Production of infectious hepatitis C virus in primary cultures of human adult hepatocytes.
OBJECTIVE
Although hepatitis C virus (HCV) can be grown in the hepatocarcinoma-derived cell line Huh-7, a cell-culture model is needed that supports its complete, productive infection cycle in normal, quiescent, highly differentiated human hepatocytes. We sought to develop such a system.


METHODS
Primary cultures of human adult hepatocytes were inoculated with HCV derived from Huh-7 cell culture (HCVcc) and monitored for expression of hepatocyte differentiation markers and replication of HCV. Culture supernatants were assayed for HCV RNA, core antigen, and infectivity titer. The buoyant densities of input and progeny virus were compared in iodixanol gradients.


RESULTS
While retaining expression of differentiation markers, primary hepatocytes supported the complete infectious cycle of HCV, including production of significant titers of new infectious progeny virus, which was called primary-culture-derived virus (HCVpc). Compared with HCVcc, HCVpc had lower average buoyant density and higher specific infectivity; this was similar to the characteristics of virus particles associated with the very-low-density lipoproteins that are produced during in vivo infection. These properties were lost after re-culture of HCVpc in poorly differentiated Huh-7 cells, suggesting that authentic virions can be produced only by normal hepatocytes that secrete authentic very-low-density lipoproteins.


CONCLUSIONS
We have established a cell-culture-based system that allows production of infectious HCV in physiologically relevant human hepatocytes. This provides a useful tool for the study of HCV interactions with its natural host cell and for the development of antiviral therapies.
Podevin
Philippe
P
INSERM, Equipe Avenir Virologie de l'Hépatite C, Institut Cochin, Paris, France.
Carpentier
Arnaud
A
Pène
Véronique
V
Aoudjehane
Lynda
L
Carrière
Matthieu
M
Zaïdi
Sakina
S
Hernandez
Céline
C
Calle
Vanessa
V
Méritet
Jean-François
JF
Scatton
Olivier
O
Dreux
Marlène
M
Cosset
François-Loïc
FL
Wakita
Takaji
T
Bartenschlager
Ralf
R
Demignot
Sylvie
S
Conti
Filoména
F
Rosenberg
Arielle R
AR
Calmus
Yvon
Y
D000328:Adult; D002454:Cell Differentiation; D045744:Cell Line, Tumor; D016679:Genome, Viral; D016174:Hepacivirus / Q000502:physiology*; D022781:Hepatocytes / Q000821:virology*; D006801:Humans; D014779:Virus Replication