Browsing by Subject "Glucose metabolism"

Now showing 1 - 3 of 3
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    A novel role for the pro-apoptotic protein Noxa in survival and glucose metabolism
    (2012-12) Lowman, Xazmin Helen
    The studies described in this dissertation focus on a novel mechanism of post-translational control as well as a novel pro-survival role for a canonical tumor-suppressor BH3-only protein, Noxa, in human hematopoietic cells and cancers. An investigation into regulatory mechanisms underlying the constitutive expression of this pro-apoptotic Bcl-2 family member in leukemias, led to the identification of a phosphorylated serine on the protein. The phospho-modification suppressed its pro-apoptotic function and was regulated by glucose levels. Glucose deprivation promoted Noxa dephosphorylation and activated its apoptotic function. The atypical cyclin dependent kinase, Cdk5, was identified as the glucose-sensitive Noxa kinase. Further investigation pointed to a role for phospho-modified Noxa in glucose metabolism; cells over-expressing Noxa increased their uptake of glucose and promoted its utilization in anabolic metabolic pathways that are favored in proliferating cells. Finally, studies showed Noxa and its anti-apoptotic binding partner Mcl-1 associated within cytosolic multi-protein complexes in proliferating T-leukemia and activated primary T-cells. Early characterization and identification of the components point to a novel glucose-responsive signaling role for these complexes. Taken together, these studies offer fresh insights into cancer cell metabolism and the regulation of Bcl-2 proteins and are expected to contribute to novel therapeutic strategies aimed at activating Noxa's pro-apoptotic function in leukemias.
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    The Role of Hepatocyte D-Β-Hydroxybutyrate Dehydrogenase In Ketone Body Metabolism And Liver Health
    (2021-09) Stagg, David
    Throughout the last decade, interest has intensified in intermittent fasting, ketogenic diets, and exogenous ketone therapies as prospective health-promoting, therapeutic, and performance-enhancing agents. However, the regulatory roles of ketogenesis and ketone metabolism on liver homeostasis remain unclear. This thesis seeks to develop a better understanding of the metabolic consequences of hepatic ketone body metabolism by focusing on the redox-dependent interconversion of acetoacetate (AcAc) and D-β-hydroxybutyrate (D-βOHB). Using targeted and isotope tracing high-resolution liquid chromatography-mass spectrometry, dual stable isotope tracer nuclear magnetic resonance spectroscopy-based metabolic flux modeling, dietary-induced mouse models of nonalcoholic fatty liver disease (NAFLD), and complementary physiological approaches in novel cell type-specific knockout mice, the roles of hepatocyte D-β-hydroxybutyrate dehydrogenase (BDH1), a mitochondrial enzyme required for NAD+/NADH-dependent oxidation/reduction of ketone bodies, are quantified. Exogenously administered AcAc is reduced to D-βOHB, and increases hepatic NAD+/NADH ratio, reflecting hepatic BDH1 activity. Livers of hepatocyte-specific BDH1 deficient mice produced no D-βOHB, but due to extrahepatic BDH1, these mice nonetheless remained capable of AcAc/D-βOHB interconversion. Compared to littermate controls, hepatocyte specific BDH1 deficient mice, maintained on either a chow or NAFLD-inducing western-style diet, showed diminished liver tricarboxylic acid (TCA) cycle flux and impaired gluconeogenesis but normal overall hepatic energy charge. Furthermore, the livers of knockout mice maintained on a 60% high fat diet were less fibrotic, with reduced markers of oxidative stress, than littermate controls. Collectively, this thesis illustrates how ketone bodies and BDH1 activity influence liver homeostasis and health. While liver BDH1 is not required for whole body equilibration of AcAc and D-βOHB, loss of the ability to interconvert these ketone bodies in hepatocytes results in impaired TCA cycle flux and glucose production, with a beneficial effect on liver fibrosis. Therefore, BDH1 is a significant contributor to hepatic mitochondrial redox, liver physiology, and organism-wide ketone body homeostasis, and augmentation of hepatic BDH1 activity could prove beneficial in the treatment of NAFLD.
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    Triacylglycerol hydrolysis and fatty acid partitioning in the liver
    (2013-05) Tholen, Jillian Theresa
    The liver is a vital organ for the homeostatic control of both carbohydrate and lipid metabolism, functioning as a major regulatory site of energy storage, processing, and transformation. This project focused on liver lipid metabolism--it's production and trafficking of very low density density lipoproteins in particular. Specifically, we investigated whether VLDL trafficking is affected by 1) glucose and/or insulin, and 2) source of fatty acid (de novo or exogenous). As these were two distinct questions, each required its own sub-study, both of which are outlined separately below. Although it is well known that glucose and insulin influence the metabolism of fatty acids (FA) and the secretion of very low-density lipoprotein (VLDL), little is understood about how they regulate the partitioning of FA hydrolyzed from hepatic stored triacylglycerol (TAG) to different metabolic pathways. The aims of this study were to investigate the effects of differing concentrations of glucose and insulin on TAG hydrolysis and the subsequent partitioning of FA to pathways of secretion and oxidation in primary hepatocytes. We utilized primary mouse hepatocytes for pulse-chase experiments using [1-14C] oleate to examine the turnover and partitioning of the labeled FA. In addition to looking at the effects of differing glucose and insulin concentrations, we wanted to investigate whether any effects differed following acute (6 h) vs. chronic (24 h) treatment. Although we did not observe any acute effects of either glucose or insulin on TAG hydrolysis or partitioning of hydrolyzed FA, chronic insulin exposure decreased both FA oxidation and TAG secretion. These results suggest that insulin and glucose do not acutely influence TAG hydrolysis or channeling of hydrolyzed FA, but longer exposure to insulin reduces FA oxidation and secretion. There is a great deal of evidence suggesting that before being incorporated into VLDL for secretion, hepatic FA are first directed to storage in the cytosolic lipid droplet pool, and must subsequently be mobilized and channeled to assembly into VLDL. It remains unclear, however, whether the liver stores FA from different sources in distinct lipid droplet pools. Furthermore, should distinct storage pools exist, it is unknown whether they are preferentially channeled to different pathways, such as VLDL assembly. We performed experiments utilizing [1-14C] acetate and [1-14C] palmitate to represent FA derived from de novo lipogenesis and exogenous uptake, respectively. There was no difference between percentage of cellular acetate (via conversion to long chain FA) or palmitate secreted into the media at 4, 8, and 16 h in primary hepatocytes. However, we consistently saw a greater percentage of the label from acetate (de novo FA) secreted into the media after 2 h. Following the same 2 h labeling period in primary hepatocytes, we observed no difference between the partitioning of [1-14C] acetate and [1-14C] palmitate to the microsomal (ER) and lipid droplet fractions. Finally, to examine in vivo partitioning of the different FA, we injected mice with labeled FA and isolated their livers to quantify the contribution of the respective FA to cellular TAG and phospholipids. There was no difference in the incorporation of differentially sourced FA in liver TAG or phospholipid fractions. The results of these studies suggest that given the choice between FA taken up from exogenous sources and FA synthesized de novo, the liver may acutely preferentially secrete FA from de novo synthesis, however more work is needed to confirm this finding.