Thesis (Ph.D.)--University of Adelaide, Dept. of Clinical and Experimental Pharmacology, 1999. / Erratum tipped in before chapter 1. Bibliography: leaves 226-248.

Thesis (Ph. D.)--University of Hong Kong, 1992.

Thesis (M.S.)--Central Connecticut State University, 1999. / Thesis advisor: Carol A. Jones. " ... in partial fulfillment of the requirements for the degree of Master of Science in Chemistry." Includes bibliographical references (leaves 42-51).

Thesis (Ph. D.)--West Virginia University, 2005. / Title from document title page. Document formatted into pages; contains x, 88 p. : ill. Vita. Includes abstract. Includes bibliographical references.

published_or_final_version / abstract / Anatomy / Doctoral / Doctor of Philosophy

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Genes involved in xenobiotic metabolism and antioxidant signaling are enhanced in the liver of long-lived models, suggesting that a link exists between increased stress resistance and longevity assurance. The ability of certain dietary components such as electrophiles and molecules with high redox potential to induce low-dose stimulation of the endogenous cellular adaptive response is proposed to confer increased resistance to environmental stressors, and thereby present a strategy for lifespan assurance. Recently the term ‘hormetics' was applied to dietary antioxidants possessing this activity, in particular those functioning as indirect antioxidants through the induction of stress responsive pathways. Despite recognition of vitamin E as an essential micronutrient in the diet, very little is still known about its biological role, therefore further investigation into its signaling properties and those of its metabolites are required. This thesis details an investigation into the potential role of vitamin E and its long-chain metabolites in the cellular adaptive stress response in a representative hepatic cell line (HepG2), with the purpose of identifying preliminary targets to aid further comprehension of the role of vitamin E in human health and disease. A global transcriptomic approach was used to identify genes differentially regulated by alpha-tocopherol, these included candidates involved in phase I and II xenobiotic metabolism, the cellular antioxidant response and DNA damage repair. Determination of intracellular alpha-tocopherol levels indicated that up-regulation of gene targets occurred in a concentration-dependent manner. Furthermore, this induction occurred in the presence of significantly elevated levels of short-chain metabolites, alpha-carboxymethylbutyl-hydroxychroman (CMBHC) and alpha-carboxyethyl-hydroxychroman (CEHC), suggesting that metabolism of alpha-tocopherol may be important for its signaling function in HepG2 cells. The role of long-chain metabolites of alpha-tocopherol in cellular stress responsive signaling was also investigated in HepG2 cells. The long-chain metabolites were found to alter mitochondrial metabolism in a concentration-dependent manner, and subsequently to induce components of the Nrf-2 signaling pathway suggesting that these metabolites may be potential hormetic agents, and require further investigation into their role in human health.

Drug-induced liver injury (DILI) is the major cause of pharmaceutical withdrawal from the market and cholestasis is one of the most common DILI observed. Cholestasis stem from abnormalities in the activity of bile acid transporters, namely the sodium-dependent taurocholate co-transporting polypeptide, Ntcp (Slc10a1), and the bile salt export pump, Bsep (Abcb11). The nuclear receptors, pregnane X receptor (PXR) and farnesoid X receptor (FXR), have been implicated in the regulation of Ntcp and Bsep. The aim of this study was to establish the relative roles of FXR and PXR in the regulation of Ntcp and Bsep, in the rat liver and sandwich cultured rat hepatocytes (SCRH) to establish the usefulness of SCRH as a good model for bile acid transport studies. For this purpose, male Sprague Dawley rats were treated with FXR and PXR ligands and siRNAs. Intraperitoneal (IP) injection of FXR ligand, CDCA, resulted in induction in FXR mRNA levels whereas siRNA treatment knocked down FXR transcript levels. FXR induction decreased Ntcp transcript levels that were reversed in FXR knockdown animals. CDCA treatment resulted in greater than 2-fold increase in Bsep mRNA levels that was absent in FXR knockdown animals. The PXR ligand, PCN, showed an induction in PXR mRNA levels while siRNA treatment resulted in the knockdown in PXR transcript levels. PXR induction did not cause any change in Ntcp and Bsep transcript levels. Furthermore, sandwich cultured rat hepatocytes were used to ascertain if the above in vivo findings are reproducible in vitro. Treatment of hepatocytes with FXR ligand, CDCA, and PXR ligand, PCN, caused greater than 5-fold increase in the respective mRNA levels whereas respective siRNA treatment caused >79% knocked in their transcript levels. FXR induction decreased Ntcp mRNA levels that were ablated in FXR knockdown cultures. FXR induction resulted in a 6-fold increase in Bsep mRNA levels that disappeared in FXR knockdown cultures. On the contrary, PXR induction did not influence the expression levels of Ntcp and Bsep. Taken together these data demonstrate that both FXR and PXR are inducible at mRNA and protein levels in vivo and their expression can be knocked down in the rat liver. Moreover, FXR negatively regulate Ntcp and positively regulate Bsep while PXR does not influence Ntcp and Bsep expression. Finally, the data shows that these in vivo findings can be successfully reproduced in sandwich cultured rat hepatocytes. To conclude, sandwich cultured rat hepatocytes mimic the in vivo situation and provide a good model for bile acid transport studies.