Indiana University-Purdue University Indianapolis (IUPUI) / Insulin-stimulated glucose uptake into skeletal muscle cells requires translocation of the glucose transporter-4 (GLUT4) from the cell interior to the plasma membrane. Insulin-stimulated GLUT4 vesicle translocation is dysregulated in Type 2 diabetes (T2D). The Group I p21–activated kinase (PAK1) is a required element in insulin-stimulated GLUT4 vesicle translocation in mouse skeletal muscle in vivo, although its placement and function(s) in the canonical insulin signaling cascade in skeletal muscle cells, remain undetermined. Therefore, the objective of my project is to determine the molecular mechanism(s) underlying the requirement for PAK1 in the process of insulin-stimulated GLUT4 vesicle translocation and subsequent glucose uptake by skeletal muscle cells. Toward this, my studies demonstrate that the pharmacological inhibition of PAK1 activation blunts insulin-stimulated GLUT4 translocation and subsequent glucose uptake into L6-GLUT4myc skeletal myotubes. Inhibition of PAK1 activation also ablates insulin-stimulated F-actin cytoskeletal remodeling, a process known to be required for mobilizing GLUT4 vesicles to the plasma membrane. Consistent with this mechanism, PAK1 activation was also required for the activation of cofilin, another protein implicated in F-actin remodeling. Interestingly, my studies reveal a novel molecular mechanism involving PAK1 signaling to p41-ARC, a regulatory subunit of the cytoskeletal Arp2/3 complex, and its interactions with another cytoskeletal factor, N-WASP, to elicit the insulin-stimulated F-actin remodeling in skeletal muscle cells. Pharmacological inactivation of N-WASP fully abrogated insulin-stimulated GLUT4 vesicle translocation to the cell surface, coordinate with blunted F-actin remodeling. Furthermore, my studies revealed new insulin-induced interactions amongst N WASP, actin, p41-ARC and PAK1; inactivation of PAK1 signaling blocked these dynamic interactions. Taken together, the above studies demonstrate the significance of PAK1 and its downstream signaling to F-actin remodeling in insulin-stimulated GLUT4 vesicle translocation and glucose uptake, revealing new signaling elements that may prove to be promising targets for future therapeutic design.

Indiana University-Purdue University Indianapolis (IUPUI) / The extracellular matrix (ECM) governs many cellular processes through biochemical and mechanical cues. Particularly, the effect ECM mechanical properties on cells fate has been well established over the years. Many hydrogel systems have been used to mimic the dynamic stiffening processes occurring in ECM. However, changes in ECM stiffness does not fully recapitulate the mechanics of native ECM, as viscoelasticity is also a major factor contributing to ECM dynamic property. This thesis describes the design and characterization of an enzyme-crosslinked hydrogel system that is not only capable of being stiffened on demand, but also can be tuned to obtain viscoelasticity. The first objective of this thesis was to utilize horseradish peroxidase (HRP) to crosslink thiol-norbornene hydrogel and use mushroom tyrosinase (MT) to create secondary DOPA-dimer crosslinks that stiffened the hydrogel. The cytocompatibility of HRP-mediated thiol-norbornene gelation and the effect of stiffening on cell fate was evaluated. The second objective of this thesis represented the first step towards developing a hydrogel system whose viscoelasticity could be dynamically tuned. Thiol-norbornene hydrogel was designed to yield dynamically adaptable boronic ester bonds via partial enzymatic reaction. Thiol-norborne hydrogel was made to contain hydroxyl phenol as well as boronic acid residues within its network. MT, in this case was used to oxidize the hydroxy phenol moieties into DOPA, which then complexed with boronic acid, created dynamic bonds, introducing viscoelasticity to an initial elastic hydrogel.

Glycemic control in patients with type 2 diabetes (T2D) can be achieved through potentiation of the signalling by glucagon-like peptide 1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP). Both incretin hormones have been traditionally characterized to be secreted by distinct enteroendocrine cells within the gut in response to nutrients. Signalling through the incretin receptors stimulates islet hormone release by potentiating glucose-stimulated insulin secretion from the β-cell and decreasing glucagon secretion from the α-cell. However, the bioactivity of GLP-1 and GIP is controlled by post-translational, N-terminal cleavage by the widely expressed serine protease dipeptidyl peptidase 4 (DPP4). As such, DPP4 inhibitors (DPP4i) have been successfully used to treat millions of patients with T2D. DPP4i target the catalytic active site of DPP4 and prevent the cleavage of the incretin hormones, thus prolonging their action. Recently, studies in genetically modified mice have demonstrated that GLP-1 is not solely an intestinally-derived peptide hormone and proposed that islet-derived GLP-1 is required for proper glucose homeostasis. Therefore, with the current study, we sought to assess whether β-cell-derived DPP4 is an important target for the regulation of glycemia. Treatment of Glp1r/Gipr^(β-cell-/-) mice with the DPP4 inhibitor sitagliptin demonstrated that β-cell incretin receptor signaling is required to mediate the beneficial effects of this class of drugs on glucose homeostasis. Additionally, Dpp4^(-/-) mice exhibited a significant reduction in hepatic glucose production during hyperinsulinemic-euglycemic clamps. Dpp4 mRNA, DPP4 protein and activity are present in isolated mouse islets, further supporting the islet as an important potential site of DPP4i action. In this study, we show that both DPP4i-treated wildtype islets and islets isolated from Dpp4^(β-cell-/-) mice exhibit increased glucose-stimulated insulin secretion (GSIS) during perifusion after a high-fat diet feeding. Genetic elimination of Dpp4 from islet β-cells also improved oral glucose tolerance and insulin sensitivity in female mice, but had no effects on circulating DPP4 or incretin levels. Finally, eliminating Dpp4 from β-cells or the whole pancreas did not improve whole-body glucose tolerance, response to DPP4i, insulin tolerance, or body weight in male mice fed chow or a high-fat diet. Therefore, we provide evidence for islet-derived DPP4 to have a role in local hormone responses to glucose; however, its role in systemic glucose metabolism is shown to be sex-dependent.

