title: Expression of E1E2 on hepatitis C RNA-containing particles released from primary cultured human hepatocytes derived from infected cirrhotic livers.
abstract: OBJECTIVE
  To determine whether liver-derived hepatitis C RNA-containing particles express the E1E2 discontinuous antigenic determinant defined by unique monoclonal antibody (mAb) D32.10 which recognizes three highly conserved segments in E1 (aa297-306) and E2 (aa480-494 and aa613-621) envelope glycoproteins.
  
  
  METHODS
  Human hepatocytes were isolated from HCV-infected cirrhotic explanted livers. The liver-derived hepatitis C virus (HCV) particles released from three distinct cultures (genotypes 1b and 2b) were characterized. HCV RNA+ was quantified by real-time RT-PCR. The E1E2 antigenic activity was assessed by indirect ELISA and immunoblotting using D32.10. The density distributions of HCV RNA and E1E2 antigen were determined by isopycnic sucrose density gradients. HCV E1E2, E2 and core antigens were detected in the cells by immunochemical staining.
  
  
  RESULTS
  Liver-derived HCV particles contained HCV RNA (10⁶-10⁷ copies/mg of protein) and core proteins and expressed the E1E2/D32.10 epitope. HCV RNA and E1E2 cosedimented between 1.15 and 1.25 g/ml in sucrose gradients. Moreover, the mAb D32.10 detected E1E2 by immunostaining in HCV-infected hepatocytes in parallel with E2 and core antigens.
  
  
  CONCLUSIONS
  Our results provide evidence that the mAb D32.10 recognizes E1E2 envelope complexes expressed in the cell cytoplasm and on the surface of HCV RNA-containing particles released from short-term cultures of in vivo infected hepatocytes.
authors.affiliation: INSERM, U871, Lyon, France.
authors.forename: Ndiémé
authors.initials: N
authors.lastname: Ndongo
authors.forename: Subjini
authors.initials: S
authors.lastname: Selliah
authors.forename: Pascale
authors.initials: P
authors.lastname: Berthillon
authors.forename: Valérie-Ann
authors.initials: VA
authors.lastname: Raymond
authors.forename: Christian
authors.initials: C
authors.lastname: Trépo
authors.forename: Marc
authors.initials: M
authors.lastname: Bilodeau
authors.forename: Marie-Anne
authors.initials: MA
authors.lastname: Petit
mesh_terms: D000911:Antibodies, Monoclonal / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D002478:Cells, Cultured; D002500:Centrifugation, Isopycnic; D000939:Epitopes / Q000276:immunology / Q000378:metabolism; D015967:Gene Expression Regulation, Viral; D016174:Hepacivirus / Q000235:genetics* / Q000378:metabolism; D006526:Hepatitis C / Q000235:genetics / Q000821:virology; D022781:Hepatocytes / Q000378:metabolism* / Q000821:virology; D006801:Humans; D008103:Liver Cirrhosis / Q000503:physiopathology; D010455:Peptides / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D012367:RNA, Viral; D014759:Viral Envelope Proteins / Q000235:genetics* / Q000276:immunology / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; C116123:E1E2 peptide; D000939:Epitopes; D010455:Peptides; D012367:RNA, Viral; D014759:Viral Envelope Proteins