extract inhibited LPS-mediated IjB degradation and
increased the levels of IjB phosphorylation, while PFs inhibited LPSmediated IjB phosphorylation and degradation, consistent with inhibition
of NF-jB activation and anti-inflammatory effect. Conclusion: Polyphenolic compounds from Cymbopogon citratus inhibited NF-jB activation.
Therefore, Cc extracts are good candidates for the development of new
drugs to treat diseases with a strong inflammatory component.
Supported by FCT (PTDC/SAU-FCF/105429/2008 project and SFRH/
BD/46281/2008.
Paper No.: 3109
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
ESTROGEN MINIMIZES CELL DEATH CAUSED BY H2O2 IN
CELLS OF RAT GLIOMA
Lucas AM Franco(1), L Yshii(1), E Kawamoto(1), L Lima(1),
C Scavone(1), C Munhoz(1)
Universidade de São Paulo (USP), Department of Pharmacology, São
Paulo, Brazil
Evidence suggests that glial cells play a major role in neuronal signalling
and inflammatory responses in the Central Nervous System (CNS).
Chronic inflammatory responses as well as glia activation are associated
with neurodegenerative diseases, such as Parkinson’s and Alzheimer’s.
Estrogen (E2) is well known by its neuroprotective actions and the
estrogen receptors ESR1, ESR2 and GPER are very important for the
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
287
manifestation of these effects in the brain. This work investigated the
protective roles of E2 in C6 cell line of rat glioma, after damage induced
by hydrogen peroxide (H2O2). PCR, western blot and immunofluorecence assays confirmed the presence and functionality of estrogen receptors ESR1 and GPER in C6 cells. Our results also confirmed that H2O2
induced death in C6 cells. And the pre-treatment with E2 (2 hours and
24 hours) decreased the H2O2 toxicity in a dose-dependent manner.
These results highlight the involvement of E2 and its receptors in preventing cellular damage in glial cells. In addition, they also suggest that
the E2 protective effects may be associated with rapid, non-genomic
actions of E2 membrane receptors.
Paper No.: 3375
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
EFFECT OF RED-WINE POLYPHENOLIC COMPOUNDS ON
ALLERGEN INDUCED HYPERREACTIVITY OF THE
AIRWAYS
Sona Franova(1), M Joskova(1), E Novakova(2), K Adamicova(3),
G Nosalova(1), O Pechanova(4)
(1) Comenius University, Jessenius Faculty of Medicine, Department
of Pharmacology, Martin, Slovakia
(2) Comenius University, Jessenius Faculty of Medicine, Department of
Microbiology, Martin, Slovakia
(3) Comenius University, Jessenius Faculty of Medicine, Department
of Pathologcal Anatomy, Martin, Slovakia
(4) Slovak Academy of Sciences, Department of Normal and Pathological Physiology, Bratislava, Slovakia
The results of some epidemiologic studies have suggested that the consumption of the food containing polyphenolic compounds might reduce
the occurrence of asthma symptoms. The aim of our experiments was to
evaluate the effect of 21 days red-wine polyphenolic compounds (Provinol) administration on allergen (OVA) induced inflammation of the airways in experimental conditions. The allergic inflammation of the
airways was induced in guinea pigs by 21 days ovalbumin sensitization.
The tracheal smooth muscle reactivity was examined by in vitro method
to bronchoconstricting mediator histamine (10-8-10-3 mol.L-1); the
changes in airways resistance by in vivo method to nebulized histamine
(10-6 mol.L-1). The histological investigation of the tracheal tissue and
BALF levels of inflammatory cytokines IL-4, IL-5 were used as parameters of airway inflammation. 21 days administration of Provinol caused a
significant decrease of specific airway resistance after histamine nebulization and decline in tracheal smooth muscle contraction amplitude to this
bronchoconstrictor. Provinol minimized the degree of inflammation estimated on the basis of eosinophil calculation and levels of inflammatory
cytokines IL-4 and IL5. In conclusion, Provinol administration in ovalbumin-sensitized guinea pigs leads to bronchodilating and antiinflammatory
effects, suggesting their possible use in therapy of allergic inflammatory
conditions of airways.
Work was supported by Grant VEGA 1/0073/08, Center of Experimental
and Clinical Respirology –‘‘Project is co-financed from EC sources’’ –
ERDF – European Regional Development Fund
Paper No.: 1864
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
IMPACT OF WINDOW L-TYPE CA2 + INFLUX ON ACTIVITY
OF CA2 + CHANNEL BLOCKERS IN AORTIC SMOOTH
MUSCLE CELLS OF C57BL6 MICE
Introduction. L-type Ca2 + channel (CC) blockers (CCBs) reduce blood
pressure more effectively in hypertensive than in normotensive subjects
and are more effective in vascular smooth muscle (VSM) than in cardiac
muscle. Several attempts have been made to explain these unique features. The depolarization of the resting potential by hypertension and the
depolarized resting potential of VSM in comparison with the heart not
only favour the inactivated state of the CC, but also increase window
L-type Ca2 + influx. Methods. The present study investigated whether the
CC agonist BAY K8644 and whether nifedipine, verapamil or diltiazem,
representatives of different CCB classes, alone or in combination, modulated window Ca2 + influx (Fura-2 method) and isometric contraction
during depolarisation of C57Bl6 mouse aorta segments with 50 mM K+.
Results. High K+-induced contractions were biphasic. The fast force
component coincided with Ca2 + influx via a population of CC that
exhibited fast activation and then inactivation. The slow, sustained force
component was attributed to voltage-dependent, but time-independent
Ca2 + influx displaying incomplete inactivation (window) and was significantly more sensitive to CCBs than the fast component. Stimulation of
CC Ca2 + influx with 30 nM BAY K8644 decreased, whereas conditions
that favoured window Ca2 + influx increased the efficacy of CCBs to
inhibit isometric contractions. Conclusion. Results suggest that inhibition
of window L-type Ca2 + influx may play a dominant role in the antihypertensive effects of CCBs.
Paper No.: 1865
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
SIGNIFICANCE OF WINDOW L-TYPE CALCIUM INFLUX
FOR CONTRACTION OF C57BL6 MOUSE AORTIC
SEGMENTS
Paul Fransen, CE Van Hove, J van Langen, H Bult
University of Antwerp, Division of Pharmacology, Antwerp, Belgium
Recently, differential expression of spliced isoforms of the voltage-gated
calcium channel (VGCC) gene Cav1.2 was demonstrated along the vascular tree. These isoforms display cell-specific electrophysiological properties with respect to the window VGCC Ca2 + influx and sensitivity to
L-type Ca2 + channel blockers. We investigated whether the window
VGCC Ca2 + influx plays a physiological role for blood vessel constriction. Methods. Ca2 + mobilisation (Fura-2 assay) and isometric contraction were studied in aortic segments of C57Bl6 mice. The membrane
potential was clamped at fixed potentials by increasing external K+ concentration. Results: Above 20 mM, K+ induced biphasic contractions.
The fast component coincided with Ca2 + influx via a VGCC population
that exhibited fast activation and then inactivation. The slow force component was temporally related to a slow voltage-dependent, but timeindependent Ca2 + influx. The slow contraction was not due to Ca2 +
induced Ca2 + release, as it was not affected by dantrolene, ryanodine or
depletion of sarcoplasmic reticulum Ca2 + stores with cyclopiazonic acid.
By modulating external Ca2 + levels before and after depolarisation with
extracellular K+, we could attribute the sustained contraction to a population of VGCCs displaying incomplete inactivation (window). At normal
extracellular K+ concentration, this window was operative, allowing a
continuous VGCC-mediated Ca2 + influx leading to basal tone. Basal
influx and force were both suppressed upon hyperpolarization by adding
the ATP-dependent K+ channel agonist levcromakalim. It is concluded
that window L-type Ca2 + influx has a profound physiological impact on
basal tension and isometric contraction of C57Bl6 mouse aortic smooth
muscle cells.
Paul Fransen, CE Van Hove, C Michiels, J van Langen, H Bult
University of Antwerp, Division of Pharmacology, Antwerp, Belgium
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
288
Paper No.: 2112
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
ACTIVITY ASSESSMENT OF INDIVIDUAL CYP2D6 VARIANTS
USING POPULATION PHARMACOKINETICS OF
DEXTROMETHORPHAN
Uwe Fuhr(1), D Frank(1), A Gaedigk(2), T Klaassen(1),
D Tomalik-Scharte(1), Ar Jetter(1,3), U Jaehde(4), R Süverkrüp(4),
J Kirchheiner(5), K Abduljalil(1,6)
(1) University Hospital, University of Cologne, Department of
Pharmacology, Cologne, Germany
(2) The Children’s Mercy Hospital and Clinics, Division of Developmental Pharmacology & Experimental Therapeutics, Kansas City, USA
(3) University Hospital Zurich, Division of Clinical Pharmacology and
Toxicology, Department of Internal Medicine, Zürich, Switzerland
(4) University of Bonn, Institute of Pharmacy, Bonn, Germany
(5) University of Ulm, Department of Pharmacology of Natural Products
& Clinical Pharmacology, Germany
(6) SimCYP Limited, Sheffield, UK
CYP2D6 activity is rate limiting for the formation of dextrorphan from
dextromethorphan. Respective metabolic ratios in urine and plasma are
thus used for CYP2D6 phenotyping. CYP2D6 has an extensive genetic
polymorphism. To enable prediction of phenotype from genotype, currently individual alleles are often categorized in null, reduced, high, and
increased activity allele groups, which are used to derive CYP2D6 activity scores. The aim of this study was to quantify the effect of major
CYP2D6 variants on dextromethorphan metabolism. To this end, plasma
and urine concentrations of dextromethorphan and dextrorphan were
evaluated from 36 healthy Caucasian male volunteers who participated
in three studies with administration of a single oral dose of 30 mg dextromethorphan-HBr. Concentrations were quantified by LC-MS/MS. All
data were modelled simultaneously using the population pharmacokinetic
software NONMEM. A five-compartment model was able to describe
the data adequately. The clearance fraction attributable to an individual
CYP2D6*1 allele was about 2.5-fold higher than that attributable to a
CYP2D6*2 allele (5010 [95% CI 3579-6441] vs. 2020 [624-3416] L/h),
while the metabolic activity related to a CYP2D6*41 allele was much
lower (85 [63.8-106.2] L/h). Urinary pH was confirmed as a significant
covariate for dextromethorphan renal clearance. These results clearly
show that important information is lost when CYP2D6*1 and *2 are considered as equal. The lack of differential information on the activity
attributable to individual alleles may be one explanation why there is
considerable interindividual variability in the pharmacokinetics of dextromethorphan and other CYP2D6 substrates within currently used
CYP2D6 activity score levels.
Paper No.: 2146
FOCUSED CONFERENCE GROUP: P08 - DEVELOPMENTS IN
THE TREATMENT OF SEXUAL DYSFUNCTION AND
DISEASES OF THE LOWER URINARY TRACT
HYDROGEN SULFIDE PROMOTES FUNCTION OF T-TYPE
CALCIUM CHANNELS AND SECRETION OF PROSTATIC
ACID PHOSPHATASE IN NEUROENDOCRINE-LIKE
PROSTATE CANCER CELLS FROM THE
ANDROGEN-SENSITIVE LNCAP CELL LINE
Kazuki Fukami(1), R Kasamatsu(1), S Yoshida(2), F Sekiguchi(1),
A Kawabata(1)
(1) Kinki University School of Pharmacy, Division of Pharmacology and
Pathophysiology, Osaka, Japan
(2) Kinki University School of Science and Engineering, Department of
Life Science,Osaka, Japan
Prostate cancer is one of the most common malignancies among men. An
increased population of prostate cancer cells acquiring a neuroendocrine
(NE) phenotype is involved in the malignancy and androgen-independent
pathology of prostate tumors. In the process of acquisition of the NE phenotype, Cav3.2 T-type calcium channels might be up-regulated, participating in the enhancement of secretory functions. Given our latest
evidence for activation/sensitization of Cav3.2 by hydrogen sulfide
(H2S), a gasotransmitter, in the present study, we asked if H2S regulates
functions of NE-like cells originating from the androgen-sensitive prostatic adenocarcinoma cell line LNCaP. LNCaP cells were differentiated
by stimulation with dibutyryl cyclic AMP into NE-like cells characterized
by neurite outgrowth and expression of T-type calcium channel-dependent membrane currents. The NE-like LNCaP cells exhibited increased
steady-state expression of mRNA for prostate-specific antigen (PSA) and
decreased response to androgen, compared with undifferentiated cells.
NaHS, a donor of H2S, enhanced T-type currents in NE-like LNCaP cells,
an effect abolished by mibefradil, a pan-T-type channel blocker. NaHS
also caused secretion of prostatic acid phosphatase (PAP) in NE-like cells.
Expression of cystathionine-b-synthase (CBS) and cystathionine-b-lyase
(CSE), distinct H2S-forming enzymes, was detected by Western blotting
in both undifferentiated and differentiated LNCaP cells. Our data demonstrate that H2S promotes secretion of PAP in the NE-like LNCaP cells
most probably through activation of T-type calcium channels. We thus
speculate that H2S might function as a general secretagogue for potential
mitogenic factors including neuropeptides in the androgen-independent
stage, participating in the progression of prostate cancer.
Paper No.: 2200
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
ROLE OF HIF-1B IN ENDOTHELIAL CELLS IN
MONOCROTALINE-INDUCED PULMONARY HYPERTENSION
IN MICE
Yayoi Fukuhara(1), S Tomita(1), Y Imamura(1), Y Horinouchi(1),
M Imanishi(1), T Sakurada(1), T Kitagawa(2), T Tamaki(1)
(1) University of Tokushima Graduate School, Institute of Health
Biosciences, Department of Pharmacology, Tokushima, Japan
(2) University of Tokushima Graduate School, Institute of Health
Biosciences, Department of Cardiovascular Surgery, Tokushima, Japan
Pulmonary hypertension (PH) is a life-threatening disease characterized
by significant increases in pulmonary arterial pressure (PAP) and right
ventricular hypertrophy (RVH). Hypoxia inducible factor (HIF)-1 has
also been shown to be related to pulmonary vascular reModelling in PH.
However, it is unclear the function of concerned factor in each cell type.
Therefore, in this study, we investigated that HIF-1 in pulmonary aortic
endothelial cells would be important mechanisms of PH development
and vascular remodeling. Endothelial cell-specific HIF-1b-deficient (HIF1b+/)) mice were generated using a Cre-loxP system. PH was induced
by monocrotaline (MCT). Four weeks after MCT injection, HIF-1b+/)
mice showed considerably less RV hypertrophy than wild type (WT)
mice. The right ventricle to left ventricle plus septum [RV/ (LV+S)] ratio
was 23.4 ± 2.5% in HIF-1b+/) mice, whereas WT mice was
29.2 ± 3.2% (p < 0.05). Medial hypertrophy is a hallmark of pathological vascular reModelling in PH. The medial thickness of PAs was markedly increased in the MCT-treated WT mice compared with untreated
WT mice. However, MCT-treated HIF-1b+/) mice significantly inhibited
the increase in percent medial thickness (WT mice:23.7 ± 5.9%, HIF1b+/) mice:12.2 ± 4.5%, p < 0.05). MCT-induced PH is attenuated in
endothelial cell-specific HIF-1b deficient mice. These results suggest that
the development of PH may be formed via HIF-1b in endothelial cells.
