【摘要】 目的建立乙型肝炎病毒X基因-丙型肝炎病毒C基因(HBV X-HCV C)共表达HepG2细胞模型,并探讨其对血管内皮细胞生长因子表达的影响。方法双酶切质粒pXT1-X,得到完整的HBV X基因片段后,将其插入到质粒PBK CMV和PBK-HCV C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-HCV C和PBK-X-C分别导入肝癌细胞株HepG2中,G418筛选,逆转录聚合酶链反应、Western blot鉴定HBV X和HCV C蛋白表达;免疫组织化学、Western blot检测血管内皮细胞生长因子蛋白质表达。结果质粒PBK-CMV、 PBK-X、PBK-HCV C和PBK X-C在HepG2细胞中有稳定表达。共表达HBV X-HCV C蛋白的细胞血管内皮细胞生长因子蛋白质表达较转染空载体的细胞及单独表达HBV X、HCV C蛋白的细胞明显升高。结论HBV X-HCV C共表达能显著上调血管内皮细胞生长因子蛋白质表达,提示HBV、HCV可能具有协同致癌作用。更多还原

【Abstract】 Objective To establish an experimental model of HCV C-HBV X co-expression protein and explore its effect on the expression of VEGF. Methods The HBV X gene was recoverd by enzyme excision and inserted into PBK-CMV and PBK-HCVC, and recombinant plasmids PBK-X and PBK-X-C were constructed. The plasmids PBK-CMV, PBK-X, PBK-HCVC and PBK-X-C were transfected into HepG2 cells with liposomes. After being selected by G418, resistant colonies were obtained. Reverse transcription PCR and Western blot were used to show HBV X and HCV core protein expression. VEGF was analyzed using immunohistochemical methods and Western blot. Results The recombinant plasmid PBK-X-C expressed HBV X and HCV core protein efficiently under the control of the vector’s promoter. VEGF and VEGFmRNA of the cells co-expressing HCV C-HBV X proteins were higher than those cells expessing HBV X, HCV C and vector alone. Conclusion HBV X-HCV C co-expression protein can increase the expression of VEGF of HepG2 cells. It suggests that HBV and HCV have a synergic action in the carcinogenesis.更多还原