title: Vibrio cholerae Cholix Toxin-Induced HepG2 Cell Death is Enhanced by Tumor Necrosis Factor-Alpha Through ROS and Intracellular Signal-Regulated Kinases.
abstract: Cholix toxin (Cholix) from Vibrio cholerae is a potent virulence factor exhibiting ADP-ribosyltransferase activity on eukaryotic elongation factor 2 (eEF2) of host cells, resulting in the inhibition of protein synthesis. Administration of Cholix or its homologue Pseudomonas exotoxin A (PEA) to mice causes lethal hepatocyte damage. In this study, we demonstrate cytotoxicity of Cholix on human hepatocytes in the presence of tumor necrosis factor α (TNF-α), which has been reported to play a fatal role in PEA administered to mice. Compared with incubating HepG2 cells with Cholix alone, co-treatment with TNF-α and Cholix (TNF-α/Cholix) significantly enhanced the activation of caspases, cytochrome c release from mitochondria into cytoplasm, and poly-ADP-ribose polymerase (PARP) cleavage, while incubation with TNF-α alone or co-treatment with TNF-α/catalytically inactive Cholix did not. In the early stage of cell death, Cholix increased phosphorylation of mitogen-activated protein kinases (e.g., p38, ERK, JNK) and Akt, which was not affected by TNF-α alone. MAPK inhibitors (SP600125, SB20852, and U0126) suppressed PARP cleavage induced by TNF-α/Cholix. Protein kinase inhibitor Go6976 suppressed JNK phosphorylation and PARP cleavage by TNF-α/Cholix. In contrast, PKC activator PMA in the absence of TNF-α promoted Cholix-induced PARP cleavage. Reactive oxygen species (ROS) inhibitor, N-acetyl cysteine (NAC), suppressed TNF-α/Cholix-induced JNK and ERK phosphorylation, resulting in inhibition of PARP cleavage. These data suggest that ROS and JNK pathways are important mediators of TNF-α/Cholix-induced HepG2 cell death.
authors.affiliation: Department of Molecular Infectiology, Graduate School of Medicine, Chiba University, Chiba, Japan.|Pathogenic Microbe Laboratory, Research Institute, National Center for Global Health and Medicine, Tokyo, Japan.
authors.forename: Kohei
authors.initials: K
authors.lastname: Ogura
authors.affiliation: Department of Analytic Human Pathology, Nippon Medical School, Tokyo, Japan.
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Terasaki
authors.affiliation: Pathogenic Microbe Laboratory, Research Institute, National Center for Global Health and Medicine, Tokyo, Japan.
authors.forename: Tohru
authors.initials: T
authors.lastname: Miyoshi-Akiyama
authors.affiliation: Department of Analytic Human Pathology, Nippon Medical School, Tokyo, Japan.
authors.forename: Mika
authors.initials: M
authors.lastname: Terasaki
authors.affiliation: Cardiovascular and Pulmonary Branch, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892-1590.
authors.forename: Joel
authors.initials: J
authors.lastname: Moss
authors.affiliation: Department of Molecular Infectiology, Graduate School of Medicine, Chiba University, Chiba, Japan.
authors.forename: Masatoshi
authors.initials: M
authors.lastname: Noda
authors.affiliation: Department of Molecular Infectiology, Graduate School of Medicine, Chiba University, Chiba, Japan.
authors.forename: Kinnosuke
authors.initials: K
authors.lastname: Yahiro
mesh_terms: D020727:ADP-Ribosylation Factors / Q000633:toxicity*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects* / Q000276:immunology; D001427:Bacterial Toxins / Q000633:toxicity*; D002470:Cell Survival / Q000187:drug effects / Q000276:immunology; D056486:Chemical and Drug Induced Liver Injury / Q000209:etiology* / Q000276:immunology / Q000378:metabolism; D018920:Coculture Techniques; D056945:Hep G2 Cells; D006801:Humans; D008264:Macrophages / Q000187:drug effects / Q000276:immunology; D008297:Male; D008810:Mice, Inbred C57BL; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000276:immunology / Q000633:toxicity*
chemical_list: D001427:Bacterial Toxins; D017382:Reactive Oxygen Species; D014409:Tumor Necrosis Factor-alpha; D020929:Mitogen-Activated Protein Kinase Kinases; D020727:ADP-Ribosylation Factors; C529269:cholix toxin, Vibrio cholerae
keywords: ADP-ribosyltransferase activity; ROS; cell death.; toxin; tumor necrosis factor-alpha