目的 构建SiRNA靶向p63载体,稳定导入胆管癌细胞QBC939后,观察细胞增殖和侵袭情况的改变.方法 荧光定量PCR检测胆管癌细胞株QBC939细胞中p63表达.构建针对p63的shRNA慢病毒载体,转染胆管癌QBC939细胞中,建立稳定表达shRNA-p63胆管癌细胞株.Western blot检测干扰效率.噻唑蓝(MTT)和软琼脂集落形成实验检测细胞增殖的变化；Boyden小室检测细胞侵袭情况的改变.结果 p63基因显示在QBC939细胞中高表达.序列分析表明,重组到慢病毒载体中的shRNA序列正确.我们利用Western blot筛选了一个干扰p63表达效率为66.8％的单克隆细胞.p63表达降低不但能抑制细胞增殖体外能力,而且还能降低细胞侵袭能力.结论 p63在胆管癌QBC939细胞中高表达.SiRNA靶向抑制p63表达后,能明显降低细胞增殖和侵袭能力.%Objective To construct a small interfering RNA (siRNA) vector targeting p63 and observe its effect on the proliferation and invasiveness of human cholangiocarcinoma cells in vitro. Methods Real-time PCR was used to examine the expression of p63 in human cholangiocarcinoma QBC939 cells. The recombinant lentivirus shRNA-p63 vector was constructed and transfected into QBC939 cells via Lipofectamine 2000 to establish a cholangiocarcinoma cell line with stable expression of siRNA-p63. The interfering efficiency of the siRNA targeting p63 was assessed using Western blotting. MTT and soft agar colony formation assays were used to evaluate the changes in the cell proliferation, and Boyden test was employed to observe the cell invasiveness after the transfection. Results QBC939 cells showed a high expression of p63. The recombinant lentivirus shRNA-p63 vector was successfully constructed as verified by sequencing. Transfection with the vector significantly suppressed the proliferation and invasiveness of QBC939 cells. Conclusion Down-regulation of p63 can inhibit the proliferation and invasiveness of human cholangiocarcinoma QBC939 cells in vitro.
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