携TGFβ1正、反义基因载体对膀胱癌细胞体外生长的作用

Effect of retroviral vectors carried sense or antisense TGFβ1 on human bladder cancer cells in vitro

目的 :探讨表达TGFβ1正、反义基因的复制缺陷型逆转录病毒载体 ,对人膀胱癌细胞生长及细胞周期调控的作用。方法 :以逆转录病毒 pRevTRE为载体 ,构建表达TGFβ1正、反义基因的复制缺陷型逆转录病毒载体 pRevTβ和 pRevTβ AS ;分别转染膀胱癌细胞株EJ ,观察不同载体对膀胱癌细胞体外增殖、克隆形成和细胞周期时相的影响。结果 :pRevTβ、pRevTβ AS载体病毒滴度分别为 0 84、0 88× 10 5CFU/ml,对EJ细胞的整合率为10 0 % ,并有效表达 ;抑制TGFβ1表达可以降低靶细胞的生长速度和克隆形成率 ,同时出现G0 /G1期细胞比例增高和S期细胞比例降低。结论 :携TGFβ1反义基因的复制缺陷型逆转录病毒载体可以下调EJ细胞内源性TGFβ1表达 ,诱导肿瘤细胞G1期阻滞 ,抑制肿瘤细胞体外生长增殖。

Purpose:To investigate the effect of replication defective retroviral vectors carried sense or antisense TGFβ1 fragment on the cell cycle regulation and proliferation of human bladder cancer. Methods:The replication defective retroviral vectors that integrated sense or antisense bioactive fragment of transforming growth factorβ1 were constructed,and named as pRevTβ and pRevTβ-AS respectively. The influence of each vector on the cell proliferation,clone-formation and alteration of cell cycle of bladder cancer cell line EJ were observed in vitro.Results:The titre of pRevTβ and pRevTβ-AS were 0.84,0.88×10 5 CFU/ml respectively,the vectors integrated to EJ cells and expressed efficiently. Inhibition TGFβ1 gene expression reduced proliferation and clone-formation rates of EJ cells. The G 0 /G 1 stage ratios in the antisense TGFβ1-transfected EJ cells were increased,simultaneously,the S stage ratios were decreased. Conclusions:The antisense TGFβ1 vector can reduce the expression of endogenous TGFβ1 in EJ cells,induce G 1 stage arrest and inhibit proliferative growth in vitro.