Drosha通过调控微RNA-195-5P促进胃癌细胞的迁移和侵袭  

Drosha induces migration and invasion of gastr ic cancer cells via regulating microRNA-195-5P

作　　者：彭美茜 徐丽云[1] 杨丹 赵茂嘉 秦旖璐 刘水清 曾欢 柳满然[1] PENG Meixi, XU Liyun, YANG Dan, ZHAO Maojia, QIN Yilu, LIU Shuiqing, ZENG Huan, LIU Manran(Key Laboratory of Laboratory Medical Diagnostics, Ministry of Education, Chongqing Medical University, Chongqing 400016, China)

机构地区：[1]重庆医科大学检验医学院临床检验诊断学教育部重点实验室,重庆400016

出　　处：《肿瘤》2018年第10期925-932,共8页Tumor

基　　金：国家自然科学基金资助项目(编号:81172296;31671481)~~

摘　　要：目的 :探讨Drosha对胃癌MGC-803和SGC-7901细胞迁移和侵袭能力的影响及其可能的作用机制。方法:将携带有Drosha-shRNA的重组慢病毒感染胃癌MGC-803和SGC-7901细胞,以感染携带有阴性对照(negative control,NC)-shRNA的重组慢病毒作为对照。采用实时荧光定量PCR法和蛋白质印迹法检测转入Drosha-shRNA后对MGC-803和SGC-7901细胞中Drosha mRNA和蛋白表达的影响,Transwell小室迁移和侵袭实验分别检测Drosha下调对MGC-803和SGC-7901细胞迁移和侵袭能力的影响;实时荧光定量PCR检测Drosha下调的MGC-803和SGC-7901细胞中微RNA-195-5P(microRNA-195-5P,miR-195-5P)的表达水平。在Drosha下调的胃癌细胞中采用脂质体法同时转入miR-195-5P-inhibitor,实时荧光定量PCR检测miR-195-5P的下调效果,Transwell小室迁移和侵袭实验分别检测下调miR-195-5P表达对细胞迁移和侵袭能力的影响。采用脂质体法将miR-195-5P-mimics转染MGC-803和SGC-7901细胞,实时荧光定量PCR检测miR-195-5P过表达的效果,Transwell小室迁移和侵袭实验分别检测miR-195-5P过表达对胃癌细胞迁移和侵袭能力的影响。结果 :感染Drosha-shRNA重组慢病毒后,MGC-803和SGC-7901细胞中Drosha mRNA和蛋白表达水平均较对照组明显下调(P值均<0.01),细胞的迁移和侵袭能力均明显被抑制(P值均<0.05),miR-195-5P的表达水平明显上调(P值均<0.001)。成功在Drosha下调的胃癌MGC-803和SGC-7901细胞中下调miR-195-5P的表达水平(P值均<0.05),miR-195-5P表达下调可以恢复Drosha下调对胃癌细胞迁移和侵袭能力的影响(P值均<0.05)。成功将miR-195-5P-mimics瞬时转入MGC-803和SGC-7901细胞,MGC-803和SGC-7901细胞中miRNA-195-5P的表达水平明显上调(P值均<0.01),细胞的迁移和侵袭能力均明显降低(P值均<0.01)。结论 :Drosha可促进胃癌细胞的迁移和侵袭能力。Drosha与miR-195-5P的表达呈负相关性,Drosha通过调控miR-195-5P的表达促进胃癌细胞的迁移和侵袭。Objective： To investigate the effects of Drosha on the migration and invasion of gastric cancer MGC-803 and SGC-7901 cells, and to explore the possible mechanism.Methods： MGC-803 and SGC-7901 cells were infected with the recombinant lentivirus carrying Drosha-shRNA or the negative control（NC）-shRNA（as the control）. The expression levels of Drosha mRNA and protein in MGC-803 and SGC-7901 cells were detected by real-time fluorescent quantitative PCR and Western blotting, respectively. The migration and invasion abilities of MGC-803 and SGC-7901 cells with Drosha gene silencing were detected by Transwell assay, and the expression level of microRNA-195-5P（miR-195-5P） was detected by real-time fluorescent quantitative PCR. After Drosha gene silenced cells were transfected with miR-195-5P-inhibitor, the down-regulation of miR-195-5P expression was tested by real-time fluorescent quantitative PCR, then the migration and invasion of gastric cancer cells were detected by Transwell assay. After miR-195-5P-mimics or the control-mimics were transfected into MGC-803 and SGC-7901 cells, the expression level of miR-195-5P was tested by real-time fluorescent quantitative PCR, then the migration and invasion of gastric cancer cells were detected by Transwell assay.Results： The expression levels of Drosha mRNA and protein in MGC-803 and SGC-7901 cells infected with the recombinant lentivirus carrying Drosha-shRNA were down-regulated as compared with the control groups（both P〈 0.01）. The migration and invasion abilities of MGC-803 and SGC-7901 cells with Drosha gene silencing were inhibited（all P〈 0.05）, and the expression level of miR-195-5P was up-regulated（both P〈 0.001）. The miR-195-5P expression level was down-regulated in MGC-803 and SGC-7901 cells with Drosha gene silencing after transfection with miR-195-5P-inhibitor（both P〈 0.05）, then the migration and invasion abilities of gastric cancer cells were rescued（all P〈 0.05）. The miR-195-5P expression level was upregulated in MGC

关 键 词：胃肿瘤 细胞运动 基因表达 DROSHA 微RNA-195-5P 

分 类 号：R735.2[医药卫生—肿瘤]