2-Cl-MGV-1介导BDNF-TrkB通路调控脑梗死后远隔脑区神经再生并改善认知损害的研究

Secondary damage of cognitive-related remote brain regions such as the thalamus and the hippocampus is involved in the occurrence and development of post-stroke cognitive deficits after cortical infarction. However, it still lacks fundamentally effective prevention and treatment measures for this secondary damage. Our previous study and published article have found that 2-Cl-MGV-1 reduced neuronal apoptosis in remote brain regions and ameliorated cognitive deficits after cerebral infarction, but whether it also promotes post-stroke neurogenesis in these brain regions and its mechanisms remain unknown. Recently, we have observed in the preliminary experiments that 2-Cl-MGV-1 increased the expression of BDNF and its receptor TrkB, and stimulated neurogenesis in the remote brain regions after cerebral infarction. This suggested that 2-Cl-MGV-1 might promote neurogenesis through the BDNF-TrkB signal pathway and ameliorate post-stroke cognitive deficits. Therefore, the present project is designed to firstly establish cortical infarction in hypertensive rats, and then evaluate the change of post-stroke cognition, neurogenesis in the cognitive-related remote brain regions and the protein levels associated with the BDNF-TrkB signal pathway before and after administration of 2-Cl-MGV-1. We further apply 252a to block the BDNF-TrkB signal pathway, and access whether neurogenesis in the above remote brain regions and the amelioration of post-stroke cognitive deficits are inhibited. Findings from the present project would help to make clear the role of neurogenesis in remote brains regions on post-stroke cognitive deficits and further reveal the mechanisms of 2-Cl-MGV-1, which would finally provide novel ideas for the prophylaxis and treatment of post-stroke cognitive deficits.

脑皮层梗死后，认知相关远隔脑区（如丘脑和海马）继发性损害参与了卒中后认知障碍的发生发展，但对此缺乏根本有效的防治措施。我们前期研究并发表论文证实，2-Cl-MGV-1可减少脑梗死后远隔脑区神经元凋亡并改善认知，但是否同时促进局部神经再生及其机制尚不清楚。新近，我们观察到2-Cl-MGV-1可增加脑梗死后远隔脑区BDNF及其受体TrkB表达和促进神经元再生，提示其可能介导BDNF-TrkB通路，促神经再生而最终改善认知。因此，本项目拟建立高血压大鼠大脑皮层梗死模型，观察2-Cl-MGV-1干预前后认知功能、认知相关远隔脑区神经元再生及BDNF-TrkB通路相关蛋白的改变；再以252a阻断BDNF-TrkB通路，评估脑梗死后上述脑区神经再生和认知改善是否受阻，以明确远隔脑区神经再生受限是否参与脑梗死后认知障碍，并揭示2-Cl-MGV-1的作用机制，以期为脑梗死后认知障碍的防治提供新思路。

脑皮层梗死后，认知相关远隔脑区（如丘脑）继发性神经元丢失参与了卒中后认知障碍的发生发展，但对此缺乏根本有效的防治措施。促进脑皮层梗死后丘脑的血管再生有望减少其神经元丢失并改善卒中后认知障碍。新近，我们的研究发现脑皮层梗死后丘脑血管内皮上转位蛋白（translocator protein, TSPO）表达水平显著增加。TSPO配体在胶质瘤和慢性缺氧模型中显示出促血管生成作用。因此，我们推测，新型TSPO配体2-Cl-MGV-1可能促进脑皮层梗死后丘脑血管再生、减少神经元丢失并改善卒中后认知障碍。选用易卒中型肾血管性高血压大鼠，建立右侧大脑中动脉皮层支闭塞（distal Middle Cerebral Artery Occlusion, MCAO）模型，分为假手术组、溶剂组和2-Cl-MGV-1组。2-Cl-MGV-1组术后24小时经腹腔注射给予2-Cl-MGV-1（7.5 mg/kg），溶剂组给予等量二甲基亚枫，此后每天1次，连续6天或13天。MCAO术后7和14天采用改良Bederson评分和Beam-walking试验评估运动功能，术后8-14天行Morris水迷宫评估脑梗死后认知损害。MCAO术后7天和14天取脑行Nissl染色评价梗死灶体积及丘脑神经元变性，免疫荧光染色观察丘脑继发性损害、血管再生和血管TSPO表达，Western Blot检测丘脑血管再生通路相关蛋白改变。脑皮层梗死后14天大鼠出现认知功能显著下降，同侧丘脑出现神经元丢失和小胶质和星形胶质细胞增生的继发性损害。同时，脑皮层梗死后丘脑新生血管显著增多，丘脑血管内皮上TSPO表达水平明显升高。2-Cl-MGV-1干预后显著增加脑皮层梗死后同侧丘脑BrdU+/Laminin+新生血管数量，并上调PI3K-AKT-mTOR信号通路及其下游效应蛋白HIF-1α和VEGF水平。脑皮层梗死后丘脑新生血管形成增多与丘脑神经元丢失减少和胶质细胞增生受抑制呈正相关，并减轻了卒中了认知障碍。本研究发现新型TSPO配体2-Cl-MGV-1可促进脑皮层梗死后丘脑血管再生和减少神经元丢失，并最终改善卒中后认知障碍。研究结果提示TSPO有望作为脑梗死远隔损害促血管再生和治疗卒中后认知障碍重要的干预靶点。