益气活血组分中药改善脑细胞外微环境促BMSCs-exosomes脑缺血后神经修复作用的研究

Brain extracellular microenviroment (BEM) damage, namely”brain collaterals Impairmentin” traditional Chinese medicine (TCM), is the major pathological process of ischemic stroke. Qi deficiency and blood stasis are the principle pathogenesis of BEM damage, and tonyfying Qi and activating blood are their main therapy. Our Previous results showed that the anti-ischemic effect of compatibility of astragaloside and panax notoginseng saponins is closely related to regulating mechanism of BEM. BEM, as a potential target, could regulate BMSCs for targeting repairing neurological function. BMSCs-exosomes are the main active substances of BMSCs. In our project, a local cerebra ischemia rats model is as the study objective, the BEM constitutes unit of extracellular space, intercellular fluid and extracellular matrix can as entry points. The effective components of Huangqi and Sanqi compatibility in promoting BMSCs-exosomes to nerve regeneration after stroke are to studied in vivo experiments. Based on the above experiments, building the BEM model in vitro is aiming to explore the mechnism of TCM constituents reparing BEM and promoting BMSCs-exosomes after ischemia. The results of this project are to potentially enrich basis of TCM in treating cerebral ischemic diseases from”Tonyfying Qi and activating blood ,Regulating brain collateral”,and provide preventive methods for promoting the recovery of neurological function after cerebral ischemia.

脑细胞外微环境（brain extracellular microenviroment，BEM）损伤属中医“脑络损伤”，是缺血性中风的主要病理过程，气虚血瘀为主要病机，益气活血为主要治法。调节BEM有望成为脑缺血治疗新靶点。前期结果显示黄芪甲苷和三七总皂苷配伍抗脑缺血作用环节与BEM调控机制密切相关。BMSCs可靶向修复神经功能，受BEM调节，BMSCs-exosomes为其主要活性物质。故本项目以MCAO模型大鼠为研究对象，BEM组成单位脑细胞外间隙、细胞间液及细胞外基质为切入点，从整体动物实验研究组分中药促BMSCs-exosomes脑缺血后神经修复作用；并建立BEM体外模型，验证其作用机制与修复缺血后BEM，促进BMSC-exosomes分泌、存活及功能蛋白的释放有关。本项目可望丰富中医药“益气活血、从络论治”治疗脑缺血性疾病的实验依据，为促进脑缺血后神经功能恢复提供有效的防治方法。

脑细胞外微环境（ brain extracellular microenviroment，BEM）损伤是缺血性中风的主要病理过程，对其调节有望成为脑缺血治疗新靶点。BMSCs可靶向修复神经功能，受BEM调节，黄芪甲苷（ASTⅣ）和三七总皂苷（PNS）配伍抗脑缺血作用环节与BEM调控机制相关。故本项目提出假说：BMSCs-exosomes 是 BMSCs 通过旁分泌机制发挥神经修复功能的效应基础，靶向性修复脑缺血后神经功能，受脑细胞外微环境的调节；ASTⅣ配伍PNS具有促 BMSCs-exosomes 神经修复的效应，且作用机制在于修复缺血后脑细胞外微环境，促进 BMSCs-exosomes 分泌及功能。为验证假说，本研究首先建立了MCAO 3天和7天 大鼠局灶性脑缺血模型，观察不同时间点细胞外微环境变化，以及对BMSCs-exosomes移植效应的影响，结果表明与MCAO 3天比较，MCAO 7天炎症因子水平下降，脑缺血外微环境有一定的改善，有利于BMSCs、BMSCs-exosomes归巢和定向修复，说明BMSCs-exosomes可修复脑缺血后神经损伤，其作用受脑细胞外微环境的影响；然后通过整体动物实验验证ASTⅣ配伍PNS具有促 BMSCs-exosomes 神经修复的作用，结果显示ASTⅣ配伍PNS通过增强局部神经营养因子释放水平，改善细胞外基质环境，从而改善脑细胞外微环境，促进BMSCs-exosomes移植向缺血灶归巢，并增强其向神经元、星形胶质细胞和微血管的定向修复能力，减轻MCAO模型大鼠神经功能损伤；最后通过建立脑细胞外微环境损伤体外模型，通过体外实验证明其作用机制与修复缺血后脑细胞外微环境，促进 BMSCs-exosomes 分泌、存活及功能蛋白的释放有关。结果显示ASTⅣ配伍PNS可修复脑缺血细胞外微环境，促进BMSCs细胞增殖，减少细胞凋亡，促进BMSCs- exosomes释放。综上结果，我们得出以下结论：ASTⅣ配伍PNS可促进 BMSCs-exosomes 的分泌和功能，调节脑细胞外微环境，发挥对脑缺血后神经修复作用。本项目丰富了中医药“益气活血，从络论治”治疗脑缺血性疾病的实验依据，为促进脑缺血后神经功能的恢复提供有效的防治方法。