基于HIF-1α/Notch信号通路探讨塞络通胶囊活化星形胶质细胞调控缺血性脑卒中后神经修复机制研究

Ischemic stroke, kind of common disease with high morbidity, high disability rate and high recurrence rate, threatens the physical and mental health of patients seriously, however, there was little breakthrough in the treatment nowadays. Studies have shown that the HIF-1α/Notch signaling pathway could activate astrocytes in the ischemic brain to mediate the inhibition signal and may contribute to the reconstitution of the synapse and axon, thereupon then protect and repair the impaired neurons, and gradually becomes one of the crucial targets in the research and development of anti-cerebra ischemia drugs. Sailuotong Capsule (SLT), which function as tonifying Qi and activating blood, is a novel TCM compound formulation designed for the treatment of ischemic cerebrovascular diseases and exerted certain effect of promoting the role of newborn mature neurons in ischemic hemisphere, but the mechanisms and targets are not yet clear. In this study, we planned to establish a mouse in vitro astrocyte-neuron co-culture system and then induce a hypoxia-reoxygenation model, and to establish a mouse model of multiple cerebral infarctions, besides, shRNA technique will be applied to study the mechanism of action of Sailuotong Capsule on astrocyte activation induced neurogenesis and neural repairment after ischemic stroke through regulation of HIF-1α/Notch signaling pathway. This study will also provide theoretical and experimental basis for the clinical application of Sailuotong Capsule.

缺血性脑卒中致残率高、复发率高、并发症多，严重威胁患者身心健康，其治疗上目前仍无突破性的进展。已有研究证实，HIF-1α/Notch信号通路可活化脑缺血后星形胶质细胞，介导细胞的分化抑制信号，有助于神经突触和轴突的重塑，从而保护及修复受损的神经元，成为治疗缺血性脑损伤药物研究的关键。具有益气活血功效的复方中药塞络通是课题组研发的治疗缺血性脑血管病的有效复方中药，既往研究表明塞络通可表现出一定的促进缺血侧新生成熟神经元新生的作用，但其作用机制及靶点尚不明确。本研究拟通过体外构建小鼠星形胶质细胞-神经元共培养体系并建立缺氧/复氧损伤模型，以及颈内动脉注射微球血管栓塞剂法致多发性脑梗死小鼠模型，同时通过shRNA慢病毒转染等手段，从调控神经发生的HIF-1α/Notch信号通路角度，探讨SLT促进脑缺血后星形胶质细胞活化进而影响神经修复与再生的分子机制，为塞络通临床应用提供理论基础和实验依据。

缺血性脑卒中致残率高、复发率高、并发症多，严重威胁患者身心健康，目前其治疗上仍无突破性的进展。本项目通过建立多发性脑梗死小鼠模型（体内实验）及小鼠星形胶质细胞-神经元共培养缺氧/复氧损伤模型（体外实验），采用运动/认知行为学、病理形态学、分子生物学、遗传学等技术手段，考察缺血后内源性神经干细胞增殖与分化作用的动态演变规律；并通过构建HIF-lα shRNA 、Notch1 shRNA慢病毒及腺相关病毒表达载体，在体内外不同水平抑制HIF-lα或Notch1靶基因的表达，探索HIF-lα/Notch信号通路调控神经修复与神经发生的机制；此外，项目还评价了具有益气活血功效的复方中药塞络通及其有效组分对HIF-1α/Notch信号通路调控缺血后神经发生的影响，揭示了其促进缺血后神经修复的潜在机制。本项目研究结果表明，塞络通可通过激活HIF-1α/Notch信号通路，影响星形胶质细胞与小胶质细胞活化，调节S100A8/A9介导的神经炎症，促进海马区神经干细胞增殖、分化及神经网络重建，从而改善多发性脑梗塞引起的运动功能与认知功能损伤。本研究从调控神经发生的HIF-1α/Notch信号通路角度对脑缺血后神经修复与再生的分子机制进行了积极探索，为寻找治疗缺血性脑血管病的药物提供了新的策略与方向，具有一定的意义和应用前景。