miR-126和miR-128在BM-MSCs修复脑出血后血脑屏障和神经损伤中的作用及机制研究

Intracerebral hemorrhage (ICH) is global disease with high morbidity, mortality and disability rate, and there is still no breakthrough in the treatment for ICH. Our previous research indicated that bone marrow mesenchymal stem cells (BM-MSCs) ameliorate blood-brain barrier dysfunction and neurological deficits after ICH in spontaneously hypertensive rats. Evidence suggests that miR-126 promotes angiogenesis and vascular integrity, and inhibits apoptosis in vascular endothelial cells through PI3K/Akt signal pathway. miR-128 decreased the neural differentiation of BM-MSCs by binding to a specific site in the 3'UTR of Wnt3a. Accordingly we speculate that overexpressing miR-126 or silencing miR-128 enhances the reparing effect of BM-MSCs by promoting BM-MSCs differentiation into vascular endothelial cells and neurons. In this research, miR-126 overexpressed or miR-128 silenced BM-MSCs will be constructed, and the effect of genetic modification on cell proliferation, apoptosis, differentiation, homing ability and anti-inflammatory properties will be analyzed. The effect of miR-126 and miR-128 on BM-MSCs repairing blood-brain barrier and nerve damage after ICH will be verified in the rat model of ICH, and the related signal pathways will be examined to clarify the molecular mechanism. The intended results will provide new theoretical base for ICH therpy.

脑出血是一种高发病率、高致残率和高死亡率的全球性疾病，目前其治疗尚无突破性进展。课题组前期研究表明骨髓间充质干细胞（BM-MSCs）移植有助于大鼠脑出血后血脑屏障及神经损伤的修复。既往研究表明miR-126促进血管新生和血管完整性，调控PI3K/Akt通路抑制血管内皮细胞凋亡，miR-128能够靶向Wnt3a抑制干细胞向神经细胞分化，因此我们推测miR-126过表达/miR-128沉默能促进BM-MSCs向血管内皮细胞和神经细胞分化，增强BM-MSCs的修复作用。本研究拟构建miR-126过表达/miR-128沉默的BM-MSCs，观察基因修饰对BM-MSCs增殖、凋亡、分化、归巢及抗炎能力的影响，并在脑出血大鼠模型中验证miR-126/miR-128修饰的BM-MSCs对血脑屏障及神经损伤的修复作用，并通过检测相关信号通路进一步明确其作用的分子机制，为脑出血的防治提供新的理论基础。

背景：MicroRNA-126(miR-126)是内皮细胞血管生成信号的重要调控因子，在血管完整性、肿瘤生长、侵袭和血管炎症中发挥重要作用。miR-128被证实具有抑制血管生成的作用,下调BM-MSCs中miR-128水平可促进bFGF和表皮生长因子诱导向神经元样细胞分化。.主要研究内容：探讨miR-126过表达/miR-128沉默修饰后的BM-MSCs的优越性，然后探讨miR-126过表达/miR-128沉默对BM-MSCs增殖、凋亡、迁移以及向血管内皮细胞和神经细胞分化能力的影响；明确miR-126过表达/miR-128沉默增强BM-MSCs对脑出血后血脑屏障和神经损伤的修复作用机制。.重要结果及数据：rno-miR-126a-3p过表达转染组的细胞中miR-126表达明显增多，rno-miR-128-3p沉默转染组的细胞中miR-128表达明显降低。rno-miR-126a-3p过表达/rno-miR-128a-3p沉默转染组细胞迁移率明显增加,凋亡率明显降低。rno-miR-126a-3p过表达转染组和rno-miR-128a-3p沉默转染组BM-MSCs数量明显升高。miR-126a+内皮细胞诱导组和anti-miR-128-3p+内皮细胞诱导组中阳性细胞的比率也升高。miR-126a+神经元样细胞诱导组和anti-miR-128-3p+神经元样细胞诱导组中阳性细胞的比率也升高。ICH+miR-126a-3p过表达BM-MSCs移植组和ICH+miR-128-3p沉默BM-MSCs组大鼠神经功能恢复最好。miR-126和anti-miR-128修饰的BM-MSCs明显减轻了BBB的破坏。ICH+miR-126a-3p-BM-MSCs和ICH+anti-miR-128-3p-BM-MSCs组中PAR 1和MMP 9的蛋白表达与ICH+Vector-BM-MSCs组相比均有所减少，同时注射miR-126和anti-miR-128修饰的BM-MSCs后，增强了p-Akt和p-ERK1/2的表达。.本研究在ICH治疗领域提供了另一个新的靶点，随着后续的不断深入研究，可能会进一步证明miR-126过表达/miR-128沉默修饰的BM-MSCs为ICH的治疗提供一个新的途径。