title: Necrostatin-1 protects against reactive oxygen species (ROS)-induced hepatotoxicity in acetaminophen-induced acute liver failure.
abstract: Excessive acetaminophen (APAP) use is one of the most common causes of acute liver failure. Various types of cell death in the damaged liver are linked to APAP-induced hepatotoxicity, and, of these, necrotic cell death of hepatocytes has been shown to be involved in disease pathogenesis. Until recently, necrosis was commonly considered to be a random and unregulated form of cell death; however, recent studies have identified a previously unknown form of programmed necrosis called receptor-interacting protein kinase (RIPK)-dependent necrosis (or necroptosis), which is controlled by the kinases RIPK1 and RIPK3. Although RIPK-dependent necrosis has been implicated in a variety of disease states, including atherosclerosis, myocardial organ damage, stroke, ischemia-reperfusion injury, pancreatitis, and inflammatory bowel disease. However its involvement in APAP-induced hepatocyte necrosis remains elusive. Here, we showed that RIPK1 phosphorylation, which is a hallmark of RIPK-dependent necrosis, was induced by APAP, and the expression pattern of RIPK1 and RIPK3 in the liver overlapped with that of CYP2E1, whose activity around the central vein area has been demonstrated to be critical for the development of APAP-induced hepatic injury. Moreover, a RIPK1 inhibitor ameliorated APAP-induced hepatotoxicity in an animal model, which was underscored by significant suppression of the release of hepatic enzymes and cytokine expression levels. RIPK1 inhibition decreased reactive oxygen species levels produced in APAP-injured hepatocytes, whereas CYP2E1 expression and the depletion rate of total glutathione were unaffected. Of note, RIPK1 inhibition also conferred resistance to oxidative stress in hepatocytes. These data collectively demonstrated a RIPK-dependent necrotic mechanism operates in the APAP-injured liver and inhibition of this pathway may be beneficial for APAP-induced fulminant hepatic failure.
authors.affiliation: Innovation Center for Immunoregulation and Therapeutics, Graduate School of Medicine, Kyoto University, Yoshida Konoe, Sakyo-ku, Kyoto 606-8501, Japan ; Division of Hepato-Biliary-Pancreatic Surgery and Transplantation, Department of Surgery, Graduate School of Medicine, Kyoto University, 54 Kawaharacho, Syogoin, Sakyo-ku, Kyoto 606-8507, Japan.
authors.forename: Kenji
authors.initials: K
authors.lastname: Takemoto
authors.affiliation: Division of Hepato-Biliary-Pancreatic Surgery and Transplantation, Department of Surgery, Graduate School of Medicine, Kyoto University, 54 Kawaharacho, Syogoin, Sakyo-ku, Kyoto 606-8507, Japan.
authors.forename: Etsuro
authors.initials: E
authors.lastname: Hatano
authors.affiliation: Department of Target Therapy Oncology, Graduate School of Medicine, Kyoto University, 54 Kawaharacho, Syogoin, Sakyo-ku, Kyoto 606-8507, Japan.
authors.forename: Keiko
authors.initials: K
authors.lastname: Iwaisako
authors.affiliation: Innovation Center for Immunoregulation and Therapeutics, Graduate School of Medicine, Kyoto University, Yoshida Konoe, Sakyo-ku, Kyoto 606-8501, Japan.
authors.forename: Masatoshi
authors.initials: M
authors.lastname: Takeiri
authors.affiliation: Innovation Center for Immunoregulation and Therapeutics, Graduate School of Medicine, Kyoto University, Yoshida Konoe, Sakyo-ku, Kyoto 606-8501, Japan.
authors.forename: Naruto
authors.initials: N
authors.lastname: Noma
authors.affiliation: Innovation Center for Immunoregulation and Therapeutics, Graduate School of Medicine, Kyoto University, Yoshida Konoe, Sakyo-ku, Kyoto 606-8501, Japan.
authors.forename: Saori
authors.initials: S
authors.lastname: Ohmae
authors.affiliation: Division of Hepato-Biliary-Pancreatic Surgery and Transplantation, Department of Surgery, Graduate School of Medicine, Kyoto University, 54 Kawaharacho, Syogoin, Sakyo-ku, Kyoto 606-8507, Japan.
