MiRNA-25在非小细胞肺癌吉非替尼耐药细胞中的表达及其作用机制  

Expressions of Micro RNA-25 in Gefitinib-resistant Non-small Cell Lung Cancer Cells and Its Mechanism

作　　者：周建英[1] 刘震天[1] 陈颖兰[1] 

机构地区：[1]江西省肿瘤医院,330029

出　　处：《实用癌症杂志》2016年第10期1567-1571,共5页The Practical Journal of Cancer

基　　金：江西省卫生厅科技计划资助项目(编号:20131206)

摘　　要：目的探讨miRNA-25在非小细胞肺癌(NSCLC)吉非替尼耐药细胞中的表达及其作用机制。方法采用RT-PCR法检测miRNA-25在NSCLC吉非替尼耐药细胞PC9/G及亲本细胞PC9中的表达;采用体外转染法将miRNA-25模拟物或抑制物分别转染PC9和PC9/G细胞,采用CCK8实验检测转染前后细胞对吉非替尼敏感性的变化;并用Westem blot检测转染前后细胞中Bim的表达变化。采用双荧光素酶报告基因验证miRNA-25是否作用于Bim基因的3'-UTR区预测靶位。结果 miRNA-25在PC9/G细胞中的表达水平是PC9细胞的(7.15±1.26)倍(P<0.01)。PC9细胞转染miRNA-25模拟物后,吉非替尼对PC9细胞增殖的抑制率较转染前明显下降(P<0.01),细胞内Bim表达水平较转染前明显下降(P<0.05)。PC9/G细胞在转染miRNA-25抑制物后,吉非替尼对PC9/G细胞增殖的抑制率与转染前比较明显增加(P<0.01),细胞内Bim表达水平较转染前明显增高(P<0.05)。经双荧光素酶报告基因验证Bim是miRNA-25的靶基因。结论 miRNA-25在NSCLC耐药细胞PC9/G中异常高表达,下调其表达可部分逆转细胞耐药性,miRNA-25可能通过作用于Bim参与NSCLC细胞吉非替尼耐药的发生和发展。Objective To investigate the expression of microRNA-25（miRNA-25） in gefitinib-resistant non-small cell lung cancer cells and its mechanism.Methods Real-time PCR（RT-PCR） was used to detect the expression of miRNA-25 in gefitinib-resistant non-small cell lung cancer cells line PC9/G and its parent cells line PC9.miRNA-25 mimics and miRNA-25 inhibitor in vitro were transiently transferred into PC9 and PC9/G cells respectively.The expression level of miRNA-25 was detected by RT-PCR.Cell viability was analyzed by CCK8 assay.Expression of Bim protein was detected by western blot.Bioinformatics software predicted the potential target genes of miRNA-25 and dual luciferase reporter gene was used to analyze the binding between miRNA-25 and 3’-UTR of Bim.Results The expression level of miRNA-25 was up-regulated in PC9/G cells as compared with PC9 cells（P〈0.01）.Over-expression of miRNA-25 inhibited gefitinib chemosensitivity（P〈0.01）,and the up-regulation of miRNA-25 led to a significant decrease in the Bim protein levels（P〈0.05）.Low-expression of miRNA-25 promoted gefitinib chemosensitivity（P〈0.01）,and the down-regulation of miRNA-25 led to a significant increase in the Bim protein levels（P〈0.05）.Dual luciferase reporter gene assay indicated that miRNA-25 regulated Bim expression by binding to the 3’-UTR of Bim mRNA.Conclusion The expression of miRNA-25 is up-regulated in PC9/G cells.Down-regulation of the expression of miRNA-25 can partially reverse the gefitinib-resistance.miRNA-25 may play a vital role in drug resistance by targeting Bim.

关 键 词：非小细胞肺癌 吉非替尼 耐药 miRNA-25 

分 类 号：R734.2[医药卫生—肿瘤]