GLP-1通过TRPM2对胰岛素分泌调节作用的研究

It is generally known that both insulin biosynthesis and exocytosis contribute to insulin secretion, and cytosolic calcium plays an important role in this process. As a Ca2+ permeable channel expressed on beta cell, TRPM2, which can be activated by cAMP, is responsible for the regulation of insulin secretion. In response to GLP-1 treatment, insulin secretion of beta cell will be changed follow a concentration dependent manner. It means higher glucose concentration can result in stimulating release of insulin, and vice versa. Extracellular glucose concentration change will alter the level of cytosolic ATP, which influence intracellular cAMP suggest that the modulatory effect of GLP-1 on insulin secretion may be related with TRPM2. Our preliminary experiment demonstrate that knock out of TRPM2 abolished the GLP-1 induced enhancement of insulin secretion, implying the relationship between TRPM2 and GLP-1 mediated insulin secretion process. So we assume that ligation of GLP-1 and GLP-1 receptor stimulate the activation of cAMP, who bind with PKA subunit to produce insulin and storing it in the granules as well as activating TRPM2 to increase Ca2+ influx that promote exocytosis and eventually lead to augmentation of insulin secretion level. To support our hypothesis, we will record the current of TRPM2 as well as gene and protein expression level under GLP-1 treatment in different glucose concentration. We also will observe the effects of TRPM2 on insulin secretion by controlling its activity and expression, and use PKA inhibitor to confirm the relationship between and TRPM2 at a deeper level. This work put GLP-1 and TRPM2 relates in together as a new perspective to discover and reveal the mechanism of insulin secretion, and it provide us a new thinking of the prevention and theraphy for diabetes and its complications.

胰岛素分泌过程中，钙离子的调节作用必不可少。beta细胞上表达的钙离子通过性阳离子通道TRPM2可被cAMP途径激活，有研究认为它参与了葡萄糖代谢和胰岛素分泌过程。GLP-1可调节beta细胞胰岛素分泌水平，这种作用呈葡萄糖浓度依赖性。细胞外葡萄糖浓度变化引发的细胞内ATP乃至cAMP水平的变化提示GLP-1的这种调节胰岛素分泌作用可能通过TRPM2途径。前期试验表明TRPM2基因敲除后GLP-1的调节作用消失。为进一步验证，我们将对不同葡萄糖浓度水平下GLP-1刺激对TRPM2的电流变化、基因及蛋白质的表达水平，以及胰岛素分泌情况进行研究。并通过调节TRPM2的活性状态及表达水平，研究其对胰岛素分泌的影响。最后利用PKA抑制剂，进一步研究GLP-1与TRPM2的相互关系。本研究将结合GLP-1和TRPM2作为新视点探索并揭示胰岛素分泌调节的发生机制，为糖尿病及其并发症的防治开辟新的道路。

瞬时受体电位M2型（TRPM2）通道是一种非选择性的阳离子通道，钙离子可通过其流入细胞内。TRPM2可以被环磷酸腺苷（cAMP）激活，在胰岛beta细胞有表达，并且对胰岛素分泌起到了调节作用。众所周知，胰高血糖素样肽-1（GLP-1）能促进葡萄糖刺激引发的胰岛素分泌（GSIS），同时细胞外葡萄糖浓度变化会导致细胞内的三磷酸腺苷（ATP）乃至cAMP水平的变化。以上的这些研究结果促使我们假设TRPM2可能作为一个调节因素参与了GLP-1的胰岛素分泌调节作用。我们的实验结果表明，TRPM2基因静默会使GLP-1的胰岛素分泌促进作用失效，这意味着TRPM2参与了此环节。我们利用GLP-1以及不同浓度的葡萄糖刺激胰岛beta细胞，记录TRPM2的电流，并且测定胰岛素分泌水平，对结果分析后发现GLP-1的葡萄糖刺激引发的胰岛素分泌的调节效应是通过TRPM2完成的。另外，可以通过抑制TRPM2的活性或降低其表达水平影响GLP-1介导的GSIS的效果。我们利用特异性抑制剂或激活剂证明了GLP-1受体下游的两种主要通路Epac和PKA通过不同的机制影响了GLP-1的GSIS调节作用。综上所述，我们的研究揭示了TRPM2在GLP-1调节胰岛素分泌过程中的作用，为糖尿病及其并发症的预防和治疗提供了新思路。