Greater Understanding of Lipids in Caenorhabditis elegansExposed to Oxygen Deprivation

Padilla Lab, Department of Biological Sciences

Jemma AlarcónTAMSter Researcher

Health Problems Related to Oxygen DeprivationHealth Problems Related to Oxygen Deprivation

Myocardial infarctionMyocardial infarction StrokeStroke Shock due to low blood Shock due to low blood

pressure caused by pressure caused by vascular disease, vascular disease, inadequate heart function, inadequate heart function, or low blood volumeor low blood volume

Solid tumor cell resistance Solid tumor cell resistance to radiation and to radiation and chemotherapychemotherapy

Pulmonary disease and Pulmonary disease and dysfunctiondysfunction

Underdeveloped lung Underdeveloped lung function in premature function in premature babiesbabies

Anemia due to blood loss Anemia due to blood loss from severe injury or from severe injury or destruction of red blood destruction of red blood cellscells

Stroke alone accounts for 1 out of every 15 deaths in the United States and is the third leading cause of death in most developed countries.

Research Approaches to Studying OResearch Approaches to Studying O22 Deprivation Deprivation

PhysiologyPhysiology- Sea turtles- Sea turtles- Brine Shrimp- Brine Shrimp- Diving Seals- Diving Seals- Birds- Birds

Cell BiologyCell Biology- Cell culture- Cell culture

Genetic approachGenetic approach- - Drosophila melanogasterDrosophila melanogaster “Fruit flyes”“Fruit flyes”- - Caenorhabditis elegansCaenorhabditis elegans

3 http://www.acuariolasmercedes.com/Guia-de-cuidado/cria-de-artemia-salina.htm4 www.exploratorium.edu/.../ mutant_flies/ebony.gif

1 www.scubamaui.com/ turtles.htm

Anoxia-Induced Suspended Animation

Normoxia

Anoxia

Time 0 24 hours Time 0 24 hours

Anoxia (<.001kPa oxygen in environment)Padilla, 2002 Mol. Biol. Cell, May 13(5)

Embryos and larvae were exposed to either normoxic or anoxic conditions

Question and approach to Question and approach to Evaluating lipids in Evaluating lipids in C. elegansC. elegans exposed to anoxia. exposed to anoxia.

Hypothesis:Hypothesis: Oxygen deprivation affects lipid accumulation. Oxygen deprivation affects lipid accumulation.

Approach:Approach: Use the dye Nile Red to assay lipid/fat accumulation Use the dye Nile Red to assay lipid/fat accumulation in in C. elegansC. elegans. . – In In C. elegansC. elegans Nile Red is taken up and attaches to lipids in Nile Red is taken up and attaches to lipids in

intestinal cells. Lipids are visualized using fluorescent microscopy.intestinal cells. Lipids are visualized using fluorescent microscopy.

Results:Results: Increased detection of lipids in worms exposed to Increased detection of lipids in worms exposed to anoxia. anoxia. – This result leads to the question of whether lipids are important for This result leads to the question of whether lipids are important for

anoxia survival.anoxia survival.

Ashrafi, K et al, (2003). Nature 421, 268-272.

DIC NomarskiMicroscopy

Fluorescent microscopy can be used to assay lipids via Nile Red detection

Lipid Droplets

Normoxia

1 Day Anoxia

2 Days Anoxia

DIC Nile Red

Nematodes Exposed to 1 or 2 days of Anoxia Accumulate Lipids as Seen by Nile Red Detection.

Size bar equals 100µm.

Comparison of Lipid Accumulation in Wild type (N2) and Comparison of Lipid Accumulation in Wild type (N2) and daf-2 daf-2 StrainsStrains

Normoxia

1 Day Anoxia

2 Days Anoxia

N2 daf-2

Size bar equals 100µm.

Discussion and Conclusion

• Lipid accumulation- Measure intensity- RNAi experiments- Viability

• Continue with daf-2 experiments- RNAi experiments- Viability

Acknowledgments

• Project was supported in part by a grant from National Institute of Health• TAMS Summer Research Scholarship. • Special thanks to Dr. Padilla for mentoring this project.• I thank Dr. Padilla lab members for valuable input and suggestions.

Alex Mendenhall Neil Stewart Vinita Hajeri Sara Anvari Bobby LaRue Nicole Parker Jesus Trejo! Desh Mohan

Worms rule!http://www.celegans.de/

http://www.bio.unc.edu/ahmed/lab/

Dr. Padilla