In vitro infection of adult normal human hepatocytes in primary culture by hepatitis C virus.

Fournier, Carole; Sureau, Camille; Coste, J.; Ducos, J; Pageaux, Georges Philippe; Larrey, Dominique; Domergue, J; Maurel, Patrick

doi:10.1099/0022-1317-79-10-2367

Cited by 120 publications

(86 citation statements)

References 42 publications

(34 reference statements)

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“…Intracellular negative-strand RNA could be detected by a strand-specific RT-PCR method and the core protein could be detected by immunofluorescence microscopy. Fournier et al were the first to demonstrate that hepatocytes isolated from non-infected patients were sensitive to HCVser infection and permissive to viral genome replication (Fournier et al, 1998). Since then, several groups have confirmed these findings.…”

Section: Primary Human Hepatocyte Cultures (Phh)supporting

confidence: 56%

Cellular models for the screening and development of anti-hepatitis C virus agents

Gondeau

Pichard-Garcia

Maurel

2009

Pharmacology & Therapeutics

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Investigations on the biology of hepatitis C virus (HCV) have been hampered by the lack of small animal models. Efforts have therefore been directed to designing practical and robust cellular models of human origin able to support HCV replication and production in a reproducible, reliable and consistent manner. Many different models based on different forms of virions and hepatoma or other cell types have been described including virus-like particles, pseudotyped particles, subgenomic and full length replicons, virion productive replicons, immortalised hepatocytes, fetal and adult primary human hepatocytes. This review focuses on these different cellular models, their advantages and disadvantages at the biological and experimental levels, and their respective use for evaluating the effect of antiviral molecules on different steps of HCV biology including virus entry, replication, particles generation and excretion, as well as on the modulation by the virus of the host cell response to infection.

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Section: Primary Human Hepatocyte Cultures (Phh)supporting

confidence: 56%

Cellular models for the screening and development of anti-hepatitis C virus agents

Gondeau

Pichard-Garcia

Maurel

2009

Pharmacology & Therapeutics

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“…Virus-like particles have been visualized in liver biopsies of HCV ϩ individuals (3)(4)(5), and in vitro infection, albeit inefficient, of primary hepatocytes and hepatocyte cell lines has been reported (6)(7)(8). The existence of extrahepatic reservoirs of HCV is suggested by the detection of viral RNA in serum and PBMC of infected individuals (9)(10)(11).…”

Section: Discussionmentioning

confidence: 97%

“…In the past, difficulties with culturing the virus and expressing fusogenic envelope glycoproteins limited studies of HCV tropism and entry. RT-PCR-and electron microscopy-based approaches were relied on to demonstrate the presence of HCV RNA and proteins in primary hepatocytes and certain hepatoma cell lines (3)(4)(5)(6)(7)(8). Furthermore, the existence of extrahepatic HCV reservoirs was suggested by the detection of viral RNA in serum and peripheral blood mononuclear cells (PBMC) (9)(10)(11).…”

mentioning

confidence: 99%

CD81 is an entry coreceptor for hepatitis C virus

Cormier

Tsamis

Kajumo

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

256

236

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Hepatitis C virus (HCV) envelope glycoproteins E1͞E2 can pseudotype retroviral particles and efficiently mediate entry into target cells. Using this experimental system, we determined HCV tropism for different cell types. Only primary hepatocytes and one hepatoma cell line were susceptible to HCV pseudovirus entry, which could be inhibited by sera from HCV-infected individuals. Furthermore, expression of the putative HCV receptor CD81 on nonpermissive human hepatic but not murine cells enabled HCV pseudovirus entry. Importantly, inhibition of viral entry by an anti-CD81 mAb occurred at a step following HCV attachment to target cells. Our results indicate that CD81 functions as a postattachment entry coreceptor and that other cellular factors act in concert with CD81 to mediate HCV binding and entry into hepatocytes.I t is estimated that 170 million people worldwide are infected with the hepatitis C virus (HCV) and are at risk of developing chronic hepatitis or cirrhosis, the latter often leading to hepatocellular carcinoma (1, 2). In the past, difficulties with culturing the virus and expressing fusogenic envelope glycoproteins limited studies of HCV tropism and entry. RT-PCR-and electron microscopy-based approaches were relied on to demonstrate the presence of HCV RNA and proteins in primary hepatocytes and certain hepatoma cell lines (3-8). Furthermore, the existence of extrahepatic HCV reservoirs was suggested by the detection of viral RNA in serum and peripheral blood mononuclear cells (PBMC) (9-11). Recently, a major technical advance in the field has been the discovery that unmodified HCV envelope glycoproteins can pseudotype retroviral particles and mediate entry into target cells (12)(13)(14)(15). This model seems to authentically replicate the early steps of the HCV life cycle, enabling detailed studies of HCV tropism and entry into target cells.The cellular tropism of enveloped viruses is largely determined by selective interactions of viral envelope glycoproteins with specific cell-surface receptors. Entry generally proceeds by a cascade of coordinated events wherein virus binding to a host molecule triggers exposure of cryptic envelope glycoprotein domains that mediate downstream interactions and functions. We and others recently demonstrated that DC-SIGN (dendritic cell-specific ICAM-3 grabbing nonintegrin; CD209) and L-SIGN (DC-SIGNR, liver and lymph node specific; CD209L) function as HCV capture receptors but do not mediate viral entry into target cells (16,17). Candidate HCV entry receptors include CD81, scavenger receptor class B type 1, low density lipoprotein receptor, and glycosaminoglycans (18)(19)(20). CD81 is the most extensively characterized putative HCV receptor. A number of groups have demonstrated that the soluble ectodomain of HCV envelope glycoprotein E2 binds specifically and with relatively high affinity (K d Ϸ10 Ϫ8 M) to human and chimpanzee CD81 (21-24). However, CD81 is widely expressed on human cells and therefore cannot account for the restricted tropism of HCV to hepatocytes...

