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<p>Normal cellular energy metabolism is fundamentally altered in cancer cells to facilitate rapid production of new cellular components, thereby enabling uncontrolled cell growth. Specifically, cancer cells rely on glycolysis and alternative pathways such as lipid and glutamine metabolism for energy, while diverting substrates away from oxidative metabolism regardless of the prevalence of oxygen in the microenvironment. This hallmark of cancer cells is referred to as the Warburg effect, the precise regulation of which is poorly understood despite several decades of research. In comparing the global gene expression profiles of ovarian cancer cells to those that overexpress Ets-1, we have revealed that this transcription factor is involved, at least in part, to this cancer-associated metabolic switch. To support the validity of these findings, we have shown that Ets-1 functionally regulates glycolytic dependence in ovarian and breast cancer cells, while concomitantly displaying a decreased capacity for oxidative phosphorylation. Reactive oxygen species are a normal byproduct of metabolism, and are produced excessively in cancer cells leading to oxidative stress. Interestingly, our genomic pathway analyses uncovered enrichments in antioxidant pathways associated with increased Ets-1 expression. Accordingly, we have also observed that Ets-1 regulates increased intracellular glutathione levels, and induces the activity of key antioxidant enzymes under oxidative stress. Sulfasalazine, an agent that restricts cystine uptake, was shown to be effective for decreasing these high glutathione levels during oxidative stress. These results are clinically relevant because high glutathione levels are associated with iii therapeutic resistance in cancer cells. Collectively, the evidence presented has identified a novel role for the transcription factor Ets-1 in the regulation of cancer energy metabolism, as well as the response to oxidative stress. We have also described a mechanism for Ets- 1-mediated therapeutic resistance, suggesting that this transcription factor may be a promising novel target to enhance conventional cancer therapies.</p> / Doctor of Philosophy (Medical Science)

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This research first examined nitrification inhibition caused by different classes of industrially relevant chemicals on activated sludge and found that conventional aerobic nitrification was inhibited by single pulse inputs of every chemical tested, with 1-chloro-2,4-dinitrobenzene (oxidant) having the most severe impact, followed by alkaline pH 11, cadmium (heavy metal), cyanide, octanol (hydrophobic) and 2,4-dinitrophenol (respiratory uncoupler). Of the different chemicals tested, the oxidative and hydrophobic chemicals showed severe nitrification inhibition relative to other treatment processes and therefore deserved further investigation. For oxidative chemicals, we hypothesized that the more severe inhibition was because nitrifying bacteria lack one or more of the microbial stress response mechanisms used to mediate the toxic effect of oxidative chemicals. During these experiments, we showed that a rapid (minutes) antioxidant potassium efflux mechanism does not exist in two nitrifying bacteria, Nitrosomonas europaea and Nitrospira moscoviensis. Furthermore, we showed that another important antioxidant molecule, glutathione, was not oxidized as readily as in a non-nitrifying bacterium. Furthermore, we hypothesized that hydrophobic chemical-induced nitrification inhibition recovered more quickly because of the presence of membrane modification stress response mechanisms. While testing this hypothesis, we showed that N. europaea modified its cell membrane in response to hydrophobic chemicals using a long-term (hours) membrane modification mechanism that required the synthesis of new fatty acids, but it did not contain a short-term (minutes) response mechanism involving a cis/trans isomerase. Therefore, investigating these nitrifier stress responses showed that nitrifiers lack short-term stress responses that may be used to rapidly detect inhibition, indicating that conventional methods of detecting nitrification inhibition, like differential respirometry and nitrate generation rate (NGR), are still the fastest and easiest methods to use. Because several conventional methods exist, we also investigated differences between differential respirometry and a UV method we developed to measure NGR. During these tests, we showed that the UV NGR method provided a more reliable measure of nitrification inhibition than differential respirometry, and that the time to maximum nitrification inhibition depended on the properties of the chemical toxin, which implies that longer exposure times may be needed to accurately predict nitrification inhibition. / Ph. D.

No / Glutathione depletion is a consistent feature of the progression of mild to severe acute pancreatitis. In this study, we examined the temporal relationship between cysteine, homocysteine, and cysteinyl-glycine levels; total reduced erythrocyte glutathione; gamma-glutamyl transpeptidase activity; and disease severity. Initially, cysteine concentration was low, at levels similar to those of healthy controls. However, glutathione was reduced whilst cysteinyl glycine and gamma-glutamyl transpeptidase activity were increased in both mild and severe attacks. As the disease progressed, glutathione and cysteinyl glycine were further increased in mild attacks and cysteine levels correlated with homocysteine (r = 0.8, P < 0.001) and gamma-glutamyl transpeptidase activity (r = 0.75, P < 0.001). The progress of severe attacks was associated with glutathione depletion, reduced gamma-glutamyl transpeptidase activity, and increased cysteinyl glycine that correlated with glutathione depletion (r = 0.99, P = 0.01). These results show that glutathione depletion associated with severe acute pancreatitis occurs despite an adequate cysteine supply and could be attributed to heightened oxidative stress coupled to impaired downstream biosynthesis.