Reproducible high level infection of cultured adult human hepatocytes by hepatitis B virus: effect of polyethylene glycol on adsorption and penetration.
We have previously succeeded in infecting normal human hepatocyte primary cultures with hepatitis B virus (HBV). However, infection was subject to individual variations even in the presence of dimethyl sulfoxide (DMSO), which appeared to increase the amounts of viral DNA associated with the cells. In this study, we have defined conditions which enhance hepatitis B virus penetration into the cells, and we show that, under these conditions, infection of hepatocytes is always possible, regardless of their individual origin. We have found that addition of polyethylene glycol (PEG) to the cultures maintained in the presence of 2% DMSO at the time of infection markedly increased the infection process and made it highly reproducible. Moreover, both the tissue and species specificity were preserved. This increased HBV infection was correlated to increased amount of internalized HBV DNA and to enhanced attachment of the virions. From these results it may be assumed that PEG could favor a better interaction between virions and cells, resulting in an activated internalization of bound viral particles. Data also show that adult human hepatocyte primary cultures, which are not equally susceptible to HBV infection, are consistently capable of viral replication when the viral genome has entered the cells. This suggests that the main limitation of the in vitro HBV infection lies in the ability of human hepatocytes to specifically bind the viral particles.
Gripon
P
P
Inserm U49, Hôpital Pontchaillou, Rennes, France.
Diot
C
C
Guguen-Guillouzo
C
C
D000328:Adult; D015139:Blotting, Southern; D002478:Cells, Cultured; D004279:DNA, Viral / Q000235:genetics / Q000302:isolation & purification; D006514:Hepatitis B Surface Antigens / Q000032:analysis / Q000096:biosynthesis; D006513:Hepatitis B e Antigens / Q000032:analysis / Q000096:biosynthesis; D006515:Hepatitis B virus / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D007700:Kinetics; D008099:Liver / Q000166:cytology / Q000382:microbiology*; D011092:Polyethylene Glycols / Q000494:pharmacology*; D014764:Viral Proteins / Q000032:analysis / Q000096:biosynthesis; D014779:Virus Replication
D004279:DNA, Viral; D006514:Hepatitis B Surface Antigens; D006513:Hepatitis B e Antigens; D014764:Viral Proteins; D011092:Polyethylene Glycols