title: Increased NAD(P)H oxidase-mediated superoxide production in renovascular hypertension: evidence for an involvement of protein kinase C.
abstract: BACKGROUND
  Angiotensin II infusion has been shown to cause hypertension and endothelial dysfunction and to increase superoxide (O-.2) production in vascular tissue, mainly via an activation of nicotinamide adenine dinucleotide (phosphate) [NAD(P)H]-dependent oxidase, the most significant O-.2 source in endothelial and/or smooth muscle cells. With these studies, we sought to determine whether endothelial dysfunction in renovascular hypertension is secondary to an activation of these oxidases.
  
  
  METHODS
  Endothelial function in aortas from rats with two kidney-one clip (2K-1C) hypertension and age-matched controls was assessed using isometric tension studies in organ chambers. Changes in vascular O-.2 production were measured using lucigenin-enhanced chemiluminescence and electron spin resonance spectroscopy.
  
  
  RESULTS
  In hypertensive animals, relaxation to endothelium-dependent (acetylcholine) and endothelium-independent nitrovasodilators (nitroglycerin) was impaired. Constriction to a direct activator of protein kinase C (PKC) phorbol ester 12,13 dibutyrate (PDBu) was enhanced, and vascular O-.2 was significantly increased compared with controls. Vascular O-.2 was normalized by the PKC inhibitor calphostin C, by the inhibitor of flavin-dependent oxidases, diphenylene iodonium, and recombinant heparin-binding superoxide dismutase, whereas inhibitors of the xanthine oxidase (oxypurinol), nitric oxide synthase (NG-nitro-l-arginine) and mitochondrial NADH dehydrogenase (rotenone) were ineffective. Studies of vascular homogenates demonstrated that the major source of O-.2 was a NAD(P)H-dependent oxidase. Incubation of intact tissue with PDBu markedly increased O-. 2, the increase being significantly stronger in vessels from hypertensive animals as compared with vessels from controls. Endothelial dysfunction was improved by preincubation of vascular tissue with superoxide dismutase and calphostin C.
  
  
  CONCLUSIONS
  We therefore conclude that renovascular hypertension in 2K-1C rats is associated with increased vascular O-.2 leading to impaired vasodilator responses to endogenous and exogenous nitrovasodilators. Increased vascular O-.2 is likely secondary to a PKC-mediated activation of a membrane-associated NAD(P)H-dependent oxidase.
authors.affiliation: Medizinische Klinik II, Kardiologie and Nephrologie, Universit atskrankenhaus Eppendorf, Universität Hamburg, Hamburg, Germany.
authors.forename: T
authors.initials: T
authors.lastname: Heitzer
authors.forename: U
authors.initials: U
authors.lastname: Wenzel
authors.forename: U
authors.initials: U
authors.lastname: Hink
authors.forename: D
authors.initials: D
authors.lastname: Krollner
authors.forename: M
authors.initials: M
authors.lastname: Skatchkov
authors.forename: R A
authors.initials: RA
authors.lastname: Stahl
authors.forename: R
authors.initials: R
authors.lastname: MacHarzina
authors.forename: J H
authors.initials: JH
authors.lastname: Bräsen
authors.forename: T
authors.initials: T
authors.lastname: Meinertz
authors.forename: T
authors.initials: T
authors.lastname: Münzel
mesh_terms: D000109:Acetylcholine / Q000494:pharmacology; D000818:Animals; D001013:Aorta, Thoracic / Q000187:drug effects / Q000503:physiopathology; D004730:Endothelium, Vascular / Q000187:drug effects / Q000503:physiopathology; D006978:Hypertension, Renovascular / Q000378:metabolism* / Q000503:physiopathology; D066298:In Vitro Techniques; D008297:Male; D009097:Multienzyme Complexes / Q000378:metabolism*; D009243:NAD / Q000378:metabolism; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism*; D009249:NADP / Q000378:metabolism; D019255:NADPH Oxidases / Q000378:metabolism*; D005996:Nitroglycerin / Q000494:pharmacology; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D011189:Potassium Chloride / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013481:Superoxides / Q000378:metabolism*; D014664:Vasodilation / Q000187:drug effects
chemical_list: D009097:Multienzyme Complexes; D009243:NAD; D013481:Superoxides; D015240:Phorbol 12,13-Dibutyrate; D009249:NADP; D011189:Potassium Chloride; C023107:NADH oxidase; D009247:NADH, NADPH Oxidoreductases; D019255:NADPH Oxidases; D011493:Protein Kinase C; D005996:Nitroglycerin; D000109:Acetylcholine