Vascular Biology – Hemodynamics – Hypertension
Kidney International (1999) 55, 252–260; doi:10.1046/j.1523-1755.1999.00229.x
Increased NAD(P)H oxidase-mediated superoxide production in renovascular hypertension: Evidence for an involvement of protein kinase C
THOMAS HEITZER1, ULRICH WENZEL1, ULRICH HINK, DIRK KROLLNER, MIKHAIL SKATCHKOV, ROLF A K STAHL, ROLAND MACHARZINA, JAN H BRÄSEN, THOMAS MEINERTZ and THOMAS MÜNZEL
Medizinische Klinik II, Kardiologie and Nephrologie, Universitätskrankenhaus Eppendorf, Universität Hamburg, Hamburg, Germany
Correspondence: Thomas Münzel, M.D., Universitätskrankenhaus Eppendorf, Abteilung für Kardiologie, Martinistra
e 52, 20246 Hamburg, Germany. E-mail: muenzel@uke.uni-hamburg.de
1T.H. and U.W. contributed equally to the manuscript and should both be considered as first authors.
Received 1 December 1997; Revised 27 July 1998; Accepted 28 July 1998.
Abstract
Increased NAD(P)H oxidase-mediated superoxide production in renovascular hypertension: Evidence for an involvement of protein kinase C.
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.
Keywords:
arterial hypertension, angiotensin II, phosphorylation, endothelial cell, vasomotor function, nitric oxide, PKC, nitrovasodilators
Abbreviations:
ACh, acetylcholine; CP°, stable nitroxyl radicals; CPH, 1-hydroxy-3-carboxy-2,2,5,5, -tetramethylpyrrolidine; DPI, diphenylene iodinum; DTPA, diethyl-tetrapentaacetic acid; EDRF, endothelium-derived relaxing factor; ESR, electron spin resonance; HB-SOD, heparin-binding superoxide dismutase; 2K-1C, two kidney one clip; L-NNA, NG-nitro-L-arginine; NADH, nicotinamide adenine dinucleotide; NADPH, nicotinamide adenine dinucleotide phosphate; NO, nitric oxide; O-2, superoxide; PDBu, phorbol ester 12, 13-dibutyrate; PKC, protein kinase C; SOD, superoxide dismutase