Paper No.: 471
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
PHARMACOVIGILANCE BETWEEN SCIENCE AND CRAFT
Giovanni Furlan
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
289
Bracco Imaging SpA, Department of Drug Safety & Pharmacoepidemiology, S. DonatoMilanese, Italy
Nowadays, drug safety is mainly an observational, rather than experimental science. Following collection of case reports, causality assessment
is performed and aggregate case reports are analysed, mainly by looking
for confounding factors and by focusing on pathology. Whenever an
important potential risk is identified, one or more studies may be conducted to confirm or disprove the signal. Often the results of the studies
are confounded and/or contradictory. Drug safety is currently engaged in
detecting a problem once it has already occurred, once the drug is on the
market. Drug safety should be engaged in primary prevention, so to prevent or minimize important risks before the drug is marketed. The aim
should be to identify/raise hypothesis on drug potential risks in pre-clinical stage and to test these hypothesis in subsequent clinical stage so to
have a precise idea of the drug risks once it is launched. Since the drug
primary and secondary mechanisms of action are the root cause of any
adverse reaction, pharmacology is a powerful tool to forecast the drug
safety profile. Despite the ability of pharmacology to forecast type A
adverse reactions in humans, sometimes these reactions are identified
once the drug has been marketed for a relatively long period of time.
This because only subjects having other risk factors for the iatrogenic
disease experience it; furthermore, severity, signs and symptoms of the
disease vary greatly from subject to subject. Just observing the adverse
events/reactions without having an understanding of the drug’s pharmacology can be very confusing.
Paper No.: 1410
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
BIOEQUIVALENCE STUDY OF TWO FORMULATIONS OF
DESMOPRESSIN NASAL SPRAY: A COMPARISON BETWEEN
THE NOVEL KW-8008RTS TO BE STORED AT ROOM
TEMPERATURE AND THE CONVENTIONAL KW-8008 TO BE
STORED IN REFRIGERATION
Hidetoshi Furuie(1), M Yagi(1), T Yajima(2), N Sakoda(2), K Matsumoto(2), H Ando(3), Y Amano(3), T Sakurai(1)
Paper No.: 2787
FOCUSED CONFERENCE GROUP: P07 - SIMULATION AND
DATA MODELLING IN DRUG DEVELOPMENT.
MOLECULAR DYNAMICS SIMULATIONS OF SEROTONIN
AND S-CITALOPRAM IN SEROTONIN TRANSPORTER
Mari Gabrielsen, AW Ravna, K Kristiansen, I Sylte
University of Tromsø, Department of Medical Biology, Tromsø, Norway
Transporters of the monoamines serotonin (SERT), dopamine (DAT) and
noradrenaline (NET) belong to the neurotransmitter:sodium symporter
(NSS) family. These presynaptic transporters remove neurotransmitters
from the synaptic cleft, thus reducing the number of neurotransmitters
available for interaction with postsynaptic receptors. The transporters are
major targets for antidepressant drugs, the selective serotonin reuptake
inhibitors (SSRIs) and the tricyclic antidepressants (TCAs), and are also
the targets for psychostimulants such as cocaine, MDMA (ecstasy), and
d-amphetamine. The three-dimensional structure of SERT has not yet
been determined. However, the first three-dimensional structure of a NSS
family member, the Aquifex aeolicus leucine transporter, LeuT, was
recently solved (Nature 2005; 437:215-23). Based on a comprehensive
alignment of NSS family members (Mol Pharmacol 2006; 70:1630-42)
and the LeuT crystal structure, a homology model of SERT has been
generated using the ICM program package. Different tryptamine derivatives and the SSRI S-citalopram have been docked into the putative substrate binding site of SERT, yielding two possible binding modes of the
ligands. In order to discriminate between these binding modes and
between the tryptamine derivatives (substrates) and S-citalopram (inhibitor), the SERT model has been embedded in a POPC lipid bilayer, generated by using the CHARMM-GUI membrane builder (J Comput Chem
2008; 29:1859-65), and long molecular dynamics (MD) simulations of
ligand-SERT-membrane complexes have been performed.
Paper No.: 2019
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
THE EFFECT OF RESVERATROL ON ILEAL SMOOTH
MUSCLE: FUNCTIONAL INVESTIGATION IN A RAT SEPSIS
MODEL
(1) Heishinkai Medical Group Inc., OPHAC Hospital, Osaka, Japan
(2) InCROM, Osaka, Japan
(3) Kyowa Hakko Kirin Co., Ltd, Japan
N Gacar(1), SS Gocmez(2), G Gacar(1), IK Celikyurt(1), Oguz Mutlu(1),
T Utkan(1)
Introduction: Desmopressin is a synthetic analogue of vasopressin and
useful for treatment of nocturnal enuresis. The conventional desmopressin nasal spray KW-8008 is inconvenient to carry because it should be
stored in a refrigerated state. KW-8008RTS is a novel formulation of desmopressin nasal spray under development and intended to be stored at
room temperature. We planned a bioequivalence study of these two formulations. However the study has some limitations, as Joukhadar et al.
suggested that there is a high inter-subject CV of approximately 60% and
intra-subject CV of more than 30% in plasma pharmacokinetics of desmopressin. Objective: The primary objective was to evaluate the bioequivalence between KW-8008RTS and KW-8008. The secondary
objective was to evaluate the safety of these formulations. Method: The
study was carried out in 28 healthy male volunteers in an open, randomized two-way crossover design with a single dose administration of
20&micro;g (10&micro;g per nostril) desmopressin in order to study bioequivalence and safety. Wash out period was at least 1 week. To avoid
large variation in plasma pharmacokinetics, the subjects were placed in a
decubitus position with a nasal clip. Results: The 90% confidence intervals of geometric mean ratios of Cmax and AUC0-t were 89.4%-102.2%
and 80.7%-106.5%, respectively. They implement the criteria of bioequivalence. No drug-related adverse events were observed. Conclusion:
The novel KW-8008RTS storable at room temperature and the conventional KW- 8008 to be refrigerated were safe and considered to be bioequivalent.
(1) Kocaeli University Medical Faculty, Department of Pharmacology,
Kocaeli, Turkey
(2) Namýk Kemal University Medical Faculty, Tekirdag, Turkey
Sepsis, a common and significant complication of acute illness, often
leads to a progressive and catastrophic condition known as multiple
organ dysfunction. The intestine is one of the major organs that are
involved in sepsis. Gastrintestinal stasis during sepsis may be associated
with gastrointestinal smooth muscle dysfunction. Resveratrol, is a natural
phytoalexin present in grapes and red wine, which possesses a variety of
biological activities including anti-inflammatory and antioxidative activities. Therefore, we aimed to examine whether the protective effects of
resveratrol on isolated intestinal smooth muscle responses and TNF-a
levels in septic rats. Polymicrobial sepsis was induced by the cecal ligation and perforation (CLP) procedure (24 h model). Rats received resveratrol (100 mg/kg,ip) or saline immeadiately after CLP. At 24 h after
CLP, the animals were anesthetized and contractile and relaxant
responses were measured in strips of ileal smooth muscle. We also measured TNF-a levels in blood samples. Contractile responses to KCl and
carbachol and relaxant responses to EFS were significantly decreased in
the strips of the sepsis group than control group (p < 0.05). Plasma
TNF-a levels of sepsis group were higher than those of control group
(p < 0.05). The impaired contractions and relaxations of strips were
markedly improved by treatment of resveratrol. Furthermore, increased
TNF-a levels of sepsis group reversed to the controls with pretreatment
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
290
of resveratrol. The results of the present study indicate that resveratrol
significantly improved CLP-induced relaxant and contractile responses
and that the beneficial effects of resveratrol might be attributed to its antiinflammatory activity.
Paper No.: 1032
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
EFFECTS OF RESVERATROL ON LEARNING AND MEMORY
IN RATS
Nejat Gacar(1), O Mutlu(1), T Utkan(1), I Komsuoglu Celikyurt(1),
SS Gocmez(2), G Ulak(1)
(1) Kocaeli University Medical Faculty, Department of Pharmacology,
Kocaeli, Turkey
(2) Namýk Kemal University Medical Faculty, Tekirdag, Turkey
Resveratrol (3,5,4-trihidroksi-trans-stilben) is a polyphenol which is
found especially in grape and red wine and exerts biological activities. In
recent studies resveratrol is shown to protect neuronal cells with its antioxidant activity, improve memory functions in streptozotocin induced
dementia models and reverse acetylcholine esterase activity. The aim of
this study was to investigate the effect of resveratrol on emotional and
spatial memory in naive rats and also on scopolamine induced memory
impairment in passive avoidance and Morris water maze (MWM) tests.
Male Wistar-albino rats and three different doses of resveratrol (12.5, 25
and 50 mg/kg) were used. Two way ANOVA post hoc Dunnett-t test is
used for the statistical analysis. There was no significant difference
between all groups for the first day latency while scopolamine significantly shortened the second day latency compared to control group and
resveratrol reversed this effect in all doses used in the passive avoidance
test. In the MWM test, 50 mg/kg resveratrol significantly increased the
time spent in the escape platform’s quadrant in the probe test while scopolamine decreased this parameter. The effect of scopolamine was
reversed by 50 mg/kg resveratrol. There was no significant difference for
the total number of movements and the total distance travelled in the
locomotor activity test in all groups. Resveratrol reversed the effect of
scopolamine and even had improving effects in naive animals at higher
doses. The results of this study should be clarified by further studies
including different learning and memory tasks.
Paper No.: 2700
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
ANALYSIS OF N-ACETYLTRANSFERASE 2 PHENOTYPE AND
GENOTYPE IN A TUNISIAN POPULATION
E Gaı̈es, Anis Klouz, I Salouage, S Trabelsi, I Hamza, N Jebabli,
M Aouichri, M Lakhal
National Center of Pharmacovigilance, Laboratory of Clinical
Pharmacology, Tunis, Tunisia
N acetylation polymorphism has been documented as a representative
pharmacogenetic trait. It is involved in the metabolism of a large number
of drugs and carcinogens and has diverse clinical consequences. The aim
of our study is to determine the acetylation phenotype, and to compare
acetylation phenotype with NAT2 genotype in Tunisian tuberculosis
patients. It consists on a prospective study including tuberculosis patients
in 2005 and in 2009. NAT2 phenotype was determined for two populations by calculating acetylation index 3 hours after isoniazid administration, the NAT2 genotype was only analysed by PCR/RFLP for patients
included in 2005. In 2005, we found a bimodal distribution of acetylation
phenotype (75% slow acetylators (SA) and 25% rapid acetylators (RA)).
The distribution of SA and RA genotypes was respectively 56% and
44%. The 341 C-to-T transition was the predominant mutation (37,3%).
Ten NAT2 alleles were found in our population, NAT2*4 was the major
one. Thirty-two different genotypes were found. The major genotype
was NAT2*6B / NAT2*4. The concordance value was 79%. The K statistics indicated moderate agreement between genotype and phenotype with
K = 0,55. In 2009, the distribution of the acetylation phenotype was
55,4% SA and 44,6% RA indicating a good concordance with genotype.
Results obtained in 2005, make us vigilant about the collection of blood
samples by respecting exactly the delay of 3 hours after isoniazid administration. This permit to obtain the results that should be obtained in
2005. Thus we can predict easily acetylation phenotypes with a poor
error risk.
Paper No.: 2785
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
TUNISIAN CLINICAL PHARMACOLOGY EXPERIENCE
E Gaı̈es, R Sahnoun, D Bensaid, N Jebabli, I Salouage, S Trabelsi,
I Hamza, M Aouichri, Anis Klouz, M lakhal
National Center of Pharmacovigilance, Laboratory of Clinical Pharmacology, Tunis, Tunisia
Clinical pharmacology is the science of drugs and their clinical use; it
connects the gap between medical practice and laboratory science. The
main objective is to promote the safety of prescription, maximise the
drug effects and minimise the side effects. At the beginning of the Clinical Pharmacology department activity on 1976, drug monitoring concerns only few molecules by the determination of correlation between
doses and concentrations. The activity of our department has been
evolved over the years and the number of monitored drugs has significantly increased. In fact, it increased from 1777 assays in 2000 to 7885
in 2009. By the way, we developed a new approach using classic pharmacokinetic for the determination of pharmacokinetics parameters.
Which was first determined only for cyclosporine and busulfan. In order
to minimise drug toxicity due to inter and intra-patients pharmacokinetic
variability, we developed drug population pharmacokinetic for several
anti-cancerous and immunosuppressive drugs. This approach proposes a
limited sampling strategy to estimate individual pharmacokinetics parameters. In conclusion, the clinician claim is increasing; this shows that the
activity of the Tunisian Clinical Pharmacology department is important
to guide clinician to promote the efficiency of such treatment and to
avoid toxicity.
Paper No.: 3176
FOCUSED CONFERENCE GROUP:
P11 - G PROTEIN-COUPLED 7TM RECEPTORS: FROM
MOLECULAR TO PHYSIOLOGICAL FUNCTION
ANGIOTENSIN II MODULATES A1D-ADRENOCEPTORS
EXPRESSION IN VASCULAR SMOOTH MUSCLE CELLS
Itzell Gallardo-Ortiz(1), P López-Sánchez(2), JJ López-Guerrero(1),
M Ibarra(1), R Villalobos-Molina(1)
(1) UNAM, FES-Iztacala, Mexico City, Mexico
(2) ESM, I.P.N., Mexico
Angiotensin II (Ang II) plays an important function regulating systemic
arterial pressure; it provokes cardiovascular reModelling and hypertension, as well as increase in a1-adrenoceptors mRNA and protein. The
aim of this work was to evaluate the influence of Ang II on a1-adrenoceptors in isolated smooth muscle cells of Wistar rat aorta. Smooth muscle cells were incubated with Ang II (100 nM) for different periods of
time (30min-24hrs), and protein was determined by Western blot for a1adrenoceptor subtypes. Data show an important increment of a1D-adrenoceptors after 30 min of Ang II incubation, then a time-dependent diminution in the expression of protein at later measurement points. Whereas
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
291
a1A-adrenoceptors shown opposite effects, they decreased at short times
and returned to basal. a1B-adrenoceptors were not modified by the peptide. Maximal expression was blocked by losartan and inhibited by
cycloheximide. Our data suggest that stimulation of AT1 receptors by
Ang II increased a1D-adrenoceptors in isolated cells, and this effect is
due to synthesis de novo of that receptor. In addition, data support a
putative crosstalk between Ang II and a1D-adrenergic pathways, and this
effect might be related with hypertension and hypertrophy induced by
Ang II.
Supported in part by grant IN224408 PAPIIT, DGAPA, UNAM.
Paper No.: 2296
FOCUSED CONFERENCE GROUP: P12 - ION
CHANNELOPATHIES: NEW WINDOWS ON COMPLEX
DISEASE AND THERAPY
CONTRIBUTION OF TRPC1 TO THE PATHOGENESIS OF
DYSTROPHIC MICE: ANALYSIS OF DOUBLEKNOCK-OUT
TRPC1 -/- DYSTROPHIN
Chiara Gallo(1), OM Dorchies(1), G Shapovalov(1), N Zanou(2),
P Gailly(2), E Roulet(1), UT Ruegg(1)
(1) University of Geneva, School of Pharmaceutical Sciences, Laboratory
ofPharmacology, Geneva, Switzerland
(2) Université Catholique de Louvain, Institute of Neuroscience, Laboratory of CellPhysiology, Brussels, Belgium
Calcium dysregulation has been shown as one of the triggering events
causing muscular degeneration in Duchenne muscular dystrophy (DMD).
