Regulation of Elevated Postexercise Insulin-stimulated Glucose Uptake by Skeletal Muscle

运动后骨骼肌对胰岛素刺激的葡萄糖摄取升高的调节

• 批准号: 10834392
• 负责人: Gregory D. Cartee
• 金额: $ 12.48万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-07-01 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10834392
• 关键词: 5' -AMP-activated protein kinase; Address; American; Biological; Blood Glucose; Carbohydrates; Cell surface; Coupling; Defect; Enzymes; Estrogen Antagonists; Estrogen Receptors; Estrogens; Event; Exercise; Exocytosis; GLUT 4 protein; GTPase-Activating Proteins; Glucose; Glucose Transporter; Glycogen; Glycogen (Starch) Synthase; Goals; Guanine Nucleotide Exchange Factors; Health Benefit; Hormones; Insulin; Insulin Resistance; Intracellular Membranes; Knock-out; Knowledge; Mediating; Membrane; Metabolic; Methods; Microscopy; Mission; Modeling; Molecular; Muscle; Mutation; Non-Insulin-Dependent Diabetes Mellitus; Outcome; Phosphorylation; Physiological; Prediabetes syndrome; Process; Proteins; Protocols documentation; Public Health; Rattus; Receptor Inhibition; Regulation; Research; Resolution; Rest; Risk; Role; Sampling; Sex Differences; Site; Skeletal Muscle; Tertiary Protein Structure; Testing; Thinness; United States National Institutes of Health; Work; adeno-associated viral vector; delivery vehicle; design; experimental study; feeding; genetic approach; glucose disposal; glucose uptake; high risk; innovation; male; novel therapeutics; prevent; rab GTP-Binding Proteins; rational design; recruit; sedentary; sex; sexual dimorphism; small hairpin RNA; therapy design

Over 100 million Americans suffer from the devastating consequences of type 2 diabetes (T2D) or prediabetes (a condition associated with elevated risk to develop T2D). Skeletal muscle accounts for up to 85% of insulin- induced blood glucose clearance, and insulin resistance for muscle glucose uptake is an essential, and perhaps primary defect for T2D. It has been known for 40 years that one exercise bout can enhance subsequent insulin- stimulated glucose uptake (ISGU) by muscle, but the mechanisms have remained elusive. The long-range goal is to fully understand the molecular, cellular, and physiological events responsible for this significant health benefit. Recent research using a unique Akt substrate of 160 kDa-knockout (AS160-KO) rat model revealed that expression of AS160 (a key regulator of GLUT4 glucose transporter localization) is essential for the elevated postexercise ISGU. Specific Aim 1 will identify mechanisms whereby AS160 leads to greater postexercise ISGU by muscle. AS160’s canonical Rab-GAP (Rab-GTPase activating protein) domain controls ISGU under sedentary conditions. Experiments using AS160-KO rats with AAV-vector (AAV) to deliver AS160 with a mutation to selectively disable its Rab-GAP domain will test if this domain is required for elevated postexercise ISGU. Although AS160 expression is essential for elevated postexercise ISGU in both sexes, AS160 phosphorylation of key sites is required only for male rats. Experiments will test estrogen’s role in the mechanisms responsible for this important sexual dimorphism. Specific Aim 2 will elucidate the regulation of subcellular GLUT4 localization in skeletal muscle postexercise. Knowledge of AS160’s role in GLUT4 distribution is limited to insulin’s ability to elevate GLUT4 exocytosis to cell surface membranes in unexercised muscle. A powerful new microscopy-based approach (STERM, Sample Thinning Enhanced Resolution Microscopy) will examine GLUT4 distribution in 7 different myocellular compartments. Coupling STERM with the AS160-KO model will be used to test if AS160 is crucial for postexercise regulation of GLUT4 distribution in both cell surface and intracellular membrane compartments. Specific Aim 3 will ascertain the role of postexercise muscle glycogen resynthesis in the reversal of the postexercise increase in ISGU. Because the health benefit of elevated postexercise ISGU could be extended by delaying its reversal, elucidating the mechanism for reversal of elevated ISGU would be valuable. Experiments will test if muscle glycogen resynthesis is crucial for reversal of elevated postexercise ISGU by muscle with carbohydrate refeeding. The innovative approach will be to reduce muscle abundance of glycogen synthase (GS, rate-limiting enzyme for glycogen synthesis) using AAV-vector delivery of shRNA-GS to muscle. Further analysis will seek to identify mechanisms underlying the relationship between muscle glycogen and reversal of elevated postexercise ISGU. The research in this project will use rigorous and innovative methods to enable significant advances in fundamental knowledge and address a critical barrier to progress in the field by elucidating mechanisms for elevated postexercise ISGU, a major health benefit.

