Survivin基因对膀胱癌细胞体内生长的影响

目的探讨通过RNA干扰下调Survivin基因对膀胱癌细胞在体内生长的抑制作用。方法构建Survivin基因shRNA真核表达载体pRNAT-U6.1/neo-survivin,转染人膀胱癌细胞T24并获得稳定的单克隆细胞。观察裸鼠皮下移植瘤内注射靶向Survivin的siRNA及稳定转染pR- NAT-U6.1/neo—survivin对膀胱癌细胞裸鼠皮下移植瘤生长的影响,肿瘤组织行免疫组织化学检测Survivin蛋白表达。结果经鉴定成功构建靶向Survivin的shRNA真核表达载体pRNAT-U6.1/neo-survivivn,稳定转染重组质粒的T24细胞Survivin mRNA表达下调达92.86%;瘤内注射50 mg siRNA-Survivin能够明显抑制肿瘤生长,与对照比较差异有统计学意义(P<0.01);未转染组在20 -29 d均发展为体积超过2000 mm3的肿瘤,转染组在43 d时形成3个体积不超过62.5 mm3肿瘤;免疫组织化学显示转染组肿瘤Survivin蛋白表达明显降低。结论裸鼠移植瘤内注射siRNA-sur- vivin明显抑制了肿瘤的生长;靶向Survivin的shRNA持久地抑制了膀胱癌T24细胞在体内的生长; Survivin基因可作为膀胱癌基因治疗的良好靶点。

Effect of Survivin gene on bladder cancer cell growth in vivo

Objective To study the inhibitory effect of RNA interference targeting Survivin on bladder cancer cell T24 growth in vivo. Methods Survivin-targeting shRNA eukaryotic expression vector pRNAT-U6. 1/neo-survivin was constructed and identified. Human bladder cancer cell T24 was transfect-ed with pRNAT-U6. 1/neo-survivin and then stable monoclonal cells were selected. The inhibitory effect of siRNA targeting Survivin on ectopic human bladder cancer was observed by intratumoral injection of siR-NA/liposome complex. Meanwhile stably transfected and untransfected cells were inoculated respectively into subcutaneous tissue of nude mice, and tumor growth status was recorded. Survivin protein expression of tumors was detected by immunohistochemistry. Results The shRNA eukaryotic expression vector was successfully constructed. Survivin mRNA expression in stably tansfected cells was downregulated by 92. 86% as compared with untransfected cells. In vivo intratumoral injection of siRNA-survivin (50 mg) could notably prevent the growth of ectopic bladder cancer (P < 0. 01). The tumor volume in the untransfected group exceeded 2000 mm3 between 20 and 29 days after inoculation, but that in the transfected group had only three tumors no more than 62. 5 mm3 after additional 2 weeks. Survivin protein expression of the tumors in transfected group was weaker than that in untransfected group. Conclusion The application of siRNA-survivin could markedly inhibit ectopic tumor growth. The shRNA targeting Survivin permanently inhibited bladder cancer cell T24 growth in vivo. Survivin may act as a valuable target for gene therapy.