Antioxidants & Obesity in C. elegans

Manhasset Science Research

Madelyn Sayed

Need: Obesity

(American Heart Association, 2004)http://www.americanheart.org/downloadable/heart/1103834132259FS16OVR5.pdf

Figure 1: Prevalence of Obesity in adults ages 20-74 by sex. (1960-2004)

http://www.oxha.org/knowledge/backgrounders/overweight_and_obesity.gif/view

Need: Obesity (2)

Figure 2: Obese and overweight men and women in India, China, Mexico, & USA.

Need: Diabetes Statistics

Figure 2.5: Diabetes in patients over 18 by race and sex. (Non-Hispanic Whites, Non-Hispanic Blacks, and Mexican Americans).

http://www.oxha.org/knowledge/backgrounders/overweight_and_obesity.gif/view

Knowledge Base Hyperlipidemia Phytochemicals Antioxidant Free Radical Beta-carotene. Thiol Hypodermis

Figure 3: Layers of the skin in the human body are shown. The hypodermis is the lowest layer in this diagram.

http://www.daviddarling.info/images/hypodermis.jpg

Knowledge Base (2)

Figure 4: The lifecycle of C. elegans in hours at a constant temperature of 25°C.

http://thalamus.wustl.edu/nonetlab/ResearchF/elegans.html

Knowledge Base (3)

Figure 5: C. elegans at various stages in its lifecycle in hours at a constant temperature of 25°C.

http://thalamus.wustl.edu/nonetlab/ResearchF/elegans.html

Seymour et al., 1997

Rats fed low and high fat diets with and without blueberries

Rats that received the blueberry-enriched powder ‘healthier’

Also true with grapes and cherries in other experiments

Jones et al., 2009 crucial regulators of

lipid homeostasis in mammals serve similar functions in C. elegans.

fat metabolism, & feeding/food-related behaviors

intestinal and hypodermal cells in C. elegans (red)

Systems transport fats between cells and tissues (green).

Figure 6: C. elegans fat transport and storage areas.

Jones et al., 2009 (2)

(A) Nile Red(B) Sudan Black B (C) LipidTOX neutral lipid staining(D) Nile Red and fatty acid lipid conjugated BODIPY staining(E) fatty acid lipid conjugated BODIPY staining(F) Both Nile red and BODIPY florescence

Figure 7: C. elegans fat deposits shown-stained with Nile Red, Sudan Black B, and other stains.

Hildebrandt, 2004 Effect of thiol antioxidant on body fat and insulin

reactivity 2 studies: obese & hyperlipidemic/normolipidemic treated with antioxidants such as x-lipoic acid and NAC Their studies showed that the effect of NAC was

statistically significant, but the effect of creatine was not.

Figure 8

Ashrafi et al., 2003 & 2007 (1)

Figure 9:

C. elegans stained with Nile Red in C, D, and E, with Sudan Black B in F and G. Fat appears fluorescent red (N.R.) and blue-black (S.B.)

Ashrafi et al., 2003 & 2007 (2)

10

Halliwell et al., 2009

Pine bark extract on metabolism and lifespan of C. elegans

Figure 11:

Antioxidant balance in body, affect on metabolism

Purpose / Hypothesis

Purpose: to test the affect of antioxidants on fat deposition in C. elegans.

Null Hypothesis: the blueberry polyphenol compounds will not reduce fat deposition in C. elegans.

Alternate Hypothesis: the blueberry polyphenol compounds will reduce fat deposition.

Methodology

Cultured axenically Grown under monoxenic conditions Pine Bark Extract E. coli strain OP50 Stain: Sudan Black B

http://individual.utoronto.ca/proy/worms.htm

Flow ChartCaenorhabditis

elegansLifespan: (2-3 weeks) Rapid generation time Experimental flexibility

Control GroupSample Size N = ? Not fed Pine Bark Extract

Experimental GroupSample Size N = ?Fed Pine Bark Extract

Measurement:Sudan Black B fat staining

Motic Plus camera with StereomicroscopeAmount of fat (using Adobe Photoshop) deposited in the worm (cm3)

Do Ability

Fluorescent Dissection microscope is available in the lab

Pycnogenol® easily obtainable C. elegans easily cultured Sudan Black B reliable fat stain Motic plus camera with stereomicroscope for

photography Adobe Photoshop available for fat measurement

Protocols for Measurement

C. elegans will be fed E.coli OP50 Pycnogenol® 50 mg capsules Sudan Black: 0.7 gm Propylene Glycol: 100.0 ml

Figure: E. Coli bacteria

Budget

Item Vendor Catalogue # Pg. Unit Price Qty Total Price

C. elegans N2 (wild type)

Carolina Science 2009

NP-17-3500 248 $10.95

(per culture)

1

NGM Carolina Science 2009

NP-17-3520 248 $6.25

(per 135 mL bottle)

2

Sudan Black III(Fatstain Herxheimer Lab Grade)

Carolina Science 2009

NP-89-2963 821 $6.65

(120 mL bottle)

2

Pycnogenol®

(Pine Bark Extract)

Vitacosthttp://www.vitacost.com/Schiff-Pycnogenol?csrc=PPCADWLT-pycnogenol&s_kwcid=TC|3001|pycnogenol||S|p|2989626851

Item #:

SCF 5127419or020525127419

$16.53

(30 capsules) (50 mg/ capsule)

1

$40.38

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