转录因子Sp1siRNA表达质粒的构建及对Huh-7肝癌细胞增殖的作用  

Construction of transcription factor Sp1-siRNA plasmid and its effects on liver cancer Huh-7 cells

作　　者：李宁[1] 杨丽娟[1] 周翡[1] 李琦[1] 高勇[1] 王理伟[2] 

机构地区：[1]同济大学附属东方医院肿瘤科,上海200120 [2]上海交通大学附属第一人民医院肿瘤科,上海200080

出　　处：《肿瘤》2009年第6期550-553,共4页Tumor

基　　金：上海市浦江人才计划项目(编号:06PJ14096);上海市科学技术委员会资助项目(编号054119628);上海市浦东新区卫生系统重点学科项目(编号:PWZXK2007-06)

摘　　要：目的:通过转录因子Sp1的siRNA表达质粒作用于Huh-7肝癌细胞动物模型,探讨Sp1在肝癌细胞增殖方面的作用。方法:构建靶向抑制Sp1基因的siRNA表达质粒,转染表达有绿色荧光蛋白的肝癌Huh-7细胞,筛选稳定表达的siRNA转染克隆;Western印迹法检测Sp1的表达变化,CCK8法和裸鼠皮下成瘤实验分别检测转染后Huh-7细胞在体内及体外的增殖情况。结果:Sp1-siRNA表达质粒特异性抑制Huh-7细胞Sp1的表达,筛选后得到稳定的Sp1-siRNA细胞克隆,其细胞增殖能力明显降低(P<0.05)。裸鼠皮下成瘤实验显示,质粒转染细胞接种21 d时Sp1-siRNA细胞种植瘤平均体积为(82.40±8.61)mm3,明显小于对照组的(160.00±20.27)mm3。结论:Sp1对Huh-7肝癌细胞具有重要的调控作用,siRNA抑制Sp1表达后可在体内外抑制Huh-7肝癌细胞增殖。Objective： To construct transcription factor Spl-siRNA plasmid and investigate its effect on Huh-7 liver cancer cell line both in vitro and in vivo. Methods： Spl-siRNA plasmid targeting Spl gene was constructed and transfected into Huh-7 cells carrying green fluorescence protein （GFP）. The cell line with stable expression of siRNA was selected. The expression of Spl in Huh-7 cells was determined by Western blot. The proliferation of Huh-7 cells was detected by CCK8 assay in vitro and was measured by subcutaneous tumor formation test in nude mice. Results： Spl-siRNA plasmid markedly suppressed the expression of Spl in Huh-7 cells. The proliferation capacity of screened cells with stable expression of Spl-siRNA was significantly decreased compared with control cells （P 〈 0.05 ）. Subcutaneous tumor formation test in nude mice showed that on day 21 after subcutaneous transplantation, the tumor volume was significantly smaller in Sp1-siRNA group than that in control groups [ （82.40 ± 8.61 ）mm3 vs （160.00 ± 20.27） mm3]. Conclusion： Spl plays a crucial role in regulating proliferation of Huh-7 hepatoma cells. Suppression of Spl by siRNA interference inhibited the proliferation of Huh7 cells both in vitro and in vivo.

关 键 词：癌 肝细胞 Sp1转录因子 RNA 小干扰 细胞增殖 模型 动物 

分 类 号：R735.7[医药卫生—肿瘤]