miR-194靶向Bmi-1调控PI3K/Akt信号通路对脑胶质瘤细胞迁移和凋亡的影响

Effect of miR-194 targeting Bmi-1 to regulate PI3K/Akt signaling pathway on migration and apoptosis of glioma cells

目的:探讨miR-194靶向Bmi-1调控PI3K/Akt信号通路对脑胶质瘤细胞迁移和凋亡的影响及作用机制。方法:qRT-PCR检测miR-194在脑胶质瘤组织和脑胶质瘤细胞中的表达;Transwell迁移实验检测miR-194对U251细胞迁移能力的影响;流式细胞术检测miR-194对U251细胞凋亡率的影响;免疫组织化学法检测Bmi-1在脑胶质瘤组织中的表达;qRT-PCR检测Bmi-1在脑胶质瘤细胞中的表达;TargetScan预测miR-194与Bmi-1的结合位点;双荧光素酶报告基因检测miR-194与Bmi-1的相互作用关系;Transwell迁移实验检测miR-194靶向Bmi-1对U251细胞迁移能力的影响;流式细胞术检测miR-194靶向Bmi-1对U251细胞凋亡率的影响;Western blot检测miR-194靶向Bmi-1对PI3K/Akt通路相关蛋白的影响。结果:miR-194在脑胶质瘤组织和脑胶质瘤细胞中呈低表达(P<0.01);过表达miR-194,U251细胞迁移能力下降(P<0.01),凋亡率提高(P<0.05);miR-194与Bmi-1存在靶向关系,且两者相互抑制;Bmi-1在脑胶质瘤组织和脑胶质瘤细胞中呈高表达(P<0.01);过表达miR-194可逆转Bmi-1过表达引起的U251细胞迁移能力增强(P<0.01);过表达miR-194可逆转Bmi-1过表达引起的U251细胞凋亡率下降(P<0.01);过表达Bmi-1,PI3K和Akt蛋白磷酸化程度增强,过表达miR-194可逆转这一现象。结论:过表达miR-194可抑制Bmi-1表达,降低PI3K和Akt蛋白磷酸化程度,抑制脑胶质瘤细胞迁移,促进其凋亡。

Objective:To explore effect and mechanism of miR-194 targeting Bmi-1 to regulate PI3 K/Akt signaling pathway on glioma cell migration and apoptosis.Methods:qRT-PCR to detect expression of miR-194 in glioma tissues and glioma cells;Transwell migration test to detect effect of miR-194 on migration ability of U251 cells;flow cytometry to detect effect of miR-194 on apoptosis rate of U251 cells.Immunohistochemical method to detect expression of Bmi-1 of glioma tissues;qRT-PCR to detect Bmi-1 expression of glioma cells;TargetScan to predicte binding site between miR-194 and Bmi-1.Dual luciferase reporter gene to detect interaction between miR-194 and Bmi-1;Transwell migration test to detect effect of miR-194 targeting Bmi-1 on migration ability of U251 cells;flow cytometry to detect effect of miR-194 targeting Bmi-1 on apoptosis rate of U251 cells.Western blot to detect effect of miR-194 targeting Bmi-1 on PI3 K/Akt pathway-related proteins.Results:miR-194 was lowly expressed in glioma tissues and glioma cells(P<0.01).Overexpression of miR-194 reduced migration ability of U251 cells(P<0.01),and increased apoptosis rate of U251 cells(P<0.05).There was a targeting relationship between miR-194 and Bmi-1,whose effects was mutual inhibition.Bmi-1 was highly expressed in glioma tissues and glioma cells(P<0.01).Overexpression of miR-194 reversed migration ability enhancing of U251 cells caused by Bmi-1 overexpression(P<0.01).Overexpression of miR-194 can reverse apoptosis rate decreasing of U251 cells caused by Bmi-1 overexpression(P<0.01).Overexpression of Bmi-1 increased phosphorylation of PI3 K and Akt proteins,however,overexpression of miR-194 can reverse this phenomenon.Conclusion:Overexpression of miR-194 inhibited expression of Bmi-1,which decreased phosphorylation of PI3 K and Akt proteins,thereby inhibiting migration and promoting apoptosis of glioma cells.