Studies on rat islet preparations have shown melatonin inhibits MAO activity and thereby reduces glucose mediated insulin release. The objective of this work is to investigate the effects of melatonin on insulin release, and on glucose mediated insulin release in the intact and in the pinealectomized rat.

No description available.

Final article is available at "wileyonlinelibrary.com" Takako Okamoto, Masaki Mandai, Noriomi Matsumura, Ken Yamaguchi, Hiroshi Kondoh, Yasuaki Amano, Tsukasa Baba, Junzo Hamanishi, Kaoru Abiko, Kenzo Kosaka, Susan K. Murphy, Seiichi Mori, Ikuo Konishi "Hepatocyte nuclear factor-1β (HNF-1β) promotes glucose uptake and glycolytic activity in ovarian clear cell carcinoma" Molecular Carcinogenesis 54:35–49 (2013) / 京都大学 / 0048 / 新制・論文博士 / 博士(医学) / 乙第12804号 / 論医博第2076号 / 新制||医||1001(附属図書館) / 80848 / 京都大学大学院医学研究科医学専攻 / (主査)教授 野田 亮, 教授 武藤 学, 教授 稲垣 暢也 / 学位規則第4条第2項該当 / Doctor of Medical Science / Kyoto University / DFAM

京都大学 / 0048 / 新制・課程博士 / 博士(人間健康科学) / 甲第18197号 / 人健博第14号 / 新制||人健||2(附属図書館) / 31055 / 京都大学大学院医学研究科人間健康科学系専攻 / (主査)教授 藤井 康友, 教授 岡 昌吾, 教授 横出 正之 / 学位規則第4条第1項該当 / Doctor of Human Health Sciences / Kyoto University / DFAM

京都大学 / 0048 / 新制・論文博士 / 博士(農学) / 乙第12936号 / 論農博第2816号 / 新制||農||1033(附属図書館) / 学位論文||H27||N4895(農学部図書室) / 32146 / (主査)教授 伏木 亨, 教授 保川 清, 教授 安達 修二 / 学位規則第4条第2項該当 / Doctor of Agricultural Science / Kyoto University / DFAM

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第18853号 / 医博第3964号 / 新制||医||1007(附属図書館) / 31804 / 京都大学大学院医学研究科医学専攻 / (主査)教授 山田 泰広, 教授 武田 俊一, 教授 野田 亮 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

The objective of these studies was to design an optimal nutritional supplement suitable for grazing horses using fat and fiber to replace the grain and molasses in the traditional sweet feed. Thoroughbred mares and foals grazing bluegrass/clover pastures were used in these studies, twenty mares and their foals in 1994 to 1995, and twenty mares and foals in 1995 to 1996. Seasonal variation in pasture was examined, and the need for supplementation of nutrients and fibers was assessed. The nutritional status of grazing mares, foals, weanlings and yearlings, fed either a starch and sugar supplement (SS) or a fat and fiber supplement (FF), was examined using growth measurements, radiographic bone evaluations, milk composition and glucose tolerance tests. These studies suggest that fiber may be an important component of an ideal supplement for improved grass/legume pastures. Seasonal variation in pasture indicated an increase in hydrolyzable and rapidly fermed carbohydrates during periods of rapid growth. The FF supplement may have buffered seasonal changes and the increased hydrolyzable carbohydrate content in rapidly growing pasture, as evidenced by smoother growth curves in the yearlings. Young horses, after weaning until the following May, had lower estimated bone mineral content when fed the FF supplement. The lower bone mineral content in the FF supplemented horses may have been due to decreased absorption of calcium or metabolic and hormonal changes associated with adaptation to the different energy sources in the supplements. Milk composition of FF supplemented mares was influenced in ways likely to improve foal health. The FF supplemented mares had enhanced linoleic acid content, which may reduce the risk of gastric ulcers in foals, and increased immunoglobulin G concentration, which may enhance passive immunity. The carbohydrate status of mares, as assessed by glucose tolerance tests, indicated a slower glucose clearance that could be a metabolic adaptation of the mares to the SS and FF supplements. / Ph. D.