title: Nitric oxide inhibits dioxin action for the stimulation of Cyp1a1 promoter activity.
abstract: Since it is known that hypoxia increases inducible nitric oxide synthase (iNOS) gene expression through the hypoxia responsive element, it was hypothesized that nitric oxide could be a mediator of hypoxia to inhibit Cyp1a1 promoter activity. In order to test this hypothesis, we have undertaken a study to examine the effects of hypoxia and nitric oxide on Cyp1a1 promoter activity in Hepa I cells. Mouse Cyp1a1 5' flanking DNA, 1.6kb, was cloned into pGL3 expression vector in order to construct pmCyp1a1-Luc. Hepa I cells were transfected with pmCyp1a1-Luc and were treated with 10(-9)M 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in the presence or absence of various hypoxic agents such as 10(-6)-10(-4) cobalt chloride or 10(-6)-10(-4)M picolinic acid or 10(-6)-10(-4) M desferrioxamine. The luciferase activity of the reporter gene was measured from pmCyp1a1-Luc transfected Hepa I cell lysate which contains 2 microg total protein using luciferin as a substrate. Hypoxic agents such as cobalt chloride, picolinic acid, and desferrioxamine showed inhibition of luciferase activity that was induced by 10(-9) M TCDD treatment in a dose dependent manner. Concomitant treatment of 1 mM N(G)-nitro-l-arginine with 10(-6)-10(-4) M cobalt chloride or 10(-6)-10(-4) M desferrioxamine or 10(-6)-10(-4) M picolinic acid or 10(-6)-10(-4) M sodium nitroprusside recovered luciferase activity from the TCDD induced luciferase activity that was inhibited by hypoxic agents. These data demonstrated that nitric oxide might be a mediator of iron chelating agents and hypoxic agents to inhibit dioxin induced Cyp1a1 promoter activity in Hepa I cells.
authors.affiliation: College of Pharmacy, Ewha Womans University, Seoul, Korea.
authors.forename: J E
authors.initials: JE
authors.lastname: Kim
authors.forename: Y Y
authors.initials: YY
authors.lastname: Sheen
mesh_terms: D000818:Animals; D001120:Arginine / Q000494:pharmacology; D002478:Cells, Cultured; D003035:Cobalt / Q000494:pharmacology; D019363:Cytochrome P-450 CYP1A1 / Q000235:genetics* / Q000378:metabolism; D003676:Deferoxamine / Q000494:pharmacology; D004147:Dioxins / Q000037:antagonists & inhibitors* / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D017930:Genes, Reporter / Q000187:drug effects; D008070:Lipopolysaccharides / Q000494:pharmacology; D008156:Luciferases / Q000378:metabolism; D051379:Mice; D009569:Nitric Oxide / Q000378:metabolism*; D019335:Nitroarginine / Q000494:pharmacology; D009599:Nitroprusside / Q000494:pharmacology; D010848:Picolinic Acids / Q000494:pharmacology; D000072317:Polychlorinated Dibenzodioxins / Q000494:pharmacology; D011401:Promoter Regions, Genetic; D013723:Teratogens / Q000494:pharmacology; D014162:Transfection
chemical_list: D004147:Dioxins; D004791:Enzyme Inhibitors; D008070:Lipopolysaccharides; D010848:Picolinic Acids; D000072317:Polychlorinated Dibenzodioxins; D013723:Teratogens; D009599:Nitroprusside; D019335:Nitroarginine; D009569:Nitric Oxide; D003035:Cobalt; D001120:Arginine; D008156:Luciferases; D019363:Cytochrome P-450 CYP1A1; C018021:cobaltous chloride; D003676:Deferoxamine; C030614:picolinic acid