The increased Ca2 + influx could result from transient membrane lesions
or from influx through store operated channels (SOC) or stretch activated
channels (SAC). These influxes may be due to an over-activation of cationic channels belonging to the TRP family. Since TRPC1 is a candidate
for SOC function and has been shown to be involved in skeletal muscle
function, we have generated a double knock-out TRPC1-/- - dys- (mdx)
mouse to analyze the involvement of TRPC1 in Ca2 + regulation and
muscle function in the dystrophic phenotype. Characterization of
TRPC1-/- - dys- (mdx) was performed by physiological, functional,
behavioral and molecular tests including TRPC1-deficient, dystrophindeficient and normal mice as controls. SOC activity was analyzed with
Ca2 + imaging experiments (Fura-2 on fibers) and 45Ca2 + flux experiments on entire muscle, using the SERCA inhibitor, thapsigargin. SAC
activity was tested using 45Ca2 + influx under hypo-osmotic conditions.
Electrophysiology was performed to characterize TRPC1 current on isolated Flexor Digitorium Brevis muscle fibers. Isometric force and fatigue
were measured in vivo. Fatigability was also tested with a grid test. Histology was performed on Tibialis Anterior muscle. Up until now,
although TRPC1 seems to play a role in force and fatigue mechanism of
otherwise normal mice, the contribution of TRPC1 to the pathogenesis
of mdx dystrophic mice is not clear. Further investigations are in
progress.
Paper No.: 1574
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
A SIMPLE, NOVEL METHOD FOR ASSESSING MEDICATION
ADHERENCE: CAPSULE PHOTOGRAPHS TAKEN WITH
CELLULAR TELEPHONES
for how to best accomplish this goal. In this study, we looked at three
types of adherence measurement methods (capsule count, MEMS and a
novel method- pictures taken by cellular telephones) and compared their
usefulness and accuracy. Overall adherence estimated by capsule count
was 94.9% (± 13.5%), by MEMS 93.6% (± 15.0%), and by photos
76.9% (± 14.6%). Weekly photographs and MEMS agreed with weekly
capsule counts with similar frequency (36% vs. 39%, respectively; OR
1.11, p= .79). When weekly measures disagreed with capsule count,
MEMS overestimated adherence more than photographs (39% vs. 14%;
OR 3.88; P < 0.001) and photographs underestimated adherence more
than MEMS (49% vs. 22%; OR 3.48; P < 0.001). Comparing against
capsule count, the novel method was found to be as useful as MEMS.
Given the ubiquity of cellular telephone use, and the relative ease of this
adherence method, we believe it to be a useful and cost effective
approach to adherence measurement that should be utilized by providers
in the future.
Supported by NIH DA018179.
Paper No.: 953
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
CORRELATION OF CYTOCHROME P450 3A4*18 GENETIC
POLYMORPHISMS WITH POSTOPERATIVE FENTANYL
REQUIREMENTS
Siew Hua Gan, PCS Tan, SK Hassan, N Abdullah
University Sains Malaysia, Human Genome Center, Kelantan, Malaysia
Interindividual variability in drug responses may be attributable to genetically-determined alteration in enzyme activity. In this study, the association between cytochrome P450 3A4 (CYP3A4) genetic polymorphisms
and postoperative fentanyl requirements was investigated. Patients
(n = 94) scheduled for gynaecological laparotomy received IV fentanyl
infusion (3 lg/kg/hr) after induction of general anaesthesia. Postoperative
fentanyl requirements were quantified by using a patient-controlled analgesia (PCA) and the number of IV fentanyl rescue analgesia required
were recorded. Pain control was assessed using visual analogue scores
(VAS) and fentanyl’s adverse effects were documented. CYP3A4*4,
CYP3A4*5 and CYP3A4*18 alleles of cytochrome P450 3A4 were identified by polymerase chain reaction–restriction fragment length polymorphism (PCR-RFLP). Differences in fentanyl requirements and VAS
among the various genotypes were compared. No mutations were
detected for CYP3A4*4 and CYP3A4*5 alleles. Allelic frequency for
CYP3A4*18 was 2.7% where eighty nine patients (94.7%) were wildtype, five (5.3%) were heterozygous while none was homozygous. No
significant difference was demonstrated between the genotype groups in
terms of fentanyl consumptions, pain control and adverse effects. The
result showed that genetic polymorphism of CYP3A4*18 does not play
an important role in influencing postoperative fentanyl requirements.
Paper No.: 954
FOCUSED CONFERENCE GROUP: P19 - GENERAL SESSION ONCOLOGY
HPLC METHOD DEVELOPEMENT FOR THE SIMULTANEOUS
DETERMINATION OF ARABINOSIDE-C AND DOXORUBICIN
CONCENTRATIONS IN HUMAN SERUM
Suew Hua Gan, AF Mistiran, AD Abdullah
GP Galloway, JE Guillén, JR Coyle, K Flower, John Mendelson
University Sains Malaysia, Human Genome Center, Kelantan, Malaysia
California Pacific Medical Center Research Institute, Addiction and Pharmacolog Research Laboratory, San Francisco, CA, USA
Medication non-adherence is an important factor in both everyday
healthcare and research methodology. Though many types of adherence
measuring utilities have been applied, there is as yet no ‘‘gold standard’’
Arabinoside-C (Ara-C) and doxorubicin hydrochloride (DOX) are two
anticancer agents commonly used for many types of cancers. In this
study, a new HPLC method was developed for the simultaneous determination of the two drugs in human serum. The effects of 1) using different
buffer types 2) different buffer pHs 3) adding different organic modifiers
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
292
such as triethylamine and acetic acid 4) changing column temperature 5)
changing mobile phase composition 5) varying the injection volume and
6) changing UV detector wavelength were investigated. Chromatographic separation was successfully achieved on a 250 mm X 4.6 mm
(C-18) column using a mobile phase consisting of a mixture of ammonium hydrogen phosphate (0.01M) adjusted to pH 6.2 and acetonitrile
(55:45). The flow rate was 0.5 ml/min and UV detection was done at
275 nm. The column temperature was set at 30C. The proposed HPLC
method was successfully applied to the determination of the investigated
drugs with good precision and accuracy as per recommended by the
FDA.
ischemia, whereas under the conditions of experimental myocardial
infarction its effect obviously decreases and in case of combined cardiocerebral pathology it is fully absent. By contrast, Afobazol (10 mg/kg) in
comparison with intact animals substantially increases cerebral perfusion
during vascular pathology of brain or heart, and it is even more effective
under the conditions of combined cardiac and cerebral vascular pathology. Thus, in conditions of combined ischemic damage of brain and
heart the cerebrovascular effects of Afobazol significantly increase, while
Nimodipin has no effect. It indicates that under the conditions of cardioneurologic disturbances these two drugs have different mechanisms of
cerebrovascular activity.
Paper No.: 991
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
EFFECT OF TAURINE, PYRIDOXINE AND A LIPOIC ACID ON
CAFETERIA DIET FED RATS
Paper No.: 1213
FOCUSED CONFERENCE GROUP: P02 - TRANSMEMBRANE
TRANSPORT: PERSPECTIVES FOR DISEASE AND DRUG
DISCOVERY
PHARMACOKINETIC INTERACTION BETWEEN PUERARIN
AND EDARAVONE AND EFFECT OF BORNEOL ON THE
BRAIN DISTRIBUTION KINETICS OF PUERARIN IN RATS
Priya Ganpathy, R Ghosh, S Pitre, V Kadam
C Gao, X Li, T Li, L Wang, Ming Xue
Bharati Vidyapeeth’s College of Pharmacy, Department of Pharmacology, Navi-Mumbai, India
Metabolic syndrome, a condition which lacks a concise definition and
treatment attracts research for newer therapies. Increased consumption of
westernized diet worldwide has contributed to more individuals being
diagnosed with metabolic syndrome. The cafeteria diet model, which
incorporates high calorie western food items, mimics the features of
human metabolic syndrome well and is a widely used screening model
for metabolic syndrome. In the present study, we have investigated the
effect of a combination of nutritional supplements on the three major
components of metabolic syndrome viz. obesity, insulin resistance and
dyslipidemia. Male Sprague-Dawley rats were fed high fat cafeteria diet
for a period of 5 weeks. The treatment group received a combination of
taurine, pyridoxine and a lipoic acid. The results of this in-vivo study
indicates that the proposed combination shows statistically significant (p
< 0.05) improvement in the body weight, serum levels of glucose, triglycerides, total cholesterol, free fatty acids and HOMA-IR. The candidate
combination proves to be beneficial in the cafeteria diet induced weight
gain, glucose and lipid imbalances and insulin resistance. Hence, from
the results of current in-vivo study we conclude that by combining nutritional supplements based on their individual pharmacologic potentials we
can alleviate the features of metabolic syndrome.
Paper No.: 1511
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
THE DIFFERENCE IN CEREBROVASCULAR EFFECTS OF
AFOBAZOL AND NIMODIPIN UNDER THE CONDITIONS OF
COMBINED CARDIAC AND CEREBRAL VASCULAR
PATHOLOGY
Tamara Ganshina, R Mirzoyan, N Khaylov, S Seredenin
Capital Medical University School of Chemical Biology and Pharmaceutical Sciences, Department of Pharmacolgy, Beijing, PR China
The aim was to investigate the pharmacokinetic interaction between
puerarin and edaravone and the effect of borneol on the brain distribution
kinetics of puerarin in rats.A reversed-phase high performance liquid
chromatography method was developed and validated for the simultaneous determination of puerarin and edaravone in rat plasma. The detection method was successfully applied to compare the pharmacokinetic
interaction and brain distribution kinetics of puerarin and edaravone,
using in situ microdialysis sampling, in rats after intravenous administration and co-administration with a single dose. The method gave good linearity and no endogenous material interfered with the two target
compounds and I.S. peaks. The limit of detection of puerarin and edaravone was 0.03 and 0.05 ug/ml, respectively. The precision determined of
testing were all within 10%. The combination of puerarin and edaravone
reduced drug elimination rates, gave a wider distribution and the dispositions of both drugs in rats were optimized. The amount of distribution of
puerarin in brain tissues were increased significantly and the elimination
of puerarin was obviously slow with borneol pretreated. The results provide an important information for an improved combined use of puerarin
and edaravone with borneol pretreated in the clinical practice.
Keywords: Puerarin; edaravone; borneol; pharmacokinetic interaction;
brain distribution kinetics; microdialysis
Paper No.: 1787
FOCUSED CONFERENCE GROUP: PW19 - INFLUENCE OF
DEGENERATION AND REPAIR IN THE CNS AND PERIPHERY
PINOCEMBRIN PREVENTS GLUTAMATE-INDUCED
APOPTOSIS IN SH-SY5Y NEURONAL CELLS VIA DECREASE
OF BAX/BCL-2 RATIO
Mei Gao(1), W-C Zhang(2), J-J Hu(1), G-H Du(1)
Russian Academy of Medical Sciences, V.V.Zakusov Institute of Pharmacology, Moscow, Russian Federation
Combined blood supply disturbances of the two most important organs –
brain and heart – occur in clinical practice quite often. However we were
unable to find any information about the influence of drugs on cerebral
circulation on the experimental models of ischemic cardiac damage or
during combined disturbances of cerebral and coronary circulation. A
well known calcium channel antagonist Nimodipin and selective anxiolytic drug with neuroprotective activity Afobazol were chosen for our
research. The experiments have shown that Nimodipin (0,03 mg/kg)
enhances cerebral perfusion of both intact animals and during cerebral
(1) Chinese Academy of Medical Sciences & Peking Union College,
Institute of Materia Medica, National Center for Pharmaceutical Screening, Beijing, PR China
(2) Beijing Institute of Technology, School of Life Science & Technology, Beijing, PR China
Pinocembrin is the most abundant flavonoids in propolis, and has been
proven to have antioxidant, antibacterial and anti-inflammatory property.
To assess the protective effects of pinocembrin on neurons, SH-SY5Y
neuronal cells were pretreated with pinocembrin for 2 h followed by
co-treatment with glutamate (2 mM) for 12 h. Cell viability was
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
293
determined by(3,4,5-dimethylthiazol-2-yl)-2,5-diphenylte-trazolium bromide assay, and apoptosis was confirmed by cell morphology, capillary
zone electrophoresis and flow cytometry assay. Cell morphology was
evaluated with Hoechst33258/PI dye. Treatment with pinocembrin (10-5,
10-6, 10-7mol/l) increased cell viability dose-dependently, inhibited LDH
release and attenuated apoptosis. Intracellular free [Ca2 + ] was increased
after glutamate exposure, and this increase was attenuated in cells treated
with pinocembrin. bax mRNA expression increased remarkably following
glutamate exposure and pinocembrin treatment manifested a reduction
effect. bcl-2 mRNA expression changes were not detected in groups with
or without pinocembrin. Western blotting results indicated that pinocembrin treatment reduced the expression of Bax and had no effect on Bcl-2,
thus decreased the BaẍCBcl-2 ratio, which is in consistent with the gene
expression result. Pinocembrin could also down-regulate the expression
of p53 protein, and inhibit the release of cytochrome c from mitochondria
to cytosol. Thus we conclude that pinocembrin exerts its neuroprotective
effects in glutamate injury model partly by inhibiting p53 expression,
thus BaẍCBcl-2 ratio, and the release of cytochrome c.
Acknowledgments: This work was supported by National Natural
Science Foundation of China(No. 30630073) and National Science and
Technology Major Projects for Key New Drug Innovation
(No.2009ZX09302-003).
Paper No.: 988
FOCUSED CONFERENCE GROUP: P19 - GENERAL
SESSION - HEPATOLOGY
CHARACTERIZATION OF METABOLITES AND
CYTOCHROME P450 ISOFORMS INVOLVED IN THE
MICROSOMAL METABOLISM OF ACONITINE
Yue Gao
Beijing Institute of Radiation Medicine, Department of Pharmacology &
Toxicology, Beijing, PR China
Intoduction: Veratridine is a lipid-soluble alkaloid extracted from Veratrum officinale and other species of the family Liliaceae. Veratridine prevents inactivation of Na+ channel via binding the receptor site 2, causes
influx of sodium ion and depolarization and induces apoptosis of neuronal
cells. Materials: In the present study, we investigated the metabolism of
veratridine and the effects of selective cytochrome P450 (CYP) inhibitors
on the metabolism of veratridine in rat liver microsomes. The metabolites
were separated and assayed by iquid-chromatography-electrospesults:
Result showed that four CYP isoforms (CYP1A, CYP2B, CYP2E1,
CYP3A) were involved in the metabolism of veratridine in vitro and
seven metabolites of veratridine were detected incubating with rat liver
microsomes. Conclusion: Some of the metaboliteswere presumed to be
potentialmediates of neurotoxicity via protein binging. Further research in
vivo needs to link the metabolism of veratridine to its toxicity.
Paper No.: 769
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
POTENTIALLY INAPPROPRIATE MEDICATIONS IN
ELDERLY ATTENDED AT PRIMARY CARE
pharmaco-therapeutically at elderly people attended in a primary care center of Santiago, Chile. Also prevalence of PIM was determined. A crosssectional study including interviews and revision of medical records in a
representative sample of elderly was developed. Adherence to treatment,
health-related quality of life (VAS-HRQoL), and functionality were measured. PIM was defined by 2003 Beers’ criteria. Data were analyzed using
STATA 10.1. Eighty three patients were included; 62.7% were women,
mean age was 72 ± 6.2 years. Hypertension (91%) and osteo-articular diseases (64%) were the most frequent illness. Within last year 25% of
patients were hospitalized, and 35% suffered at least one fall. The 92%
were functionally independents, and their mean VAS-HRQoL was 62.5 ±
22.5. The 53% received poly-pharmacy, and the 83% was adherents to
treatment. The 30.1% received PIM, and PIM use was the only variable
statistically associated with a greater number of prescriptions (5.7 ± 2,51
versus 4.7 ± 1,70, respectively). Prevalence of PIM found in present study
could indicate that it is necessary to assess the effect of PIM use over
health resources utilization, and HRQoL of older adults. However it is
necessary to do additionally studies including a greater number of
patients, representative of the national older population.
