microRNA-7对人肝癌细胞MHCC-97H上皮间质转化的影响及作用机制

目的 探讨microRNA-7(miR-7)对肝癌细胞株MH-CC-97H上皮-间质转化(EMT)的影响及作用机制.方法 将构建的miR-7真核载体单独及分别联合野生型或突变型 EGFR 3'-UTR真核载体共转染MHCC-97H细胞,采用Real-time PCR、Western blot 检测 EMT 标识蛋白 E-cadherin、β-catenin、N-cadherin 和 Vimentin 的表达;细胞划痕、Transwell实验检测细胞侵袭和迁移能力的变化;双荧光素酶实验分析 miR-7对EGFR的调控作用.结果 与正常组相比,转染 miR-7真核载体组细胞E-cadherin、β-catenin表达均显著升高(P＜0. 01),N-cadherin、Vimentin 表达则明显降低(P＜0. 01);同时,细胞的侵袭、迁移能力均明显减弱(P＜0.01);分别与转染GV272 + EGFR 3'-UTR野生型和转染GV272/ miR-7 + EGFR 3'-UTR 突变型相比,转染 GV272/miR-7 + EGFR 3'-UTR野生型组萤火虫荧光素酶活性显著降低(P＜0.01).结论 miR-7可通过与EGFR 3'-UTR结合而抑制 EGFR信号通路,降低MHCC-97H细胞侵袭、迁移能力,进而抑制MHCC-97H细胞EMT.

Effect and mechanism of mircoRNA-7 on epithelial-mesenchymal transition of human hepatocellular carcinoma cell

Objective To investigate the effect and mechanism of microRNA-7 (miR-7) on epithelial-mesenchymal transition of MHCC-97H cell. Methods MiR-7 eukaryotic vector was co-transfected into wild-type or mutant EGFR 3'-UTR eukaryotic vector, respectively. The expression of E-cadherin, β-catenin, N-cadherin and Vimentin were detected by real-time PCR and Western blot. Meanwhile, cell scratch test and transwell assay was used to detect the invasion and migration ability of cells. Dual luciferase assay was used to analyze the effect of miR-7 on EGFR of 3'-UTR. Results Compared with normal group, the expression of E-cadherin and β-catenin was significantly increased in GV268/miR-7 group (P ＜0. 01 ), but the expression of N-cadherin and Vimentin was significantly decreased (P ＜0. 01). Meanwhile, the invasion and migration ability was also significantly decreased (P ＜0. 01). Compared with EGFR 3'-UTR wild-type group and GV272/miR-7 + EGFR 3'-UTR mutation group respectively, the luciferase activity of GV272/miR-7 + EGFR 3'-UTR wild-type group was significantly higher (P ＜0. 01). Conclusion miR-7 could inhibit the EGFR signaling pathway by binding to EGFR 3'-UTR, decrease the invasion and migration of MHCC-97H cells and inhibit the EMT of MHCC-97H cells.