Primary Hepatocyte Culture Supports Hepatitis C Virus Replication: A Model for Infection-Associated Hepatocarcinogenesis
Primary Hepatocyte Culture Supports Hepatitis C Virus Replication: A Model for Infection-Associated Hepatocarcinogenesis
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DOI：
10.1002/hep.23616
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发表时间：
2010-06-01
期刊：
影响因子：
13.5
通讯作者：
Kumar, Ajit
中科院分区：
文献类型：
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作者：
Banaudha, Krishna;Orenstein, Jan M.;Kumar, Ajit
要分析丙型肝炎病毒感染后导致肝癌发生的肝脏基因表达的进行性变化，需要对正常分化的原代肝细胞进行长期培养。我们报告了一种支持传染性丙型肝炎病毒高效复制的原代肝细胞培养系统。通过逆转录聚合酶链式反应扩增培养物中的总细胞RNA来分析肝功能，这些培养物在无血清限定的培养液中保持长达190天。通过白蛋白、甲胎蛋白、细胞色素P4502E1、细胞角蛋白-18、1型胶原、转化生长因子-β1、基质金属蛋白酶-2、基质金属蛋白酶-13和干扰素α受体1和2的表达来评估持续的肝功能。正常分化的人原代肝细胞支持1a、1b和2a型丙型肝炎病毒感染性克隆的高效复制；抗CD81病毒进入因子抗体抑制病毒感染。病毒释放到感染丙型肝炎病毒的原代肝细胞的培养上清液中，反复传代至未成熟的肝细胞。三种丙型肝炎病毒的复制表明，在自然感染中观察到了干扰素敏感性。结论：生理性宿主细胞的持续培养可促进丙型肝炎病毒感染和进展性肝病宿主反应的研究。(《肝病》2010；51：1922-1932)
Analysis of progressive changes in hepatic gene expression that underlie hepatocarcinogenesis following hepatitis C virus (HCV) infection require examination of long-term cultures of normally differentiating primary human hepatocytes. We report a culture system of primary hepatocytes that support productive replication of infectious HCV. Hepatic functions were analyzed by reverse-transcription polymerase chain reaction amplification of total cell RNA from cultures maintained in serum-free defined medium for up to 190 days. Sustained hepatic function was assessed by expression of albumin, alpha-fetoprotein, cytochrome P4502E1, cytokeratin-18, type-1 collagen, transforming growth factor-beta 1, matrix metalloproteinase-2 (MMP-2), MMP-13, and interferon alpha-receptors 1 and 2. Normally differentiated human primary hepatocytes supported productive replication of infectious clones of HCV genotypes 1a, 1b, and 2a; virus infection was inhibited by antibodies against CD81 virus entry factor. Virus released into the culture media of HCV-infected primary hepatocytes repeatedly passage to naive hepatocytes. Replication of the three HCV genotypes shows interferon sensitivity observed in natural infections. Conclusion: Sustained cultures of physiologic host cells for the propagation of infectious HCV strains should accelerate studies of host response to HCV infection and progressive liver disease. (HEPATOLOGY 2010;51:1922-1932)