Paper No.: 891
FOCUSED CONFERENCE GROUP: P01 - CLINICAL
PHARMACOLOGY IN THE EMERGING COUNTRIES
ESTIMATION OF TRANSFER OF NIFURTIMOX, FOR
CHAGAS DISEASE, INTO BREASTMILK. POPULATION
PHARMACOKINETICS AND SIMULATION ANALYSIS
Facundo Garcia-Bournissen(1), J Altcheh(2), A Panchaud(3), S Ito(1)
(1) Hospital for Sick Children, Division of Clinical Pharmacology,
Toronto, Ontario, Canada
(2) Servicio de Parasitologia, Hospital de Niños ‘R. Gutierrez’, Buenos
Aires, Argentina
(3) Swiss Teratogen Information Service, Division of clinical Pharmacology and Toxicology, University Hospital, Lausanne, Switzerland
Introduction: Women with Chagas disease receiving treatment with nifurtimox are discouraged from breastfeeding. Many patients who would
receive treatment with nifurtimox live in extreme poverty, and have limited access to resources such as clean water and baby formula and may
not have safe alternatives to breast milk. We aimed to estimate, using
limited available pharmacokinetics data, potential infant exposure to nifurtimox through breast milk. Methods: Original nifurtimox plasma concentrations were obtained from published studies. Pharmacokinetic
parameters were estimated using nonlinear mixed effect Modelling with
NONMEM version VI. One thousand nifurtimox plasma-concentration
profiles were simulated and used to calculate the amount of drug that an
infant would be exposed to, if breastfed 150 ml/kg/day. Results and Conclusion: Estimation of bBreast milk concentrations on the basis of peak
plasma levels (1,361 ng/ml) and milk-plasma ratio were estimated. We
calculated infant nifurtimox exposure of a breastfed infant of a mother
treated with this drug to be below 10% of the maternal weight-adjusted
dose, even if milk – plasma ratio were overestimated. Simulation led to
similar estimates. Risk for significant infant exposure to nifurtimox
through breast milk seems small and below the level of exposure of
infants with Chagas disease receiving nifurtimox treatment. This potential degree of exposure may not justify discontinuation of breastfeeding.
Jose Luis Garcia Fuentes(1), M Jiron(1), I Ruiz(1), M Santibañez(2)
(1) Universidad de Chile, Departamento de Ciencias y Tecnologia
Farmaceutica, Santiago, Chile
(2) Departamento de Salud, Comuna de San Ramon, Santiago, Chile
Quality of drugs prescription in elderly has been estimated applying some
clinical tools. In Chile, older adults are principally attended at the primary
health care system, and until now there are no studies for detecting
the use of potentially inappropriate medicines (PIM) in these subjects.
The purpose of this study was characterized socio-demographic and
Paper No.: 3407
FOCUSED CONFERENCE GROUP: PW19 - INFLUENCE OF
DEGENERATION AND REPAIR IN THE CNS AND PERIPHERY
FAVORABLE STUDENT ATTITUDES TOWARDS HOSPITAL
TRAINING IN PHARMACOLOGY IN NURSING DEGREE
Cristina Garcı́a-Cabanes, V Maneu, MM Palmero, J Formigós-Bolea
University of Alicante, Department of Optics, Pharmacology & Anatomy, Alicante, Spain FARMAGITE
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
294
Pharmacology is a crucial discipline for students of nursing. Since
2000, the Area of Pharmacology of the University of Alicante has
been developing clinical training in Pharmacology as a part of the
hospital training. The objective of our study was to assess student’s
opinion about the hospital training in Pharmacology as a model for
strengthening learning objectives from Pharmacology theoretical classes. The implementation of this study in the subject ‘‘Pharmacology’’
of the second year of the Degree in Nursing, began the academic
year 2008-2009 and it has continued during the academic year 20092010. A total of 394 students did the hospital training during the second semester distributed in 9 hospitals. During the hospital training
the students had to report the pharmacological treatment of 3 patients:
prescribed drugs, therapeutic use, dosage prescribed, routes of administration and adverse reactions. A teacher of pharmacology visited
them once a month in order to solve doubts. At the end of the clinical training, evaluation was accomplished with an anonymous survey
through an online questionnaire self-administrated in the intranet. On
a scale of 1 to 10, results showed that students are aware of the
importance of the Pharmacology to nurses activity. Regarding Pharmacology clinical training, students think that acquired knowledge is relevant to their professional work. Respondents think that it should be
maintained in the new EHEA curricula. Student attitude was assessed
by noting their degree of agreement using a Likert scale. The overall
student attitude towards hospital training on pharmacology was positive.
Paper No.: 3277
FOCUSED CONFERENCE GROUP: P19 - GENERAL
SESSION - ONCOLOGY
CYTOTOXIC ACTIVITY OF BIOACTIVE FRACTIONS OF
BIOACTIVE FRACTIONS OF ASPARAGUS OFFICINALIS L.
Mª Dolores Garcı́a-Gimenez, R De la Puerta, MA Fernández-Arche,
MT Saenz-Rodrı́guez
University of Sevilla, Faculty of Pharmacy, Department of Pharmacology, Sevilla, Spain
Background: Triguero asparagus from Huetor - Tajar is a popular vegetable consumed in many part of the world but this product also has a
wide spectrum of bioactive compounds as polyphenols, dietary fiber
and saponins. Objetive: In this work, we report the cytotoxic activity
of bioactive fractions of Asparagus officinalis provided by Huetor-Tajar
(Granada) (flavonoids, fibers, saponins and freeze-dried asparagus).
Methods: The HT-29 human colon adenocarcinoma cell line was used.
The cytotoxic activity was evaluated using the sulforhodamine B
(SRB) assay following protocols established by the National Cancer
Institute, National Institutes of Health, Bethesda, MD. This colorimetric
assay estimates cell number indirectly by staining total cellular protein
with the dye SRB. HT-29 cells were seeded in 96-well plates (5000
cells/well) and, after 24 h, the cells were treated for 48 h with the fractions. The protein-bound dye was dissolved in 10 mM Tris base solution for optical density determination at 492 nm using a microplate
reader.Cells viability was expressed as porcentage in relation to controls. Data were averaged from al least three independent experiments
and are expressed as means ± SEM. The working concentrations used
were 1,10,25,50,100 and 1000 lM. Results: Our results show that the
flavonoids and saponins fractions inhibit the proliferation of HT-29
human colon adenocarcinoma cells in concentration-dependent manners.
IC50 values were 60 ± 4.68 and 73.80 lM respectively. 5-FU was
used a positive control (IC50:6.35 ± 4.32lM). Cell viability induced
by the flavonoids and saponins fractions was not reduced in the presence of the antioxidants NAC and MnTMPyP. This suggests that reactive oxygen species are not involved in the cytotoxic activity of these
fractions.
This sudy was supported by MCYT-AGL 2007-63703/ALI.
Paper No.: 1760
FOCUSED CONFERENCE GROUP: P14 - ADDICTION AND
DOPING: NEUROBIOLOGICAL AND CLINICAL
EFFECT OF METHYLENEDIOXYMETHAMPHETAMINE
(MDMA) ON CYTOSOLIC CALCIUM LEVELS IN PC 12
CELLS. ROLE OF NICOTINIC RECEPTORS
Sara Garcia-Rates, E Escubedo, J Camarasa, D Pubill
University of Barcelona, Department of Pharmacology and Therapeutic
Chemistry, Barcelona, Spain
Previous work from our group pointed to a role of a7 nicotinic acetylcholine receptors (nAChR) in the neurotoxicity induced by methamphetamine (MA)and MDMA because methyllycaconitine (MLA), a specific
a7 nAChR antagonist, attenuated neurotoxic effects such as oxidative
stress and dopaminergic damage. Also, we demostrated that MA and
MDMA displaced specific nicotinic receptor radioligands indicating that
they can directly interact with nAChR. Once demonstrated the affinity
we studied the effect of preincubation with MDMA on nAChR density
and, as nicotinic ligands such as nicotine do, MDMA induced binding
upregulation of both a7 and heteromeric nAChR types after incubation
times ranging from 6 to 48 h. We have now studied the effect of MDMA
on the activation of nAChR subtypes and the consequent calcium mobilization using fluorimetry in Fluo-4-loaded PC12 cells. MDMA produced
a rapid and sustained increase in fluorescence without reaching AChinduced maximum effect, and inhibited the ACh- induced response.
These results suggest that MDMA acts as a partial agonist on a7 nAChRs as the response was almost completely inhibited by MLA (10 nM)
and a-bungarotoxin (100 nM) but not by the b2 antagonist dihydro-berythroidine. Subsequently, calcium-induced Ca2 + release from endoplasmic reticulum and entry through voltage-operated calcium channels
are also implicated. Also, treatment with MDMA for 24 h significantly
increased basal Ca2 + levels and induced an increase in a-spectrin breakdown products indicating calpain and caspase 3 activation. Moreover,
pretreatment with MDMA induced functional up-regulation of calcium
responses to specific agonists of both heteromeric and a7 nAChR suggesting a nicotine-like regulatory effect.
Paper No.: 1951
FOCUSED CONFERENCE GROUP: P17 - NEW APPROACHES
AND TARGETS IN PSYCHIATRY
OXYTOCIN ANTAGONIST PREVENTS PRO-COGNITIVE
EFFECT OF ANGIOTENSIN IV
Paul Gard, C Naylor
University of Brighton, Centre for Biomedical and Health Science
Research, Brighton, UK
Angiotensin IV (AIV) is a hexapeptide fragment of angiotensin II which
has been shown to enhance memory consolidation and recall in rat and
mouse models. The mechanism of this effect is unknown. AIV has been
shown to act via insulin-regulated aminopeptidase (IRAP), which previously has been known as placental leucine aminopeptidase and oxytocinase; AIV inhibits the aminopeptidase activity of this enzyyme. Low doses
of oxytocin have similarly been shown to enhance aspects of memory in
animal models. The possibility therefore exists that the effects of AIV on
memory are mediated by an accumulation of endogenous oxytocin consequent to inhibition of oxytocinase. Using novel object recognition, memory was quantified as the proportion of time spent exploring the novel
object compared with the familiar object (D score). The effects of subcutaneous injection of 4.7lg.kg-1 AIV on novel object recognition in male
and female C57/BL6 mice were determined in the presence and absence
of 10 lg.kg-1 d(CH2) 5 [Tyr(Me)2, Orn8-AVT], an oxytocin receptor
antagonist. In comparison to saline control, AIV significantly enhanced
novel object recognition in both male and female mice. Prior administration of the oxytocin antagonist reduced the effects of the AIV such that
they were not significantly different from saline control, without having a
deleterious effect on the saline treated animals. Neither the AIV nor the
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
295
oxytocin antagonist significantly influenced mouse locomotor activity.
These results raise the possibility that the pro-cognitive effects of AIV are
mediated by accumulation of endogenous oxytocin although the selectivity of the antagonist actions warrant futher investigation.
Paper No.: 2052
FOCUSED CONFERENCE GROUP: P14 - ADDICTION AND
DOPING: NEUROBIOLOGICAL AND CLINICAL
INTRANASAL ADMINISTRATION OF THE DOPAMINE D3
RECEPTOR ANTAGONIST SB-277011A INHIBITS COCAINE
SELF-ADMINISTRATION AND COCAINE-ENHANCED
BRAIN-STIMULATION REWARD IN RATS
Eliot Gardner(1), Xiao-Qing Peng(1), Xia Li(1), Charles Ashby(2),
Zheng-Xiong Xi(1)
(16%) two days after the testosterone injection (p = 0.007). This is the
first time a perturbation in the lipoprotein profile is observed after only a
single dose of testosterone. The circulatory level of total testosterone was
associated with the total cholesterol levels on day 2, indicating that the
observed effect is directly associated with testosterone and not an artificial
effect of the injection i.e. stress. Moreover, HMGCR specific mRNA level
in HepG2 cells and protein expression in blood of the volunteers was
induced by testosterone. Our results may provide a partial molecular
explanation for the vasculatory adverse side effects of AAS abuse.
Paper No.: 3265
FOCUSED CONFERENCE GROUP: PW19 - INFLUENCE OF
DEGENERATION AND REPAIR IN THE CNS AND PERIPHERY
NEUROPROTECTIVE EFFECT OF DIMEBON IN RATS WITH
INTRACEREBRAL POST-TRAUMATIC HEMATOMA
(HEMORRHAGIC STROKE)
(1) National Institute on Drug Abuse, Intramural Research Program, Baltimore, MD, USA
(2) Saint John’s University, Jamaica, New York, USA
TL Garibova(1), TA Voronina(1), SA Litvinova(1), VV Grigoriev(2),
SO Bachurin(2), Tatiana S Kalinina(1)
SB-277011A is the most well characterized selective DA D3 receptor
antagonist. Extensive research during the past decade demonstrates that
systemic administration of SB-277011A consistently inhibits the actions
of cocaine or other addictive drugs in most animal models relating to
drug addiction. However, further development of SB-277011A has been
halted by GlaxoSmithKline Pharmaceuticals, due to unexpectedly poor
bioavailability (~2%) and a very short half-life (<20 min) in primates.
Recent research indicates that a wide variety of therapeutic compounds
with poor bioavailability can be delivered intranasally from nose into the
brain. In the present study, we investigated whether intranasal microinjections of SB-277011A inhibit intravenous cocaine self-administration
or cocaine-enhanced brain-stimulation reward (BSR) similar to that produced after intraperitoneal administration. We found that intranasal microinjections of SB-277011A produced a dose-dependent reduction in
break-point level for intravenous cocaine self-administration under progressive-ratio reinforcement conditions. Intranasal administration of SB277011A also produced a significant attenuation of cocaine-enhanced
BSR in rats. These findings suggest that intranasal administration of SB277011A is as effective in attenuating cocaine’s rewarding efficacy as
intraperitoneal SB-277011A administration. Such data support the potential use of intranasal SB-277011A in the treatment of drug addition in
humans.
Supported by NIDA IRP.
(1) Russian Academy of Medical Science, Zakusov Institute of Pharmacology RAMS, Moscow, Russian Federation
(2) Institute of Physiologically Active Compounds RAS, Moscow,
Russian Federation
Paper No.: 1834
FOCUSED CONFERENCE GROUP: P14 - ADDICTION AND
DOPING: NEUROBIOLOGICAL AND CLINICAL
TESTOSTERONE ENANTHATE INDUCES THE EXPRESSION
OF HMGCR AND CHOLESTEROL LEVELS
Paper No.: 1023
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
THE PROTECTIVE EFFECT OF ATORVASTATIN AGAINST
PROLIFERATIVE ALTERATIONS PRODUCED BY CROTON
OIL ON MOUSE EPIDERMIS
Nina Gårevik, C Skogastierna, A Rane, L Ekström
Karolinska Institute, Division of Clinical Pharmacology, Department of
Laboratory Medicine, Stockholm, Sweden
Background: The rate-limiting step in the cholesterol synthesis is catalysed by HMG-CoA reductase (HMGCR). It is well established that anabolic-andogenic steroids induce deleterious alterations in lipid
metabolism which may increased the risk of coronary artery disease. In
this study we investigated whether a single dose of testosterone enanthate
affect the cholesterol biosynthesis and the expression of the HMGCR
Study Design and Methods: 55 healthy male volunteers were given 500
mg testosterone enanthate as a single intramuscular dose of Testoviron—Depot. Total cholesterol levels prior to, and two days after testosterone administration were analysed. Protein expression of HMGCR in
whole blood was investigated by Western blotting. In order to study
whether testosterone regulates the mRNA expression of HMGCR, in vitro
studies were performed in a human liver cell-line (HepG2). Results and
Discussion: The total level of cholesterol was significantly increased
This investigation was carried out with purpose to examine the neuroprotective properties of dimebon (2,3,4,5-tetrahydro-2,8-dimethyl-5-(2(6-methyl-3-pyridyl)ethyl)-1H-pyrido(4,3-b)indole), an agent with neuroprotective activity, a positive modulator of NMDA and AMPA subtype
of glutamatergic receptors. The study used new models of intracerebral
post-traumatic hematoma (hemorrhagic stroke). Hemorrhagic stroke (HS)
was produced in animals by cerebral tissue destruction in the internal
capsule region. Dimebon in dose of 0.1 mg/kg was administered i.p. 3-4
hrs following the operation. It was established that the agent diminished
HS-induced neurological deficit, movement co-ordination disturbances.
