自噬在凝血酶预处理诱导脑缺血耐受过程中的作用及机制研究

项目名称： 自噬在凝血酶预处理诱导脑缺血耐受过程中的作用及机制研究

项目编号： No.81501007

项目类型： 青年科学基金项目

立项/批准年度： 2016

项目学科： 医药、卫生

项目作者： 胡枢坤

作者单位： 复旦大学

项目金额： 17.5万元

中文摘要： 脑缺血后继发性脑损伤是病情加重以及影响预后的重要原因。在中枢神经系统，缺血、缺氧等预处理都可以诱导脑缺血的耐受。缺血预处理的脑保护作用可以通过诱导自噬实现。凝血酶预处理也有脑保护作用，也可以诱导自噬，我们设想，凝血酶预处理诱导的脑保护作用也是通过自噬实现的。.本研究采用动物实验随机对照的方法，应用自噬活性药物，调节自噬的活性，评估自噬活性变化对凝血酶预处理脑保护效应的影响。在此基础上，深一步研究其中的病理机制。凝血酶预处理减轻脑缺血后脑损伤过程中上调了热休克蛋白27和32，P44/42 MAPK和JNK细胞信号传导通路也在此过程中被激活。那么，自噬调节凝血酶预处理脑保护作用的过程是否和上述蛋白有关？是否通过上述细胞信号传导通路起作用？需要我们进一步的验证。通过本研究，我们预期：自噬是凝血酶预处理诱导脑缺血耐受的关键因素；自噬调节凝血酶预处理脑保护作用的过程是通过相关细胞信号传导通路实现的。

中文关键词： 脑缺血；脑损伤；凝血酶；自噬

英文摘要： Due to secondary brain injury, ischemic stroke could cause severe functional defects like hemiparesis. Pretreatment with a low dose of thrombin attenuates the brain edema and significantly reduces the infarct size in the rat middle cerebral artery occlusion model. Interestingly, autophagy occurs after thrombin treatment in the brain. .Our preliminary data showed that cortical co-injection of thrombin and 3-methyladenine (3MA, autophagy inhibitor) aggravated the cerebral infarction. In contrast, co-injection of thrombin and rapamycin (autophagy agonist) reduced the infarct size after cerebral ischemia. These results indicated that the autophagy activities were negatively correlated with the extent of ischemic brain damages. We thereby hypothesized that the protective effect of thrombin preconditioning after ischemic stroke was achieved through the autophagy..In this study, we will investigate the relations between autophagy activities and the extent of brain damage after thrombin preconditioning. The autophagy activities, regulated by 3MA and rapamycin, respectively, will be assessed by monitoring LC3 I/II conversion rate, beclin1 activities, and cellular ultrastructure. Infarct size, brain edema and neurological deficits will be applied to evaluate brain damage. Meanwhile, we will measure the changes of the heat shock protein (HSP)27 and HSP32, activated P44/42 mitogen-activated protein kinase (MAPK) and c-Jun amino-terminal kinase (JNK), which were reported to be up-regulated by thrombin preconditioning. Furthermore, to clarify the roles of MAPK and JNK cell signaling pathways in thrombin preconditioned ischemia, we will evaluate the brain damages after the level of P44/42 MAPK and/or JNK are inhibited..Our studies will provide novel insights into the role and mechanisms of the autophagy in thrombin preconditioning induced cerebral ischemic tolerance and thereby shed lights on the studies of clinical treatment and therapeutic targets for ischemic stroke.

英文关键词： brain ischemia;brain injury;thrombin;autophagy