title: Hepatocyte Nicotinamide Adenine Dinucleotide Phosphate Reduced Oxidase 4 Regulates Stress Signaling, Fibrosis, and Insulin Sensitivity During Development of Steatohepatitis in Mice.
abstract: OBJECTIVE
  Reactive oxidative species (ROS) are believed to be involved in the progression of nonalcoholic steatohepatitis (NASH). However, little is known about the sources of ROS in hepatocytes or their role in disease progression. We studied the effects of nicotinamide adenine dinucleotide phosphate reduced oxidase 4 (NOX4) in liver tissues from patients with NASH and mice with steatohepatitis.
  
  
  METHODS
  Liver biopsy samples were obtained from 5 patients with NASH, as well as 4 patients with simple steatosis and 5 patients without steatosis (controls) from the University of California, Davis Cancer Center Biorepository. Mice with hepatocyte-specific deletion of NOX4 (NOX4(hepKO)) and NOX4(floxp+/+) C57BL/6 mice (controls) were given fast-food diets (supplemented with high-fructose corn syrup) or choline-deficient l-amino acid defined diets to induce steatohepatitis, or control diets, for 20 weeks. A separate group of mice were given the NOX4 inhibitor (GKT137831). Liver tissues were collected and immunoblot analyses were performed determine levels of NOX4, markers of inflammation and fibrosis, double-stranded RNA-activated protein kinase, and phospho-eIF-2α kinase-mediated stress signaling pathways. We performed hyperinsulinemic-euglycemic clamp studies and immunoprecipitation analyses to determine the oxidation and phosphatase activity of PP1C.
  
  
  RESULTS
  Levels of NOX4 were increased in patients with NASH compared with controls. Hepatocyte-specific deletion of NOX4 reduced oxidative stress, lipid peroxidation, and liver fibrosis in mice with diet-induced steatohepatitis. A small molecule inhibitor of NOX4 reduced liver inflammation and fibrosis and increased insulin sensitivity in mice with diet-induced steatohepatitis. In primary hepatocytes, NOX4 reduced the activity of the phosphatase PP1C, prolonging activation of double-stranded RNA-activated protein kinase and phosphorylation of extracellular signal-regulated kinase-mediated stress signaling. Mice with hepatocyte-specific deletion of NOX4 and mice given GKT137831 had increased insulin sensitivity.
  
  
  CONCLUSIONS
  NOX4 regulates oxidative stress in the liver and its levels are increased in patients with NASH and mice with diet-induced steatohepatitis. Inhibitors of NOX4 reduce liver inflammation and fibrosis and increase insulin sensitivity, and might be developed for treatment of NASH.
authors.affiliation: Department of Nutrition, University of California Davis, Davis, California.
authors.forename: Ahmed
authors.initials: A
authors.lastname: Bettaieb
authors.affiliation: Department of Medicine, Gastroenterology and Hepatology, University of California Davis, and VA Medical Center, Sacramento, California.
authors.forename: Joy X
authors.initials: JX
authors.lastname: Jiang
authors.affiliation: Department of Medicine, Gastroenterology and Hepatology, University of California Davis, and VA Medical Center, Sacramento, California.
authors.forename: Yu
authors.initials: Y
authors.lastname: Sasaki
authors.affiliation: Department of Medicine, Gastroenterology and Hepatology, University of California Davis, and VA Medical Center, Sacramento, California.
authors.forename: Tzu-I
authors.initials: TI
authors.lastname: Chao
authors.affiliation: Department of Medicine, Gastroenterology and Hepatology, University of California Davis, and VA Medical Center, Sacramento, California.
authors.forename: Zsofia
authors.initials: Z
authors.lastname: Kiss
authors.affiliation: Department of Medicine, Gastroenterology and Hepatology, University of California Davis, and VA Medical Center, Sacramento, California.
authors.forename: Xiangling
authors.initials: X
authors.lastname: Chen
authors.affiliation: Department of Medicine, Gastroenterology and Hepatology, University of California Davis, and VA Medical Center, Sacramento, California.
authors.forename: Jijing
authors.initials: J
authors.lastname: Tian
authors.affiliation: Kyoto Prefectural University of Medicine, Kyoto, Japan.
authors.forename: Masato
authors.initials: M
authors.lastname: Katsuyama
authors.affiliation: Kyoto Prefectural University of Medicine, Kyoto, Japan.
authors.forename: Chihiro
authors.initials: C
authors.lastname: Yabe-Nishimura
authors.affiliation: Department of Nutrition, University of California Davis, Davis, California.
authors.forename: Yannan
authors.initials: Y
authors.lastname: Xi
authors.affiliation: Genkyotex, Geneva, Switzerland.
authors.forename: Cedric
authors.initials: C
authors.lastname: Szyndralewiez
authors.affiliation: Wolfgang Goethe University, Frankfurt am Main, Germany.
authors.forename: Kathrin
authors.initials: K
authors.lastname: Schröder
authors.affiliation: King's College London British Heart Foundation Centre, London, UK.
authors.forename: Ajay
authors.initials: A
authors.lastname: Shah
authors.affiliation: Wolfgang Goethe University, Frankfurt am Main, Germany.
authors.forename: Ralph P
authors.initials: RP
authors.lastname: Brandes
authors.affiliation: Department of Nutrition, University of California Davis, Davis, California.
authors.forename: Fawaz G
authors.initials: FG
authors.lastname: Haj
authors.affiliation: Department of Medicine, Gastroenterology and Hepatology, University of California Davis, and VA Medical Center, Sacramento, California. Electronic address: njtorok@ucdavis.edu.
authors.forename: Natalie J
authors.initials: NJ
authors.lastname: Török
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D001706:Biopsy; D004032:Diet / Q000379:methods; D004195:Disease Models, Animal; D005234:Fatty Liver / Q000139:chemically induced / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism; D022781:Hepatocytes / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D007333:Insulin Resistance; D015227:Lipid Peroxidation / Q000187:drug effects; D008099:Liver / Q000166:cytology / Q000473:pathology; D008103:Liver Cirrhosis / Q000188:drug therapy* / Q000473:pathology; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D009249:NADP / Q000008:administration & dosage / Q000494:pharmacology*; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases / Q000378:metabolism*; D009765:Obesity / Q000188:drug therapy / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects*; D054645:Protein Phosphatase 1 / Q000378:metabolism; D011720:Pyrazoles / Q000378:metabolism; D047069:Pyrazolones; D011725:Pyridines / Q000378:metabolism; D011728:Pyridones; D013312:Stress, Physiological / Q000187:drug effects
chemical_list: D015415:Biomarkers; D011720:Pyrazoles; D047069:Pyrazolones; D011725:Pyridines; D011728:Pyridones; C576694:setanaxib; D009249:NADP; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases; C498910:NOX4 protein, human; D054645:Protein Phosphatase 1
keywords: Inflammation; Mouse Model; Obesity; Stress Signaling