title: Gliotoxin-mediated apoptosis of activated human hepatic stellate cells.
abstract: BACKGROUND
  Activated hepatic stellate cells (HSCs) play a central role in liver fibrogenesis, and apoptosis of activated HSCs might be essential to clear HSCs from injured liver. Gliotoxin induces apoptosis of activated human and rat HSCs by an unknown mechanism.
  
  
  OBJECTIVE
  This study investigated the role of reactive oxygen species (ROS) and membrane permeability transition (MPT) in gliotoxin-induced apoptosis of activated human HSCs.
  
  
  METHODS
  Primary and immortalized human HSCs were analyzed using confocal microscopy for ROS with dichlorodihdrofluorescence diacetate (DCFH-DA) fluorophore and for the mitochondrial membrane potential (MMP) using tetramethylrhodamine methylester (TMRM).
  
  
  RESULTS
  Gliotoxin at higher concentrations (> or =7.5 microM) markedly increased ROS formation, and ROS production was also evident at concentrations of gliotoxin causing necrotic cell death (> or =32.5 microM). Gliotoxin rapidly (begins about 20 min at 1.5 microM and 10 min at 7.5 microM) disrupts MMP at a concentration as low as 300nM. MMP disruption was followed by cytochrome c release and caspase-3 activation. The MPT inhibitors, cyclosporine A (5 microM) plus trifluoperazine (12.5 microM), blocked depolarization of the mitochondrial membrane and release of cytochrome c, but did not block apoptosis in HSCs.
  
  
  CONCLUSIONS
  Gliotoxin (0.3-7.5 microM) induces apoptosis of activated human HSCs with induction of MPT, cytochrome c release and caspase-3 activation, whereas at higher doses (>32.5 microM), it induces necrosis. However, gliotoxin also activates a mitochondrial independent pathway.
authors.affiliation: Division of Digestive Diseases and Nutrition, Department of Medicine, CB# 7038, Glaxo Research Bldg. Rm. 156, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
authors.forename: Young-Oh
authors.initials: YO
authors.lastname: Kweon
authors.forename: Yong-Han
authors.initials: YH
authors.lastname: Paik
authors.forename: Bernd
authors.initials: B
authors.lastname: Schnabl
authors.forename: Ting
authors.initials: T
authors.lastname: Qian
authors.forename: John J
authors.initials: JJ
authors.lastname: Lemasters
authors.forename: David A
authors.initials: DA
authors.lastname: Brenner
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D017304:Annexin A5 / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D002461:Cell Line, Transformed; D016572:Cyclosporine / Q000494:pharmacology; D045304:Cytochromes c / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D018492:Dopamine Antagonists / Q000494:pharmacology; D005912:Gliotoxin / Q000494:pharmacology*; D006801:Humans; D007166:Immunosuppressive Agents / Q000494:pharmacology*; D008099:Liver / Q000166:cytology*; D008103:Liver Cirrhosis / Q000378:metabolism / Q000473:pathology; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D009336:Necrosis; D018384:Oxidative Stress / Q000187:drug effects; D011485:Protein Binding / Q000187:drug effects; D017382:Reactive Oxygen Species / Q000378:metabolism; D014268:Trifluoperazine / Q000494:pharmacology
chemical_list: D017304:Annexin A5; D018492:Dopamine Antagonists; D007166:Immunosuppressive Agents; D016328:NF-kappa B; D017382:Reactive Oxygen Species; D014268:Trifluoperazine; D005912:Gliotoxin; D016572:Cyclosporine; D000255:Adenosine Triphosphate; D045304:Cytochromes c; C505407:CASP3 protein, human; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases