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SERUM OXIDATIVE STRESS PARAMETERS
AND HOMOCYSTEINE LEVELS IN PATIENTS
WITH RENAL TRANSPLANT
ANGELA ANTONESCU1, MARIANA MUREŞAN1*, OTILIA
MICLE1, LIANA MICLE1, LUCIANA DOBJANSCHI1, LAURA
VICAŞ1, M. DOROFTEIU2
1
Faculty of Medicine and Pharmacy Oradea
University of Medicine and Pharmacy „Iuliu Haţieganu” Cluj Napoca,
Faculty of Pharmacy, no. 6 Louis Pasteur st.
*corresponding author: marianamur2002@yahoo.com
2
Abstract
The aim of our study was to determine the level of homocysteine, oxidativ stress
markers and the lipidic profile in pacients with renal transplant.
The researches included 25 patients with renal transplant followed up four
months (from March until June 2007). It was evaluated the levels of lipid peroxides (a
colorimetric method with thiobarbituric acid) and ceruloplasmin (Ravin method). The
concentration of homocysteine was assessed using an enzymatic method.
We also measured the concentration of cholesterol and tryglicerides. All the
results were compared with a control group.
The patients presented an oxidative stress reflected by a high level of
malondialdehide (MDA) compared to the control group. The concentration of
ceruloplasmin was reduced. They also showed a hiperhomocysteinemia. Cholesterol level
was in a normal range, and the triglycerides increased.
Rezumat
Scopul lucrării a fost acela de a doza homocisteina, unii markeri ai stresului
oxidativ precum şi evaluarea profilului lipidic şi a implicaţiilor lor în procesul aterosclerotic
la pacienţii cu transplant renal.
Cercetările noastre au inclus 25 de pacienţi cu transplant renal pe o perioadă de
patru luni (din martie până în iunie 2007). A fost evaluat nivelul de peroxizi lipidici
(metoda colorimetrică cu acid tiobarbituric) şi ceruloplasmina (metoda Ravin). Pentru
punerea în evidenţă a homocisteinei am utilizat metoda enzimatică.
De asemenea s-au determinat concentraţiile de colesterol şi trigliceride.
Toate rezultatele au fost comparate cu un lot martor. Pacienţii transplantaţi sunt
supuşi unei agresiuni oxidative reflectate printr-un nivel crescut de malonildialdehidă
(MDA) comparativ cu lotul martor şi concentraţie scăzută a ceruloplasminei. Lotul studiat a
prezentat valori ridicate ale homocisteinei. Colesterolul s-a încadrat în limite normale, în
schimb s-a înregistrat o hipetrigliceridemie.