authors.forename: Kan
authors.initials: K
authors.lastname: Toriguchi
authors.affiliation: Division of Hepato-Biliary-Pancreatic Surgery and Transplantation, Department of Surgery, Graduate School of Medicine, Kyoto University, 54 Kawaharacho, Syogoin, Sakyo-ku, Kyoto 606-8507, Japan.
authors.forename: Kazutaka
authors.initials: K
authors.lastname: Tanabe
authors.affiliation: Division of Hepato-Biliary-Pancreatic Surgery and Transplantation, Department of Surgery, Graduate School of Medicine, Kyoto University, 54 Kawaharacho, Syogoin, Sakyo-ku, Kyoto 606-8507, Japan.
authors.forename: Hirokazu
authors.initials: H
authors.lastname: Tanaka
authors.affiliation: Division of Hepato-Biliary-Pancreatic Surgery and Transplantation, Department of Surgery, Graduate School of Medicine, Kyoto University, 54 Kawaharacho, Syogoin, Sakyo-ku, Kyoto 606-8507, Japan.
authors.forename: Satoru
authors.initials: S
authors.lastname: Seo
authors.affiliation: Division of Hepato-Biliary-Pancreatic Surgery and Transplantation, Department of Surgery, Graduate School of Medicine, Kyoto University, 54 Kawaharacho, Syogoin, Sakyo-ku, Kyoto 606-8507, Japan.
authors.forename: Kojiro
authors.initials: K
authors.lastname: Taura
authors.affiliation: Molecular Microbiology and Immunology, Keck School of Medicine, University of Southern California, 1975 Zonal Ave, Los Angeles, CA 90033, USA.
authors.forename: Keigo
authors.initials: K
authors.lastname: Machida
authors.affiliation: Department of Cardiovascular Medicine, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8655, Japan.
authors.forename: Norihiko
authors.initials: N
authors.lastname: Takeda
authors.affiliation: Department of Target Therapy Oncology, Graduate School of Medicine, Kyoto University, 54 Kawaharacho, Syogoin, Sakyo-ku, Kyoto 606-8507, Japan.
authors.forename: Shigehira
authors.initials: S
authors.lastname: Saji
authors.affiliation: Division of Hepato-Biliary-Pancreatic Surgery and Transplantation, Department of Surgery, Graduate School of Medicine, Kyoto University, 54 Kawaharacho, Syogoin, Sakyo-ku, Kyoto 606-8507, Japan.
authors.forename: Shinji
authors.initials: S
authors.lastname: Uemoto
authors.affiliation: Innovation Center for Immunoregulation and Therapeutics, Graduate School of Medicine, Kyoto University, Yoshida Konoe, Sakyo-ku, Kyoto 606-8501, Japan.
authors.forename: Masataka
authors.initials: M
authors.lastname: Asagiri
keywords: ABTS, 2,2′-azino-bis (3-ethylbenzothiazoline)-6-sulfonic acid; ALF, acute liver failure; ALT, alanine aminotransferase; APAP, acetaminophen; AST, aspartate aminotransferase; Acetaminophen; Acute liver failure; CM-H2DCFDA, 5-(and-6)-chloromethyl-2′,7′-dichlorodihydrofluorescein diacetate, acetyl ester; CXCL1, chemokine (C-X-C motif) ligand 1; CYP2E1, cytochrome P450 2E1; DMSO, dimethyl sulfoxide; Drp1, dynamin-related protein 1; FBS, fetal bovine serum; GSH, glutathione; Hepatocytes; LDH, lactate dehydrogenase; NAPQI, N-acetyl-p-benzoquinone; NO, nitric oxide; Nec-1, necrostatin-1; Necroptosis; PGAM5, phosphoglycerate mutase family member 5; PI, propidium iodide; RIPK, receptor-interacting protein kinase; RIPK-dependent necrosis; ROS, reactive oxygen species; Reactive oxygen species; SNAP, S-nitroso-N-acetyl-dl-penicillamine; WST-8, 2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium; bFGF, basic fibroblast growth factor; λPP, lambda protein phosphatase