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Resveratrol and Decreased Fat Deposition in C. elegans. Alexandra Golway- Period 3 Manhasset Science Research. Need. No Data
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Resveratrol and Decreased Fat Deposition in C. elegans
Alexandra Golway- Period 3
Manhasset Science Research
1998
Obesity Trends* Among U.S. Adults1990, 1998, 2007
(*BMI 30, or about 30 lbs. overweight for 5’4” person)
2007
1990
No Data <10% 10%–14% 15%–19% 20%–24% 25%–29% ≥30%
Figure 1: Obesity trends among U.S. adults
Source: Center for Disease Control & Prevention [http://www.cdc.gov/nccdphp/dnpa/obesity/trend/maps/obesity_trends_2007.ppt#487,5,Slide 5]
Need
Knowledge Base
Figure 2: Chemical structure of Nile Red (A) & Sudan Black (B) staining. Nile Red staining in wild-type and mutant C. elegans (C-E). Sudan Black staining in wild-type (F) & mutant (G) organisms.Source: (Ashrafi, 2007) [http://www.wormbook.org/chapters/www_obesity/obesity.html]
Resveratrol
Figure 3: chemical structure of resveratrol (C14H12O3)
[http://www.3dchem.com/molecules.asp?ID=277]
• Chemical found in grapes; red wine
• Popular dietary supplement- advertised as anti-aging substance
•May explain “French paradox”
Literature ReviewRNAi
Mutant C. elegans strains
Nile Red Staining ingested
Wild-type C. elegans
Fat deposition monitored as RNAi silences genes
Results
305 gene silences cause reduced fat storage
112 gene silences cause increased fat storage
Figure 4: RNAi was used to silence certain genes to determine which ones regulated fat in C. elegans (Ashrafi, et al, 2003) [chart made by author using materials from article]
Literature Review
• C. elegans in dauer need AMP-activated protein kinase (AMPK) to survive– Dauer- larval “hibernation”
stage in which metabolic rate is slowed; fat deposits increase to sustain organism
– AMPK- fat regulatory enzyme
• aak2 mutant dauers lack AMPK destroy fat
• (Roy, et al, 2009)
C. Elegans dauerorganisms
Wild-type N2 strain-Has AMPK
aak-2 mutant strainNo AMPK
Sudan Blackstaining
Sudan Blackstaining
Fat reservesRationed at
Constant level
Fat reservesdestroyed
Organisms died
Figure 5: The enzyme AMPK is necessary for C. elegans in dauer to survive and regulate fat deposition (Roy, et al, 2009) [chart created by author]
Literature Review• Resveratrol
improves health and survival of mice on a high-calorie diet
• (Baur, et al, 2006)
Positive effects ofResveratrol
in mice
Increased survival
Activation of AMPK Increased insulin
Sensitivityless fatty acid
syntase
More mitochondriaLess damage to
Liver and pancreas
Figure 6: Resveratrol caused many positive changes in the overall health of mice similar to the effects of caloric restriction
(Baur, et al, 2006) [chart made by author adapted from article]
Literature Review
Human preadipocytes
prevented from becomingfat cells
Decreasedfat storage
Stimulates adiponectin
Anti-obesity properties
of resveratrol
Figure 7: In human preadipocytes (fat cell precursors), resveratrol in extremely high doses (3-5 g) caused positive changes in the cells. (Fischer-Posovsky, et al, 2008) [chart made by author]
Purpose/Hypothesis
• Purpose: to observe the effects of resveratrol on fat deposition in C. elegans and compare it to caloric restriction
• Null hypothesis: Resveratrol will cause no change in fat deposition in C. elegans
• Alternate hypothesis: Resveratrol will cause a decrease in fat deposition and mimic the effects of caloric restriction
Methodology
C. ElegansN= 75
25= control- no resveratrol
or caloric restriction
25= experimental- caloric restriction
25= experimental- Resveratrol
( 5uM,10uM, 20uM per day)
Sudan Black StainingInserted into E. coli food-
Ingested by organism
Fluorescent microscopeExamination- every 3 days
Statistical analysis-Using SPSS program
Protocols
• C. elegans placed in petri dishes
• Nematode Growth Medium (NGM) used
• E. coli is food source• Resveratrol and
Sudan Black will be ingested by organisms Picture by Victoria Wei
Budget
Vendor Order # Quantity Description Unit Price Total Price
Caenorhabditis Genetic Center TJ3014 10 Wild-type C. elegans N2 $7.00 $70.00
Sigma-Aldrich R5010-100mg 1 Resveratrol- 100 mg $81.50 $81.50
Carolina Biological 2009 NP-17-3500 2 Sudan III Stain $26.85 $53.70
Total Price: $205.20
Do-ability
• Resveratrol studies have been done in previous years
• C. elegans is a useful model organism that is easily obtained
• Sudan Black staining and fluorescent microscope examination are possible
Bibliography• Ashrafi, K. Obesity and the regulation of fat metabolism (March 9, 2007), , ed. The Research Community, WormBook,
doi/10.1895/wormbook.1.130.1, <http://www.wormbook.org/chapters/www_obesity/obesity.html>.• Ashrafi, Kaveh, Francesca Y. Chang, Jennifer L Watts, Andrew G. Fraser, Ravi S. Kamath, Julie Ahringer, and Gary Ruvkin. “Genome-
wide RNAi analysis of Caenorhabditis elegans fat regulatory genes.” Nature (2003): 268-72• Baur, Joseph A., Kevin J. Pearson, Nathan L. Price, Hamish A. Jamieson, Charles Lerin, Avash Kalra, Vinayakumar V. Prabhu, Joanne Sl
Allard, Guillermo Lopez-Lluch, Kaitlyn Lewis, Paul J. Pistell, Suresh Poosala, Kevin G. Becker, Olivier Boss, Dana Gwinn, Mingyi Want, Sharan Ramaswamy, Kenneth W. Fishbein, Richard G. Spencer, Edward G. Lakatta, David Le Couteur, Reuben J. Shaw, Placido Navas, Pere Puigserver, Donald K. Ingram, Rafael de Cabo & David A. Sinclair. “Resveratrol improves health and survival of mice on a high-calorie diet.” Nature (2009) Vol. 444.
• Dasgupta, Biplab and Jeffrey Milbrandt. “Resveratrol stimulates AMP kinase activity in neurons.” Proceedings of the National Academy of Sciences of the United States of America. 2007, April 24: 104(17): 7217-7222
• The Endocrine Society. "Red Wine's Resveratrol May Help Battle Obesity." ScienceDaily 17 June 2008. 4 June 2009 <http://www.sciencedaily.com /releases/2008/06/080616115850.htm>.
• Roy, Richard and Patrick Narbonne. “Caenorhabditis elegans dauers need LKB1/AMPK to ration lipid reserves and ensure long-term survival.” Nature (2009) Vol. 457.
• Stiernagle, Theresa. “Maintenance of C. elegans.” February 11, 2006), , ed. The Research Community, WormBook, doi/10.1895/wormbook.1.101.1, <http://www.wormbook.org/chapters/www_obesity/obesity.html>.