电压门控钾通道Kv1.5反义核酸对胃癌细胞SGC7901生长的抑制作用  

Voltage gated potassium channel Kv1.5 in the retardation of growth of gastric cancer cells

作　　者：韩者艺[1] 吴开春[1] 蒋海萍[1] 申慧琴[1] 时永全[1] 韩英[1] 刘长江[1] 薛妍[1] 刘娜[1] 郭长存[1] 乔泰东[1] 樊代明[1] 

机构地区：[1]第四军医大学西京医院

出　　处：《解放军医学杂志》2004年第9期773-775,共3页Medical Journal of Chinese People's Liberation Army

基　　金：国家自然科学基金创新群体基金 (编号 30 0 30 1 4 0 );国家自然科学基金(编号 30 1 0 0 0 79)资助课题

摘　　要：目的　构建电压门控钾通道Kv1 5分子的反义核酸 (Kv1 5AS)的真核表达载体 ,建立Kv1 5AS转染的胃癌SGC790 1细胞亚系 ,探讨Kv1 5分子对胃癌细胞生长的抑制作用。方法　通过DNA重组技术 ,将Kv1 5全长cDNA片段从 pBK Kv1 5载体反向亚克隆至真核表达载体pcDNA3 1 ,构建表达Kv1 5反义核酸的重组载体 pcDNA3 1 Kv1 5AS。借助脂质体将pcDNA3 1 Kv1 5AS转导入胃癌细胞SGC790 1。通过MTT实验绘制细胞生长曲线 ,肿瘤细胞集落形成实验检测集落形成率 ,流式细胞仪进行细胞周期分析 ,并利用Westernblot检测Kv1 5和CyclinD1蛋白在基因转染细胞中的表达。结果　限制性酶切鉴定结果提示 ,重组载体 pcDNA3 1 Kv1 5AS构建成功。Westernblot结果表明 ,转染pcDNA3 1 Kv1 5AS后 ,Kv1 5蛋白在SGC790 1细胞中的表达显著降低。与转染空载体的对照细胞相比 ,转染Kv1 5AS表达载体后 ,SGC790 1细胞生长变缓 ,集落形成率显著降低 ,细胞发生了G1 期阻滞。Westernblot结果证实 ,CyclinD1蛋白在Kv1 5AS转染细胞中表达降低。结论　电压门控钾通道Kv1 5反义核酸可抑制胃癌细胞SGC790Objective To investigate the retardation effect of voltage gated potassium channel Kv1 5 on growth of gastric cancer cells SGC7901. Methods By using restriction enzymes of Hind Ⅲ and Kpn I, cDNA encoding Kv1 5 was reversely constructed into eukaryotic expression vector pcDNA3 1. SGC7901 cells were transfected with the recombinants using LipofectAMINE2000. Stable clones of transfectants were obtained after selection by G418 The growth of cells was monitored by cell growth curve and cell colony formation. The effect of Kv1 5 protein on cell cycle was examined by flow cytometry. Expression of Cyclin D1 protein was detected by Western blot. Results The antisense was found to effectively inhibit the expression of Kv1 5 protein in the transfectants. The growth and colony formation of transfectants were significantly reduced as compared with controls. The cell cycle review showed retardation of transfectants with Kv1 5 antisense in the G 1 phase. Expression of Cyclin D1 protein was decreased in Kv1 5 antisense transfectants. Conclusion The antisense of Kv1 5 has effect of retardation on growth of gastric cancer cells SGC7901

关 键 词：胃癌细胞 反义核酸 SGC7901细胞 转染 表达载体 pcDNA3 电压门控钾通道 限制性酶切 亚克隆 DNA片段 

分 类 号：R735.2[医药卫生—肿瘤] R737[医药卫生—临床医学]