title: Toxic bile salts induce rodent hepatocyte apoptosis via direct activation of Fas.
abstract: Cholestatic liver injury appears to result from the induction of hepatocyte apoptosis by toxic bile salts such as glycochenodeoxycholate (GCDC). Previous studies from this laboratory indicate that cathepsin B is a downstream effector protease during the hepatocyte apoptotic process. Because caspases can initiate apoptosis, the present studies were undertaken to determine the role of caspases in cathepsin B activation. Immunoblotting of GCDC-treated McNtcp.24 hepatoma cells demonstrated cleavage of poly(ADP-ribose) polymerase and lamin B1 to fragments that indicate activation of effector caspases. Transfection with CrmA, an inhibitor of caspase 8, prevented GCDC-induced cathepsin B activation and apoptosis. Consistent with these results, an increase in caspase 8-like activity was observed in GCDC-treated cells. Examination of the mechanism of GCDC-induced caspase 8 activation revealed that dominant-negative FADD inhibited apoptosis and that hepatocytes isolated from Fas-deficient lymphoproliferative mice were resistant to GCDC-induced apoptosis. After GCDC treatment, immunoprecipitation experiments demonstrated Fas oligomerization, and confocal microscopy demonstrated DeltaFADD-GFP (Fas-associated death domain-green fluorescent protein, aggregation in the absence of detectable Fas ligand mRNA. Collectively, these data suggest that GCDC-induced hepatocyte apoptosis involves ligand-independent oligomerization of Fas, recruitment of FADD, activation of caspase 8, and subsequent activation of effector proteases, including downstream caspases and cathepsin B.
authors.affiliation: Division of Gastroenterology and Hepatology, Mayo Medical School, Clinic, and Foundation, Rochester, Minnesota 55905, USA.
authors.forename: W A
authors.initials: WA
authors.lastname: Faubion
authors.forename: M E
authors.initials: ME
authors.lastname: Guicciardi
authors.forename: H
authors.initials: H
authors.lastname: Miyoshi
authors.forename: S F
authors.initials: SF
authors.lastname: Bronk
authors.forename: P J
authors.initials: PJ
authors.lastname: Roberts
authors.forename: P A
authors.initials: PA
authors.lastname: Svingen
authors.forename: S H
authors.initials: SH
authors.lastname: Kaufmann
authors.forename: G J
authors.initials: GJ
authors.lastname: Gores
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D001647:Bile Acids and Salts / Q000494:pharmacology*; D020169:Caspases / Q000378:metabolism*; D002401:Cathepsin B / Q000378:metabolism*; D002460:Cell Line; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D053938:DNA Fragmentation / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D005999:Glycochenodeoxycholic Acid / Q000494:pharmacology; D007700:Kinetics; D034921:Lamin Type B; D034882:Lamins; D008099:Liver / Q000187:drug effects* / Q000378:metabolism; D051379:Mice; D009687:Nuclear Proteins / Q000378:metabolism; D009842:Oligopeptides / Q000378:metabolism; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D018124:Receptors, Tumor Necrosis Factor / Q000235:genetics; D015843:Serpins / Q000235:genetics; D014764:Viral Proteins; D019014:fas Receptor / Q000378:metabolism*
chemical_list: D001647:Bile Acids and Salts; D015853:Cysteine Proteinase Inhibitors; D034921:Lamin Type B; D034882:Lamins; D009687:Nuclear Proteins; D009842:Oligopeptides; D012333:RNA, Messenger; D018124:Receptors, Tumor Necrosis Factor; D015843:Serpins; D014764:Viral Proteins; D019014:fas Receptor; D005999:Glycochenodeoxycholic Acid; C074641:interleukin-1beta-converting enzyme inhibitor; D011065:Poly(ADP-ribose) Polymerases; D020169:Caspases; D002401:Cathepsin B