Red wine polyphenolic compounds (RWPCs) are potent inducers of endothelium-dependent relaxations of coronary arteries, which involve both nitric oxide and endothelium-derived hyperpolarizing factor (EDHF). The EDHF-mediated relaxation to RWPCs is critically dependent on the formation of reactive oxygen species by a flavin-dependent enzyme. The aim of the present study was to determine the role of redox-sensitive protein kinases including p38 MAPK, ERK1/2 and PI3-kinase/Akt in RWPCs-induced EDHF-mediated relaxation. Porcine coronary artery rings were suspended in organ chambers for measurement of changes in isometric tension. Confluent cultures of porcine coronary artery endothelial cells were used to determine the phosphorylation level of p38 MAPK, ERK1/2 and Akt by Western blot analysis. All experiments were performed in the presence of indomethacin and Nomega-nitro-L-arginine. RWPCs caused pronounced endothelium-dependent relaxations, which were significantly reduced by wortmannin and LY294002, two inhibitors of PI3-kinase, and not affected by PD98059 (an inhibitor of ERK1/2 kinase kinase) and SB203580 (an inhibitor of p38 MAPK). In contrast, wortmannin did not affect relaxations to bradykinin or levcromakalim. RWPCs elicited within minutes a sustained and concentration-dependent phosphorylation of p38 MAPK, ERK1/2 and Akt in endothelial cells. The phosphorylation of Akt in response to RWPCs was abolished by wortmannin and LY294002, and by the membrane-permeant analogue of superoxide dismutase Mn(III)tetrakis(1-methyl-4-pyridyl)porphyrin. The present findings demonstrate that RWPCs cause EDHF-mediated relaxations of coronary arteries; these responses are critically dependent on the redox-sensitive activation of the PI3-kinase/Akt pathway in endothelial cells.
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