靶向沉默c-FLIPs基因表达对膀胱癌细胞增殖及凋亡的影响及机制

摘要

目的 探讨抑制c-FLIPs基因表达对膀胱癌细胞增殖及凋亡的影响及机制.方法 RT-PCR及Western blot分别检测膀胱癌组织及细胞中c-FLIPs基因的表达;si-c-FLIPs转染人膀胱癌T24细胞,同时转染正常对照组(normal)和阴性对照组(si-control),48 h后Western blot检测c-FLIPs、Cleaved caspase 8、β-链蛋白(β-catenin)、细胞周期蛋白(Cyclin D1)蛋白表达;细胞计数试剂盒-8(CCK8)实验检测细胞增殖;流式细胞仪检测细胞凋亡.结果 膀胱癌组织及细胞中c-FLIPs基因的表达均显著高于癌旁组织(P<0.01).T24细胞中c-FLIPs基因的表达最高,选择作为研究对象;转染si-c-FLIPs后FLIPs的表达显著降低;si-c-FLIPs组细胞存活率及β-catenin和Cyclin D1蛋白表达显著低于normal组,细胞凋亡率及Cleaved caspase 8蛋白表达显著高于normal组(P<0.01).结论 抑制膀胱癌中c-FLIPs基因表达可降低癌细胞的增殖及诱导细胞的凋亡,其机制与下调Wnt/β-catenin信号通路有关.%Objective To investigate the effects of silenced c-FLIPs gene on proliferation and apoptosis of bladder cancer cells and its mechanism.Methods RT-PCR and Western blot were used to detect the expression of c-FLIPs in bladder cancer tissues;si-c-FLIPs.transfected human bladder cancer T24 cells, and transfected normal control group (normal) and negative control group (Si-Control), the expression of c-FLIPs, Cleaved caspase8, β-catenin, Cyclin D1 protein were detected after 48 h by Western blot.Cell proliferation was detected by CCK8 experiment;cell apoptosis were determined by flow cytometry.Results The expression of c-FLIPs in bladder cancer tissues and cells was significantly higher than that in adjacent tissues (p<0.01).The highest c-FLIPs gene expression in T24 cells, and selected as the research object;the expression of FLIPs after transfected si-c-FLIPs significantly decreased;the survival rate and c-FLIPs, β-catenin, Cyclin D1 protein expression in si-c-FLIPs group was significantly lower than those in normal group, cell apoptosis rate and Cleaved caspase8 protein expression was significantly higher than those in normal group (p<0.01).Conclusion The inhibition of c-FLIPs gene's expression in bladder cancer can reduce the proliferation of cancer cells and induce apoptosis, which is related to the down-regulation of Wnt/β-catenin signal pathway.