The Effect of Centrophenoxine on Parkinson’s Disease Victoria Wei

Need

Taken from Rajput AH, Offord KP, Beard CM, Kurland LT. Epidemiology of parkinsonism: incidence, classification, and mortality. Ann Neurol. 1984;16:278-282.

Figure 1 The amount of Parkinson’s disease cases per 100,000 people in the United States as age increases

Knowledge Base

Parkinson’s disease is a brain disorder involving the nerves.

http://www.spinstudios.co.uk/sa/pa3.jpg

Figure 2 The effects of Parkinson’s disease

Knowledge Base

Figure 3 The life cycle of C. eleganshttp://www.wormatlas.org/handbook/fig.s/IntroFIG6.jpg

Knowledge Base

Figure 4 Lipofuscin in neurons of the human brain. http://www.innovitaresearch.org/news/res/06042501_01.jpg

Knowledge Base

http://commons.wikimedia.org/wiki/File:Centrophenoxine.svg

Figure 5 Molecular structure of centrophenoxine

Literature ReviewCaldwin, et. al. (2008)

Figure 7 Both images taken from Caldwin, Guy A.; K.A. Caldwell. “Traversing a wormhole to Combat Parkinson’s disease.” Disease Models and Mechanisms. Volume 1. pp.000-000. 2008.

Figure 6

Literature ReviewSutphin, et. al. (2009)

Sutphin, George; M. Kaeberlein. “Measuring Caenorhabditis elegans Life Span on Solid Media” JOVE. 2009.

Figure 8Auto fluorescent pigments present in Day 4 and Day 8 C. elegans

Literature ReviewGerstbrein, et. al.

(2008)

Figure 9 Auto fluorescent pigments present in Day 7 and Day 12 C. elegans.

Gerstbrein, Beate; G. Stamatas; N. Kollias; M. Driscoll. “In viv spectrofluorimetry reveals endogenous biomarkers that report healthspan and dietary restriction in Caenorhabditis elegans.

Literature ReviewApplication of centrophenoxine to the C. elegans

decreased the rate of lipofuscin accumulation by an average of 41.3%. (Shulkin, et. al., 1978)

Figure 10 Table displaying effect of centrophenoxine on lipofuscin in C. elegans

Shulkin, D.J.; B.M. Zuckerman. “Spectrofluorometric analysis of the effect of centrophenoxine on lipofuscin accumulation in the nematode C. elegans.” Age. Volume 5. Pp. 50-53. 1982.

PurposeThe purpose of the experiment is to observe the effects

of centrophenoxine on Parkinson’s disease in C. elegans

HypothesisNull- the symptoms of Parkinson’s disease will remain the same with

or without the application of centrophenoxine.Alternate- the symptoms of Parkinson’s disease will lessen with the

application of centrophenoxine.

The Effects of Centrophenoxine on the development of Parkinson’s disease in C. elegans

Wild type C. elegans and ham-1 (ot339) C. elegans obtained from the Caenorhabditis Genetics Center

N=80

Wild type C. elegans: N=40 ham-1(ot339) C. elegans: N=40

6.8 mM centrophenoxine will be applied to the Nematode Growth Media for 21 days.

Use of 4',6-diamidino-2-phenylindole (DAPI) to observe the amount of auto fluorescent pigment- lipofuscin- in C. elegans. GFP filters may also be used to observe whether the C. elegans DA neurons will be affected by the application of centrophenoxine

Statistical analysis

Given 6.8 mM centrophenoxineN=20

Control N=20

Given 6.8 mM centrophenoxineN=20

Control N=20

ProtocolC. elegans are grown

in petri dishes containing Nematode Growth Media (NGM) from Carolina Biological and fed U.V. killed Escherichia coli.

Figure 11 Culturing the C. elegans in Petri dishes

Picture by author

ProtocolBoth Ampicillin and 5-Fluoro-2 -deoxyuridin will be ′

used with NGM in the petri dishes with C. elegans

http://upload.wikimedia.org/wikipedia/commons/b/b6/Ampicillin_structure.svg

http://www.sigmaaldrich.com/structureimages/30/mfcd00006530.gif

Figure 12 Ampicillin Figure 13 FUDR

E.coli + NGM with centrophenoxine + Ampicillin + FUDR using the DAPI and GFP filter to observe amount of lipofuscin and fluorescence present in both C. elegans groups

Protocol

Sutphin, George; M. Kaeberlein. “Measuring Caenorhabditis elegans Life Span on Solid Media” JOVE. 2009.

Figure 14 Age synchronization of C. elegans

Protocol

Figure 15 4',6-diamidino-2-phenylindole (DAPI)

http://upload.wikimedia.org/wikipedia/commons/7/7a/DAPI.png

Figure 16 Fluorescence microscopehttp://www.bcgsc.ca/people/msleumer/htdocs/fact_sheet/image004.jpg

Figure 17 GFP expressed in C. elegans http://www.ion.ucl.ac.uk/images/Fluorescence-Microscope.jpg

BudgetVendor Cat# Item Qty. Unit $ Total $

Caenorhabditis Genetics Center GS1214 ham-1(ot339) C. elegans 1 $7 $7

Caenorhabditis Genetics Center AB1 Wild type C. elegans 1 $7 $7

Sigma D9542-5MG DAPI 1 $51.60 $51.60 Sigma S2002 Sodium azide 1 $21.20 $21.20

Sigma F0503-100MG FUDR 1 $117 $117

Sigma SLC5377-25G Centrophenoxine Hydrochloride 1 $97.82 $97.82

Carolina Biological 741270 Petri dishes 10 $6.45 $64.50

Carolina Biological 216880 Ampicillin dry powder 1 $43.25 $43

Carolina Biological 173520 Nematode Growth Agar 2 $6.25 $12.50

Carolina Biological OP50 E. coli 1 $7 $7

Invitrogen D21490 DAPI 1 $116.00 $116.00

Nova-tech 1482FLi Fluorescence Microscope 1 $3,760 $3,760

Tritech MINJ-F-EFB GFP filter set 1 $938.38 $938 Total cost $5,243

Do-abilityAvailable for Purchase:

ham-1(ot339) and wild type C. elegans strains from CGC DAPI and Sodium Azide from SigmaNGM and OP50 E.coli from Carolina BiologicalCentrophenoxine purchasable from Science Lab.com

Equipment already Acquired:DAPI filter, GFP filter, fluorescent microscope

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elegans.” Age. Volume 5. Pp. 50-53. 1982. Sutphin, George; M. Kaeberlein. “Measuring Caenorhabditis elegans Life Span on Solid Media” JOVE. 2009. “What is Parkinson’s?” American Parkinson Disease Association West Coast Office. <“http://www.apdawest.org/WhatIsParkinsons.html#2”>.

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