HepG2 cells support viral replication and gene expression of hepatitis C virus genotype 4 in vitro

AIM： TO establish a cell culture system with longterm replication of hepatitis C virus （HCV） genome and expression of viral antigens in vitro. METHODS： HepG2 cell line was tested for its susceptibility to HCV by incubation with a serum from a patient with chronic hepatitis C. Cells and supernatant were harvested at various time points during the culture. Culture supernatant was tested for its ability to infect na＇ive cells. The presence of minus （antisense） RNA strand, and the detection of core and E1 antigens in cells were examined by RT-PCR and immunological techniques （flow cytometry and Western blot） respectively. RESULTS： The intracellular HCV RNA was first detected on d 3 after infection and then could be consistently detected in both cells and supernatant over a period of at least three months. The fresh cells could be infected with supernatant from cultured infected cells. Flow cytometric analysis showed surface and intracellular HCV antigen expression using in house made polyclonal antibodies （anti-core, and anti-E1）. Western blot analysis showed the expression of a cluster of immunogenic peptides at molecular weights extended between 31 and 45 kDa in an one month old culture of infected cells whereas this cluster was undetectable in uninfected HepG2 cells. CONCLUSION： HepG2 cell line is not only susceptible to HCV infection but also supports its replication in vitro. Expression of HCV structural proteins can be detected in infected HepG2 cells. These cells are also capable of shedding viral particles into culture media which in turn become infectious to uninfected cells.

瞄准：与丙肝的长期的复制建立一个房间文化系统病毒(HCV ) 染色体和病毒的抗原的表示在试管内。方法：HepG2 房间线被孵化与长期的丙肝从一个病人与浆液为它的危险性测试到 HCV。房间和上层清液在文化期间在各种各样的时间点被收获。文化上层清液为它感染天真的房间的能力被测试。存在减(反感觉) 在房间的核心和 E1 抗原的 RNA 海滨，和察觉被 RT-PCR 和免疫学的技术(流动血细胞计数和西方的污点) 分别地检验。结果：细胞内部的 HCV RNA 首先在 d 上被检测 3 在感染以后然后能一致地在至少三个月的一个时期上在房间和上层清液被检测。新鲜房间能从有教养的感染的房间感染上层清液。流动 cytometric 分析证明表面和在房子里使用的细胞内部的 HCV 抗原表示使 polyclonal 成为了抗体(反核心，和 anti-E1 ) 。西方的污点分析证明在分子量的产生免疫性的肽的簇的表示在一个月内在 31 和 45 kDa 之间延长了感染的房间的旧文化而这簇在 uninfected HepG2 房间是无法发现的。结论：HepG2 房间线产生 HCV 感染而且支持它的复制在试管内不仅。HCV 结构的蛋白质的表示能在感染的 HepG2 房间被检测。这些房间也能够流病毒的粒子进接着对 uninfected 房间变得传染的培养基。