Caffeic Acid and C.
elegans’ Protection
Against Toxin Induced
Neurodegeneration
By: Michelle Chyn
Need

Degree of PD in Patients
Average person
DM
US $
pounds
3,920
2,590
1,690

Fluctuations in
performance
6,010
3,970
2,580
More stable condition
2,700
1,780
1,160
Hoehn and Yahr scale
stage I
2,230
1,470
9,60
Hoehn and Yahr scale
stage V
11,870
7,830
5,100
Parkinson’s Disease
(PD)- 500,000 people
in U.S.
$6,000,000,000 spent
annually for PD
treatment
National Institute of Neurological
Disorders and Stroke
Figure 1: Cost of PD treatment per person annually
Dodel RC, Pepperl S, Köhne-Volland R, Szucs T, Werhahn KJ, Noachtar S, Oertel WH. “Costs of drug treatment of neurologic diseases: Parkinson disease,
dystonia, epilepsy.” European Journal of Neurology. Vol. 7. Pp. 479-85 1991.
Need Cont.

Coffee consumption in North America and Europe = 1/3 of the tap water intake
http://www.e-importz.com/images/coffee_consumption.gif
Figure 3: Coffee Consumption
http://www.parkinsonsdecisionaid.eu.com/images/ac/2008/incidence.gif
Figure 2: PD in European Countries
Knowledge Base
Figure 4:
http://en.wikipedia.org/wiki/Basal_ganglia
 C.
elegans biosynthesize dopamine in the corpus
stratum (basil ganglia) (Braugart, 2004)
Knowledge Base Cont.

Figure 5: Nerve Synapse
http://upload.wikimedia.org/wikipedia/en/a/a6/Chemical_synapse_schema.jpg
Copper is a neurotoxin
when consumed in
excess amounts
(Gaggelli et al., 2006)
Literature Review


Antioxidants in coffee decreases risk of development of agerelated diseases (Eskelinen et. al, 2009)
Caffeine intake doesn’t show health benefits (Lopez-Garcia,
et. al, 2008)
Figure 6: Antioxidant
http://pubchem.ncbi.nlm.nih.gov/summary/summary.cgi?cid=689043&loc=ec_rcs
Literature Review Cont.

Caenorhabditis elegans MPP+ model used as PD
model for testing anti-PD drugs (Braungart, et al.
2004)
Figure 7: Doseresponse effect of
MPP+ treatment
on the mobility of
wild-type C.
elegans
animals.
http://content.karger.com/ProdukteDB/produkte.asp?Aktion=ShowPDF&ArtikelNr=80983&Ausgabe=230458&ProduktNr=229093&filename=80983.pdf
Literature Review Cont.

Green Florescent Protein (GFP) florescence
shown in dopaminergic neurons of transgenic
strains of C. elegans (Braungart, et al., 2004)
http://content.karger.com/ProdukteDB/produkte.asp?Aktion=ShowPDF&ArtikelNr=80983&Ausgabe=230458&ProduktNr=229093&filename=80983.pdf
Figure 8: GFP expression in dopaminergic neurons of untreated
and MPP+ treated worms
Literature Review Cont.

Direct addition of dopamine-2-c did not help
increase noradrenaline presence in PD patients
(Goodall and Alton, 1969)
Figure 9: % noradrenaline
recovery after dopamine2-c infusion
http://www.pubmedcentral.nih.gov/picrender.fcgi?artid=297487&blobtype=pdf
Purpose

Observing the effect of caffeic acid on neurotoxininduced degeneration in C. elegans
Hypothesis
H0- caffeic acid will not affect the degeneration
rate in the C. elegans
 HA- caffeic acid will decrease degeneration in the
C. elegans.

Methodology
Compounds in Coffee and C. elegans Protection Against Parkinson’s Disease
Caenorhabditis elegans egIs1[Pdat-1::GFP] strain from
Caenorhabditis Genetics Center (CGC)
N=90
Group 2
Control
addition of copper oxide
addition of copper oxide,
concentratin according to LD 50
caffeic acid- lower concentration
according to LD 50
N=30
N=30
Group 3
addition of copper oxide
caffeic acid- higher concentration
according to LD 50
N=30
C. elegans cultured at 20°C in NGM plates in normal oxygen environment.
Caffeic acid and MMP+ concentrations inserted in food of E. coli for 3 days at
L1 stage. Organisms then moved out into normal NGM plates.
Data collected through stereo florescent microscope viewing of GFP
florescence on a scale of normal, weak, and none, and area through
Adobe Photoshop on the 3rd day.
Statistical analysis using SPSS and ANOVA
Protocols
Figure 10: Culturing
of C. elegans on a
petri dish
Picture drawn by Victoria Wei

Eggs laid overnight at L4 stage on fresh NGM
plates (Mehta, et al, 2009)
Protocols Cont.
N=30 per group, total
N=90
 Group 1: Copper oxide
 Group 2: Copper oxide
and Caffeic acid
 Group 3: Copper oxide
and Caffeic acid

http://www.scq.ubc.ca/wp-content/uploads/2006/07/wormcycle.gif
Figure 11: C. elegans growth stages
Protocols Cont.
Data collect on scaled brightness of GFP
expression (Braungart, et al., 2004) and area of
florescence on Photoshop
 Data analysis using SPSS

Protocols
Do-Ability

Equipment available:
GFP filter and florescent microscope
 UV lamp
 Petri dishes
 Photoshop


Equipment needed:
C. elegans pdat-1::GFP strain from CGC
 E. coli OP50 strain from CGC
 Copper, sodium azide, and caffeic acid from Sigma
 NGM from Carolina Biological

Budget
Vendor
Cat #
Pg.
Item
Qty.
Description
Caenorhabditis
Genetics Center BZ555
C. elegans pdat1::gfp
Sigma
Caffeic Acid
15 g
Nematode Growth
Agar
2 135 mL
C0625
Carolina
Biological
173520
Caenorhabditis
Genetics Center
741270
S2002
Carolina
Biological
Sigma
Sigma
Total $
110
$7.00
$7.00
$53.30
$53.30
$6.25
$12.50
$7.00
$7.00
$6.45
$64.50
E. coli OP50
Carolina
Biological
Sigma
Unit $
216880
F0503
203122
petri dishes
10 100 x 15 mm
Sodium azide
1 25 g
$21.20
$21.20
ampicillin
15 g
$43.25
$43.25
fluorodeoxyurodine
1 100 mg
$117.00
$117.00
copper
1 10 g
$146.50
$146.50
Lab
Fluorescent
Microscope
Lab
GFP filter
$3,760.00
$938.38
Total Cost
$5,052.43
Bibliography
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