DIABETIC NEUROPATHY: RECEPTORS AND SIGNALING MECHANISM

糖尿病神经病：受体和信号传导机制

• 批准号: 3464494
• 负责人: LILIANA N BERTI-MATTERA
• 金额: $ 10.06万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-09-30 至 1997-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3464494
• 关键词: G protein; SDS polyacrylamide gel electrophoresis; Schwann cells; adenylate cyclase; adrenergic receptor; biological signal transduction; bradykinin; cell line; cyclic AMP; diabetic neuropathy; glucose; immunochemistry; laboratory rat; membrane transport proteins; muscarinic receptor; neuropeptide receptor; phospholipase C; potassium; receptor coupling; sodium; sodium potassium exchanging ATPase; stimulant /agonist; tissue /cell culture; western blottings

Diabetic neuropathy, one of the major complications of chronic diabetes is characterized by several biochemical alterations including a reduction in myo-inositol levels, decrease in Na+K+-ATPase activity and abnormalities in phosphoinositide (PPI) metabolism and adenylyl cyclase activity. The current hypothesis regarding the pathophysiology of diabetic neuropathy assign a central role to the decrease in myo-inositol uptake-dependent phosphoinositide turnover and hence availability of second messengers, like diacylglycerol which modulate Na+K+-ATPase activity through activation of protein kinase C. While it is generally accepted that the reduction in Na+K+-ATPase plays a key role in the development of neuropathy, its precise regulation is far from clear. Recent evidence from our and other labs suggests that peripheral nerve possesses muscarinic, bradikynin, purinergic and beta-adrenergic receptors which seem to be linked to stimulation of the breakdown of PPI or adenylyl cyclase. We have recently identified several types of guanine-nucleotide binding (G) proteins, that are known to be involved in other tissues in coupling between the agonist-receptor complexes and the adenylyl cyclase or phospholipase C systems. In nerve and myelin membranes from experimental diabetic animals we demonstrated that some of these signalling pathways are altered. With this evidence, we propose to: 1) characterize the relative levels of G-proteins in total and subcellular fractions of sciatic nerve from diabetic rats by ADP- ribosylation with bacterial toxins and immunoblotting; 2) investigate the receptor-linked signalling pathways that modulate the activity of phospholipase C and adenylyl cyclase and their influence on Na+K+-ATPase activity in sciatic nerve slices or membranes by: a)activation of phospholipase C through muscarinic, purinergic and bradikynin receptors b) modulation of adenylyl cyclase activity through activation of beta- adrenergic and muscarinic receptors: 3) evaluate how the alterations in Na+K+-ATPase in nerves from diabetic animals correlate with changes in the different components of the above mentioned signalling pathway and 4) determine if in cultured Schwann cells the above mentioned agonists are linked to cAMP and phospholipase C and if these pathways are altered under conditions that mimic diabetes like high glucose and myo-inositol free culture conditions.

糖尿病神经病变是慢性糖尿病的主要并发症之一 其特征在于几种生化改变，包括减少 在肌醇水平中，Na+K+-ATP酶活性降低， 磷酸肌醇（PPI）代谢和腺苷酸环化酶异常 活动 目前的假设，关于病理生理学的 糖尿病性神经病变对肌醇减少起中心作用 摄取依赖性磷酸肌醇周转， 第二信使，如调节Na+K+-ATP酶的甘油二酯 通过激活蛋白激酶C的活性。 虽然人们普遍 认为Na+-K+-ATP酶的减少在细胞凋亡中起着关键作用。 神经病变的发展，其精确的调节还远不清楚。 我们和其他实验室的最新证据表明，外周神经 具有毒蕈碱，bradikynin，嘌呤能和β-肾上腺素能 似乎与PPI分解刺激有关的受体 或腺苷酸环化酶。 我们最近发现了几种类型的 鸟嘌呤核苷酸结合（G）蛋白，已知涉及 在其他组织中，激动剂-受体复合物之间的偶联， 腺苷酸环化酶或磷脂酶C系统。 在神经和髓磷脂中 从实验糖尿病动物的膜，我们证明，一些 这些信号通路的改变。 有了这些证据，我们建议 1）表征总G蛋白的相对水平， ADP-2对糖尿病大鼠坐骨神经亚细胞组分的影响 核糖基化与细菌毒素和免疫印迹; 2）研究 受体相关的信号通路，调节的活动， 磷脂酶C和腺苷酸环化酶及其对Na ~+-K ~+-ATP酶影响 坐骨神经切片或膜中的活性，通过：a）激活 磷脂酶C通过毒蕈碱、嘌呤能和bradikynin受体 B）通过激活β-腺苷酸环化酶来调节腺苷酸环化酶活性， 肾上腺素能受体和毒蕈碱受体：3）评估 糖尿病动物神经Na+K+-ATP酶与糖尿病大鼠神经细胞膜Na+K+-ATP酶活性变化的关系 上述信号通路的不同组分， 4)确定在培养的雪旺细胞中，上述激动剂 与cAMP和磷脂酶C相连，如果这些途径被改变， 在类似糖尿病的条件下，如高葡萄糖和肌醇 自由培养条件。