title: Oxidative stress plays a major role in chlorpromazine-induced cholestasis in human HepaRG cells.
abstract: Drugs induce cholestasis by diverse and still poorly understood mechanisms in humans. Early hepatic effects of chlorpromazine (CPZ), a neuroleptic drug known for years to induce intrahepatic cholestasis, were investigated using the differentiated human hepatoma HepaRG cells. Generation of reactive oxygen species (ROS) was detected as early as 15 minutes after CPZ treatment and was associated with an altered mitochondrial membrane potential and disruption of the pericanalicular distribution of F-actin. Inhibition of [3H]-taurocholic acid efflux was observed after 30 minutes and was mostly prevented by N-acetyl cysteine (NAC) cotreatment, indicating a major role of oxidative stress in CPZ-induced bile acid (BA) accumulation. Moreover, 24-hour treatment with CPZ decreased messenger RNA (mRNA) expression of the two main canalicular bile transporters, bile salt export pump (BSEP) and multidrug resistance protein 3 (MDR3). Additional CPZ effects included inhibition of Na+ -dependent taurocholic cotransporting polypeptide (NTCP) expression and activity, multidrug resistance-associated protein 4 (MRP4) overexpression and CYP8B1 inhibition that are involved in BA uptake, basolateral transport, and BA synthesis, respectively. These latter events likely represent hepatoprotective responses which aim to reduce intrahepatic accumulation of toxic BA. Compared to CPZ effects, overloading of HepaRG cells with high concentrations of cholic and chenodeoxycholic acids induced a delayed oxidative stress and, similarly, after 24 hours it down-regulated BSEP and MDR3 in parallel to a decrease of NTCP and CYP8B1 and an increase of MRP4. By contrast, low BA concentrations up-regulated BSEP and MDR3 in the absence of oxidative stress.
  
  
  CONCLUSIONS
  These data provide evidence that, among other mechanisms, oxidative stress plays a major role as both a primary causal and an aggravating factor in the early CPZ-induced intrahepatic cholestasis in human hepatocytes.
authors.affiliation: Inserm UMR991, Liver Metabolisms and Cancer, Rennes, France.
authors.forename: Sébastien
authors.initials: S
authors.lastname: Anthérieu
authors.forename: Pamela
authors.initials: P
authors.lastname: Bachour-El Azzi
authors.forename: Julie
authors.initials: J
authors.lastname: Dumont
authors.forename: Ziad
authors.initials: Z
authors.lastname: Abdel-Razzak
authors.forename: Christiane
authors.initials: C
authors.lastname: Guguen-Guillouzo
authors.forename: Bernard
authors.initials: B
authors.lastname: Fromenty
authors.forename: Marie-Anne
authors.initials: MA
authors.lastname: Robin
authors.forename: André
authors.initials: A
authors.lastname: Guillouzo
mesh_terms: D018435:ATP Binding Cassette Transporter, Subfamily B / Q000378:metabolism; D000074020:ATP Binding Cassette Transporter, Subfamily B, Member 11; D018528:ATP-Binding Cassette Transporters / Q000378:metabolism; D000199:Actins / Q000378:metabolism; D006528:Carcinoma, Hepatocellular / Q000378:metabolism / Q000473:pathology*; D045744:Cell Line, Tumor; D002746:Chlorpromazine / Q000009:adverse effects* / Q000494:pharmacology; D002779:Cholestasis / Q000139:chemically induced* / Q000378:metabolism / Q000503:physiopathology*; D006801:Humans; D066298:In Vitro Techniques; D008113:Liver Neoplasms / Q000378:metabolism / Q000473:pathology*; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D018384:Oxidative Stress / Q000187:drug effects / Q000502:physiology*; D017382:Reactive Oxygen Species / Q000378:metabolism; D013656:Taurocholic Acid / Q000378:metabolism
chemical_list: C093491:ABCB11 protein, human; D018435:ATP Binding Cassette Transporter, Subfamily B; D000074020:ATP Binding Cassette Transporter, Subfamily B, Member 11; D018528:ATP-Binding Cassette Transporters; D000199:Actins; D017382:Reactive Oxygen Species; D013656:Taurocholic Acid; C095122:multidrug resistance protein 3; D002746:Chlorpromazine