Dimebon was found to prevent the animal death caused by HS. The animals which were learned the passive avoidance reflex prior to HS on
Day 3 after operation demonstrated disturbances in the retrieval of conditioned passive avoidance reflex (PAR). With dimebon PAR retrieval
disorders were much less pronounced. The results obtained from these
studies provided fair evidence of neuroprotective and memory-restoring
properties of dimebon in a new model of hemorrhagic.
Alireza Garjani(1), Y Doustar(2), H Rezazadeh(1), S Andalib(1), N
Maleki-Dizaji(1)
(1) Tabriz University of Medical Sciences, School of Pharmacy, Department of Pharmacology, Tabriz, Iran
(2) Islamic Azad University Faculty of Veterinary Medicine, Tabriz, Iran
A growing body of preclinical data indicates that statins may have antineoplastic properties, but some studies raise the possibility that statins
may possess a carcinogenic potential. A single application of croton oil
to mouse skin induces an ordered sequence of ultrastructural changes in
the cells of epidermis, which correlate with changes in thickness, number
of nucleated cell layers, and mitotic index of the interfollicular epidermis.
To evaluate the effect of atorvastain on ultrastructural and histological
alterations produced by croton oil, Swiss albino mice were received atorvastatin (3, 6, 9 mg/kg/day; orally) for a period of 2 weeks prior to single
topical application of croton oil (0.5% in 200ll acetone/mouse) to the
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
296
shaved backs of the animals. After 24 hour animals were killed under
deep anesthesia and the skin was excised and fixed in Stieve’s solution
and then processed for histological examination. All parameters: disturbance of cell polarity, inflammatory response, nucleated cell layer, thickness of epidermal layer, and mitotic index were inhibited dose
dependently (P < 0.001) by pre-treatment with atorvastatin. The number
of cells in mitosis in 2500 lm2 of interfollicular epidermis was reduced
from 3 ± 0.3 in untreated group to 2 ± 0.2, 1 ± 0.2 (P < 0.01), and 0.6 ±
0.2 (P < 0.001) by 3, 6, and 9 mg/kg of atorvastatin, respectively. The
results of this study show strong anti-inflammatory and anti-proliferative
effects of atorvastatin and suggest that statins may suppress events associated with carcinogenesis.
Paper No.: 2733
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
A JERTE VALLEY CHERRY-BASED PRODUCT ACTS AS AN
ENHANCER OF SLEEP, IMPROVES MOOD AND POSSESSES
ANTIOXIDANT PROPERTIES
Maria Garrido(1), D Gonzalez-Gomez(2), M Lozano(2), MT Hernandez(2), MS Salcedo(3), C Barriga(1), SD Paredes(1), AB Rodriguez(1)
(1) University of Extremadura Faculty of Science, Department of Physiology, (Neuroimmunophysiology and Chrononutrition Research Group)
Badajoz, Spain
(2) Technological Institute of Food and Agriculture (Intaex), Badajoz,
Spain
(3) Perpetuo Socorro Hospital, SES, Children’s Psychiatry Unit, Badajoz,
Spain
Tryptophan, serotonin and melatonin are implicated in the regulation of
sleep. Melatonin directly, or indirectly by supplementation with its precursor tryptophan, functions as a powerful antioxidant. Jerte Valley cherries are rich in tryptophan (Cubero J et al, Food Anal. Methods 2009;
doi 10.1007/s12161-009-9084-1), serotonin and melatonin (GonzálezGómez D et al, Eur. Food Res. Technol. 2009; 229:223-229). The study
was aimed at evaluating the effect of Jerte Valley cherry product intake
on the sleep-wake cycle and mood of middle-aged volunteers as well as
determining the participants’ urine levels of total antioxidant capacity,
cortisol, 5-hydroxyindolacetic acid, and 6-sulfatoxymelatonin. Volunteers
consumed 28 g of this product twice a day for 5 days. Actigraphic monitoring was used to record and display the temporal patterns of the individuals’ activity and rest during 5 days before the beginning of the assay
(basal), 5 days of assay (assay) and 5 days afterwards (post-assay). Psychological tests were performed in basal, assay and post-assay conditions. Total antioxidant capacity, cortisol, 5-hydroxyindolacetic acid and
6-sulfatoxymelatonin were analyzed using colorimetric or ELISA assay
kits in first-void morning and 21:00 h urines collected on the third and
fifth day in basal, assay and post-assay conditions. The ingestion of this
product improved nocturnal rest and mood, decreased cortisol and
increased total antioxidant capacity, 5-hydroxyindolacetic acid and 6-sulfatoxymelatonin urinary levels. The product may be employed as a
source of antioxidants and be useful in states where sleep and mood disorders have been reported.
Funded by PDT0A008-Junta de Extremadura.
Paper No.: 1370
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
EFFECT OF AT1 RECEPTOR BLOCKADE ON THE
ANTIOXIDANTS ENZYMES ACTIVITIES IN AN
EXPERIMENTAL MODEL OF PERIODONTAL DISEASE
INDUCED BY LIPOPOLYSACCHARIDE IN THE RAT
In addition to its vasoactive actions, angiotensin II (AngII) has been
implicated in the release of pro-inflammatory cytokines, activation of
mitogenic factors, oxidative stress and modulation of nitric oxide synthesis, through stimulation of AT1 receptor (AT1R).AT1R stimulation
increases oxidative stress through activation of NAD(P)H oxidase.
Increased reactive oxygen species (ROS) production leads to tissue damage associated with activation of the inflammatory process. Local administration of bacterial endotoxin (lipopolysaccharide, LPS) in the gum of
the rats produces an inflammatory response that progress in periodontal
disease. AT1R blockade reverse the AngII mediated pro-inflammatory
effects in the vasculature system and in chronic hypertension. The aim of
the present study was to assess the role of AT1R blockade in the inflammatory reaction and ROS generation in an experimental model of LPSinduced periodontal disease in the rat. Male Sprague-Dawley rats (220240g) were divided in four groups: Control, LPS, VAL: Valsartan (20
mg/kg, p.o, 7 and 14 days) and LPS+VAL. Rats were anesthetized with
10% ketamine (60 mg7kg) and euthanized by decapitation, the maxilla
dissected and the connective tissue collected in the appropriate buffer.
CAT, SOD, GPx activity was determined by spectrophotometric analysis
and NOS activity was assayed by monitoring the conversion of radiolabelled L-arginine to L-citrulline. LPS significantly increased tissue CAT,
SOD, GPx and NOS activity. VAL blunted the LPS-increased antioxidant
enzymes and NOS activity. Our results indicate a role AngII and AT1
receptor in the pathogenesis of the LPS-induced periodontal disease.
(Misión Ciencia, Proyecto ECCV No. 2007001585).
Paper No.: 2305
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
NOVEL SMALL MOLECULAR WEIGHT CCL2 INHIBITORS
REDUCE INFLAMMATORY RESPONSE IN MICE
Beatrice Garrone, A di Matteo, G Mangano, C Bartella, C Apicella,
N Cazzolla, G Furlotti, A Guglielmotti
Angelini Research Center-ACRAF, Department of Pharmacology, Rome,
Italy
Prevention of inflammation through blockade of the chemokines/chemokine receptors system is a major target for pharmacological intervention.
Angelini has an ongoing development program on CCL2 inhibition
based on an original small molecule, bindarit, which is a selective inhibitor of CCL2 synthesis. Bindarit has recently completed a phase II clinical
study in patients with diabetic nephropathy and is currently tested in coronary in-stent restenosis. Based on bindarit promising results, a group of
derivatives was selected from in-house library and screened in order to
select new CCL2 synthesis inhibitors. Activity of compounds on CCL2
production was evaluated in vitro on LPS-stimulated MonoMac6 cells.
CCL2 accumulation was quantified by ELISA. In addition, the effect on
specific CCL2 mRNA transcripts was also assessed. In vivo effects were
studied in mice measuring CCL2 plasma levels in LPS-treated animals
as well as thioglycollate-induced peritoneal macrophages recruitment. In
the MonoMac6 model, compounds showed CCL2 inhibition with an
IC50 in the range 20-329 lM. The most active compounds (IC50 lower
than 100 lM) were tested in vivo. Intraperitoneal administration of AF6,
AF11 and AF13 significantly reduced CCL2plasma levels with 39, 42
and 51% inhibition, respectively. Moreover, AF1, AF13and AF14
showed a significant effect on thioglycollate-induced peritonitis by
decreasing macrophages recruitment in the range 30-43%. Present results
and potential advantages of CCL2 synthesis inhibitors versus CCL2-specific mAbs and receptor antagonists call for further characterization
of these novel small molecular weight compounds in inflammatory
conditions.
Maria del Rosario Garrido, MG Matos, JA Silva, Y Mathison, A Israel
University of Venezuela, Schook of Pharmacy, Laboratory of Neuropeptides, Caracas, Venezuela
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
297
Paper No.: 2352
FOCUSED CONFERENCE GROUP: P04 - PHARMACOEPIDEMIOLOGY, CURRENT CONTROVERSIES AND OPPORTUNITIES
QUALITY OF ANTIBIOTIC PRESCRIBING IN INFANT IN
AMBULATORY CARE
Kristina Garuoliene, J Gulbinovic
Department of Pathology, Forensic Medicine & Pharmacology, Vilnius,
Lithuania
Inappropriate antibiotic prescribing in infants increases antimicrobial
resistance, unnecessary health care costs and may influence infants’
health status in the future life. Quality of antibiotic prescribing in infants
in ambulatory care in Lithuania during 2003-2008 was analysed. The
data on antibiotic dispensing was obtained from the population based
electronic database of the Lithuanian Compulsory Health Insurance
Information system. This database provides the complete prescription
medication history of patients, diagnosis according IDC-10, identifies
prescriber and dispensing pharmacy. The study included all infants (0-12
months of age), who claimed at least one dispensing for any systemic
antibacterial (ATC code: J01) during 2003-2008. The number of dispensed prescriptions per 1000 infants decresed from 1047 in 2003 to 719
in 2008. The prevalence rate of infants treated with antibiotics decresed
from 56,7% to 43,8% during study period. The percentage of infants
who received multiple courses of antibiotic treatment (more that 3 per
year) decresed from 18,5% in 2003 to 15,2% in 2008. The most common
indications for antibiotic prescribing were acute bronchitis, acute pharyngitis, and common cold which made up 29%, 24% and 13% out of all
antibiotic prescriptions, respectively. Broad spectrum penicillins with or
without b lactamase inhibitors were the most often used antibiotics and
made up 70% out of all prescriptions. Three fold differences in antibiotic
prescribing were found between different regions. Obtained results suggest wide unnecessary prescribing of antibiotics in infants.
Paper No.: 1974
FOCUSED CONFERENCE GROUP:
P11 - G PROTEIN-COUPLED 7TM RECEPTORS: FROM
MOLECULAR TO PHYSIOLOGICAL FUNCTION
THE EFFECT OF NOCICEPTIN ON UTERINE
CONTRACTIONS IN TERM-PREGNANT RATS IN VITRO
Robert Gaspar(1), A Klukovits(1), K Tekes(2), O Gunduz(3),
S Benyhe(3), A Borsodi(3), BH Deak(2), J Hajagos-Tóth(1), J Verli(1),
G Falkay(1)
(1) University of Szeged Faculty of Pharmacy, Department of Pharmacodynamics and Biopharmacy, Szeged, Hungary
(2) Semmelweis University, Department of Pharmacodynamics,
Budapest, Hungary
(3) Hungarian Academy of Sciences, Institue of Biochemistry, Biological
Research Centre, Szeged, Hungary
The actions of the endogenous peptide nociceptin (N/OFQ) on the myometrium have not been investigated previously. Our aim was to study
the presence and functional role of N/OFQ in the modulation of uterine
contractility in pregnant rats at term.The presence of N/OFQ and its
receptors (NOP) in the uterus were detected by radioimmunoassay (RIA)
and radioligand-binding experiments. The N/OFQ-stimulated G-protein
activation was assessed by a [32S]GTPcS-binding technique. The effects
of N/OFQ in uterine rings precontracted with KCl or oxytocin were also
tested in vitro. Uterine levels of cAMP were measured by enzyme immunoassay (EIA). The K+ channel blockers tetraethylammonium and paxilline were used to study the role of K+ channels in mediating the uterine
effects of N/OFQ. Both N/OFQ and NOP were present in the uterus. N/
OFQ revealed a maximum contraction inhibition of ~30%, which was
increased to 40% by naloxone. Naloxone and pertussis toxin significantly
attenuated the G-protein-stimulating effect of N/OFQ. The uterine cAMP
levels were found to be elevated by N/OFQ and naloxone, and after
preincubation with pertussis toxin. Tetraethylammonium and paxilline
reduced the contraction-inhibiting effect of N/OFQ and naloxone to ~10
and 15%. We presume that N/OFQ plays a role in regulating uterine contractility at term. Its effect is mediated partly by Gs-proteins coupled to
NOP receptors and elevated cAMP levels, and also by Ca2 + -dependent
K+ channels. Our results demonstrate a novel action and signalling pathway for N/OFQ that might be a potential drug target.
Paper No.: 1260
FOCUSED CONFERENCE GROUP: P06 - THE HEART GONE
WRONG; STABILIZATION OF CARDIAC FUNCTION
EFFECT OF GRAPEFRUIT JUICE ON DRONEDARONE PHARMACOKINETICS IN HEALTHY SUBJECTS
Christine Gaud(1), A Brunet(2), E Sultan(2), J Galleyrand(3), T Duvauchelle(4)
(1) Sanofi-Aventis R&D, Department of Clinical and Exploratory Pharmacology,Montpellier, France
(2) Sanofi-Aventis R&D, Department of Global Metabolism and Pharmacokinetics,Montpellier, France
(3) Sanofi-Aventis R&D, Department Biostatistics and Programming,
Montpellier,France
(4) Institut ASTER, Paris, France
Dronedarone is a multichannel blocker developed for the treatment of
patients with atrial fibrillation. It is highly metabolized by CYP3A4 with
a first-pass metabolism. This study evaluated the effect of multiple ingestions (tid) of 300mL double-strength grapefruit juice (GFJ), on pharmacokinetics (PK) of dronedarone and its active metabolite, SR35021. This
randomized, open-label, non–placebo-controlled, 2-treatment (dronedarone + water or dronedarone + GFJ), 2-period crossover study was conducted in 24 healthy young men. Each period had 3 phases: Phase 1 and
Phase 2 with single doses of dronedarone in fasted and fed conditions
(data not presented), and Phase 3 with multiple doses of dronedarone
400 mg bid in fed condition for 10 days. GFJ was ingested for 20 days.