MDA
ceruloplasmin
homocysteine


renal transplant
atherosclerosis
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INTRODUCTION
Epidemiological studies have shown that high levels of
homocysteine in the blood are related to a higher risk of coronary heart
disease, stroke, osteoporosis and peripherial vascular disease. In kidney
transplant recipients, total homocysteine has been shown to be associated
with atherosclerotic events [5]. Homocysteine levels may increase in a
number of ways, which are correlated to both folic acid and vitamin B 12
metabolism [12]. This substance is a sulfur aminoacid formed during the
conversion of methionine to cysteine. Homocysteine is either metabolized to
cysteine via the transsulfuration pathway or remethylated to methionine via
the remethylation pathway [13].
In our study we have examined the level of reactive oxygen species
(ROS) and homocysteine in patients with renal transplant and their
implication in the atherosclerotic process.
MATERIALS AND METHODS
A group of 25 patients with renal transplant were studied between
May and June 2007. In order to establish the metabolic profile, we have
determined the level of cholesterol and triglycerides. The amount of ROS
was established by evaluating malondialdehyde (MDA) serum levels with
thiobarbituric acid (TBA), a colorimetric method.
The antioxidant resistance of the organism was evaluated by
measuring the plasmatic ceruloplasmin levels using Ravin method. Serum
homocysteine concentration was estimated using an enzymatic method
(total serum homocysteine was measured using the enzymatic homocysteine
assay, cat no. FHER100, on Hitachi 912 instrument).
All the results were compared to the control group.
RESULTS AND DISCUSSION
The patients with renal transplant have an increased level of MDA
4.34 ± 1.06 nmoles/ ml in comparison to the control group in which the
concentration is 1.91 ± 0.33 nmoles/ml (p < 0,001) (fig. 1).
The concentration of ceruloplasmin in the serum of these patients is
low, comparing to the control group (32.11 ± 1.62 mg % in the control
group vs. 26.22 ± 3.45 mg% in the patients with renal transplant.) (p> 0,1)
(fig. 2). The level of serum homocysteine increased from 7.97 ± 1.57
µmoles/l in the reference group, to 19,8 ± 5,62 µmoles/l in patients with
renal transplant.
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MDA
nmol/ml
p<0.001
reference
patients with
renal transplant
4.34 + 1.06
1.91 + 0.33
Figure 1
The values of serum malondialdehyde in patients with
renal transplant in comparison to the control group
Ceruloplasmin
mg %
p>0.1
reference
patients with
renal transplant
32.11 + 1.62
26.22 +3.45
Figure 2
The values of ceruloplasmin in the patients
with renal transplant in comparison to the control group
Homocysteine
mol/l
p<0.05
reference
patients with
renal transplant
19.8 + 5.62
7.97 + 1.57
Figure 3
The values of homocysteine in patient
with renal transplant in comparison to the control group
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We also assessed the levels of serum cholesterol, triglycerides
which are considered cardiovascular disease risk factors (table I).
Table I
Serum values of cholesterol and triglycerides in patients with renal transplant
Parametres
Normal value
March
April
May
June
Cholesterol
(mg%)
Triglyceride
(mg%)
200-239 mg%
185±36.99
159±46.48
152.85±43.62
260.27±36.35
150-199 mg%
160.57±86.92
184.3±74.99
260.89±96.44
223.05±97.14
Our results demonstrated that in the third month of the study,
cholesterol level increased, whereas triglycerides were high every month of
the study.
Hyperlipemia is a common and an important risk factor after renal
transplant. A retrospective study during the first 3 years after the kidney
transplant showed that the patients had an increased level of cholesterol,
LDL and HDL and a decreased level of triglycerides [8].
The renal transplantation of our patients was performed 2-5 years
ago, and received without interruption immunosupressant drugs. In 2005,
Marubayashi et.al. demonstrated in a study that the patients with kidney
transplant which were treated with immunosupressant drugs had a
significant increased level of cholesterol, and tryglicerides [9].
Our results are similar to other articles. Some studies suggest that
hiperhomocysteinemia may be a cardiovascular risk factor in renal
transplant [10, 6, 5]. The three main metabolic pathways of homocysteine
are: transmethylation pathway – the production of homocysteine from
methionine; transsulfuration, breakdown of homocysteine and production of
cysteine and taurine; remethylation of homocysteine to methionine
including the folate cycle, which acts as the methyl donor and provides the
carboxyl group necessary to convert homocysteine into methionine. An
alternative remethylation pathway in the liver and kidney uses betaine
instead of folate [7, 14]. Compound of metabolised homocysteine is
hydrogen sulfide. This is derived from homocysteine transformed in Lcysteine under the influence of two enzymes: cistationine β sintetaze and
cistationine γ-sintetaze. The function of H2S is similar but not identical to
the NO and CO. It is considered like gasotransmitter [2].
The patients with renal transplant in our research had a high level
of homocysteine. A high level of homocysteine contributes to the
progression of renal transplant disfunction [15]. Treatment with folic acid
and other B vitamines (B12; B6) proved to be effective in lowering or even
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normalizing the homocysteine plasma level, an easy and inexpensive way to
reduce the rate of renal injuries in these patients [15]. Homocysteine may
damage cells directly, by promoting oxidative stress.
We found an increased concentration of MDA (p< 0,001) and a low
level of ceruloplasmin (p>0.1) in the patients with renal transplant.
Hiperhomocysteinemia may cause endothelial disfunction. One of
the most important endothelial cell products is nitric oxid (NO), which is
synthetized from L-arginine by the enzyme endothelial nitric oxide synthase
(eNOS) [11]. NO may modulate homocysteine concentration directly by
inhibiting methionine synthase, the enzyme that synthesizes methionine
from homocysteine and 5-methytetrahydrofolate [3]. eNOS inhibition can
accelerate atherosclerosis in animal models and abnormalities of the
endothelial NO pathway [1].
Free radicals enhance lipid peroxidation and oxidized LDL
represents a source of further damage to endothelial cells. Therefore we
believe that hiperhomocysteinemia may induce an oxidative stress.
Increased concentration of MDA has been described in individuals
following a methionine load, which can lead to hiperhomocysteinemia [4].
Our results demonstrated the presence of an oxidative stress in the
patients with renal transplant. The concentration of ceruloplasmin was low,
which proves a reduced protective reaction against the oxidative stress.
CONCLUSIONS





The level of MDA in patients with renal transplant is increased in
comparison with the control group and the concentration of
ceruloplasmin is reduced which means that an oxidative stress is
present
The patients with renal transplant developed hiperhomocysteinemia
We suggest an antioxidant treatment with folic acid and vitamin B6,
to reduce the level of homocysteine acid and also to suppress the
progresion of homocysteine - induced atherosclerosis
Renal transplant is associated with a high concentration of
tryglicerides and a normal range of cholesterol
It is also important to control the lipidic profile in order to prevent
the post transplant hiperlipidemia and atherosclerosis.
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