Date of Award

Fall 2019

Document Type

Open Access Dissertation

Department

College of Pharmacy

First Advisor

Hippokratis Kiaris

Abstract

The endoplasmic reticulum is the site where integral membrane and secreted proteins are synthesized and folded, and is also the site of synthesis of steroids, lipids, and other macromolecules. While ER is found in most cell types, hepatocytes in particular contain large amounts of both rough and smooth ER to facilitate their tasks, which include lipoprotein assembly and secretion, cholesterol biosynthesis, and lipid metabolism. As such, stress in the ER – and the cells’ ability to respond to it – plays a key role in disease pathogenesis. In particular, hepatic steatosis, or fatty liver, is linked to ER stress. However, if and how inherent variations in the resulting unfolded protein response (UPR) affect the predisposition to ER stress-associated metabolic conditions, including hepatic steatosis, remains to be established. By using genetically diverse deer mice (Peromyscus maniculatus) as a model, we show that the profile of tunicamycininduced UPR in fibroblasts isolated at puberty varies between individuals and predicts deregulation of lipid metabolism and diet induced hepatic steatosis later in life. Among the different UPR targets tested, CHOP more consistently predicted elevated plasma cholesterol and hepatic steatosis. Compared to baseline levels or inducibility, the maximal intensity of the UPR following stimulation best predicts the onset of pathology. Differences in the expression profile of the UPR recorded in cells from different populations of deer mice correlate with the varying response to ER stress in altitude adaptation. Upon analysis of gene expression in the livers, no differences in the expression levels of various UPR-associated genes between steatotic and nonsteatotic vi livers were recorded. Then we asked if there is a change in the relative levels of these genes as compared to each other. While we did not note any changes to the coordination of UPR genes among themselves, we did note that there was a loss of coordination between ER stress genes and autophagy and metabolic genes, particularly hepatic lipase (HL), but only in the livers of mice who had developed steatosis. Additionally, using rat hepatoma cells which naturally express HL, we demonstrated that induction of ER stress leads to the downregulation of HL alongside the upregulation of ER stress-induced genes. Together, this data demonstrates the ability of tunicamycin-treated fibroblasts isolated early in life to predict the propensity of an individual to develop hepatic steatosis; this approach might also be useful for other conditions associated with ER stress, both metabolic and otherwise. The results of the liver analysis imply that ER stress plays a role in the expression of HL and potentially other important metabolic genes, and that the loss of coordination of ER stress response with other cellular responses could be a more important factor in the transition to disease than the levels of individual transcripts alone.

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