Plasma dronedarone/SR35021 concentrations were determined using a
validated liquid chromatography-mass spectrometry method. Non-compartmental PK parameters were calculated. Concomitant oral ingestions
of double-strength GFJ 300 mL tid increased maximal plasma concentration (Cmax) and area under the plasma curve (AUC0-12) of dronedarone
after 400 mg bid multiple doses in fed condition by2.5- and 3.0-fold,
respectively, and did not change SR35021 Cmax and AUC. For electrocardiography (ECG) parameters, mean changes from baseline were
+19.4 msvs. +5.7 ms in PR and +21.5 ms vs. +7.8 ms in QTc during
dronedarone + GFJ and dronedarone + water co-administrations, respectively. In very stringent conditions, GFJ increased dronedarone exposure
by approximately 3-fold, an increase that was associated with consistent
changes of dronedarone in ECGparameters. Patients should, therefore,
avoid GFJ beverages while taking dronedarone.
Paper No.: 1261
FOCUSED CONFERENCE GROUP: P06 - THE HEART GONE
WRONG; STABILIZATION OF CARDIAC FUNCTION
EFFECTS OF DRONEDARONE ON THE
PHARMACOKINETICS AND PHARMACODYNAMICS OF
WARFARIN IN HEALTHY SUBJECTS
Christine Gaud(1), J-P Duret(2), A Brunet(2), E Sultan(2),
R Ebrahimi(3)
(1) Sanofi-Aventis R&D, Department of Clinical and Exploratory Pharmacology, Montpellier, France
(2) Sanofi-Aventis R&D, Department of Global Metabolism and Pharmacokinetics, Montpellier, France
(3) Sanofi-Aventis R&D, Department Biostatistics and Programming,
Montpellier, France
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
298
Dronedarone is a multichannel blocker developed for the treatment of
patients with atrial fibrillation (AF). This study assessed the effects of
multiple doses of dronedarone on the single dose pharmacokinetics (PK)
and pharmacodynamics(PD) of warfarin, a frequently co-administered
drug in patients with AF. Following a randomized 2-treatment, 2sequence, crossover design, 17 healthy young men received a single dose
of warfarin 30 mg in Period 1 and a 14-day multiple administration of
dronedarone 600 mg bid co-administered with a single dose of warfarin
30 mg on Day 8 of Period 2. Plasma was analyzed for R- and S-warfarin
(2 chiral forms) using a validated high-performance liquid chromatography/ultraviolet method. Non-compartmental PK parameters and PD
activity (international normalized ratio [INR]) were determined. Ratios of
geometric means (warfarin + dronedarone/warfarin alone) with 90% confidence intervals (CI) were: R- warfarin, 1.04 (0.98–1.11) for maximal
plasma concentration (Cmax) and 1.11 (1.05–1.18) for area under the
plasma curve (AUClast); S- warfarin, 1.07 (1.00–1.14) for Cmax and
1.19 (1.13–1.26) for AUClast. INR ratios of geometric means (warfarin
+ dronedarone/warfarin alone) with 95% CI were 1.06 (0.99–1.13) at
peak and 1.07 (1.02–1.12) for AUC. Dronedarone at a higher dose than
the proposed therapeutic one (400 mg bid) had a very limited effect on
PK of S-warfarin and no effect on R-warfarin. There were no clinically
relevant effects on INR, suggesting that monitoring should be done in
accordance with the warfarin label.
Paper No.: 1262
FOCUSED CONFERENCE GROUP: P06 - THE HEART GONE
WRONG; STABILIZATION OF CARDIAC FUNCTION
EFFECTS OF DRONEDARONE ON THE
PHARMACOKINETICS OF DIGOXIN IN HEALTHY SUBJECTS
Christine Gaud(1), F Hurbin(2), A Brunet(2), E Sultan(2),
J Galleyrand(3), E Guénolé(4)
(1) Sanofi-Aventis R&D, Department of Clinical and Exploratory
Pharmacology, Montpellier, France
(2) Sanofi-Aventis R&D, Department of Global Metabolism and Pharmacokinetics, Montpellier, France
(3) Sanofi-Aventis R&D, Department Biostatistics and Programming,
Montpellier, France
(4) Therapharm Recherches, Caen, France
Dronedarone is a multichannel blocker developed for the treatment of
patients with atrial fibrillation. In vitro, it has a potential to inhibit
digoxin secretion mediated by P-glycoprotein (P-gp). This study assessed
the effects of multiple doses of dronedarone on multiple-dose pharmacokinetics (PK) of digoxin and the tolerability of co-administration. A placebo-controlled, 2-treatment crossover study was conducted in 20
healthy young men receiving digoxin 0.25 mgqd for 10 days (loading
dose, 0.75 mg), alone or co-administered with dronedarone 400 mg bid
for 10 days in fed condition. Plasma was analyzed for digoxin with a validated immunoassay method. Non-compartmental PK parameters were
calculated. ECG and vital signs were regularly performed. Ratios of geometric means (digoxin + dronedarone/digoxin alone) with 90% confidence intervals were 1.75 (1.58–1.93) for maximal plasma concentration
(Cmax), 2.57(2.21–2.98) for area under the plasma curve (AUC0-24),
0.57 (0.46–0.69) for renal clearance (ClR0-24), and 1.45 (1.19–1.78) for
urinary excretion (Ae0-24).The tolerability of the co-administration was
satisfactory. No prolonged QTc were observed. Dronedarone at therapeutic dosing regimen significantly increased steady state digoxin exposures
by 2.6-fold, due to decreased digoxin renal clearance and increased
absorption through P-gp inhibition at both levels. When digoxin is coadministered with dronedarone, it is recommended that digoxin dose
should be reduced by 50% and serum digoxin levels, clinical signs and
ECG should be closely monitored according to label.
Paper No.: 1263
FOCUSED CONFERENCE GROUP: P06 - THE HEART GONE
WRONG; STABILIZATION OF CARDIAC FUNCTION
PHARMACOKINETIC AND PHARMACODYNAMIC
INTERACTION OF DRONEDARONE AND VERAPAMIL IN
HEALTHY SUBJECTS
Christine Gaud(1), S Doroumian(2), A Brunet(2), E Sultan(2),
J Galleyrand(3)
(1) Sanofi-Aventis R&D, Department of Clinical and Exploratory
Pharmacology, Montpellier, France
(2) Sanofi-Aventis R&D, Department of Global Metabolism and Pharmacokinetics, Montpellier, France
(3) Sanofi-Aventis R&D, Department Biostatistics and Programming,
Montpellier France
Dronedarone is a multichannel blocker developed for the treatment of
patientswith atrial fibrillation. It is highly metabolized by, and has a
potential to inhibit, CYP3A4. This study assessed the pharmacokinetic
(PK) and pharmacodynamic (PD) interactions of multiple doses of dronedarone and the calcium-channel blocker verapamil (CYP3A4 substrate).
A randomized crossover study with 13-day washouts was conducted in
21 healthy young men receiving dronedarone 400 mg bid alone, verapamil 240 mg qd alone and co-administration for 14 days in fed condition.
Plasma was analyzed for dronedarone and its active metabolite,
SR35021, using a validated liquid chromatography mass spectrometry/
mass spectrometry method, and for verapamil/norverapamil using a
validated high-performance liquid chromatography/ultraviolet method.
Non-compartmental PK analyses were performed. Effects on electrocardiography (ECG) parameters were evaluated. Ratio of geometric means
(verapamil +dronedarone/verapamil alone) with 90% confidence intervals
(CI) were 1.30(1.14–1.48) and 1.29 (1.15–1.44) for verapamil and norverapamil area under the plasma curve (AUC0-24), respectively. Ratio of
geometric means (dronedarone +verapamil/dronedarone alone) with 90%
CI were 1.48 (1.38–1.58) and 0.98(0.92–1.04) for dronedarone and
SR35021 AUC0-12, respectively. Co-administration increased the PR
and QTcF (Fridericia’s correction) greater than each product alone (mean
changes compared with dronedarone alone: +10.2 ms and +11.9 ms,
respectively; compared with verapamil alone: +15.8 ms and +31.9
ms,respectively). Dronedarone increased verapamil exposure 1.3-fold,
and verapamil increased dronedarone exposure 1.5-fold. Due to alterations in ECG values, verapamil should be initiated at low dose and
up-titration should be done only after ECG assessment.
Paper No.: 1264
FOCUSED CONFERENCE GROUP: P06 - THE HEART GONE
WRONG; STABILIZATION OF CARDIAC FUNCTION
PHARMACOKINETICS AND TOLERABILITY OF
DRONEDARONE IN PATIENTS WITH MODERATE HEPATIC
IMPAIRMENT
Christine Gaud(1), A Brunet(2), E Sultan(2), J Galleyrand(3),
K Lasseter(4), T Marbury(5)
(1) Sanofi-Aventis R&D, Department of Clinical and Exploratory
Pharmacology, Montpellier, France
(2) Sanofi-Aventis R&D, Department of Global Metabolism and Pharmacokinetics, Montpellier, France
(3) Sanofi-Aventis R&D, Department Biostatistics and Programming,
Montpellier,France
(4) Clinical Pharmacology Associates, Miami, FL, USA
(5) Clinical Research Center, Orlando, FL, USA
Dronedarone is a multichannel blocker developed for the treatment of
patients with atrial fibrillation. It is highly metabolized by CYP3A4 and
highly bound to plasma protein. This study assessed the pharmacokinetics (PK) of dronedarone and its active metabolite (SR35021), and the
pharmacodynamics (PD) and tolerability of dronedarone in patients with
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
299
hepatic impairment. Thirteen patients with moderate hepatic impairment
(Child-Pugh class B) were globally matched by sex, age, and weight
with 13 healthy subjects. After determining that PK changes were limited
(<2-fold increase in exposure) and safety results were satisfactory in 4
patients and 4 subjects receiving 400 mg qd for 7 days, the study proceeded with 400 mg bid (therapeutic dose) for 7 days in 8 patients and 8
subjects. Total and unbound plasma concentrations of dronedarone/
SR35021 were assessed using a validated liquid chromatography-mass
spectrometry method, and a non-compartmental analysis was performed.
Steady state was reached within 3 to 6 days in patients and subjects for
dronedarone and SR35021. In patients as compared to subjects, total and
unbound steady state exposures increased by 1.3- and 1.9-fold for dronedarone, respectively, and decreased by ~ 53 % and ~62% for SR35021.
For electrocardiography (ECG) parameters, mean changes from baseline
were +16.77 ms vs. +7.97 ms in PR and +8.58 ms vs. +5.37 ms in QTcF
in patients and subjects, respectively. Some patients experienced mild
gastrointestinal disorders. Based on mild to moderate exposure increases,
ECG effects and tolerability, dronedarone may be used at the therapeutic
dose in patients with moderate hepatic impairment.
Paper No.: 2242
FOCUSED CONFERENCE GROUP:
P11 - G PROTEIN-COUPLED 7TM RECEPTORS: FROM
MOLECULAR TO PHYSIOLOGICAL FUNCTION
STEREOSELECTIVITY OF THE RECOMBINANT AND NATIVE
HISTAMINE H3 RECEPTOR ISOFORMS: THE
H3-AUTORECEPTOR IS A SHORT ISOFORM
Florence Gbahou, A Rouleau, S Morisset, J-M Arrang
INSERM U894 Centre de Psychiatrie et Neurosciences, Sainte-Anne,
Paris, France
The histamine H3 receptor (H3R) was pharmacologically characterized as
an autoreceptor regulating histamine synthesis and release in the rat
brain. Activation of histaminergic neurons promotes arousal, attention,
improves learning and, has been proposed as a symptomatic therapeutic
approach in human attentional and ageing disorders such as Alzheimer’s
disease. Since its cloning in 1999, the molecular studies of the receptor
have shown that the H3R belongs to the G-protein-coupled receptor family, displays constitutive activity, and has many isoforms generated by
deletion of a pseudo-intron in the third intracellular loop. In spite of the
length of this loop, only minor pharmacological differences have yet
been reported between H3R isoforms. In this study, we hypothesized that
a deletion in this important loop would promote some modifications in
the 3D structure of the H3R. To support our hypothesis, we used stereoenantiomers which are molecules of identical composition but with different three-dimensional structures. Two pairs of R(-) and S(+) agonist
and antagonist enantiomers displaying a high stereoselectivity at native
H3-autoreceptors were studied in various functional tests at native H3Rs
and recombinant isoforms including the non-deleted H3R isoform and a
short isoform common to all species. Our data show strong differences
between the stereoselectivity, i.e. conformations, of the isoforms, and
show that a short isoform plays the role of H3-autoreceptor.
Paper No.: 2988
FOCUSED CONFERENCE GROUP: P19 - GENERAL SESSION ONCOLOGY
CHARACTERIZATION OF A LIPID KINASE INVOLVED IN
THE SYNTHESIS OF ALKYL LYSOPHOSPHATIDIC ACID
Amanda Gellett, Y Kharel, K Lynch
to the glycerol backbone via an acyl, an alkyl, or an alkenyl linkage.
Acyl LPA is the most abundant of the three classes, however, alkyl LPA
has been shown to contribute to the aggressiveness of ovarian cancer and
platelet aggregation. There are multiple potential pathways for alkyl LPA
synthesis. Intracellularly, alkyl LPA could be formed by the esterase,
KIAA1363, deacetylating 2-acetyl MAGE (monoalkylglycerol ether) followed by phosphorylation by a heretofore unidentified lipid kinase. An
alternate intracellular pathway would involve phosphorylation of alkyl acylglycerols by DGK (diacylglycerol kinase), with subsequent cleavage
of the acyl fatty acid by phospholipase A2. The alkyl acylglycerol, 2-acetyl-MAGE, is phosphorylated by human ovarian cancer cell (SKOV-3)
lysates and mouse liver homogenates. Conversely, direct phosphorylation
of MAGE to form alkyl LPA could not be demonstrated in these extracts.
Forced expression of various isoforms of mammalian DGK in HEK293T
cells resulted in increased phosphorylation of 2-acetyl-MAGE in cell
extracts. Thus phosphorylation of 2-acetyl-MAGE by DGK followed by
deacetylation is a potential route for the intracellular synthesis of alkyl
LPA. Sphingosine and analogs such as the immunomodulatory prodrug,
FTY720, increase Type I DGK activity in broken cell assays. Ongoing
studies include determining whether stimulation of whole SKOV-3 cells
by sphingosine analogs increases production of alkyl LPA.
This research was supported by NIH grants R01 GM067958 and T32
GM007055.
Paper No.: 3140
FOCUSED CONFERENCE GROUP: P09 - NFLAMMATION AND
IMMUNOPHARMACOLOGY: NEW TOOLS FOR OLD
DISEASES
DICLOFENAC SODIUM SUPPRESSES RAT CALVARIAL BONE
DEFECT REGENERATION
T Georgopoulou-Karanikola(1), Stergios Tsartsalis(2), H Kalekou(3),
AA Veis(1), B Kokkas(2), AT Tsirlis(1), M Mironidou-Tzouveleki(2)
(1) Aristotle University of Thessaloniki School of Dentistry, Department
of Alveolar Surgery, Implantology and Radiology, Thessaloniki, Greece
(2) Aristotle University of Thessaloniki School of Medicine, A’ Laboratory of Pharmacology, Thessaloniki, Greece
(3) Aristotle University of Thessaloniki School of Dentistry, Department
of Oral Medicine and Oral Pathology, Thessaloniki, Greece
Introduction: Diclofenac sodium is a non-steroidal anti-inflammatory
drug, inhibiting both cyclooxygenase-1 and cyclooxygenase-2, which is
frequently prescribed for analgesic therapy. However, its effect on bone
regeneration has raised doubts about its use after several medical procedures in medicine and dentistry. The purpose of this study was to examine the effect of diclofenac on rat calvarial bone defect regeneration.
