Expression of paramyxovirus V proteins promotes replication and spread of hepatitis C virus in cultures of primary human fetal liver cells

Abstract

Here, we demonstrate that primary cultures of human fetal liver cells (HFLC) reliably support infection with laboratory strains of HCV, although levels of virus replication vary significantly between different donor cell preparations and frequently decline in a manner suggestive of active viral clearance. To investigate possible contributions of the interferon (IFN) system to control of HCV infection in HFLC, we exploited the well-characterized ability of paramyxovirus (PMV) V proteins to counteract both IFN induction and antiviral signaling. The V proteins of measles virus (MV) and parainfluenza virus 5 (PIV5) were introduced into HFLC using lentiviral vectors encoding a fluorescent reporter for visualization of HCV-infected cells. V protein-transduced HFLC supported enhanced (10–100 fold) levels of HCV infection relative to un-transduced or control vector-transduced HFLC. Infection was assessed by measurement of virus-driven luciferase, by assays for infectious HCV and viral RNA, and by direct visualization of HCV-infected hepatocytes. Live cell imaging between 48 and 119 hrs post-infection demonstrated little or no spread of infection in the absence of PMV V protein expression. In contrast, V protein-transduced HFLC showed numerous HCV infection events. V protein expression efficiently antagonized the HCV-inhibitory effects of added IFNs in HFLC. In addition, induction of the type III IFN, IL29, following acute HCV infection was inhibited in V protein-transduced cultures. Conclusions These studies suggest that the cellular IFN response plays a significant role in limiting the spread of HCV infection in primary hepatocyte cultures. Strategies aimed at dampening this response may be key to further development of robust HCV culture systems, enabling studies of virus pathogenicity, and the mechanisms by which HCV spreads in its natural host cell population.

Modeling hepatitis C virus infection using human induced pluripotent stem cells

“…Thus, iHLCs support the complete HCV life cycle, including replication and release of infectious virions. Therefore, iHLCs sustain the entire HCV viral life cycle of at least genotype 2a, in accordance with prior reports showing that human fetal hepatocytes are capable of sustaining the HCV life cycle (22,23). Future work toward a fully personalized in vitro model of HCV infection would incorporate both personalized hepatocyte-like cells and isolates from HCV patients, including the most common HCV genotype in the United States, genotype 1a.…”

Characterization of nonprimate hepacivirus and construction of a functional molecular clone

“…Primary EFLCs were obtained from an equine fetus at 80 d of gestation, which is an earlier stage of development than that used to establish HCV permissive HFLCs (16-24 wk). EFLCs of hepatocytes, fibroblasts, and presumably other cells were established, and the presence of hepatocytes was confirmed by morphology (23) and high levels of miR-122 (Fig. 4 B and D).…”

“…Given our finding that NPHV is a hepatotropic equine virus, the narrow host and tissue tropism of HCV, and the lack of equine liver cell lines, we attempted to establish equine fetal liver cultures (EFLCs) similar to human fetal liver cultures (HFLCs) known to support HCV replication (23). Primary EFLCs were obtained from an equine fetus at 80 d of gestation, which is an earlier stage of development than that used to establish HCV permissive HFLCs (16-24 wk).…”

Entry and replication of recombinant hepatitis C viruses in cell culture

“…Fetal PHH Long-lived as compared to adult PHH [87][88][89][90]. Structures looking like bile canaliculi [87].…”

“…Structures looking like bile canaliculi [87]. Inhibition of innate immune response promotes HCVcc productive infection and spread [87] [87]…”