1. The Effect of Centrophenoxine on Parkinson’s Disease
Victoria Wei

2. Need
Taken from Rajput AH, Offord KP, Beard CM, Kurland LT. Epidemiology of parkinsonism: incidence, classification, and mortality. Ann Neurol. 1984;16:
Figure 1 The amount of Parkinson’s disease cases per 100,000 people in the United States as age increases

3. Knowledge Base
Parkinson’s disease is a brain disorder involving the nerves.
Figure 2 The effects of Parkinson’s disease

4. Knowledge Base Figure 3 The life cycle of C. elegans
Figure 3 The life cycle of C. elegans

5. Knowledge Base Figure 4 Lipofuscin in neurons of the human brain.
Figure 4 Lipofuscin in neurons of the human brain.

6. Knowledge Base Figure 5 Molecular structure of centrophenoxine
Figure 5 Molecular structure of centrophenoxine

7. Literature Review Caldwin, et. al. (2008) Figure 7 Figure 6
Notes that I will need about this slide
Figure 6
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

8. Literature Review Sutphin, et. al. (2009) Figure 8
Auto fluorescent pigments present in Day 4 and Day 8 C. elegans
Notes that I will need about this slide
Sutphin, George; M. Kaeberlein. “Measuring Caenorhabditis elegans Life Span on Solid Media” JOVE

9. Literature Review Gerstbrein, et. al. (2008)
Figure 9 Auto fluorescent pigments present in Day 7 and Day 12 C. elegans.
Notes that I will need about this slide
Gerstbrein, Beate; G. Stamatas; N. Kollias; M. Driscoll. “In viv spectrofluorimetry reveals endogenous biomarkers that report healthspan and dietary restriction in Caenorhabditis elegans.

10. Literature Review
Application 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
Notes that I will need about this slide
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

11. Purpose
The purpose of the experiment is to observe the effects of centrophenoxine on Parkinson’s disease in C. elegans
Hypothesis
Null- 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.

12. Methodology
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
Given 6.8 mM centrophenoxine
N=20
Control
N=20
Given 6.8 mM centrophenoxine
N=20
Control
N=20
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

13. Protocol
C. elegans are grown in petri dishes containing Nematode Growth Media (NGM) from Carolina Biological and fed U.V. killed Escherichia coli.
Picture by author
Figure 11 Culturing the C. elegans in Petri dishes

14. Protocol
Both Ampicillin and 5-Fluoro-2′-deoxyuridin will be used with NGM in the petri dishes with C. elegans
Figure 13 FUDR
Figure 12 Ampicillin
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

15. Protocol Figure 14 Age synchronization of C. elegans
Sutphin, George; M. Kaeberlein. “Measuring Caenorhabditis elegans Life Span on Solid Media” JOVE
Figure 14 Age synchronization of C. elegans

16. Protocol Figure 15 4',6-diamidino-2-phenylindole (DAPI)
Figure 15 4',6-diamidino-2-phenylindole (DAPI)
Figure 17 GFP expressed in C. elegans
Figure 16 Fluorescence microscope

17. Budget Vendor Cat# Item Qty. Unit $ Total $
Caenorhabditis Genetics Center
GS1214
ham-1(ot339) C. elegans 1 $7
AB1
Wild type C. elegans
Sigma
D9542-5MG
DAPI
$51.60
S2002
Sodium azide
$21.20
F MG
FUDR
$117
SLC G
Centrophenoxine Hydrochloride
$97.82
Carolina Biological
741270
Petri dishes 10
$6.45
$64.50
216880
Ampicillin dry powder
$43.25
$43
173520
Nematode Growth Agar 2
$6.25
$12.50
OP50
E. coli
Invitrogen
D21490
$116.00
Nova-tech
1482FLi
Fluorescence Microscope
$3,760
Tritech
MINJ-F-EFB
GFP filter set
$938.38
$938
Total cost
$5,243

18. Do-ability Available for Purchase: Equipment already Acquired:
ham-1(ot339) and wild type C. elegans strains from CGC
DAPI and Sodium Azide from Sigma
NGM and OP50 E.coli from Carolina Biological
Centrophenoxine purchasable from Science Lab.com
Equipment already Acquired:
DAPI filter, GFP filter, fluorescent microscope

19. Bibliography
 "About Parkinson Disease." National Parkinson Foundation. <”
Braungart, Evelyn; Gerlach, Manfred; Riederer; Peter, Baumeister, Ralf; and Hoener, Marius C. “Caenorhabditis elegans MPP+ Model of Parkinson’s Disease for High-throughout Drug Screening.” Neurodegenerative Disease Volume 1: pgs
Caldwin, Guy A.; K.A. Caldwell. “Traversing a wormhole to Combat Parkinson’s disease.” Disease Models and Mechanisms. Volume 1. pp
Colleta, Susan. Introduction to C. elegans. Waksman Student Scholars. < 2009
Gerstbrein, Beate; G. Stamatas; N. Kollias; M. Driscoll. “In viv spectrofluorimetry reveals endogenous biomarkers that report healthspan and dietary restriction in Caenorhabditis elegans.
Hall, D. H.; Z. F. Altun. “C. elegans Atlas.” Genetics Research, 90 , pp
Hunt, Sara S. The Aging Process. Washington D.C. April 2004.
Kenyon, Cynthia. “Environmental Factors and Gene Activities That Influence Life Span” C. elegans II. Cold Spring Harbor Press
Kisiel, Marion J.; B. Zuckerman. “Effects of Centrophenoxine on the Nematode Caenorhabditis Briggsae” Age. Volume 1. Pp January 1978.
Mc Naught, KS; P. Jenner. “Proteasomal function is impaired in substantia nigra in Parkinson's disease “ Neuroscience Letters. Volume 297. pp
O'Riordan ; A.M. Burnell. Intermediary metabolism in the dauer larva. II. The glyoxylate cycle and fatty acid oxidation. Comp. Biochem. Physiol. Volume 95. pp
Rajput AH, Offord KP, Beard CM, Kurland LT. Epidemiology of parkinsonism: incidence, classification, and mortality. Ann Neurol. 1984;16:
Schneider, Howard F.; C. Nandy. “Effects of Centrophenoxine on Lipofuscin Formation in Neuroblastoma Cells in Culture” Journal of Gerontology. Volume 32. Pp
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
Sutphin, George; M. Kaeberlein. “Measuring Caenorhabditis elegans Life Span on Solid Media” JOVE
“What is Parkinson’s?” American Parkinson Disease Association West Coast Office. <“