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目的探讨沉默细胞凋亡蛋白抑制剂1(cellular inhibitor of apoptosis protein 1,c IAP-1)基因对膀胱癌细胞株T24及EJ细胞化疗敏感性的影响。方法体外培养T24及EJ细胞,分别接受4.6、9.2、18.4、36.8、73.6μmol/L的表柔比星处理。构建p GCsi-H1-shRNA(沉默c IAP-1基因)及p GCsi-H1-control(对照)两种质粒载体,并分别对处于对数生长期的T24及EJ细胞进行转染。未转染的细胞作为阴性对照。采用荧光标记法及CCK8试验检测细胞凋亡情况,Western blot及实时定量聚合酶链反应(qRT-PCR)分别检测c IAP-1蛋白及mRNA的相对表达量,流式细胞术检测细胞周期的变化情况。结果转染p GCsi-H1-shRNA的T24及EJ细胞较转染p GCsi-H1-control的细胞及未转染的细胞c IAP-1 mRNA和蛋白的表达水平显著降低,差异均具有统计学意义(P均<0.05)。不同浓度表柔比星处理条件下,与未沉默c IAP-1的细胞比较,沉默c IAP-1的细胞生存率均明显降低,其细胞周期明显被阻滞在G1/G0期,细胞凋亡率显著增加(P均<0.05)。结论沉默c IAP-1基因表达可提高膀胱癌细胞对表柔比星化疗敏感性。
Objective To investigate the effect of c IAP-1 gene on chemosensitivity of bladder cancer cell line T24 and EJ. Methods T24 and EJ cells were cultured in vitro and treated with epirubicin 4.6, 9.2, 18.4, 36.8 and 73.6 μmol / L, respectively. Two plasmids, p GCsi-H1-shRNA (silenced c IAP-1 gene) and p GCsi-H1-control (control), were constructed and T24 and EJ cells in logarithmic growth phase were transfected respectively. Non-transfected cells served as negative controls. Cell apoptosis was detected by fluorescent labeling and CCK8 assay. The relative expression of c IAP-1 protein and mRNA was detected by Western blot and qRT-PCR respectively. The changes of cell cycle were detected by flow cytometry Happening. Results The expression of IAP-1 mRNA and protein in T24 and EJ cells transfected with p-GCsi-H1-shRNA was significantly lower than that in transfected p GCsi-H1-control cells and untransfected cells, the differences were statistically significant (P <0.05). Compared with the cells without silencing c IAP-1, the cell viability of silencing c IAP-1 was significantly decreased under different concentrations of epirubicin treatment, and the cell cycle was arrested in G1 / G0 phase and apoptosis Rate increased significantly (all P <0.05). Conclusion Silencing c IAP-1 gene expression may enhance the sensitivity of bladder cancer cells to chemotherapy with epirubicin.