人胎肝干细胞体外感染乙型肝炎病毒模型的建立  

IN VITRO INFECTION OF HUMAN FETAL HEPATIC STEM CELLS WITH HEPATITIS B VIRUS

作　　者：王姝[1] 金敏[1] 郭向飞[1] 王新为[1] 邱志刚[1] 谌志强[1] 李君文[1] 

机构地区：[1]军事医学科学院卫生学环境医学研究所天津市环境与食品安全风险监控技术重点实验室,天津300050

出　　处：《解放军预防医学杂志》2011年第3期168-172,共5页Journal of Preventive Medicine of Chinese People's Liberation Army

摘　　要：目的建立一种稳定的基于人胎肝干细胞的维持乙型肝炎病毒（HBV）复制和传代的体外细胞培养模型,为深入研究乙肝病毒的生命周期和发病机制提供必要的工具。方法原代分离12-20周胎龄的胎肝干细胞,用乙肝病毒阳性血清感染胎肝干细胞;荧光定量PCR追踪检测细胞上清和细胞内乙肝病毒DNA及细胞内乙肝病毒复制中间体—闭合环状双链DNA（cccDNA）;化学发光法检测HBsAg、HBeAg分泌情况;免疫细胞化学法鉴定HBcAg表达情况;原位杂交检测细胞内病毒DNA。结果在经HBV感染的细胞上清液中可持续检测出乙肝病毒DNA、HBsAg和HBeAg,细胞内可检测出HBV-cccDNA和HBcAg。结论乙肝病毒可感染胎肝干细胞,并在细胞内增殖、分泌、传代,这一模型可模拟乙肝病毒自然感染人体的病理生理过程,为HBV感染机制研究和抗病毒药物的研发提供了必要的工具。Objective To establish a stable in vitro cell culture model to keep hepatitis B virus（HBV） replication and passage,in order to provide a useful tool for studying the life cycle of HBV and pathogenesis of HBV infection.Methods Human fetal hepatic stem cells were isolated from aborted fetuses of 12~20 weeks,and infected with sera from HBV-infected patients.Then HBV DNA,and cccDNA were detected by real-time PCR.Chemiluminescent method and immunohistochemical staining were used to examine HBsAg,HBeAg,HBcAg expression,and in situ cytohybridization of infected cells to detect the HBV DNA in the cells.Results HBV DNA,HBsAg,HBeAg in the culture medium and HBcAg,HBV-ccc DNA in the cells could be detected continually during the time of HBV infection.Conclusion A stable HBV cell culture infection system based on the human fetal liver stem cells was established,in which infection,replication and secretion of virus in the cells could occur.This model could simulate the pathophysiology of natural infection with HBV and provide a useful tool for the development of effective antiviral drug.

关 键 词：乙型肝炎病毒 胎肝干细胞 细胞培养 感染 复制 表达 

分 类 号：R394.4[医药卫生—医学遗传学] R373.2[医药卫生—基础医学]