Hepatitis B virus infection and replication in primarily cultured human fetal hepatocytes
Hepatitis B virus infection and replication in primarily cultured human fetal hepatocytes
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DOI：
10.3748/wjg.v13.i7.1027
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发表时间：
2007-02-21
影响因子：
4.3
通讯作者：
Chen, Mo-Rui
中科院分区：
文献类型：
--
作者：
Lin, Min;Chen, Qun;Chen, Mo-Rui
目的：探讨乙型肝炎病毒（HBV）在原代培养的人胎儿肝细胞（HFHs）中的感染和复制情况。方法：在无血清培养基中培养人胎肝细胞，加入HBV阳性血清，研究肝细胞对HBV感染的易感性。每天收集上清进行HBsAg、HBeAg ELISA检测，HBV-DNA荧光定量PCR检测。免疫组化法检测培养肝细胞白蛋白、HBcAg、CK8、CK18的表达。测定乳酸脱氢（LDH）的含量，以确定细胞膜的完整性。结果：建立了稳定的肝细胞培养体系。HBV可感染肝细胞并复制，肝细胞样细胞免疫组化检测HBcAg表达。HBV感染后第2 ~ 18天上清液中均可检测到HBV- dna，第3 ~ 18天HBsAg和HBeAg均呈阳性。培养基中的HBV从第0天到第6天增加，随后随着细胞逐渐失去其肝细胞表型而减少。结论：HBV可感染人胎儿肝细胞并进行复制。该体外模型允许对人类HBV进入细胞和随后复制相关的早期事件进行详细研究。(c) 2007年WIG出版社。版权所有。
AIM: To investigate the infection and replication of hepatitis B virus (HBV) in primarily cultured human fetal hepatocytes (HFHs).METHODS: The human fetal hepatocytes were cultured in serum-free medium, HBV-positive serum was added into the medium to study the susceptibility of hepatocytes to HBV infection. The supernatant was collected for ELISA assay of HBsAg and HBeAg, and quantitative fluorescence PCR for HBV-DNA assay daily. Albumin and HBcAg, CK8 and CK18 expressions were detected by immunohistochemistry in cultured hepatocytes. Content of lactate dehydrogenate (LDH) was measured to find out the integrity of the cell membrane.RESULTS: A stable hepatocyte culture system was established. HBV could infect the hepatocytes and replicate, and HBcAg expression could be detected by immunohistochemistry in hepatocyte-like cells. HBV-DNA in the supernatant could be detected from d 2 to d 18 and HBsAg and HBeAg were positive on d 3-d 18 after HBV infection. HBV in medium increased from d 0 to d 6 and subsequently decreased as the cells were progressively loosing their hepatocyte phenotypes.CONCLUSION: HBV could infect human fetal hepatocytes and replicate. This in vitro model allowed a detailed study on early events associated with human HBV entry into cells and subsequent replication. (c) 2007 The WIG Press. All rights reserved.