siRNA沉默胃癌SGC-7901细胞HO1基因的细胞系的建立及其生物学行为  

Construction and functional analysis of siRNA expression vector of HO1 gene in gastric cancer SGC-7901 cells

作　　者：程力[1] 李乐平[1] 张黎[1] 苗瑞政[1] 靖昌庆[1] 郭晓波[1] 

机构地区：[1]山东大学附属省立医院胃肠外科,山东济南250021

出　　处：《基础医学与临床》2012年第11期1259-1263,共5页Basic and Clinical Medicine

基　　金：国家自然科学基金青年基金(81101858);山东省自然科学基金(Y2007C127;ZR2011HM041)

摘　　要：目的构建HO1基因的真核干扰表达载体,评估其转染人胃癌细胞系SGC-7901细胞后对HO1基因的干扰效果及其功能。方法将外源性重组真核干扰表达载体HO1基因(pS/HO1)转染到人胃癌细胞系SGC-7901内,经G418筛选并建立siRNA表达载体稳定沉默胃癌SGC-7901细胞HO1基因的细胞系,分为SGC-7901-pS/HO1组,转染空质粒细胞(SGC-7901-pS)组和未处理细胞(SGC-7901)组;用实时荧光定量PCR和蛋白印迹验证HO1基因在各组细胞中的表达,并通过CCK-8和克隆形成实验分别观察HO1基因被干扰后细胞的生物学行为。结果与SGC-7901-pS组相比,SGC-7901-pS/HO1细胞中HO1基因蛋白表达明显减少,降低了5.58倍(0.321±0.051 vs 1.675±0.153,P<0.05);与对照组相比较,SGC-7901-pS/HO1实验组较SGC-7901-pS对照组细胞增殖数量明显减少(P<0.001);与转染pS空载体的SGC-7901-pS细胞对照组相比,SGC-7901-pS/HO1细胞的克隆形成明显减少,降低了3.45倍(8.32±1.142 vs 2.32±0.362,P<0.05)。结论 HO1基因真核siRNA表达载体筛选成功,为继续深入的研究HO1基因在胃癌中的功能提供了依据。Objective To investigate the effect and function of HO1 gene in its human gastric cancer SGC-7901 cell line by constructing the eukaryotic siRNA expression vector of HO1 gene and to explore its potential role in gastric carcinogenesis.Methods A recombinant plasmid carrying siRNA specific for HO1 gene expression vector was constructed and transfected into SGC-7901 cells by lipofectin-mediated method.Cells（SGC-7901-pS/HO1） stably expressing siRNA specific for HO1 gene were selected using G418.SGC-7901 cells untransfected and those transfected with empty pS plasmid（SGC-7901-pS） were used as controls.The expression levels of HO1 gene were detected by real-time PCR and Western blot in stably transfected-SGC-7901（SGC-7901-pS/HO1） cells,biological function of HO1 gene was analyzed by cell growth activity and soft agar colony formation assay.Results Compared with SGC-7901-pS cells,the expression of HO1 protein of SGC-7901-pS/HO1 cells was an average 5.58-fold decrease（0.321±0.051 vs 1.675±0.153,P0.05）.The colony formation rate of SGC-7901-pS/HO1 cells was significantly reduced as compared to that of control group（P0.05）.The cell growth activity showed same tendency in SGC-7901-pS/HO1 group,SGC-7901-pS cells group and SGC-7901 cells group.Conclusion The eukaryotic expression vector of siRNA specific for HO1 gene was constructed successfully,which might contribute to probe the functions of HO1 in human gastric cancer in depth.

关 键 词：胃癌 HO1基因 干扰载体 SGC-7901细胞 

分 类 号：R735.2[医药卫生—肿瘤]