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“…Hepatocytes are the primary target cells for HCV infection (6)(7)(8). Virus-like particles have been visualized in liver biopsies of HCVϩ individuals (9)(10)(11), and in vitro infection, albeit inefficient, of primary hepatocytes and hepatoma cells has been documented (12)(13)(14). The existence of extrahepatic reservoirs of HCV is suggested by the detection of viral RNA in serum and peripheral blood mononuclear cells of HCVϩ individuals (15)(16)(17).…”

Section: H Epatitis C Virus (Hcv) Is the Etiologic Agent Of Non-a Non-bmentioning

confidence: 99%

L-SIGN (CD209L) and DC-SIGN (CD209) mediate transinfection of liver cells by hepatitis C virus

Cormier

Durso

Tsamis

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

188

163

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Target cell tropism of enveloped viruses is regulated by interactions between viral and cellular factors during transmission, dissemination, and replication within the host. Binding of viral envelope glycoproteins to specific cell-surface receptors determines susceptibility to viral entry. However, a number of cell-surface molecules bind viral envelope glycoproteins without mediating entry. Instead, they serve as capture receptors that disseminate viral particles to target organs or susceptible cells. We and others recently demonstrated that the C type lectins L-SIGN and DC-SIGN capture hepatitis C virus (HCV) by specific binding to envelope glycoprotein E2. In this study, we use an entry assay to demonstrate that HCV pseudoviruses captured by L-SIGN؉ or DC-SIGN؉ cells efficiently transinfect adjacent human liver cells. Virus capture and transinfection require internalization of the SIGN-HCV pseudovirus complex. In vivo, L-SIGN is largely expressed on endothelial cells in liver sinusoids, whereas DC-SIGN is expressed on dendritic cells. Capture of circulating HCV particles by these SIGN؉ cells may facilitate virus infection of proximal hepatocytes and lymphocyte subpopulations and may be essential for the establishment of persistent infection.H epatitis C virus (HCV) is the etiologic agent of non-A non-B hepatitis in humans (1, 2). Only Ϸ15% of infected individuals clear the virus, and Ϸ170 million people worldwide are persistently infected with HCV (3, 4). These individuals may remain asymptomatic or may develop chronic hepatitis or cirrhosis, the latter often leading to hepatocellular carcinoma (5). Hepatocytes are the primary target cells for HCV infection (6-8). Virus-like particles have been visualized in liver biopsies of HCVϩ individuals (9-11), and in vitro infection, albeit inefficient, of primary hepatocytes and hepatoma cells has been documented (12)(13)(14). The existence of extrahepatic reservoirs of HCV is suggested by the detection of viral RNA in serum and peripheral blood mononuclear cells of HCVϩ individuals (15)(16)(17). Both B and T lymphocytes appear to be infected in vivo, which is supported by in vitro infection of B and T cell lines (7,8,18). One study, however, shows that replicating forms of HCV RNA are restricted to hepatocytes, whereas only nonreplicating forms are present in B lymphocytes, and none are in T lymphocytes (6).HCV envelope glycoproteins E1 and E2 mediate entry into target cells. We and others recently demonstrated that unmodified E1E2 heterodimers reach the cell surface and are incorporated into retroviral pseudoparticles, which can infect primary hepatocytes and some hepatoma cell lines (19 -22). Use of the soluble E2 ectodomain as a surrogate model for studying HCV interactions with cell-surface molecules has identified several potential HCV entry receptors, including CD81, scavenger receptor class B type 1, low-density lipoprotein receptor, and glycosaminoglycans (22-24). Several groups, including ours, have shown that CD81 is necessary but not sufficient for HCV pseu...

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In vitro infection of adult normal human hepatocytes in primary culture by hepatitis C virus.

Cited by 120 publications

References 42 publications

Cellular models for the screening and development of anti-hepatitis C virus agents

Cellular models for the screening and development of anti-hepatitis C virus agents

CD81 is an entry coreceptor for hepatitis C virus

L-SIGN (CD209L) and DC-SIGN (CD209) mediate transinfection of liver cells by hepatitis C virus

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