miR-1过表达对鼻咽癌细胞的调控作用及其机制

Regulation and mechanism of miR-1 overexpression on nasopharyngeal carcinoma cells

目的探讨miR-1过表达对鼻咽癌细胞的调控作用及其机制。方法将CNE2细胞随机分为对照组与转染组,分别转染negative control miRNA mimics、miR-1 mimics,采用CCK-8法检测两组细胞增殖能力、流式细胞术检测两组细胞周期、细胞划痕实验检测两组细胞迁移能力、Transwell实验检测两组细胞侵袭能力,采用qRT-PCR法和Western blotting法检测c-met mRNA和蛋白表达。采用miRNA靶基因数据库预测miR-1的潜在靶点,荧光素酶报告基因法检测miR-1与其潜在靶点的结合情况。结果与对照组比较,转染组细胞增殖、迁移及侵袭能力均明显降低(P均<0. 05)。与对照组比较,转染组G0/G1期细胞比例升高(P <0. 05),S期细胞比例降低(P <0. 05),G2期细胞比例变化不明显(P> 0. 05)。与对照组比较,转染组c-met mRNA及蛋白表达均明显降低(P均<0. 05)。经miRNA靶基因数据库预测,c-met是miR-1的潜在靶点之一; miR-1 mimics可明显抑制c-met野生型质粒荧光素酶活性,而对突变型质粒荧光素酶活性无明显影响。结论 miR-1过表达可抑制鼻咽癌细胞增殖、迁移及侵袭,其作用机制可能与降低细胞内c-met表达有关。

Objective To investigate the regulatory effect and mechanism of microRNA-1 overexpression on nasopharyngeal carcinoma cell line(CNE2).Methods CNE2 cells were randomly divided into the control group and transfection group,which were transfected with negative control microRNA mimics and microRNA-1 mimimics,respectively.CCK8 assay was used to detect the proliferation of CNE2 cells,flow cytometry was used to detect the growth cycle of CNE2 cells,Scratch assay was used to detect the migration ability of CNE2 cells,Transwell assay was used to detect the invasion ability of CNE2 cells,qRT-PCR and Western blotting were used to detect the effect of microRNA-1 on the mRNA and protein expression of c-met gene.We used microRNA target gene database to predict whether c-met was one of the potential targets of microRNA-1.Luciferase reporter gene assay was used to detect whether microRNA-1 could bind to 3′-UTR of c-met.Results The cell proliferation,migration and invasion abilities of the transfection group were lower than those of the control group(all P<0.05).Compared with the control group,the proportion of cells in the G0/G1 phase was higher,while the proportion of cells in the S phase was lower(P<0.05),and there was no significant difference in the proportion of cells in the G 2 phase(P>0.05).Compared with the control group,the mRNA and protein expression of c-met gene in the transfection group was lower(P<0.05).Mir-1 mimics significantly inhibited the luciferase activity of wild-type plasmid c-met,but had no significant effect on the luciferase activity of mutant plasmid c-met.Conclusion Overexpression of microRNA-1 can inhibit the proliferation,migration and invasion of nasopharyngeal carcinoma cells,and its mechanism may be related to the reduction of the expression of c-met in cells.