乙型肝炎病毒X基因-丙型肝炎病毒C基因融合表达蛋白对肝癌细胞胰岛素样生长因子1受体表达的影响

Effect of hepatitis B virus X gene and hepatitis C virus C gene coexpression protein on IGF-1R of HepG2 cells

目的建立HBVX-HCVC融合蛋白细胞表达模型,并探讨其对细胞胰岛素样生长因子-1的影响。方法双酶切质粒pXT1-X,得到完整的HBVX基因片段后,将其插入到质粒PBK-CMV和PBK-HCVC的相应酶切位点,得到重组质粒PBK-X和PBK-X-C;再将质粒PBK-CMV、PBK-X、PBK-HCVC和PBK-X-C分别导入肝癌细胞株HepG2中,G418筛选,RT-PCR、蛋白印迹鉴定HBVX和HCVC蛋白表达。流式细胞仪、蛋白印迹检测IGF-1受体蛋白表达。结果质粒PBK-CMV、PBK-X、PBK-HCVC和PBK-X-C在HepG2细胞中有稳定表达。表达融合蛋白的细胞的胰岛素样生长因子-1受体活性较转染空载体的细胞及单独表达HBVX、HCVC蛋白的细胞明显升高。结论HBVX-HCVC融合蛋白能显著上调胰岛素样生长因子-1受体活性,提示HBV、HCV可能具有协同致癌作用。

Objective To establish an experimental model of HCV C-HBV X coexpression protein and to explore its effect on insulin-like growth factor 1 receptor（IGF-1R）. Methods The HBV X gene was recoverd by enzyme excision, and cloned into PBK- CMV and PBK-HCV C for getting the recombine plasmids PBK-X and PBK-X-C. The plasmids PBK-CMV, PBK-X, PBK-HCV C and PBK-X-C were transfected into HepG2 cells with liposome, After selected with G418, resistant colonies were obtained. The reverse transcription PCR and Western blot was analyzed to show HBV X and HCV core protein expressed. IGF-1R were analyzed by reverse transcription PCR and Western blot, Results The recombinant plasmid PBK-X-C could express HBV X and HCV core protein efficiently under the control of vector＇s promoter, IGF-1R of the cells coexpressed HCV C-HBV X protein was higher than that of the cells expessed HBV X, HCV C and vector along, Conclusion HBV X -HCV C coexpression protein can increase the expression of IGF-1R in HepG2 cells. It suggests that HBV and HCV can cooperate with carcinogenesis,