超过 1 亿美国人遭受 2 型糖尿病 (T2D) 或糖尿病前期的毁灭性后果 （一种与罹患 T2D 风险升高相关的疾病）。骨骼肌占胰岛素的85% 诱导血糖清除，而胰岛素抵抗对于肌肉葡萄糖摄取是至关重要的，也许 T2D 的主要缺陷。 40 年来，人们都知道一次锻炼可以增强随后的胰岛素水平 肌肉刺激葡萄糖摄取（ISGU），但其机制仍然难以捉摸。长远目标 是要充分了解导致这一重要健康的分子、细胞和生理事件 益处。最近使用 160 kDa 敲除 (AS160-KO) 大鼠模型的独特 Akt 底物进行的研究表明 AS160（GLUT4 葡萄糖转运蛋白定位的关键调节因子）的表达对于升高 运动后 ISGU。具体目标 1 将确定 AS160 导致更大运动后 ISGU 的机制 通过肌肉。 AS160 的规范 Rab-GAP（Rab-GTP 酶激活蛋白）结构域控制 ISGU 久坐的条件。使用带有 AAV 载体 (AAV) 的 AS160-KO 大鼠来传递带有突变的 AS160 的实验 有选择地禁用其 Rab-GAP 域将测试该域是否是提升的运动后 ISGU 所必需的。 尽管 AS160 表达对于男女运动后 ISGU 的升高至关重要，但 AS160 磷酸化 仅雄性大鼠需要关键位点。实验将测试雌激素在相关机制中的作用 对于这一重要的性别二态性。具体目标 2 将阐明亚细胞 GLUT4 的调节 运动后骨骼肌的定位。关于 AS160 在 GLUT4 分布中的作用的知识仅限于 胰岛素在未运动的肌肉中提高 GLUT4 细胞表面膜胞吐作用的能力。一个强大的新 基于显微镜的方法（STERM，样品稀释增强分辨率显微镜）将检查 GLUT4 分布在7个不同的肌细胞区室中。 STERM 与 AS160-KO 模型的耦合将用于 测试 AS160 是否对于运动后调节细胞表面和细胞内的 GLUT4 分布至关重要 膜隔室。具体目标 3 将确定运动后肌糖原再合成在 运动后 ISGU 增加的逆转。因为运动后 ISGU 升高对健康有益 可以通过延迟其逆转来延长，阐明升高的 ISGU 逆转的机制将是 有价值的。实验将测试肌糖原再合成对于逆转运动后升高是否至关重要 ISGU 通过肌肉补充碳水化合物。创新方法是减少肌肉丰度 使用 shRNA-GS 的 AAV 载体递送糖原合成酶（GS，糖原合成的限速酶） 到肌肉。进一步的分析将寻求确定肌肉之间关系的潜在机制。 糖原和运动后 ISGU 升高的逆转。该项目的研究将采用严格和 创新方法，使基础知识取得重大进展，并解决关键障碍 通过阐明运动后 ISGU 升高的机制（一项主要的健康益处），该领域取得了进展。