Date of Award

1-1-2016

Embargo Period

1-1-2019

Document Type

Thesis - MUSC Only

Degree Name

Master of Science (MS)

Department

Biochemistry and Molecular Biology

College

College of Graduate Studies

First Advisor

L. Ashley Cowart

Second Advisor

J. Alan Diehl

Third Advisor

Besim Ogretman

Fourth Advisor

Don C. Rockey

Fifth Advisor

Joe Blumer

Abstract

Nonalcoholic fatty liver disease is defined as the accumulation of excess lipids in the liver of patients who drink little to no alcohol. Often times this disease can go undiagnosed during the benign stage of simple steatosis. Inflammation develops in approximately 20% of diagnoses with fatty liver disease, however the mechanism of this development remains to be fully understood. Using a customized obesogenic diet from our lab, mice were subjected to high saturated fat feeding for approximately 18 weeks to develop NAFLD. It was observed that sphingosine kinase 1 (SphK1) was mRNA was increased with HSFD. TNFalpha and MCP-1 cytokines were also increased in livers with HSFD. Sphingosine 1 phosphate (S1P), the product of sphingosine kinase 1, was determined to increase expression of these cytokines in a receptor-dependent manner. These findings were further supported with cultured primary hepatocytes treated with the saturated fatty acid, palmitate. The regulation of SphK1 and inflammation in NAFLD remained elusive even with these findings. Recent literature suggests a potential link between endoplasmic reticulum stress and inflammation. We formed the hypothesis that saturated fatty acid overload leads to endoplasmic reticulum stress in hepatocytes, which promotes induction of sphingosine kinase 1, and its product S1P, to initiate a pro- inflammatory response, perhaps in an attempt to resolve ER stress. Cell culture techniques for isolated primary mouse hepatocytes were used to determine the role 5 of ER stress in regulating SphK1. Both palmitate, which is known to induce ER stress, and thapsigargin, a potent inducer of ER stress were found to increase SphK1 protein. We determined that this regulation was not through the IRE1alpha branch of ER stress. A dual role of S1P enhancing and repressing ER stress was observed to affect the PERK/CHOP pathway of ER stress in an S1P receptor-dependent manner. To translate these results to an in vivo model, a hepatocyte specific SphK1-/- mouse was generated. Future work remains necessary to fully understand the complex mechanism of sphingolipid metabolism regulating ER stress and inflammation in NAFLD.

Rights

All rights reserved. Copyright is held by the author.

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