EZH2调节E-Cadherin对胶质瘤细胞生物学行为影响的实验研究

Effect of EZH2 on proliferation,invasion and migration of glioma cells through regulating E-Cadherin

目的探讨染色体调节因子zeste增强子同源物2(EZH2)在胶质瘤U87细胞增殖、侵袭、迁移的影响以及可能的调控机制。方法体外培养胶质瘤细胞株U87,转染EZH2 RNA干扰质粒作为siEZH2组,另外设置空质粒转染(NC)组和空白对照(Blank)组,采用实时荧光定量PCR(QPCR)法检测干扰效率。采用MTT法、Transwell小室法、划痕实验检测各组细胞的增殖、侵袭与迁移活性。采用Western blotting法和免疫荧光染色法检测EZH2、E-Cadherin、α-Catenin、α-Tubulin蛋白表达和共定位情况。结果siEZH2组EZH2 mRNA的表达量为0.41±0.06,低于NC组和空白对照组(P<0.05)。siEZH2组U87细胞的增殖活性较NC组和空白对照组明显减弱(P<0.05)。siEZH2组U87细胞侵袭率为(56.85±12.93)%,迁移率为(22.93±7.45)%,明显低于NC组和空白对照组(P<0.05)。siEZH2组E-Cadherin蛋白和α-Catenin蛋白表达量分别为1.74±0.19和1.22±0.16,高于NC组和空白对照组(P<0.05)。而siEZH2组、NC组、空白对照组组α-Tubulin蛋白的表达量几乎无变化(P>0.05)。经细胞免疫荧光染色法证实,U87细胞中内源性EZH2与E-Cadherin、α-Catenin蛋白存在分布上的共定位。结论沉默EZH2表达可抑制脑胶质瘤细胞U87的增殖、侵袭和迁移活性,其可能的作用机制与上调E-Cadherin、α-Catenin蛋白表达有关。

Objective To investigate the effect and mechanism of enhancer of zeste homolog 2(EZH2)on proliferation,invasion and migration of glioma cells.Methods Human glioma cell line U87 was cultured in vitro,and transfected with EZH2 RNA interference plasmid as siEZH2 group.Vector plasmid was treasfected into U87 cells as NC group and U87 cells with out any treatment as Blank control group.QPCR was performed to detect the expression level of EZH2 and the interference efficiency.The cell proliferation,invasion and migration activities of U87 cells were detected by MTT assays,transwell invasion assay and scratch assay.Western blotting and immunofluorescence staining were used to detected the expressions and co-location of EZH2,E-Cadherin,α-Catenin,α-Tubulin proteins in U87 cells.Results The expression of EZH2 mRNA in siEZH2 group was 0.41±0.06,lower than that in NC group and Blank group(P<0.05).The proliferation of U87 cells in siEZH2 group was inhibited,and there was statistical significance among siEZH2 group,NC group and Blank group(P<0.05).The invasion rate and migration rate of U87 cells in si-EZH2 group were(56.85±12.93)%and(22.93±7.45)%,which were both lower than these in NC group and Blank group(P<0.05).The expressions of E-Cadherin andα-Catenin proteins of U87 cells in si-EZH2 group were respectively(1.74±0.19)and(1.22±0.16),higher than those in NC group and Blank group(P<0.05).There were co-localization between endogenous EZH2 with E-Cadherin orα-Catenin proteins in U87 cells by immunofluorescence staining.Conclusion Silencing EZH2 RNA of U87 cells would inhibit the proliferation,invasion and migration of glioma cells through up-regulating E-Cadherin orα-Catenin levels.