Undergraduate Honors Thesis Projects

Date of Award

Spring 4-5-2017

Document Type

Honors Paper

Degree Name

Biochemistry and Molecular Biology-BS

Department

Biochemistry and Molecular Biology

Advisor

Dr. John Tansey

First Committee Member

Dr. David Sheridan

Second Committee Member

Dr. Meredith Frey

Keywords

perilipin 5a, protein, lipid droplet, mouse, tissue

Subject Categories

Biochemistry | Biology | Cell Anatomy | Cell and Developmental Biology | Laboratory and Basic Science Research | Molecular Biology | Other Biochemistry, Biophysics, and Structural Biology

Abstract

The steep rise in both childhood and adult obesity over the past three decades has moved to the forefront of public consciousness in recent years. This development has generated a marked increase in general health awareness and lifestyle changes for a vast number of individuals, most notably in the form of increased physical activity and diet alterations. The latter point is especially salient in a biochemical context, because of the myriad factors that can result in “fat accumulation”. Chief among these factors is the Perilipin 5A gene, (known as PLIN5A) which encodes the protein Perilipin 5A of the Perilipin family of lipid associated proteins. Perilipin 5A, along with Adipose-differentiation related protein (ADRP) and TIP-47 make up what is known as the PAT family of proteins that interact with fat containing lipid droplets within the cell. They are instrumental in regulating fat (triglyceride or triacylglycerol) storage at the cellular level. Perilipin 5A is key to lipolysis in white and brown adipose, heart, liver, and some muscle tissue. This research endeavor delineated the role that Perilipin 5A expression plays in fat mobilization for normal diet fed and high fat fed mice in each of 5 target mouse tissues: white adipose, brown adipose, liver, cardiac, and brown tissue. To test this, two groups of mice were put on either a “normal” or “high fat” diet for a period of weeks, during which glucose levels and weights are measured. After being terminated, the tissue of interest (brown adipose, white adipose, cardiac, liver, and muscle tissue) were thoroughly analyzed using thin layer chromatography, and western blotting to verify how Perilipin 5A expression affected lipid storage patterns in each tissue type. The correlation between plasma glucose levels and lipid storage was also further explored with blood glucose analysis prior to, during, and after diets were administered. This was done by first performing thin layer chromatography to measure triacylglycerol content in multiple tissue types for each mouse. Thin layer chromatography (TLC) was employed to measure fatty content triacylglycerol (TAG) in target mice tissue and compare lipid levels in both high fat diet fed and normal diet fed mice. The mice with higher fat stores also had a higher blood glucose, on average. A western blotting procedure was used to analyze the presence of target proteins. In particular, Perilipin 5A levels were analyzed for each of the five tissue types. The western blot for each of the tissue showed some varied PLIN5A expression for high fat diet and normal diet mice, though the extent to which expression varied is unclear. Greater expression of Perilipin 5A was evident in the increased blood glucose readings with the high fat diet mice as the study progressed, indicative of greater glucose production by the liver, as a result of lipolytic stimulation causing free glycerol to enter the bloodstream. This study further characterized the extent to which Perilipin 5A regulates both lipid storage and lipolysis, and the data provided insight into how increased Perilipin 5A expression affects certain tissue types. Summarily, this research sought to understand how greatly Perilipin 5A expression was impacted by diet, and subsequently which tissue types were most affected by increased Perilipin 5A expression. The results may help put into better context the extent to which lipid synthesis and lipolysis exert their effects in various tissue, and the extent to which diet can impact both.

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