title: Cytochrome P450 expression-induction profile and chemically mediated alterations of the WIF-B9 cell line.
abstract: BACKGROUND
  WIF-B9 is a hybrid cell line obtained by fusion of rat hepatoma cells (Fao) and human fibroblasts (WI38). It exhibits the structural and functional characteristics of differentiated hepatocytes, including active bile canaliculi. The aim of the present study was to characterize the WIF-B9 cell line as a model for analysing drug-induced hepatic effects. The drug metabolism potential of WIF-B9 cells was identified by studying the rat and human CYP (cytochrome P450) mRNA constitutive expression profile and induction potential after exposure to reference inducers. The morphological alterations provoked by chemical entities were also characterized.
  
  
  RESULTS
  Competitive reverse transcriptase-PCR revealed that four rat (1A1, 2B1/2, 2E1 and 4A1) and four human (1A1, 2Cs, 2D6 and 2E1) CYP mRNA isoforms were constitutively expressed in WIF-B9 cells. The rat CYP forms were expressed at levels 2-4 orders of magnitude higher than the human forms. Exposure for 20-72 h to increasing concentrations of CYP reference inducers (beta-naphthoflavone, 3-methyl cholanthrene, dexamethasone, phenobarbital, clofibrate and pregnenolone 16alpha-carbonitrile) revealed that the rat CYP 1A1, 1A2, 3A1, 3A2 and 4A1 and human CYP 1A1 and 2Cs mRNAs were inducible. Rat CYP 1A1 and 1A2 were the most inducible isoforms since they were overexpressed up to 100-fold after 20-48 h of treatment with beta-naphthoflavone. Human CYP 1A1 and 2Cs mRNAs were induced 3-fold after 48 h of treatment with phenobarbital. Other mechanisms involved in hepatotoxicity were explored using microscopy and immunofluorescence. The WIF-B9 cell line exhibited fragmentation and dilatation of bile canaliculi upon exposure to erythromycin, and to isoniazid and cytochalasins, respectively. Monensin promoted cell depolarization and cytoplasmic granulation. Ethionine promoted cytoplasmic vacuolation and dilatation of the Golgi structures.
  
  
  CONCLUSIONS
  These results indicate that the CYP expression and induction profiles and the morphological features of WIF-B9 cells allow prediction in vitro of the induction and hepatotoxicity profiles of chemical entities.
authors.affiliation: Pfizer Global R&D, Safety Sciences Europe, ZI Pocé sur Cisse, Amboise, France. Christine.biagini@pfizer.com
authors.forename: Christine
authors.initials: C
authors.lastname: Biagini
authors.forename: Virginie
authors.initials: V
authors.lastname: Bender
authors.forename: Françoise
authors.initials: F
authors.lastname: Borde
authors.forename: Elodie
authors.initials: E
authors.lastname: Boissel
authors.forename: Marie-Claire
authors.initials: MC
authors.lastname: Bonnet
authors.forename: Marie-Thérèse
authors.initials: MT
authors.lastname: Masson
authors.forename: Doris
authors.initials: D
authors.lastname: Cassio
authors.forename: Stephan
authors.initials: S
authors.lastname: Chevalier
mesh_terms: D000818:Animals; D001648:Bile Canaliculi / Q000187:drug effects / Q000502:physiology; D045744:Cell Line, Tumor; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D003593:Cytoplasm / Q000187:drug effects; D004790:Enzyme Induction; D022781:Hepatocytes / Q000201:enzymology*; D006801:Humans; D006822:Hybrid Cells; D007527:Isoenzymes / Q000378:metabolism; D008854:Microscopy, Electron; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D007527:Isoenzymes; D003577:Cytochrome P-450 Enzyme System