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目的 MicroRNAs(miRNAs)是内源性表达长度约为22个核苷酸的小RNA分子,通过结合到靶mRNA上而抑制基因的表达。近年的研究表明某些miRNAs可以调节肿瘤细胞的增殖与迁移,然而miRNA在子宫内膜癌中的作用有待于进一步探索。本研究旨在探讨miR-34c在人子宫内膜癌中的表达规律、功能和分子机制。方法通过荧光定量PCR技术检测20例人子宫内膜癌组织标本中miR-34c的表达。通过阳离子脂质体介导的方法将miR-34c转染入人子宫内膜癌细胞RL95-2中,应用MTS法检测子宫内膜癌细胞的增殖,克隆形成实验检测子宫内膜癌细胞的生长,流式细胞术检测细胞周期,Transwell实验检测子宫内膜癌细胞的迁移。生物信息学和荧光素酶报告基因实验确认miR-34c作用的靶基因。RNAi技术下调c-Met的表达。通过蛋白印迹检测miR-34c对cMet蛋白以及细胞内重要信号通路与细胞周期相关蛋白的影响。结果 miR-34c在子宫内膜癌中表达明显降低。miR-34c能使子宫内膜癌细胞RL95-2细胞周期滞留在G1期,显著抑制细胞的增殖。细胞移形实验发现miR-34c能够显著抑制RL95-2细胞迁移。生物信息分析表明c-Met是miR-34c作用的靶基因,进一步通过荧光素酶报告基因实验得以证实。蛋白印迹分析发现miR-34c能下调RL95-2细胞中的c-Met蛋白、p-Akt蛋白以及p-ERK1/2蛋白表达情况,同时还能够下调CDK4、CDK6蛋白以及p-Rb蛋白的表达水平。RNAi技术下调c-Met的表达能够明显抑制子宫内膜癌细胞的增殖与迁移。结论本研究结果证明miR-34c通过作用于靶基因c-Met而抑制子宫内膜癌细胞的增殖与迁移,说明miR-34c是子宫内膜癌中的重要抑癌基因。
MicroRNAs (miRNAs) are small RNA molecules that endogenously express about 22 nucleotides in size and inhibit gene expression by binding to target mRNAs. Recent studies have shown that certain miRNAs can regulate the proliferation and migration of tumor cells, however, the role of miRNA in endometrial cancer remains to be further explored. This study aimed to investigate the expression, function and molecular mechanism of miR-34c in human endometrial carcinoma. Methods The expression of miR-34c in 20 specimens of human endometrial carcinoma was detected by fluorescence quantitative PCR. MiR-34c was transfected into human endometrial carcinoma RL95-2 by cationic liposome-mediated method. The proliferation of endometrial carcinoma cells was detected by MTS assay. The growth of endometrial carcinoma cells was detected by colony formation assay , Flow cytometry to detect cell cycle, Transwell assay of endometrial cancer cell migration. Bioinformatics and luciferase reporter assays confirmed the role of miR-34c as a target gene. RNAi technology down-regulates the expression of c-Met. The effect of miR-34c on cMet protein and important intracellular signaling pathways and cell cycle related proteins was detected by Western blotting. Results The expression of miR-34c in endometrial carcinoma was significantly lower. miR-34c can endometrial cancer cells RL95-2 cell cycle retention in the G1 phase, significantly inhibited cell proliferation. Cell migration assay found miR-34c can significantly inhibit RL95-2 cell migration. Bioinformatics analysis showed that c-Met is the target gene of miR-34c and further confirmed by luciferase reporter assay. Western blot analysis showed that miR-34c down-regulated the expression of c-Met, p-Akt and p-ERK1 / 2 in RL95-2 cells as well as down-regulated the expression of CDK4, CDK6 and p-Rb . The down-regulation of c-Met expression by RNAi technology can significantly inhibit the proliferation and migration of endometrial cancer cells. Conclusion The results of this study demonstrate that miR-34c inhibits the proliferation and migration of endometrial cancer cells by acting on the target gene c-Met, indicating that miR-34c is an important tumor suppressor gene in endometrial cancer.