体外RNA干扰抑制RegⅣ基因表达对胃癌细胞增殖及凋亡的影响  

Influence of down-regulation of RegⅣ expression by small interfering RNA on proliferation and apoptosis of gastric cancer cells

作　　者：刘兵[1] 项明[1] 于颖彦[1] 杨秋蒙[1] 蔡劬[1] 陈雪华[1] 李建芳[1] 刘炳亚[1] 朱正纲[1] 

机构地区：[1]上海交通大学医学院附属瑞金医院普外科上海消化外科研究所,上海市200025

出　　处：《世界华人消化杂志》2009年第6期549-553,共5页World Chinese Journal of Digestology

基　　金：国家科技部"十一五""863"重大专项基金资助项目;No.2006AA02A402;No.2006AA02A301;国家自然科学基金资助项目;No.30572127;No.30770961;No.30670939;上海市浦江人才计划基金资助项目;No.PJ200700367;上海市科委重点基础研究基金资助项目;No.05JC14013~~

摘　　要：目的:拟探讨RegⅣ基因在胃癌细胞中的表达及干扰该基因对胃癌细胞增殖和凋亡的影响.方法:应用实时定量PCR方法检测九株胃癌细胞株中RegⅣ基因的mRNA表达.针对RegⅣ基因设计三条小干扰RNA片段(siRNA1、siRNA2、siRNA3),瞬时转染高表达胃癌细胞株,实时定量PCR方法检测转染后RegⅣ基因的mRNA的表达水平,CCK-8法检测细胞增殖能力,流式细胞仪检测细胞凋亡.结果:以永生化正常胃黏膜细胞株GES-1作为参照,除MKN-45和SNU-1胃癌细胞株外,其余胃癌细胞株中RegⅣ表达水平均升高5倍以上,尤以SNU-16细胞株明显,其RegⅣ的表达水平高出GES-1细胞数千倍,遂选用SNU-16细胞进行RNA干扰.合成的三对siRNA对RegⅣ基因表达均有明显抑制作用,抑制率分别为79.3%、77.4%和60.4%.选用siRNA1干扰SNU-16细胞株后96h和120h,其细胞增殖率受到明显抑制(P=0.0057,0.0173).流式细胞仪检测显示72h细胞凋亡率明显增加(P=0.0231).结论:干扰RegⅣ基因具有抑制胃癌细胞增殖,促进凋亡的作用,RegⅣ基因可能成为胃癌基因靶向治疗的分子靶点.AIM： To investigate RegIV expression level in human gastric cancer cell lines and the effect of RNA interfering of RegIV on proliferation and apoptosis of gastric cancer cells. METHODS： The expression levels of ReglV in nine gastric cancer cell lines were examined by real time PCR, and 3 small interfering RNA （siR- NA1, siRNA2, siRNA3） targeting ReglV were designed and transfected into gastric cancer cell lines. The changes of ReglV mRNA expression level after siRNA interfering were detected by real time PCR. The proliferation was assayed by CCK-8 method. Flow cytometry was used to de- tect apoptosis of gastric cancer cells. RESULTS： Compared with gastric mucosa cell line GES-1, the expression level of ReglV in gastric cancer cells was 5-fold or higher except MKN-45 and SNU-1. The expression level of RegIV in SNU-16 was the highest and was several thousand-fold higher than that in GES-1. SNU-16 was used for siRNA experiment. Three siRNAs showed notably downegulated expres- sion of RegIV mRNA levels with inhibitory rate of 79.3%, 77.4% and 60.4%, respectively in comparison with that in control group. So siRNA1 was used to do cell proliferation assay. After 96 hours＇ and 120 hours＇ transfection of siRNA1, the proliferation of SNU-16 cell significantly decreased compared with the control group （P = 0.0057, 0.0173, respectively）. The results of flow cytometry revealed that 72 h after transfection with siRNA1, the apoptosis rate of SNU-16 significantly increased. CONCLUSION： Interfering and down-regulating ReglV gene can inhibit proliferation and promote apoptosis of gastric cancer cells, indicating that RegIV gene is probably a target for gastric cancer gene therapy.

关 键 词：胃癌 REGIV RNA干扰 生长抑制 凋亡 

分 类 号：R735.2[医药卫生—肿瘤] R733.71[医药卫生—临床医学]