Peroxide generation by p47phox-Src activation of Nox2 has a key role in protein kinase C-induced arterial smooth muscle contraction

• Published in: American journal of physiology. Heart and circulatory physiology 2009-04, Vol.296 (4), p.H1048-H1057
• Main Authors: Gupte, Sachin A, Kaminski, Pawel M, George, Shimran, Kouznestova, Lioubov, Olson, Susan C, Mathew, Rajamma, Hintze, Thomas H, Wolin, Michael S
• Format: Article
• Language: English
• Subjects: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid - pharmacology; Animals; Aorta - drug effects; Aorta - metabolism; Cattle; Coronary Vessels - drug effects; Coronary Vessels - metabolism; Membrane Glycoproteins - metabolism; Mice; Muscle Contraction - drug effects; Muscle Contraction - physiology; Muscle, Smooth, Vascular - drug effects; Muscle, Smooth, Vascular - metabolism; NADPH Oxidase 2; NADPH Oxidases - metabolism; Nitroglycerin - pharmacology; Peroxides - metabolism; Phorbol 12,13-Dibutyrate - pharmacology; Protein Kinase C - metabolism; Reactive Oxygen Species - metabolism; Receptors, Thromboxane - agonists; src-Family Kinases - metabolism; Superoxides - metabolism; Vasoconstriction - drug effects; Vasoconstriction - physiology; Vasoconstrictor Agents - pharmacology; Vasodilator Agents - pharmacology
• ISSN: 0363-6135; 1522-1539
• DOI: 10.1152/ajpheart.00491.2008

Departments of 1 Physiology, 2 Biochemistry & Molecular Biology, and 3 Pediatrics, New York Medical College, Valhalla, New York Submitted 11 May 2008 ; accepted in final form 13 January 2009 Protein kinase C (PKC) stimulation of NAD(P)H oxidases (Nox) is an important component of multiple vascular disease processes; however, the relationship between oxidase activation and the regulation of vascular smooth muscle contraction by PKC remains poorly understood. Therefore, we examined the signaling cascade of PKC-elicited Nox activation and the role of superoxide and hydrogen peroxide in mediating PKC-induced vascular contraction. Endothelium-denuded bovine coronary arteries showed a PKC-dependent basal production of lucigenin (5 µM)-detected Nox oxidase-derived superoxide, which was stimulated fourfold by PKC activation with 10 µM phorbol 12,13-dibutyrate (PDBu). PDBu appeared to increase superoxide generation by Nox2 through both p47 phox and peroxide-dependent Src activation mechanisms based on the actions of inhibitors, properties of Src phosphorylation, and the loss of responses in aorta from mice deficient in Nox2 and p47 phox . The actions of inhibitors of contractile regulating mechanisms, scavengers of superoxide and peroxide, and responses in knockout mouse aortas suggest that a major component of the contraction elicited by PDBu appeared to be mediated through peroxide derived from Nox2 activation stimulating force generation through Rho kinase and calmodulin kinase-II mechanisms. Superoxide generated by PDBu also attenuated relaxation to nitroglycerin. Peroxide-derived from Nox2 activation by PKC appeared to be a major contributor to the thromboxane A 2 receptor agonist U46619 [GenBank] (100 nM)-elicited contraction of coronary arteries. Thus a p47 phox and Src kinase activation of peroxide production by Nox2 appears to be an important contributor to vascular contractile mechanisms mediated through activation of PKC. hydrogen peroxide; NADPH oxidase; protein kinase C; Src kinases; coronary artery Address for reprint requests and other correspondence: M. S. Wolin, Dept. of Physiology, New York Medical College, Valhalla, NY 10595 (e-mail: mike_wolin{at}nymc.edu )