HCV核心基因转染的胆管癌细胞中p53的表达及意义

目的：探讨丙肝病毒感染在胆管癌中的致癌机理。方法：通过分子克隆技术构建HCV－C基因重组质粒（PBK－HCVc)，经酶切鉴定后，将其转染到胆管癌细胞（QBC939）中，经G418选择培养，免疫细胞学，Western blotting鉴定其表达，透射电镜观察细胞形态学改变。并用免疫细胞学检测p53的表达。结果：构建的PBK－HCVc质粒在QBC939细胞中有稳定表达，透射电镜见胞质空泡内球型病毒颗粒，病毒颗粒直径50－80nm，有外膜结构。HCV核心基因转染后QBC939细胞的p53表达比转染前明显增多（P＜0．01）。结论：本实验为进一步探讨丙肝病毒感染与肝门部胆管癌中的致癌机理提供了理想的细胞株模型，HCV核心基因可诱发p53突变，可能与丙肝病毒感染的胆管细胞癌变有关。

Expression of p53 in HCV core gene transfected cholangiocarcinoma and its significance

Objective To establish a model to explore the role of HCV in development of cholangiocarcinoma. Methods Recombinant plasmid of HCV-C gene was constructed by molecular cloning technique and identified by restricted enzyme map. Then it was transfected into QBC939 cells with lipofection. Resistant colonies were obtained after the plasmid was selected with G418 made by the necomycin and kanamycin resistance gene. The colonies were analyzed by immunohistochemistry and Western blotting. The morphology was observed with electron microscopy (EM) and the expression of p53 with immunohistochemistry. Results The recombinant plasmid could effectively express HCV-C protein in QBC939 cells. The HCV-like, spherical particles with a diameter of 50-80 nm possessing outer membrane were seen in the cytoplasm by EM. There was statistical difference in expression of p53 between HCV-core-transfected cells and non-transfected ones ( P <0.01). Conclusions Since HCV-C gene can steadily express in cholangiocarcinoma cells, the transfected tumor cells (QBC939-HCVc) can be used to study the effect of HCV on development of cholangiocarcinoma. The infection of HCV might be related to p53 protein expression and carcinogenesis.