外源性p16 基因导入对人膀胱癌细胞的作用  

Multiple inhibitory mechanisms of wild type p16 gene transfer into human bladder cells by retroviral vector

作　　者：武玉东[1] 马腾骧[1] 吴海[1] 张文颖[1] 吴长利[1,2] 

机构地区：[1]天津市泌尿外科研究所,300211 [2]河南医科大学第一附属医院泌尿外科

出　　处：《中华外科杂志》1999年第8期456-458,共3页Chinese Journal of Surgery

基　　金：天津市卫生局科研基金

摘　　要：目的　通过外源性野生型ｐ１６ 基因导入内源ｐ１６ 基因纯合性缺失及具有内源性ｐ１６ 基因表达的膀胱癌细胞系２５３ Ｊ和 Ｅ Ｊ，观察对肿瘤细胞的生长抑制作用，并探讨其作用机制。　方法　构建ｐ１６ 逆转录病毒表达载体，经包装细胞 Ｐ Ａ３１７ 包装，转染膀胱癌细胞系；应用 Ｎｏｒｔｈｅｒｎ 杂交及免疫细胞化学方法检测外源ｐ１６ 基因的表达；流式细胞仪检测细胞周期；原位细胞凋亡检测及透射电镜观察细胞凋亡；并对导入外源ｐ１６ 基因的细胞进行裸鼠致瘤能力及病理学特性分析。　结果　转染外源ｐ１６ 基因的肿瘤细胞生长速率明显下降，细胞被抑制在 Ｇ０ 、 Ｇ１ 期，裸鼠致瘤能力明显降低。其中转基因的 Ｅ Ｊ细胞 Ｈｒａｓ 基因ｍ Ｒ Ｎ Ａ 表达明显降低；而转基因的２５３ Ｊ细胞出现细胞凋亡。　结论　外源ｐ１６ 基因导入可通过不同的分子机制抑制膀胱肿瘤细胞的恶性增殖和促进细胞的分化。Objective To investigate the inhibitory mechanisms of wild type p16 gene transfer into human bladder cancer cells with or without endogenous p16 gene expression. Methods p16 recombinant retrovirus vector was constructed and transfected in human bladder cancer cell EJ and 253J. Northern blot analysis and immunocytochemistry were used to detect the expression of exogenous p16 gene. Flow cytometry analysis was used to detect tumor cell cycle.Electronic microscope and in situ labelling apoptotic DNA fragment detection (TUNEL) was used to detect tumor cell apoptosis after gene transfection. Tumorigenicity was observed in nude mice. Results The growth rate of the transfected EJ cells was significantly retarded, the majority of transfected EJ cells were arrested on G 0+G 1 phases of cell cycle, tumorigenicity of the transfected cells was reduced on nude mice. H ras gene expression in transfected EJ cells reduced as compared with nontransfection.Apoptotic cells were observed in transgenic 253J cells. Conclusions Malignant proliferation of bladder cancer cells can be inhibited by exogenous p16 gene transfection via different molecular pathway.

关 键 词：膀胱癌细胞 P16基因 外源性基因导入 

分 类 号：R737.140.2[医药卫生—肿瘤]