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

Effects of silenced c-FLIPs gene on proliferation and apoptosis of bladder cancer cells and its mechanism

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.