miR-134通过下调叉头盒蛋白M1抑制肝细胞癌细胞增殖

发布时间：2018-02-02 12:50

  本文关键词： 微小RNA- 叉头盒蛋白M 肝细胞癌 细胞增殖　出处：《第三军医大学学报》2017年01期 　论文类型：期刊论文

【摘要】：目的探讨miR-134下调叉头盒蛋白M1(forkhead box M1,FOXM1)的机制及其影响肝细胞癌(hepatocellular carcinoma,HCC)细胞增殖的作用。方法应用CCK-8法检测miR-134对HCC细胞增殖的作用;经Western blot和Real-time PCR检测人正常肝细胞L02和5种HCC细胞系中FOXM1和miR-134的表达;用miR-134模拟物(miR-134 mimic)或miR-134抑制剂(miR-134 inhibitor)转染HCC细胞后,经Western blot和Real-time PCR检测FOXM1及其下游靶基因Cyclin D1的表达;应用生物信息学分析miR-134在人FOXM1 3'-UTR的可能结合位点,通过报告基因检测实验分析miR-134与FOXM1的3'-UTR的特异性结合;将miR-134 mimic和FOXM1表达质粒(无3'-UTR)共转染于HCC细胞后,经CCK-8法检测细胞的增殖情况。结果 miR-134可显著抑制HCC细胞增殖(P0.05);与人正常肝细胞L02相比,miR-134在HCC细胞中的表达明显降低(P0.05),而FOXM1蛋白表达明显增强,二者存在负相关(R2=0.705,P0.05);miR-134可显著下调FOXM1蛋白及其下游靶基因Cyclin D1的表达(P0.05);报告基因检测实验证实miR-134可特异性结合于FOXM1 mRNA的3'-UTR(P0.01);细胞增殖实验检测结果显示,过表达缺失3'-UTR的FOXM1可显著减弱miR-134对HCC细胞增殖的抑制效应(P0.05)。结论 miR-134通过靶向结合于FOXM1的3'-UTR而下调FOXM1蛋白的表达,从而抑制HCC细胞的增殖。
[Abstract]:Objective to investigate the down-regulation of forkhead box M1 by miR-134. FOXM1) and its effect on hepatocellular carcinoma. Methods CCK-8 assay was used to detect the effect of miR-134 on the proliferation of HCC cells. The expression of FOXM1 and miR-134 in L02 and 5 HCC cell lines were detected by Western blot and Real-time PCR. HCC cells were transfected with miR-134 mimic or miR-134 inhibitor HCC. The expression of FOXM1 and its downstream target gene Cyclin D1 was detected by Western blot and Real-time PCR. Bioinformatics was used to analyze the possible binding sites of miR-134 in human FOXM1 3- UTR. The specific binding of miR-134 to FOXM1 was analyzed by reporter gene detection. MiR-134 mimic and FOXM1 expression plasmids were co-transfected into HCC cells. The proliferation of HCC cells was detected by CCK-8. Results miR-134 could significantly inhibit the proliferation of HCC cells. Compared with human normal hepatocytes L02, the expression of miR-134 in HCC cells was significantly decreased, while the expression of FOXM1 protein was significantly increased. There was a negative correlation between them. MiR-134 could significantly down-regulate the expression of FOXM1 protein and its downstream target gene Cyclin D1. The reporter gene assay confirmed that miR-134 could specifically bind to FOXM1 mRNA. The results of cell proliferation test showed that. Overexpression of FOXM1 with deletion of 3H-UTR significantly attenuated the inhibitory effect of miR-134 on the proliferation of HCC cells (P0.05). Conclusion miR-134 can down-regulate the expression of FOXM1 protein by targeting the 3- UTR binding to FOXM1. Thus, the proliferation of HCC cells was inhibited.
【作者单位】： 第三军医大学基础医学部生物化学与分子生物学教研室;
【基金】：国家自然科学基金面上项目(81672377,31470066)~~
【分类号】：R735.7
【正文快照】： 生物化学与分子生物学教研室)specific binding of miR-134 with 3'-UTR of FOXM1 mRNA.After co-transfected with miR-134 mimic orFOXM1-expressing plasmids(without 3'-UTR),the HCC cell proliferation was detected by CCK-8 assay.Results miR-134 dramatically suppr，

本文编号：1484550