基于针刺对脑出血神经重塑作用的内质网应激与自噬稳态调控机制研究

A large number of neurons after cerebral hemorrhage missing outcomes directly affect the prognosis of the disease, how to actively and effectively repair damaged neurons, restore nerve function , is the key to prevention and treatment of disease. Recent studies have shown that autophagy as a new way of programmed cell death, "live" and "destruction" in cells is of great significance in the switch. The endoplasmic reticulum stress -Africa unfolded protein response (UPR) -induced autophagy is an important signaling pathways. Based on a new scientific discovery, and the early stage of the research evidence about the clinical and laboratory studies of acupuncture treatment of cerebral hemorrhage, this project intends to duplicate the cerebral hemorrhage rat model, using real-time fluorescent quantitative PCR, western blot, electron microscope technology, through the dynamic observation of acupuncture on cerebral hemorrhage after the induction of autophagy and the effect of endoplasmic reticulum stress from UPR two signal transduction pathways PERK/elF2 alpha, IRE1 molecular pathways into the mechanism of action of autophagy after cerebral hemorrhage and acupuncture reduce bleeding in the brain injury may be material foundation and effective targets, and then verify "acupuncture stimulating - positive feedback or negative feedback regulation of UPR - induced the moderate activation of autophagy - nerve cells endoplasmic reticulum stress and the steady state reconstruction of autophagy - autologous nerve cells to repair start is acupuncture promoting cerebral hemorrhage nerve repair and reshape one of the key mechanisms of" scientific hypothesis.

脑出血后如何积极有效地修复损伤的神经元，恢复其神经功能，是防治该病的关键。新近研究表明，自噬作为新的程序性细胞死亡方式，在细胞"存活"与"毁灭"切换中的意义重大。而内质网应激中非折叠蛋白质反应(UPR)是诱导细胞自噬的重要信号通路。基于这一新的科学发现，以及课题组前期有关针刺治疗脑出血的临床与实验室研究证据，本项目拟复制脑出血大鼠模型，采用实时荧光定量PCR、western blot、电镜等技术，通过动态观察针刺对脑出血后内质网应激诱导自噬的影响，从UPR两条信号转导通路PERK/elF2α、IRE1的分子作用途径深入阐释脑出血后自噬的作用机制以及针刺减轻脑出血损伤的可能物质基础与有效作用靶点，进而验证"针刺刺激-正反馈或负反馈调控UPR-诱导神经细胞自噬的适度激活-内质网应激与自噬的稳态重构-神经细胞自体修复启动是针刺促进脑出血神经修复与重塑的关键作用机制之一"的科学假说。

自噬作为新的程序性细胞死亡方式在细胞"存活"与"毁灭"切换中意义重大，而内质网应激中非折叠蛋白质反应(UPR)是诱导细胞自噬的重要分子机制。基于这一新的科学发现以及课题组前期有关针刺治疗脑出血的临床与实验室研究证据，本项目制备脑出血大鼠模型，采用实时荧光定量PCR、western blot技术，通过动态观察针刺对脑出血后内质网应激诱导自噬的影响，从UPR两条信号转导通路PERK/elF2α、IRE1的分子作用途径深入阐释脑出血后自噬的作用机制以及针刺减轻脑出血损伤的可能物质基础与有效作用靶点。结果表明：（1）PERK/EIF2通路的正反馈因子相关蛋白PERK、p-EIF2、ATF4、XBP1在脑出血后高表达，PERK/EIF2通路对脑出血大鼠脑组织细胞自噬的有影响；（3）IRE-1信号通路IRE1、TRAF2蛋白相对表达量呈随时间降低趋势，7d时相对值升高并为高值，IRE1信号通路对大鼠脑出血后内质网应激诱导神经细胞自噬发生有一定的调控作用。（4）内质网应激相关蛋白 GRP78、Beclin1相对表达量呈随时间降低趋势，7d时相对值升高并为高值。（5）针刺头部腧穴可促进大鼠脑出血后脑组织中PERK/EIF2、IRE1信号通路路关键因子的蛋白/mRNA 表达，激活细胞内PERK/EIF2、IRE1通路，调节下游靶基因转录，进而影响机体自噬相关蛋白GRP78、Beclin1在脑出血大鼠脑组织的表达。进而验证“针刺刺激-正反馈或负反馈调控UPR-诱导神经细胞自噬的适度激活-内质网应激与自噬的稳态重构-神经细胞自体修复启动是针刺促进脑出血神经修复与重塑的关键作用机制之一”的科学假说。为针刺治疗脑出血提供了新的科学依据，为脑出血的治疗研究拓宽了思路。