Factors that modify insulin action

改变胰岛素作用的因素

• 批准号: 7996761
• 负责人: MARIA G BUSE
• 金额: $ 9.52万
• 依托单位: MEDICAL UNIVERSITY OF SOUTH CAROLINA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-01-01 至 2010-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7996761
• 关键词: 1-Phosphatidylinositol 3-Kinase; Accounting; Actins; Address; Adenoviruses; Adipocytes; Affect; Alanine; Animal Model; Binding; Blood Vessels; Cells; Chronic; Complications of Diabetes Mellitus; Data; Development; Diabetes Mellitus; Disease; Distal; Dose; Enzymes; Epidemic; Event; Fatty acid glycerol esters; Fructose; Functional disorder; GRB2 gene; Glucocorticoids; Glucose; Glutamine; Goals; Hepatic; Hexosamines; Human; Hyperglycemia; In Vitro; Insulin; Insulin Receptor; Insulin Resistance; Insulin-Dependent Diabetes Mellitus; Insulin-Like Growth Factor I; Insulin-Like-Growth Factor I Receptor; Knock-out; Lead; Leptin; Liver; MAPK8 gene; Mass Spectrum Analysis; Mediating; Metabolic syndrome; Metabolism; Modeling; Modification; Modus; Molecular Target; Morbidity - disease rate; Mus; Muscle Fibers; Mutate; Non-Insulin-Dependent Diabetes Mellitus; Nonesterified Fatty Acids; Nutrient; O-GlcNAc transferase; Obesity; PTEN gene; PTEN protein; PTPN11 gene; Pathway interactions; Patients; Phosphorylation; Phosphorylation Site; Play; Polycystic Ovary Syndrome; Post-Translational Protein Processing; Prevalence; Process; Production; Protein phosphatase; Proteins; Rattus; Receptor Signaling; Role; Septicemia; Signal Transduction; Sirolimus; Site; Testing; Time; Toxic effect; UDP-glucosamine; Uremia; base; design; diabetic patient; glucose transport; human FRAP1 protein; improved; in vivo; insight; insulin signaling; insulin tolerance; interest; knock-down; liver metabolism; mortality; mutant; new therapeutic target; novel; overexpression; phosphatidylinositol 3,4,5-triphosphate; prevent; protein expression; public health relevance; research study; response; small hairpin RNA

DESCRIPTION (provided by applicant): "Glucose toxicity" accounts for insulin resistance in patients with uncontrolled type 1 diabetes and contributes to it in type 2 diabetes. It contributes to the vascular complications, the major causes of morbidity and mortality in diabetic patients. Sustained exposure of cells to high glucose increases flux via the hexosamine synthesis pathway, enhancing the production of UDP-N-acetyl glucosamine (UDP-GlcNAc), the substrate of O-GlcNAc transferase (OGT). OGT catalyzes the reversible, single addition of O-GlcNAc to specific Ser/Thr residues. O-GlcNAcylation and O-phosphorylation are often reciprocal. We have recently identified, for the first time, four sites of O-GlcNAcylation on IRS-1 (as well as 11 novel Ser/Thr phosphorylation sites) by mass spectrometry. Preliminary data suggest that O-GlcNAc may affect IRS-1 signal transduction. Our major objective in Spec. Aim 1 is to firmly establish whether the O-GlcNAc modification alters IRS-1 signaling. In in vitro studies Ser will be mutated to Ala at the four O-GlcNAc sites, singly and in combination, wild type and mutated IRS-1 will be expressed in HepG2 cells and in intact liver via adenovirus, and their interactions with IRS-1 binding partners in response to insulin or to IGF-1 studied. In in vivo studies endogenous mouse IRS-1 will be knocked out with shRNA adenovirus and substituted with wild type or mutant IRS-1-expressing adenovirus. The effect of these manipulations on glucose and insulin tolerance tests and the expression of hepatic gluconeogenic enzymes will be studied. In Spec Aim 2 studies will be continued in a model of high glucose/ low dose insulin-induced insulin resistance of glucose transport and Akt activation in 3T3-L1 adipocytes. Insulin signaling to PI(3)K is largely maintained, but Akt activation is markedly impaired. Recent data indicate that PTEN protein expression is increased and insulin stimulated PtdIns(3,4,5)P3 is diminished. Stimulation of PTEN expression is inhibited by rapamycin. mTORC-1 activation clearly plays a role, cPKC may contribute, but JNK does not. The mechanism of enhanced PTEN expression, mTORC-1 action and the role of cPKC will be investigated. Several mechanisms which may synergize with or be mediated by mTOR will be addressed, including dysregulation of actin dynamics and possible activation of a phosphoprotein phosphatase(s). The analysis of the modus operandi in this model will be contrasted with the insulin resistance of glucose transport elicited by exposing cells to FFA. Insights gained from this model will be applied to L-6 myotubes and to intact rats made insulin resistant by chronic hyperglycemia. Understanding how different excess nutrients modify insulin's signaling may lead to the rational development of novel therapeutic targets.

描述（由申请人提供）：“葡萄糖毒性”是1型糖尿病患者的胰岛素抵抗，并在2型糖尿病中促成它。它导致血管并发症，这是糖尿病患者发病和死亡率的主要原因。细胞持续暴露于高葡萄糖会增加通过己胺合成途径的通量，从而增强了O-GLCNAC转移酶（OGT）的底物（OGT）的UDP-N-乙酰基葡萄糖胺（UDP-GLCNAC）的产生。 OGT将O-GlCNAC单一添加到特定的SER/THR残基中催化可逆的单一添加。 O-Glcnacylation和O磷酸化通常是相互的。我们最近首次确定了四个站点 IRS-1（以及11个新型Ser/Thr磷酸化位点）上的O-Glcnacylation通过质谱法。初步数据表明O-GLCNAC可能会影响IRS-1信号转导。我们的主要目标。 AIM 1是牢固确定O-GLCNAC修改是否会改变IRS-1信号传导。在体外研究中，SER将在四个O-GLCNAC位点突变为ALA，单独，组合，野生型和突变的IRS-1将在HEPG2细胞和完整的肝脏中通过腺病毒来表达，以及它们与IRS-1的相互作用，以及与IRS-1的相互作用，响应于胰岛素或对IGF-1研究的响应。在体内研究中，内源性小鼠IRS-1将用shRNA腺病毒敲除，并用野生型或突变体取代 表达IRS-1的腺病毒。这些操纵对葡萄糖和胰岛素耐受性测试的影响以及肝糖原性酶的表达。在SPEC AIM 2中，将在3T3-L1脂肪细胞中的高葡萄糖/低剂量胰岛素诱导的胰岛素耐药性和AKT激活的模型中继续进行研究。胰岛素对PI（3）K的信号在很大程度上维持，但AKT激活显着受损。最近的数据表明，PTEN蛋白表达增加，胰岛素刺激的PTDINS（3,4,5）P3减少。雷帕霉素抑制PTEN表达的刺激。 MTORC-1激活显然起着作用，CPKC可能会贡献，但JNK却没有。将研究增强PTEN表达，MTORC-1作用和CPKC作用的机制。将解决可能与MTOR协同或介导的几种机制，包括肌动蛋白动力学的失调以及可能激活磷酸蛋白磷酸酶（S）。在该模型中对操作方法的分析将与通过将细胞暴露于FFA引起的葡萄糖转运的胰岛素抵抗形成对比。从该模型中获得的见解将应用于L-6肌管，并通过慢性高血糖制成胰岛素抗药性的完整大鼠。了解不同的过量营养如何改变胰岛素的信号传导可能会导致新型治疗靶标的合理发展。