MiR-4739通过下调MIEN1表达抑制胃癌NCI-N87细胞恶性生长和转移的机制

MiR-4739 inhibits malignant growth and metastasis of gastric cancer NCI-N87 cells through down-regulating the expression of MIEN1

目的:研究miR-4739对胃癌NCI-N87细胞恶性生长和转移的影响及其机制。方法:采用qRT-PCR检测胃黏膜上皮GES-1细胞和胃癌HGC-27、NCI-N87、HS-746T细胞中miR-4739表达水平。在胃癌NCI-N87细胞中转染miR-4739 mimics后,采用CCK-8检测细胞增殖、流式细胞术检测凋亡、Transwell小室检测迁移和侵袭、蛋白质印迹法检测剪切的含半胱氨酸的天冬氨酸蛋白水解酶3(C-caspase-3)、波形蛋白(Vimentin)和上皮性钙黏附素(E-cadherin)的表达。采用在线靶基因预测软件预测miR-4739的靶基因,随后采用荧光素酶报告系统鉴定二者的靶向关系。在胃癌NCI-N87细胞共转染miR-4739 mimics和迁移侵袭增强因子1(MIEN1)过表达载体pcDNA-MIEN1后,检测细胞增殖、凋亡、迁移和侵袭能力以及C-caspase-3、Vimentin和E-cadherin表达情况。结果:胃癌HGC-27、NCI-N87、HS-746T细胞中miR-4739表达水平均低于胃黏膜上皮GES-1细胞,差异均有统计学意义(P<0.05)。胃癌NCI-N87细胞转染miR-4739 mimics后,与转染miR-mimics-NC相比,细胞增殖、迁移、侵袭能力降低,细胞凋亡率升高,E-cadherin、C-caspase-3蛋白表达水平升高,Vimentin蛋白表达水平下降,差异均有统计学意义(P<0.05)。miR-4739可靶向抑制MIEN1表达。胃癌NCI-N87细胞共转染miR-4739 mimics和pcDNA-MIEN1后,与共转染miR-4739 mimics和pcDNA相比,细胞增殖、迁移、侵袭能力升高,细胞凋亡率降低,E-cadherin、C-caspase-3蛋白表达水平减少,Vimentin蛋白表达水平升高,差异均有统计学意义(P<0.05)。结论:MiR-4739可通过靶向抑制MIEN1表达阻碍胃癌NCI-N87细胞的恶性生长和转移潜能。

Objective:To study the effects and mechanism of miR-4739 on malignant growth and metastasis of gastric cancer NCI-N87 cells.Methods:The expression level of miR-4739 in gastric epithelial GES-1 cells and gastric cancer HGC-27,NCI-N87,HS-746T cells were detected by qRT-PCR.After transfection of miR-4739 mimics into gastric cancer NCI-N87 cells,CCK-8 was used to detect cell proliferation,flow cytometry was used to detect apoptosis,Transwell chamber was used to detect migration and invasion,and Western blot was used to detect the expression of cleaved cysteinyl aspartate specific proteinase 3(C-caspase-3),Vimentin and epithelical cadherin(E-cadherin).The online target gene prediction software was used to predict the target gene of miR-4739,and then the luciferase reporter system was used to identify their targeting relationship.After co-transfection of miR-4739 mimics and migration and invasion enhancer 1(MIEN1)overexpression vector pcDNA-MIEN1 into gastric cancer NCI-N87 cells,the cell proliferation,apoptosis,migration and invasion,and the expression of C-caspase-3,Vimentin and E-cadherin were detected.Results:The expression levels of miR-4739 in gastric cancer HGC-27,NCI-N87 and HS-746T cells were significantly lower than that in gastric epithelial GES-1 cells(P<0.05).After transfection of miR-4739 mimics into gastric cancer NCI-N87 cells,compared with the cells transfected with miR-mimics-NC,the cell proliferation,migration and invasion ability were decreased,the cell apoptosis rate was increased,the protein expression levels of E-cadherin and C-caspase-3 were increased,and the protein expression level of Vimentin was decreased,and the differences were statistically significant(P<0.05).miR-4739 could targeted inhibit the expression of MIEN1.After co-transfection of miR-4739 mimics and pcDNA-MIEN1 into gastric cancer NCI-N87 cells,compared with the cells co-transfected with miR-4739 mimics and pcDNA,the cell proliferation,migration and invasion ability were increased,the cell apoptosis rate was decreased,the protein expression levels of E-cadherin and C-caspase-3 were decreased,and the protein expression level of Vimentin was increased,and the differences were statistically significant(P<0.05).Conclusion:MiR-4739 can block the malignant growth and metastatic potential of gastric cancer NCI-N87 cells through targeting inhibition of MIEN1’s expression.