Materials: Adult male Wistar rats (250-300g) underwent a surgical procedure in which two artificial defects 6-mm in diameter were made on each
of the parietal bones by means of a specific trephine bar. Rats were
divided in two groups, A and B, of 8 rats in each group. The animals of
group A received daily intramuscular injections of diclofenac sodium
(5mg/kg) for 4 weeks, while the non-treated animals of group B served
as control. Calvarial bone samples from the defect area were decalcified
in EDTA in acid buffer and embedded in paraffin. H+E stained sections
from the sagittal diameter of the defect area were subjected to histological and histomorphometric analysis. Results: At 4 weeks a thin layer of
fibrous connective tissue had formed in the defect area closing the defect.
Partial bone regeneration in the defect area was observed in both groups.
Histomorphometric analysis revealed a lower percentage of new bone in
group A (15,55 ± 10,34) compared to group B (29,34 ± 13,47) (p <
0,05). Conclusion: Our results suggest that diclofenac has an inhibitory
effect on the process of bone regeneration in the rat calvarial defect
model.
University of Virginia, Department of Pharmacology, Charlottesville,
VA, USA
Lysophosphatidic acid (LPA) designates a group of phosphoglycerides
divided into three groups based on whether the carbon chain is connected
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
300
Paper No.: 2438
FOCUSED CONFERENCE GROUP: P05 - TRANSLATIONAL
SCIENCE IN THE METABOLIC SYNDROME: BASIC AND
CLINICAL PHARMACOLOGY
CARBOPLATIN-INDUCED TOXICITY INCREASED TO
GRANULOCYTE-MACROPHAGE PROGENITORS (CFU-GM)
IN OTSUKA-LONG-EVANS-TOKUSHIMA FATTY (OLETF)
RATS
rats increased the latency time during learning, short and long memory
retention tests. In hot-plate analgesic test and in analgesy-meter test and
ketamine-treated rats did not change the latency of reaction compared to
controls. Our results permitted us to suggest that ketamine improved
learning and memory processes and has weak effect on nociception in
doses applied.
References: 1. Riedel G. et al. Behav. Brain Res, 2003, 140 (1-2):1-47.
2. Fürst Z. Neuropsychopharmacol. Hung, 2008, 10:127-30.
Krisztina Géresi(1), K Benkõ(2), B Szabó(3), A Megyeri(1), B Peitl(1),
Z Szilvássy(1), I Benkõ(1)
(1) Medical and Health Science Center, University of Debrecen, Departmant of Pharmacology and Pharmacotherapy, Debrecen, Hungary
(2) Euromedic Diagnostics, Medical and Health Science Center, Univesity of Debrecen, Debrecen, Hungary
(3) Heart Center, Semmelweis University, Budapest, Hungary
OLETF rats have a congenital defect in the expression of CCK-1 receptor gene. Because of the increased food intake a metabolic syndrome has
developed early. In our other experiments we found an impaired granulopoiesis in Zucker obese rats, probably by metabolic changes due to insulin resistance. We study the toxicity of carboplatin, an alkylating-related
cytotoxic drug on granulocyte-macrophage progenitors (CFU-GM) in
OLETF and in their healthy counterparts, LETO rats. At first sight granulopoiesis, as measured by cellularity, the frequency of CFU-GM
and the CFU-GM content of the femoral bone marrow &#8211; did not
differ in obese OLETF and LETO rats. However, testing the vulnerability
of CFU-GM cells cultured them in the presence of carboplatin, we
detected an increased damage in the CFU-GM progenitors obtained from
OLETF compared with LETO. After in vivo administration of a CCK-1
selective antagonist, lorglumide, we could establish a similarly increased
toxicity on the progenitors in LETO rats as in CCK-1 deficient OLETF
rats. Lorglumide might have a direct effect on progenitor cells, because it
could inhibit colony formation of CFU-GM cells of LETO rats also in vitro. As far as we know, this is the first finding concerning CCK-1 receptorial effects on CFU-GM hemopoietic progenitors. Obesity may alter
the function of CFU-GM cells by different mechanisms. As obesity has
become pandemic the higher risk for myelotoxicity of anticancer treatment has a growing importance. The CCK-1 receptor deficiency maybe
also some of the reason of the increased risk of anticancer chemotherapy
in some obese patients.
Paper No.: 1550
FOCUSED CONFERENCE GROUP: P03 - ION CHANNELS IN
ANALGESIA & ANAESTHESIA
EFFECTS OF KETAMINE ON NOCICEPTION, LEARNING
AND MEMORY IN RATS
Damianka Getova, N Doncheva
Medical University Plovdiv, Department of Pharmacology, Sofia, Bulgaria
Ketamine is intravenous anesthetic acting on NMDA-glutamate receptors
(1), which are involved in nociception and memory functions (2). We
study the effects of ketamine using two nociceptive tests and active and
passive avoidance tests. Rats (n = 10) were chronically treated: 1st group
with saline, 2nd, 3rd and 4th groups with ketamine 10, 15 and 20 mg/kg.
Active avoidance test an automatic reflex conditioner was done with
learning and memory tasks and the number of avoidances and escapes
were counted. Step-through and step-down passive avoidance tests were
done with learning, short and long memory retention task. It was measured the latency of reaction. The nociceptive tests were done hot-plate
and alangesy-meter and the reaction time were measured. In active
avoidance test the control rats learned the tasks and increased the number
of avoidances on learning session as well as on memory retention test.
Ketamine in all doses used increased the number of avoidances on learning and on memory retention test. The number of escapes was not changed. In both passive avoidance tests the controls and ketamine-treated
Paper No.: 2495
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
OXALIPLATIN-INDUCED NEUROPATHY: GLIAL
ACTIVATION IN SPINAL CORD AND BRAIN
Carla Ghelardini(1), L Di Cesare Mannelli(1), L Bonaccini(2),
A Pacini(2), A Bartolini(1)
(1) University of Florence, Department of Pharmacology, Florence, Italy
(2) University of Florence, Department of Anatomy, Florence, Italy
Oxaliplatin, unlike other platinum derivatives, does not result in significant renal impairment or ototoxicity, and it has only mild hematological
and gastrointestinal toxicity. On the other hand the limiting side effect is
its neurotoxicity that acts as bases for a neuropathic syndrome. A tangled
panel of symptoms may be disabling for these patients, adversely affecting activities of daily living and thereby quality of life. Chronic pain as
well as the finding of nerve hyperexcitability suggest an involvement of
the central nervous system. In a rat model of painful oxaliplatin-induced
neuropathy (2,4 mgkg-1 intraperitoneally, daily injected for 21 days), we
investigated the profile of glia activation in the spinal cord and in brain
areas involved in pain sensation. In the central nervous system a general
glia activation was provoked by oxaliplatin administration. In particular,
microglia (Iba1 antigen) showed a round-shaped body, indicative of a
differentiation toward a macrophagic phenotype. Moreover, the astrocytes (GFAP antigen) showed an increased density and signs of activation, such as increase in the body size, thickening of the processes and
labelling intensity. These modifications concerned the dorsal horn of the
spinal cord, as evidenced by DAB staining. Astrocytes and microglia
examined by confocal microscopy in different brain districts revealed an
increasing activation in amygdala, raphe nuclei, thalamic nuclei, somatosensorial area 1 and periacqueductal grey matter (PAG). Further understanding of the molecular mechanisms that underlie the effects of glia on
pain processing should lead to the development of new and more efficient approaches for the treatment of pain.
Paper No.: 2948
FOCUSED CONFERENCE GROUP:
P11 - G PROTEIN-COUPLED 7TM RECEPTORS: FROM
MOLECULAR TO PHYSIOLOGICAL FUNCTION
INTERACTIONS OF SDF-1 AND ADP IN PLATELET
ACTIVATION
Olof Gidlöf, C Högberg, B Olde, D Erlinge
Lund University, Department of Molecular Cardiology, Lund, Sweden
Introduction: Apart from playing a crucial role in maintaining hemostasis, platelets are also increasingly recognized as important mediators of
inflammation. Platelets interact with endothelial cells and leukocytes
through an extensive repertoire of inflammatory molecules. Several studies have shown that chemokines such as stromal cell-derived factor one
(SDF-1) are capable of inducing platelet aggregation. The mechanism by
which SDF-1 activates platelets is not yet fully understood and current
reports are partly contradictory. The aim of this study was to elucidate
the signalling pathways involved in SDF-1 mediated platelet activation.
Materials: Whole blood, platelet rich plasma and washed platelets were
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
301
obtained from healthy individuals. Results: Platelet aggregation induced
by 10 nM SDF-1b was reduced by 80% with the addition of 10 lM of
the P2Y12 receptor agonist AZD6140 (p < 0,01). In addition, cAMP levels in PGE1-treated platelets were decreased by 14% when adding 10
nM SDF-1b (p < 0,05). 5 lM of ADP caused a 12% upregulation of the
SDF-1 receptor CXCR4 on platelets (p < 0,05). Conclusion: We show
that SDF-1b mediated platelet activation is dependent on P2Y12 receptor
signalling. Treating platelets with the chemokine results in a decrease in
cAMP levels, confirming that activation requires Gai-coupled signalling.
We also demonstrate that stimulating platelets with ADP results in an
increased surface expression of the SDF-1 receptor, CXCR4. Taken
together, these data suggest possible signalling crosstalk between SDF-1
and ADP in platelet activation.
Paper No.: 1602
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
PHARMACOLOGICAL BASIS FOR THE MEDICINAL USE OF
PSYLLIUM HUSK IN CONSTIPATION AND DIARRHEA
Anwar-ul-Hassan Gilani(1), MH Mehmood(1,2)
(1) Aga Khan University, Department of Biological and Biomedical
Sciences, Karachi, Sind, Pakistan
(2) University of Karachi, Sind, Pakistan
The psyllium husk was studied to rationalize its medicinal use in gut
motility disorders using both the in-vivo and in-vitro assays. The aqueous
methanolic extract (Pop.Cr) caused laxative effect in mice at 100 and 300
mg/kg. The laxative effect was partially sensitive to atropine or
SB203186 (5-HT4 receptor antagonist). At higher doses (500 and 1000
mg/kg), it exhibited antisecretory and antidiarrheal activities. The antidiarrheal effect of Pop.Cr was increased when repeated in mice pretreated
with L-NAME (nitric oxide synthase inhibitor). When tested in guinea
pig-ileum, Pop.Cr caused a concentration-dependent stimulant effect at 110 mg/mL, which was partly inhibited in the presence of atropine or/
SB203186, suggesting partially cholinergic and 5-HT4 receptor-mediated
effect. In spontaneously contracting rabbit jejunum, Pop.Cr caused partially atropine-sensitive stimulatory effect followed by slight relaxation at
the highest tested concentration. The stimulatory effect was increased
when repeated in the presence of L-NAME or methylene blue and
Pop.Cr relaxed the K+ (80 mM)-induced contraction, indicating the
involvement of Ca++ channels and NO pathway. The relaxation was
potentiated with atropine pretreatment while inhibited in tissue pretreated
with L-NAME or methylene blue. This study indicates that psyllium
husk possesses a gut stimulant effect mediated through multiple pathways, which might be complementing the laxative effect attributed to its
fiber contents. Moreover, the presence of gut relaxant components
explains its medicinal use in diarrhea and perhaps also meant by nature
to offset the excessive stimulant effect which could be harmful otherwise.
Paper No.: 1771
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
ANTIDEPRESSANT ACTIVITY OF ORCHIS MASCULA
Anwar-ul Hassan Gilani(1), S Khan(1), HS Siddiqi(1,2)
(1) Aga Khan University, Department of Biological and Biomedical
Sciences, Karachi, Pakistan
(2) University of Karachi, Karachi, Pakistan
Depression is most popular among the mental disorders and is one of the
leading causes of disability and morbidity around the world. Herbal medicines have been used by every human society to cure or treat variety of
health problems. Orchis mascula (O. mascula) has been used in folk
medicine for various aliments including neurological disorders. It is also
used as tonic for nervous disability and reported to be nutritious for
neurasthenia. In the present study O. mascula root methanolic extract
was evaluated for its antidepressant properties using forced-swim test
(FST), tail suspension test (TST), yohimbine potentiation test (YPT) and
locomotor test (LMT). Phenelzine, imipramine and fluoxetine were used
as a reference drugs. Our results showed that O. mascula extract (10 100 mg/kg) caused dose-dependent reduction in the immobility time of
rodents using FST (EC50 = 47 ± 5.1 mg/kg rats) and TST (EC50 = 31 ±
3.7 mg/kg mice) indicating antidepressant property of extract. Data also
showed that the pretreatment of animals with extract exhibited marked
reduction in the immobility time which does not correlate with the spontaneous activity recorded by LMT e.g. at the dose of 30 mg/kg 51%
decrease (control counts: 7206 ± 534 and test counts: 3508 ± 421) was
found in the locomotor counts of mice, thus further confirming the present findings. In conclusion, the O. mascula root extract possesses antidepressant activity. However, further studies are required to investigate the
active principle responsible for the antidepressant property using bioassay directed fractionation method.
Paper No.: 1772
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
PHARMACOLOGICAL USE OF BORAGO OFFICINALIS
Anwar-ul-Hassan Gilani
Aga Khan University, Department of Biological and Biomedical
Sciences, Karachi, Pakistan
Borago officinalis Linn. (family Boraginaceae), commonly known as
Borage, originally indigenous to Mediterranean region and now is found
all over the world. Different parts of the plants particularly the seed oil is
used for medicinal purpose. Borage is popular for its wide medicinal use,
such as, gastrointestinal (colic, cramps, diarrhea), airways (asthma, bronchitis), cardiovascular, (cardiotonic, antihypertensive and blood purifier),
urinary (diuretic and kidney/bladder disorders), anti-dyslipidemic and
other metabolic and sexual disorders. Moreover, it is used as nerve tonic,
demulcent, while its oil is used as anti-arthritic, anti-inflammatory and
anti-neurodermatitis. Borage contains multiple chemicals, such as coumarins, tannins, flavonoids, polyphenolics including vanillic acid, p-coumaric acid phenolic acid, caffeic acid, rosmarinic acid, chlorogenic acid and
scopoletin, alkaloids including amabiline, lycopsamine and supinidine,
hydrocarbons mainly tetracosane and heptacosane, alcohols including
cis-3-hexenol and hexanol and predominant fatty acids including a-linolenic, stearidonic, c-linolenic, palmitic and linoleic acids other than niacin, thiamine, riboflavin, ascorbic acid, salicylic acid. It is reported to
possess antioxidant, renal protective and cytotoxic. We observed that its
antispasmodic, bronchodilator and antihypertensive activities are mediated possibly due to calcium antagonist constituents. Borage oil, the main
source of c-linolenic possesses antihypertensive, anti-dyslipidemic and
immunomodulator activities by raising the level of 6-omega polyunsaturated fatty acids in the various tissues. Thus, Borage has a lot of therapeutic potential particularly in inflammatory and metabolic disorders.
Borage is generally considered safe with minor side-effects except in
pregnancy; however, seed oil also contains pyrrolizidine alkaloids, thus
requiring further studies to establish the safety of Borage oil.
