人肝癌细胞系7721丙型肝炎病毒体外感染模型的建立

目的　建立接近体内自然感染状态并能稳定支持丙型肝炎病毒（ＨＣＶ） 体外长期复制的感染细胞模型。方法　将人肝癌细胞系７７２１ 与慢性丙型肝炎患者血清共温育８ 小时后,分别以逆转录 聚合酶链反应、免疫组化、原位杂交检测细胞和／ 或培养上清中的ＨＣＶ ＲＮＡ、ＨＣＶ 抗原表达及ＨＣＶＲＮＡ 在感染细胞内的定位。结果　感染血清和细胞共同温育后的第２ ～６６ 天,从细胞内和培养上清中均可间断地检测出ＨＣＶ 正、负链ＲＮＡ,即使其间细胞传代６ 次,ＨＣＶ ＮＳ３ 、ＮＳ５ 抗原在细胞内能稳定表达,原位杂交显示ＨＣＶ ＲＮＡ 阳性物质多位于细胞浆。结论　７７２１ 细胞不但对ＨＣＶ 易感,而且可以稳定地支持ＨＣＶ 的体外长期复制,此模型可以用于ＨＣＶ 感染、复制机制的研究、抗病毒药物的评价及保护性抗体／ 疫苗的初步筛选。

Establishment of HCV infection model in vitro using a human liver cell line 7721

Objective To establish a cell model that is close to the HCV replication in vivo and can support long-term HCV replication in vitro.Methods A human liver cell line 7721 was inoculated with serum from a chronic hepatitis C patient for 8 hours.After incubation,the presence of HCV RNA,the expression of HCV antigen and the location of HCV RNA in cells and/or supernatant were detected by RTPCR,immunohistochemistry and in situ hybridization respectively.Results It was found that plus-and minus-strand of HCV RNA could be discontinuously detected in both cells and supernatant as late as 66 days after inoculation even if cells had been subcultured for 6 times.HCV NS3、NS5 antigens could be expressed in cells and HCV RNA was mainly located within cytoplasm.Conclusion The above results suggested that 7721 cell line was not only susceptible to HCV but also could support its long-term replication in vitro.This HCV infection model in vitro was proved to be a useful tool for studying the mechanism of HCV infection and replication,as well as evaluating the antiviral compounds and screening the protective antibodies/vaccines.