title: Vanadate-induced activation of activator protein-1: role of reactive oxygen species.
abstract: The present study was undertaken to test the hypothesis that the toxicity and carcinogenicity of vanadium might arise from elevation of reactive oxygen species leading to activation of the transcription factor activator protein-1 (AP-1). The AP-1 transactivation response has been implicated as causal in transformation responses to phorbol esters and growth factors. To investigate the possible activity of vanadium in the activation of AP-1, we treated mouse epidermal JB6 P+ cells stably transfected with an AP-1 luciferase reporter plasmid with various concentrations of vanadate. This resulted in concentration-dependent transactivation of AP-1. Superoxide dismutase (SOD) and catalase inhibited AP-1 activation induced by vanadate, indicating the involvement of superoxide anion radical (O2-*), hydroxyl radical (*OH) and/or H2O2 in the mechanism of vanadate-induced AP-1 activation. However, sodium formate, a specific *OH scavenger, did not alter vanadate-induced AP-1 activation, suggesting a minimal role for the *OH radical. NADPH enhanced AP-1 activation by increasing vanadate-mediated generation of O2-*. N-acetylcysteine, a thiol-containing antioxidant, decreased activation, further showing that vanadate-induced AP-1 activation involved redox reactions. Calphostin C, a specific inhibitor of protein kinase C (PKC), inhibited activation of AP-1, demonstrating that PKC is involved in the cell signal cascades leading to vanadate-induced AP-1 activation. Electron spin resonance (ESR) measurements show that JB6 P+ cells are able to reduce vanadate to generate vanadium(IV) in the presence of NADPH. Molecular oxygen was consumed during the vanadate reduction process to generate O2-* as measured by ESR spin trapping using 5,5-dimethyl-L-pyrroline N-oxide as the spin trapping agent. SOD inhibited the ESR spin adduct signal, further demonstrating the generation of O2-* in the cellular reduction of vanadate. These results provide support for a model in which vanadium, like other classes of tumor promoters, transactivates AP-1-dependent gene expression. In the case of vanadium, AP-1 transactivation is dependent on the generation of O2-* and H2O2, but not *OH.
authors.affiliation: Pathology and Physiology Research Branch, Health Effects Laboratory Division, National Institute for Occupational Safety and Health, Morgantown, WV 26505, USA.
authors.forename: M
authors.initials: M
authors.lastname: Ding
authors.forename: J J
authors.initials: JJ
authors.lastname: Li
authors.forename: S S
authors.initials: SS
authors.lastname: Leonard
authors.forename: J P
authors.initials: JP
authors.lastname: Ye
authors.forename: X
authors.initials: X
authors.lastname: Shi
authors.forename: N H
authors.initials: NH
authors.lastname: Colburn
authors.forename: V
authors.initials: V
authors.lastname: Castranova
authors.forename: V
authors.initials: V
authors.lastname: Vallyathan
mesh_terms: D000111:Acetylcysteine / Q000494:pharmacology; D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D002374:Catalase / Q000494:pharmacology; D002460:Cell Line / Q000187:drug effects; D002471:Cell Transformation, Neoplastic / Q000139:chemically induced / Q000235:genetics; D004578:Electron Spin Resonance Spectroscopy; D004791:Enzyme Inhibitors / Q000494:pharmacology; D000078404:Epidermal Cells; D005561:Formates / Q000494:pharmacology; D016166:Free Radical Scavengers / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects*; D017930:Genes, Reporter; D006861:Hydrogen Peroxide / Q000378:metabolism; D017665:Hydroxyl Radical / Q000378:metabolism; D008156:Luciferases / Q000096:biosynthesis / Q000235:genetics; D051379:Mice; D009249:NADP / Q000494:pharmacology; D009281:Naphthalenes / Q000494:pharmacology; D010084:Oxidation-Reduction; D010101:Oxygen Consumption; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000502:physiology; D017382:Reactive Oxygen Species / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D013482:Superoxide Dismutase / Q000494:pharmacology; D013481:Superoxides / Q000378:metabolism; D018808:Transcription Factor AP-1 / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation / Q000187:drug effects*; D014162:Transfection; D014638:Vanadates / Q000494:pharmacology* / Q000633:toxicity
chemical_list: D000975:Antioxidants; D004791:Enzyme Inhibitors; D005561:Formates; D016166:Free Radical Scavengers; D009281:Naphthalenes; D017382:Reactive Oxygen Species; D011993:Recombinant Fusion Proteins; D018808:Transcription Factor AP-1; C030544:formic acid; D013481:Superoxides; D017665:Hydroxyl Radical; D014638:Vanadates; D009249:NADP; D006861:Hydrogen Peroxide; D002374:Catalase; D008156:Luciferases; D013482:Superoxide Dismutase; D011493:Protein Kinase C; C058819:calphostin C; D000111:Acetylcysteine