GLUCOSE TOXICITY IN SKELETAL MUSCLE

与糖尿病相关的高血糖加重了胰岛素抵抗 通过葡萄糖对骨骼肌的直接毒性作用。这反过来 导致血糖控制恶化， 发病率葡萄糖的毒性作用机制 骨骼肌是未知的，但最终结果是降低敏感性， 肌肉Glut 4葡萄糖转运蛋白转化为胰岛素。一系列转基因小鼠 已经开发了一种结构型Glut 1葡萄糖转运蛋白， 在骨骼肌中特异性过表达。基础葡萄糖转运是 在这些小鼠的肌肉中， 肌膜Glut 1过表达。增加的基础通量 葡萄糖进入肌肉模拟高血糖症的影响， Glut 4的脱敏作用。这些小鼠提供了一个独特的模型系统， 研究肌肉葡萄糖毒性在体内的直接影响， 无高血糖和循环改变引起的继发性效应 胰岛素水平最近的证据表明，葡萄糖流量的增加 通过葡萄糖胺代谢途径可能是负责葡萄糖- 诱导的骨骼肌胰岛素抵抗。长期目标是 项目是利用这种转基因小鼠模型来帮助阐明 葡萄糖诱导的骨骼肌胰岛素抵抗的机制。到 为实现这一目标，我们提出以下具体目标： L.为了确定胰岛素和收缩诱导的易位是否 Glut 4到质膜的缺陷是肌肉中的Glut 1 用定量免疫金电子显微镜观察转基因小鼠。 2.为了确定体内细胞内游离葡萄糖的升高 影响骨骼肌基因表达模式， 可能导致葡萄糖毒性的新基因。这将是 通过使用逆转录差异mRNA展示来完成 Glut 1转基因骨骼肌的分析。 3.为了确定葡萄糖胺代谢的改变是否涉及 Glut 1转基因小鼠中Glut 4活化缺陷。这将是 通过研究氨基葡萄糖对Glut 4的直接影响 易位在肌肉中，测量葡萄糖胺代谢物的水平， 肌肉的Glut 1转基因小鼠，并检查葡萄糖胺的作用 对葡萄糖调节的候选基因表达的影响， 具体目标#2。过表达谷氨酰胺：果糖-6-磷酸转基因小鼠 肌肉中的磷酸酰胺转移酶将产生并表征 为了直接检验葡萄糖胺增加 代谢是葡萄糖诱导的胰岛素抵抗的原因， 骨骼肌

The hyperglycemia associated with diabetes exacerbates insulin resistance via a direct toxic effect of glucose on skeletal muscle. This in turn contributes to the worsening of glycemic control and the development of morbidity. The mechanism by which glucose exerts a toxic effect on skeletal muscle is unknown, but the end result is decreased sensitivity of the muscle Glut4 glucose transporter to insulin. A line of transgenic mice has been developed in which the constitutive Glut1 glucose transporter is overexpressed specifically in skeletal muscle. Basal glucose transport is dramatically elevated in the muscle of these mice via the constitutive overexpression of Glut1 in the sarcolemma. The increased basal flux of glucose into muscle mimics the effect of hyperglycemia and results in the desensitization of Glut4. These mice provide a unique model system for studying the direct effects of muscle glucose toxicity in vivo in the absence of secondary effects due to hyperglycemia and altered circulating insulin levels. Recent evidence suggests that increased flux of glucose through the glucosamine metabolic pathway may be responsible for glucose- induced insulin resistance in skeletal muscle. The long-term goal of this project is to exploit this transgenic mouse model to help elucidate the mechanism of glucose-induced insulin resistance in skeletal muscle. To accomplish this goal, we propose the following specific aims: l. To determine whether insulin and contraction-induced translocation of Glut4 to the plasma membrane are defective in the muscle of Glut1 transgenic mice using quantitative immunogold electron microscopy. 2. To determine how elevation of intracellular free glucose in vivo affects the pattern of skeletal muscle gene expression and to identify novel genes that may contribute to glucose toxicity. This will be accomplished by the use of reverse transcription differential mRNA display analysis of the Glut1 transgenic skeletal muscle. 3. To determine whether altered glucosamine metabolism is involved in the defect in Glut4 activation in the Glut1 transgenic mice. This will be explored by investigating the direct effect of glucosamine on Glut4 translocation in muscle, measuring levels of glucosamine metabolites in muscle of Glut1 transgenic mice, and examining the effect of glucosamine on the expression of glucose-regulated candidate genes identified in specific aim #2. Transgenic mice overexpressing glutamine:fructose-6- phosphate amidotransferase in muscle will be generated and characterized in order to directly test the hypothesis that increased glucosamine metabolism is responsible for glucose-induced insulin resistance in skeletal muscle.