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)或糖尿病前期的破坏性后果 (这种情况与患T2D的风险增加有关)。骨骼肌占胰岛素的85%- 诱导血糖清除和胰岛素抵抗对于肌肉葡萄糖摄取是必不可少的，而且可能 T2D的主要缺陷。40年来，人们一直知道，一次锻炼可以提高随后的胰岛素水平。 肌肉刺激葡萄糖摄取(ISGU)，但其机制仍不清楚。远景目标 是充分了解导致这种重大健康状况的分子、细胞和生理事件 利益。最近使用160 kDa基因敲除(AS160-KO)大鼠模型的独特Akt底物的研究表明， GLUT4葡萄糖转运体定位的关键调节因子AS160的表达对 运动后ISGU。具体目标1将确定AS160导致运动后ISGU更大的机制 用肌肉。AS160‘S Rab-Gap(Rab-GTP酶激活蛋白)结构域控制ISGU 久坐的状态。携带AAV载体的AS160-KO大鼠携带突变AS160的实验研究 为了选择性地禁用其RAB-GAP结构域，将测试运动后ISGU是否需要该结构域。 尽管AS160的表达是运动后男女ISGU升高的关键，但AS160的磷酸化 只有雄性老鼠才需要关键部位的数量。实验将测试雌激素在相关机制中的作用 因为这一重要的性二态。特异性目标2将阐明亚细胞GLUT4的调节 运动后骨骼肌的定位。对AS160‘S在GLUT4分发中的作用的了解仅限于 胰岛素在未经锻炼的肌肉中将GLUT4胞吐提升到细胞表膜的能力。一个强大的新技术 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 分布于7个不同的肌细胞室。将STERM与AS160-KO型号相结合将用于 测试AS160是否对运动后细胞表面和细胞内GLUT4分布的调节起关键作用 隔膜隔间。具体目标3将确定运动后肌肉糖原再合成在 运动后ISGU增加的逆转。因为运动后高强度运动对健康的益处 可以通过延迟其逆转来延长，阐明ISGU升高的逆转机制将是 很有价值。实验将测试肌肉糖原重新合成是否对运动后升高的逆转至关重要 ISGU由肌肉与碳水化合物重新摄取。创新的方法将是减少肌肉丰度 糖原合成酶(GS，糖原合成限速酶)利用AAV载体递送shRNA-GS 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 糖原与运动后ISGU升高的逆转。本项目的研究将使用严谨和 创新方法，使基础知识取得重大进展，并解决 通过阐明运动后ISGU升高的机制在该领域的进展，ISGU是一种主要的健康益处。