mirPS细胞与内皮祖细胞移植共同促进脑缺血损伤修复

Transplantation of induced pluripotent stem cells (iPS) and endothelial progenitor cells (EPC) was reported to be neuroprotective after cerebral ischemia. But the strategy needed to be improved for their unsatisfactory efficacy and safety. The mechanism involved in the differentiation process of pluripotent into neural cells was not precisely understood. miR-302s could be transfected into HEK293 cells to generate pluripotent stem cells which were named as mirPS cells. MicroRNA (miR) contributed to the modulation of stem cell differentiation into neural cells and endothelial cells. Thus we proposed a novel strategy to ameliorate cerebral ischemia injury through combination of neural stem cell transplantation and endothelial progenitor cell transplantation, in order to exploit both of the promising therapeutic channels, i.e., promoting compensation of neural cellular function by neural stem cells and enhancing angiogenesis by endothelial progenitor cells. In brief, miR-302s and miR-145 were electroporated simultaneously to induce HEK293 cells into mirPS cells. After the mirPS cells were differentiated in vitro into neural stem cells, the stem cells were mixed with EPC and transplanted into lateral ventricle of mice with cerebral ischemia.

移植诱导性多能干细胞或内皮祖细胞（EPC）可在缺血性脑损伤后发挥脑保护作用。但这两种疗法尚不完善，多能干细胞向神经细胞分化的调控机制尚不清楚。有报道指出可用miR-302s制备多能干细胞（mirPS）。我们前期研究有3个初步发现：1.共转染miR-145有助于制备mirPS；2.经侧脑室及外周循环移植小鼠自体EPC可改善缺血性脑损伤恢复效果；3.anti-miR-107可促进移植的EPC向血管内皮细胞分化。因此我们考虑共转染miR-302s和miR-145，制备mirPS，并向神经干细胞诱导分化，将神经干细胞、EPC及anti-miR-107混合后移植入脑缺血小鼠侧脑室，比较该策略的脑保护作用是否高于单独输注，从促进脑损伤后神经细胞修复和增强脑损伤部位血管修复再生两方面，改善脑缺血损伤后的恢复效果。

移植诱导性多能干细胞或内皮祖细胞（EPC）可在缺血性脑损伤后发挥脑保护作用。但这两种疗法尚不完善，多能干细胞向神经细胞分化的调控机制尚不清楚。有报道可用miR-302s制备多能干细胞（mirPS），小RNA（miR）在干细胞向神经细胞和血管内皮细胞分化过程中发挥调控作用。我们发现上调miR-145利于获得mirPS，小鼠侧脑室EPC移植联合anti-miR-107可促进脑缺血损伤部位血管生成。因此我们考虑共转染miR-302s和miR-145，制备mirPS，并向神经干细胞诱导分化，将神经干细胞和EPC混合移植入脑缺血小鼠侧脑室，比较该策略的脑保护作用是否高于单独输注，同时探讨miR在多能干细胞向神经细胞分化过程中的角色，尝试通过调节miR水平，促进mirPS和EPC向神经细胞和血管内皮细胞分化成熟，从促进脑损伤后神经细胞修复和增强脑损伤部位血管修复再生两方面，改善脑缺血损伤后恢复。重要结果：导入miR-302s载体和miR-145载体可将HEK293细胞诱导去分化为mirPS，并进而诱导分化为神经干细胞；microRNA芯片初筛发现mirPS和神经干细胞之间差异表达的小RNA（miR-219、 miR-128和miR-765）；明确miR-219、miR-128及其靶分子在mirPS和神经干细胞差异表达；确认miR-219及miR-128对于mirPS细胞再分化的作用；发现mirR-219可能通过靶向作用于SNX27分子，促进mirPS细胞再分化；发现mirR-765可能通过靶向作用于Hes1分子，促进mirPS细胞再分化；比较侧脑室移植EPC和静脉注射EPC方法对缺血性脑损伤的修复效果，发现这两种方法均对脑缺血损伤可产生修复效果，在主要指标上无显著统计学差异；确定EPC移植联合anti-miR-107对脑缺血损伤的治疗效应；观察神经干细胞+EPC双通道策略的疗效，发现 mirPS诱导分化产生的神经干细胞与EPC联合移植对小数脑缺血损伤可起到治疗效应。