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Caffeic Acid and C. elegans’ Protection Against Toxin Induced Neurodegeneration By: Michelle Chyn

Caffeic Acid and C. elegans’ Protection Against Toxin Induced Neurodegeneration

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Caffeic Acid and C. elegans’ Protection Against Toxin Induced Neurodegeneration. By: Michelle Chyn. Need. 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. - PowerPoint PPT Presentation

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Page 1: Caffeic Acid and  C. elegans’  Protection Against Toxin Induced Neurodegeneration

Caffeic Acid and C. elegans’ Protection Against Toxin Induced NeurodegenerationBy: Michelle Chyn

Page 2: Caffeic Acid and  C. elegans’  Protection Against Toxin Induced Neurodegeneration

Degree of PD in Patients DM US $ pounds

Average person 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

Need 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

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.

Figure 1: Cost of PD treatment per person annually

Page 3: Caffeic Acid and  C. elegans’  Protection Against Toxin Induced Neurodegeneration

http://www.parkinsonsdecisionaid.eu.com/images/ac/2008/incidence.gif

http://www.e-importz.com/images/coffee_consumption.gif

Coffee consumption in North America and Europe = 1/3 of the tap water intakeNeed Cont.

Figure 2: PD in European Countries

Figure 3: Coffee Consumption

Page 4: Caffeic Acid and  C. elegans’  Protection Against Toxin Induced Neurodegeneration

Knowledge Base

C. elegans biosynthesize dopamine in the corpus stratum (basil ganglia) (Braugart, 2004)

Figure 4:

http://en.wikipedia.org/wiki/Basal_ganglia

Page 5: Caffeic Acid and  C. elegans’  Protection Against Toxin Induced Neurodegeneration

Knowledge Base Cont.

Copper is a neurotoxin when consumed in excess amounts (Gaggelli et al., 2006)

Figure 5: Nerve Synapsehttp://upload.wikimedia.org/wikipedia/en/a/a6/Chemical_synapse_schema.jpg

Page 6: Caffeic Acid and  C. elegans’  Protection Against Toxin Induced Neurodegeneration

Literature Review Antioxidants in coffee decreases risk of development

of age-related diseases (Eskelinen et. al, 2009) Caffeine intake doesn’t show health benefits (Lopez-

Garcia, et. al, 2008)

http://pubchem.ncbi.nlm.nih.gov/summary/summary.cgi?cid=689043&loc=ec_rcs

Figure 6: Antioxidant

Page 7: Caffeic Acid and  C. elegans’  Protection Against Toxin Induced Neurodegeneration

Literature Review Cont. Caenorhabditis elegans MPP+ model used as

PD model for testing anti-PD drugs (Braungart, et al. 2004)

Figure 7: Dose-response effect of MPP+ treatmenton the mobility of wild-type C. elegansanimals.

http://content.karger.com/ProdukteDB/produkte.asp?Aktion=ShowPDF&ArtikelNr=80983&Ausgabe=230458&ProduktNr=229093&filename=80983.pdf

Page 8: Caffeic Acid and  C. elegans’  Protection Against Toxin Induced Neurodegeneration

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

Page 9: Caffeic Acid and  C. elegans’  Protection Against Toxin Induced Neurodegeneration

Literature Review Cont. Direct addition of dopamine-2-c did not help

increase noradrenaline presence in PD patients (Goodall and Alton, 1969)

http://www.pubmedcentral.nih.gov/picrender.fcgi?artid=297487&blobtype=pdf

Figure 9: % noradrenaline recovery after dopamine-2-c infusion

Page 10: Caffeic Acid and  C. elegans’  Protection Against Toxin Induced Neurodegeneration

Purpose Observing the effect of caffeic acid on

neurotoxin-induced degeneration in C. elegans

H0- caffeic acid will not affect the degeneration rate in the C. elegans

HA- caffeic acid will decrease degeneration in the C. elegans.

Hypothesis

Page 11: Caffeic Acid and  C. elegans’  Protection Against Toxin Induced Neurodegeneration

MethodologyCompounds in Coffee and C. elegans Protection Against Parkinson’s Disease

Caenorhabditis elegans egIs1[Pdat-1::GFP] strain from Caenorhabditis Genetics Center (CGC)

N=90

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.

Control

addition of copper oxide, concentratin according to LD 50

N=30

Group 2

addition of copper oxide

caffeic acid- lower concentration according to LD 50

N=30

Group 3

addition of copper oxide

caffeic acid- higher concentration according to LD 50

N=30

Statistical analysis using SPSS and ANOVA

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.

Page 12: Caffeic Acid and  C. elegans’  Protection Against Toxin Induced Neurodegeneration

Protocols

Eggs laid overnight at L4 stage on fresh NGM plates (Mehta, et al, 2009)

Picture drawn by Victoria Wei

Figure 10: Culturing of C. elegans on a petri dish

Page 13: Caffeic Acid and  C. elegans’  Protection Against Toxin Induced Neurodegeneration

Protocols Cont.

http://www.scq.ubc.ca/wp-content/uploads/2006/07/wormcycle.gif

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

Figure 11: C. elegans growth stages

Page 14: Caffeic Acid and  C. elegans’  Protection Against Toxin Induced Neurodegeneration

Protocols Cont. Data collect on scaled brightness of GFP

expression (Braungart, et al., 2004) and area of florescence on Photoshop

Data analysis using SPSS

Page 15: Caffeic Acid and  C. elegans’  Protection Against Toxin Induced Neurodegeneration

Protocols

Page 16: Caffeic Acid and  C. elegans’  Protection Against Toxin Induced Neurodegeneration

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

Page 17: Caffeic Acid and  C. elegans’  Protection Against Toxin Induced Neurodegeneration

BudgetVendor Cat # Pg. Item Qty. Description Unit $ Total $

Caenorhabditis Genetics Center BZ555

C. elegans pdat-1::gfp 110 $7.00 $7.00

Sigma C0625 Caffeic Acid 15 g $53.30 $53.30

Carolina Biological 173520

Nematode Growth Agar 2135 mL $6.25 $12.50

Caenorhabditis Genetics Center E. coli OP50 $7.00 $7.00

Carolina Biological 741270 petri dishes 10100 x 15 mm $6.45 $64.50

Sigma S2002 Sodium azide 125 g $21.20 $21.20

Carolina Biological 216880 ampicillin 15 g $43.25 $43.25

Sigma F0503 fluorodeoxyurodine 1100 mg $117.00 $117.00

Sigma 203122 copper 110 g $146.50 $146.50

LabFluorescent Microscope $3,760.00

Lab GFP filter $938.38

Total Cost $5,052.43

Page 18: Caffeic Acid and  C. elegans’  Protection Against Toxin Induced Neurodegeneration

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Bibliography Cont. Kiontke, Karin and Sudhaus, Walter. “Ecology of Caenorhabditis species.” WormBook The online review of C.

elegans Biology. <http://www.wormbook.org/chapters/www_ecolCaenorhabditis/ecolCaenorhabditis.htm> Lagasse, Paul. Columbia Encyclopedia. “Coffee”. Columbia University Press. May 2, 2009. Lopez-Garcia, Esther; van Dam, Rob M.; Li, Tricia Y.; Rodriguez-Artalejo, Fernando; and Hu, Frank B. “The

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