miR-485-5p靶向调控Survivin基因对人结直肠癌HCT116细胞侵袭性的影响

doi:10.6040/j.issn.1671-7554.0.2015.083

摘要/Abstract

摘要： 目的探讨miR-485-5p对人结直肠癌HCT116细胞中Survivin基因的靶向调控作用以及对细胞侵袭性的影响。方法培养HCT116细胞,应用免疫荧光检测Survivin基因在细胞中的表达。采用生物信息学方法对miR-485-5p和Survivin基因的靶向匹配关系进行预测,并应用双荧光素酶报告基因系统鉴定。脂质体2000转染miR-485-5p模拟物后,Real-time PCR检测miR-485-5p和Survivin基因的表达,Western blotting检测Survivin蛋白的表达,Transwell小室检测HCT116细胞体外的侵袭性。结果倒置相差显微镜下可见细胞均匀分布,形态规则均一;细胞免疫荧光显示胞质内有Survivin蛋白的表达。靶基因预测软件miRanda和TargetScan显示miR-485-5p和Survivin基因匹配良好,双荧光素酶报告基因系统鉴定发现miR-485-5p能够结合Survivin mRNA 3'UTR,并有效抑制其表达。Real-time PCR和Western blotting检测结果显示,过表达miR-485-5p能够导致Survivin基因表达的下降。Transwell实验结果显示,miR-485-5p的过表达能够抑制HCT116细胞的侵袭。结论 miR-485-5p通过靶向作用于Survivin mRNA 3'UTR能够负性调控Survivin基因的表达,并能够抑制人结直肠癌HCT116细胞的侵袭。

关键词: 微小RNA, 结直肠肿瘤, 生存素, HCT116细胞

Abstract: Objective To identify the role of miR-485-5p in human colorectal cancer HCT116 cell invasion. Methods HCT116 cells were cultured in vitro, and cellular morphology was observed with phase contrast microscope. The miR-485-5p was predicted with bioinformatics and identified with dual luciferase report system. Expressions of miR-485-5p and survivin were determined with Real-time PCR and Western blotting after transfection of miR-485-5p mimics. The invasion of HCT116 cells was detected in vitro with transwell chamber. Results Survivin expression was positive as detected by immunofluorescence cytochemistry. Dual luciferase reporter system indicated that miR-485-5p inhibited survivin expression by binding to 3'UTR of survivin mRNA. Real-time PCR and Western blotting showed that over-expression of miR-485-5p down-regulated survivin expression. The invasion of HCT116 cells was suppressed by over-expression of miR-485-5p. Conclusion The miR-485-5p may negatively regulate survivin expression by binding to its 3'UTR in human colorectal cancer HCT116 cells and inhibit cell invasion.

Key words: Colorectal neoplasm, MiRNA, Survivin, HCT116 cells

中图分类号:

R735.3

曾仁仁, 张俊华, 张银旭. miR-485-5p靶向调控Survivin基因对人结直肠癌HCT116细胞侵袭性的影响[J]. 山东大学学报（医学版）, 2015, 53(6): 23-27.

ZENG Renren, ZHANG Junhua, ZHANG Yinxu. The role of miR-485-5p in human colorectal cancer HCT116 cell invasion by regulating survivin expression[J]. JOURNAL OF SHANDONG UNIVERSITY (HEALTH SCIENCES), 2015, 53(6): 23-27.

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