微小RNA-106b参与内皮细胞介导的血管新生作用机制研究

Involvement of miR-106b in endothelial cells-mediated angiogenesis

目的研究微小RNA(microRNA,miR)-106b是否参与动脉粥样硬化相关的血管新生,明确miR-106b在内皮细胞中参与血管新生的作用机制。方法内皮细胞培养并转染miR-106b,分为miR-106b组、空白对照组和阴性对照组。提取RNA、反转录及实时定量PCR检测miR-106b相对表达量以明确转染效率。观察各组内皮细胞在凝固的基质胶中管腔形成情况。TUNEL法检测转染后细胞凋亡状况并进行靶基因的预测,采用Western blot法检测筛选的蛋白相对表达量变化。结果与空白对照组和阴性对照比较,miR-106b组在基质胶中管腔形成明显减少;与阴性对照组比较,miR-106b组管腔比值、信号转导及转录激活因子3mRNA相对表达量及蛋白表达明显降低(P<0.05,P<0.01)。阴性对照组与miR-106b组细胞凋亡率比较,差异无统计学意义(1.19%vs 3.39%,P>0.05)。结论 miR-106b在内皮细胞抑制血管新生可能是通过下调信号转导及转录激活因子3,与血管内皮生长因子无直接关系。

Objective To study whether miR-106b is involved in endothelial cells-mediated angiogenesis .Methods miR-106b cultured and transfected with endothelial cells was divided into miR-106b group ,blank control group and positive control group .RNA was extracted from miR-106b .The transfection efficiency was confirmed by inverse transcription .Endothelial cell tubes formed in matrigel were observed .Apoptosis of transfected miR-106b was assayed by TUNEL assay .Target genes of miR-106b were detected .Expressions of miR-106b and candidate genes were detected by RT-PCR and Western blot ,respectively .Results The number of endothelial cell tubes formed in matrigel was significantly less in miR-106b group than in blank control group and positive control group .The ratio of formed tubes ,signal transduction and mRNA expression level were significantly lower in miR-106b group than in positive control group （ P〈0.05 ,P〈0.01） .No significant difference was found in apoptosis of transfected miR-106b among the 3 groups （1.19% vs 3 .39% ,P〉0 .05） .Conclusion miR-106b inhibits endothelial cells-mediated angiogenesis by down-regulating the signal transduction and STAT 3 ,which is not directly related with VEGFA .