Paper No.: 3288
FOCUSED CONFERENCE GROUP:
P11 - G PROTEIN-COUPLED 7TM RECEPTORS: FROM
MOLECULAR TO PHYSIOLOGICAL FUNCTION
FROM PHYLOGENY TO PHARMACOLOGY;
CHEMOGENOMIC PHARMACOLOGICAL GPCR
CLUSTERING AND LIGAND-INFERENCE
David Gloriam(1), S Garland(2)
(1) University of Copenhagen, Department of Medicinal Chemistry,
Copenhagen, Denmark
(2) GlaxoSmithKline Pharmaceuticals PLC, Stevenage, UK
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
302
Conventional phylogenetic analysis utilises the whole receptor sequence.
This has limitations; however, from a ligand design perspective because
the majority of residues are not involved in ligand binding and thus evolutionary patterns dominate over ligand-binding properties. Clustering on
only the binding residues generate a pharmacological clustering that
resembles more closely experience from cross-screening of ligands. We
defined Family A ligand accessible residues (TM bundle inward-facing)
from all receptor with experimentally determined structures and compared with datasets extracted from literature[1]. Specific motifs of ligandbinding residues (‘‘interaction fingerprints’’) were identified and linked to
privileged structures. By comparing such motifs data could be inferred
and used in lead generation by cross-target ligand inference[2].
References: 1. Gloriam, D.E. et al.; Definition of the G protein-coupled
receptor...; PM:19537715 2. Gloriam DE, Garland SL.; Chemogenomic
analysis can rationalise and predict the binding profile of privileged
structures at GPCRs; In preparation
Paper No.: 1303
FOCUSED CONFERENCE GROUP: P13 - MAXIMISING
BENEFITS AND MINIMIZING HARMS FROM DRUGS
A RANDOMIZED CLINICAL TRIAL UTILIZING THE DRUG
BURDEN INDEX TO REDUCE EXPOSURE TO
ANTICHOLINERGIC AND SEDATIVE MEDICATIONS IN
OLDER PEOPLE
D Gnjidic(1), S Hilmer(1), D Le Couteur(1,2), D Abernethy(3)
(1) University of Sydney, and Royal North Shore Hospital, Royal North
Shore Hospital, Departments of Clinical Pharmacology and Aged Care,
Sydney, NSW,Australia
(2) Concord Hospital, Centre for Research and Education on Ageing,
Sydney, NSW, Australia
(3) Food and Drug Administration, Silver Spring, Maryland, USA
The DBI is a pharmacological model that measures individual’s exposure
to anticholinergic and sedative drugs. Higher DBI has been associated
with functional impairment in observational studies of older people
(Hilmer SN et al, Arch Intern Med 2007; 167: 781-7; Cao YJ et al, Clin
Pharmacol Ther 2008; 83:422-9; Gnjidic D et al, Br J Clin Pharmacol
2009; 68:97-105). The aim of this study was to assess the impact of providing information about DBI to General Practitioners (GPs) on prescribing for older people. Participants were people aged ‡70 years, living in
self-care retirement villages in Sydney, Australia. The intervention
involved a letter and phone call to GPs, using DBI to prompt them to
consider cessation or dose reduction of anticholinergic and sedative medications. A total of 115 participants were enrolled. At baseline, 19 out of
57 participants in the intervention and 31 out of 58 participants in the
control group had a DBI >0 (p < 0.05). At follow up, DBI change was
observed in 16 participants. DBI decreased in 12 participants, six in
intervention (32%), and six in control group (19%). GPs identified the
following barriers to reducing anticholinergic and sedative drugs: uncomfortable altering prescriptions initiated by specialists; unable to influence
patient’s attitudes; unaware of patient’s medications and strong clinical
indication. The intervention targeting GPs’ prescribing practices was less
effective than anticipated in reducing anticholinergic and sedative drugs,
and barriers were identified. Future studies should trial multidisciplinary
interventions, engaging patients, specialists, GPs and pharmacists.
Paper No.: 782
FOCUSED CONFERENCE GROUP: PW12 - EXPERIMENTAL
PAIN METHODS IN DEVELOPMENT AND VALIDATION OF
ANALGESICS
EVATUATION OF THE EFFECTS INDUCED BY NICORANDIL
AND ITS IMMEDIATE PRECURSOR IN THE MODELS OF
FORMALDEHYDE-INDUCED NOCICEPTIVE RESPONSE AND
CARRAGEENAN-INDUCED EDEMA IN MICE
Federal University of MInas Gerais, Department of Pharmacology &
Pharmaceutic Products, Belo Horizonte, Brazil
NO-releasing compounds have been used as therapeutic agents
(MILLER, M.R. et al.; Br. J. Pharmacol.; 2007; v. 151: p. 305). Nicorandil [N-(2-nitroxyethyl) nicotinamide] has both the ability to release NO
and open ATP-dependent K+ channels. The lack of information about the
antinociceptive properties of nicorandil prompted us to synthesize it and
its immediate precursor [N-(2-hidroxyethyl) nicotinamide] and investigate their effects in models of nociceptive and inflammatory pain as well
as edema. Nicorandil (50, 100 or 150 mg/kg) or its precursor (50, 100,
150, 250, 500 or 1000 mg/kg) were administered per os 1 h prior the
injection of formalin, or the induction of paw edema with carrageenan,
in female Swiss mice. The motor activity was investigated using rotarod. The results were analyzed by one-way Anova followed by Newman-Keuls test (p < 0.05). Nicorandil (50, 100 or 150 mg/kg) inhibited
both phases of the formalin model. Only the highest dose (1000 mg/kg)
of its precursor inhibited the second phase of this response. Neither nicorandil nor its precursor inhibited carrageenan-induced edema. The motor
activity of mice was not impaired by the compounds. Nicorandil, but not
its immediate precursor, presented antinociceptive activity that was not
associated to impairment of motor activity. None of them exhibited antiinflammatory activity, suggesting that the antinociceptive activity may be
more related to a central action. These results demonstrate the role of nicotinamide derivatives or NO-releasing compounds as pain modulators.
The role of NO in the inflammatory processes is still under debate (WEI,
XM, Int. Immunopharmacol.; 2003; v. 3: p. 1581).
Paper No.: 2996
FOCUSED CONFERENCE GROUP:
P11 - G PROTEIN-COUPLED 7TM RECEPTORS: FROM
MOLECULAR TO PHYSIOLOGICAL FUNCTION
AUTOCRINE EFFECTS OF EXTRACELLULAR CYCLIC AMP
IN SKELETAL MUSCLE
Rosely Godinho, E Aparecida-Santos, AL Andrade-Lopes, T Chiavegatti
University Federal of São Paulo, Department of Pharmacology, São
Paulo, Brazil
Cyclic AMP (cAMP)-mediated pathway regulates several skeletal muscle
processes such as muscle contraction and expression of synaptic proteins.
We have showed that AC activation mediated by GsPCR leads to extracellular accumulation and subsequent conversion of cAMP into adenosine. (Chiavegatti et al., Br J Pharmacol, 2008; 53:1331-1340). In the
present study, we evaluate possible existence of this pathway in vivo and
the autocrine action of extracellular cAMP on skeletal muscle. Firstly,
intra (cAMPi) and extracellular cAMP (cAMPe) were analyzed in rat
EDL muscles. 30-min treatment of muscle with forskolin or ß-adrenoceptor agonist isoproterenol increased by 4-13 fold the basal cAMPi and by
10-36 fold the cAMPe. The inhibitor of organic anion transporters probenecid (100 lM) attenuated by 60% the cAMPe rise, without interference in cAMP synthesis. In vivo, 2h-treatment of adult rats with ß2adrenoceptor agonist fenoterol (3-10 mg/kg) increased by 84% plasma
cAMP content, an effect abolished by pre-treatment with probenecid.
Autocrine actions of cAMP were studied in L6 skeletal muscle cultures.
The membrane impermeable cAMP (1 or 10 lM) increased by 56% and
215% the cAMPi, which was inhibited by non-selective adenosine receptor CGS 14953. In summary, our results demonstrate the cAMP efflux
through organic anion transporters in rat skeletal muscle, which might
contribute to the increment in plasma cAMP induced by fenoterol. More
importantly, the cAMPe is able to regulate cellular function in an autocrine fashion via its metabolite adenosine, activating receptors coupled to
Gas, which provides a positive feedback response to intracellular cAMP
signalling.
Adriana Godin, W Ferreira, F Oliveira, L Rocha, R Vieira,
E Nascimento Jr, J Seniuk, A de Fátima, M Coelho
Journal compilation 2010 Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 107 (Suppl. 1), 162–692
303
Paper No.: 1177
FOCUSED CONFERENCE GROUP:
P12 - ION CHANNELOPATHIES: NEW WINDOWS ON
COMPLEX DISEASE AND THERAPY
THE ANTIVASOCONSTRICTOR EFFECT OF POTASSIUM
CHANNEL OPENER P1075 IN THE HUMAN VASCULAR
GRAFTS
Ljiljana Gojkovic-Bukarica(1), A Novakovic(2), B Beleslin-Cokic(1),
M Peric(1), J Markovic-Lipkovski(1), S Cirovic(1), V Kanjuh(3)
(1) Belgrade University Faculty of Medicine, Department of Pharmacology, Belgrade, Republic of Serbia
(2) Belgrade University Faculty of Pharmacy, Belgrade, Republic of
Serbia
(3) Republic of Serbian Academies of Sciences and Arts, Belgrade,
Republic of Serbia
The human internal mammary artery (HIMA) and saphenous vein (HSV)
are used in coronary artery bypass grafting, but spasm of grafts may
occur. In order to find agent that can prevent spasm, the aim of our study
was to evaluate the antivasoconstrictor effect of K-channel opener,
P1075 (N-cyano-N’-(1,1-dimethylpropyl)-N’’-3-pyridylguanidine), on the
contractions of HIMA and HSV evoked by exogenously applied noradrenaline (NA) or by NA realized during electric field stimulation (EFS,
20 Hz, adrenergic origin). P1075 induced a concentration-dependent
inhibition of both EFS-contractions, and contractions evoked by exogenous NA of the HSV (pEC50 values of 7.06 and 6.70, P > 0.05) and of
the HIMA (6.83 and 6.02, P < 0.05). Glibenclamide (1 microM), a selective blocker of ATP-sensitive K (KATP)-channels completely antagonized
the effect of P1075 on the EFS-contractions of HIMA and HSV and contractions evoked by exogenous NA on the HSV. However, 10 microM of
glibenclamide had to be used for antagonism of P1075 effect on exogenous NA contractions in HIMA. Immunomorphological study confirmed
presence of different subtypes of KATP channels (Kir6.1 and Kir6.2) in
the smooth muscle of HIMA and Kir6.2 in HSV. Thus, P1075 exhibits
potent inhibitory effect on NA-evoked contractions of the HIMA and
HSV. It seems that in the antivasoconstrictor effect of P1075 in the
HIMA and HSV are involved different subtypes of KATP channels.
Paper No.: 2308
FOCUSED CONFERENCE GROUP: P16 - NATURAL
PRODUCTS: PAST AND FUTURE?
THE EFFECTS OF LAVANDULA STOECHAS EXTRACT ON
CHOLESTEROL LEVELS AND CARDIAC FUNCTIONS ON
ISCHEMIA-REPERFUSION MODEL IN ISOLATED RAT
HEARTS
Sule Gok, E Olmez, A Sonmez, K Vural, E Darýverenli, M Soylar
Celal Bayar University, School of Medicine, Department of Pharmacology, Manisa,Turkey
Introduction: Lavandula stoechas (LS) has been commonly used as cholesterol reducing and vessel dilating in traditional medicine in Turkey. In
this study, we investigated the effects of LS extract on serum lipid profile
and myocardial ischemia-reperfusion injury of isolated rat hearts with
cholesterol-fed. Materials/Animals: Rats were fed with 2% cholesterolenriched or standard chow for 8 weeks. In treated groups, the extract of
LS (100 and 500 mg/kg/day, p.o.) or atorvastatin (20 mg/kg/day, p.o), a
HMG CoA reductase inhibitor, were also administered. At the end of 8
weeks, the hearts of anaesthetized rats were removed and perfused by
Langendorff system. After 30 min, the hearts were subjected to 30 min
global ischemia followed by 30 min reperfusion. Lipid profiles of rat serums were also analyzed. Results: Total and LDL-cholesterol levels of
cholesterol-fed rats were significantly higher than all other groups. Treatment with LS or atorvastatin significantly reduced cholesterol. In hearts,
after ischemia, left ventricular developed pressure abolished and did not
recover by reperfusion in all groups. Left ventricular end diastolic pres-
sure significantly elevated in ischemia. This increase was more in reperfusion in all groups, and significanty greater in the higher dose of LS
and atorvastatin treated rats with cholesterol-fed. Perfusion pressure was
significantly high in reperfusion compared to the preischemic period in
all groups. Ventricular tachycardia and ventricular fibrillation were not
prevented by LS and atorvastatin. Conclusion: These results suggest that
LS and atorvastatin reduces cholesterol levels in rats, but not provide
recovery on cardiac functions and arrhythmias in this model.
This Project was supported by TUBITAK (No:108S082)
Paper No.: 768
FOCUSED CONFERENCE GROUP: P15 - ENDOTHELIUM IN
HEALTH AND DISEASE
ABCB1 C3435T GENETIC POLYMORPHISM ALTERS
CLINICAL RESPONSE TO LOSARTAN IN PATIENTS WITH
HYPERTENSION
MT Goktas(1), F Pepedil(2), G Sain-Guven(2), B Cakir(3), Melih
O Babaoglu(1), U Yasar(1), A Bozkurt(1)
(1) Hacettepe University, Faculty of Medicine, Department of Pharmacology, Ankara, Turkey
(2) Hacettepe University,Faculty of Medicine, Department of Internal
Medicine, Ankara, Turkey
(3) Hacettepe University, Faculty of Medicine, Department of Public
Health, Ankara, Turkey
Losartan is a selective angiotensin II receptor antagonist used in hypertension treatment. Losartan has been shown to be a substrate for the
drug-efflux transporter ATP-binding cassette sub-family B member 1
(ABCB1, MDR1). ABCB1 is a genetically polymorphic transporter. The
ABCB1 3435T allele has been shown to be associated with altered transporter functions, leading to change in systemic bioavailability and clinical efficacy of its substrates. The aim of this study was to investigate the
effects of the ABCB1 C3435T genetic polymorphism on clinical efficacy
of losartan in hypertensive patients. Patients (n = 74) diagnosed to have
mild (Grade I) hypertension were included in the study. Genomic DNA
was extracted from peripheral blood lymphocytes. Genotyping was performed using polymerase chain reaction and endonuclease digestion
methods. Blood pressure measurements were recorded before and 6
weeks after losartan (100 mg/day) administration. Blood pressure
changes in patients with wild-type genotype as compared to those with
variant genotypes were analysed by using Student’s t test. The average
(mean ± SD) decrease in systolic blood pressure in patients carrying the
T allele (11.6 ± 9.7 mmHg, n = 55) was significantly higher when compared to the patients with homozygous wild-type (CC, 7.6 ± 9.4 mmHg,
n = 19) genotype (P = 0.03). These results suggest for the first time in literature that MDR1 C3435T genetic polymorphism may play an important role in clinical response to losartan in hypertension.
(The study was supported by Hacettepe University Research Centre
(0801101009) and Turkish Academy of Sciences, Young Scientist Award
MOB: TUBA-GEBIP/ 2007-06)
Paper No.: 843
FOCUSED CONFERENCE GROUP: P17 - NEW APPROACHES
AND TARGETS IN PSYCHIATRY
THE INVOLVMENT OF ERK-DEPENDENT PR