title: Superoxide anion radicals activate hepatic stellate cells after entry through chloride channels: a new target in liver fibrosis.
abstract: It is generally accepted that reactive oxygen species (ROS) play an important role in the pathogenesis of liver fibrosis. ROS, however, constitute a group of species with varying properties making it likely that their contribution to the pathological mechanism varies. LX-2 hepatic stellate cells (HSCs) were exposed to superoxide anion radicals (O2(·-)) generated by xanthine and xanthine oxidase. To rule out that the activation of HSCs is due to hydrogen peroxide derived from O2(·-), control incubations with copper, zinc-superoxide dismutase and tempol were studied as well. Influx of O2(·-) activated HSCs, evidenced by the expression of α-smooth muscle actin and the secretion of transforming growth factor β1 and collagen. We further found that blockade of chloride channels with 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS), 5-nitro-2-(3-phenylpropyl-amino) benzoic acid (NPPB) or indanyloxyacetic acid (IAA-94) prevented the increase of intracellular O2(·-) levels as well as the activation of HSCs. These findings suggest that O2(·-) is involved in the development of liver fibrosis and that entry of O2(·-), through chloride channels, in stellate cells is critical for their activation. This study provides new insight into the mechanism by which ROS induce liver fibrosis. Furthermore, our data suggest that chloride channels constitute a potential target for new anti-fibrotic drugs.
authors.affiliation: Maastricht University, Faculty of Health, Medicine and Life Sciences, Department of Toxicology, Maastricht, The Netherlands. Electronic address: gj.denhartog@maastrichtuniversity.nl.
authors.forename: Gertjan J M
authors.initials: GJ
authors.lastname: den Hartog
authors.affiliation: Maastricht University, Faculty of Health, Medicine and Life Sciences, Department of Toxicology, Maastricht, The Netherlands.
authors.forename: Shufan
authors.initials: S
authors.lastname: Qi
authors.affiliation: Maastricht University, Faculty of Health, Medicine and Life Sciences, Department of Human Biology, Maastricht, The Netherlands.
authors.forename: Jonathan H O
authors.initials: JH
authors.lastname: van Tilburg
authors.affiliation: Department of Internal Medicine, Division of Gastroenterology and Hepatology, Maastricht University Medical Centre, Maastricht, The Netherlands.
authors.forename: Ger H
authors.initials: GH
authors.lastname: Koek
authors.affiliation: Maastricht University, Faculty of Health, Medicine and Life Sciences, Department of Toxicology, Maastricht, The Netherlands.
authors.forename: Aalt
authors.initials: A
authors.lastname: Bast
mesh_terms: D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000199:Actins / Q000235:genetics; D002460:Cell Line; D018118:Chloride Channels / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006016:Glycolates / Q000494:pharmacology; D055166:Hepatic Stellate Cells / Q000378:metabolism*; D006801:Humans; D008103:Liver Cirrhosis / Q000378:metabolism*; D009579:Nitrobenzoates / Q000494:pharmacology; D013481:Superoxides / Q000378:metabolism*; D053773:Transforming Growth Factor beta1 / Q000378:metabolism; D019820:Xanthine / Q000494:pharmacology; D014969:Xanthine Oxidase / Q000494:pharmacology
chemical_list: C541116:ACTA2 protein, human; D000199:Actins; D018118:Chloride Channels; D006016:Glycolates; D009579:Nitrobenzoates; D053773:Transforming Growth Factor beta1; D013481:Superoxides; D019820:Xanthine; C058176:5-nitro-2-(3-phenylpropylamino)benzoic acid; C015765:MK 473; D014969:Xanthine Oxidase; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid
keywords: 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid disodium salt hydrate (DIDS, PubChem CID: 4294801); 5-Nitro-2-(3-phenylpropyl-amino) benzoic acid (NPPB, PubChem CID: 4549); Collagen; Hepatic stellate cells; Indanyloxyacetic acid (IAA-94, PubChem CID: 656717); Superoxide anion radicals; Tempol (PubChem CID: 137994); Transforming growth factor-β; α Smooth muscle actin