eNOS activation induced by a polyphenol-rich grape skin extract in porcine coronary arteries.
OBJECTIVE
Drinking red wine is associated with a decreased mortality from coronary heart diseases. This study examined whether polyphenols contained in a grape skin extract (GSE) triggered the endothelial formation of nitric oxide (NO) and investigated the underlying mechanism.


METHODS
Vascular reactivity was assessed in organ chambers using porcine coronary artery rings in the presence of indomethacin (a cyclooxygenase inhibitor) and charybdotoxin plus apamin (inhibitors of endothelium-derived hyperpolarizing factor-mediated responses). The phosphorylation level of Src, Akt and endothelial NO synthase (eNOS) were assessed by Western blot analysis, and the formation of reactive oxygen species (ROS) was investigated using dihydroethidine and dichlorodihydrofluorescein.


RESULTS
GSE-induced endothelium-dependent relaxations were abolished by N(G)-nitro-L-arginine (an eNOS inhibitor) and ODQ (a soluble guanylyl cyclase inhibitor), and they were reduced by MnTMPyP, polyethyleneglycol catalase, PP2 (an inhibitor of Src kinase) and wortmannin (an inhibitor of phosphoinositide 3-kinase). GSE caused phosphorylation of Src, which was prevented by MnTMPyP. It also caused phosphorylation of Akt and eNOS, which were prevented by MnTMPyP, polyethyleneglycol catalase, PP2, wortmannin and LY294002. GSE elicited the formation of ROS in native and cultured endothelial cells, which was prevented by MnTMPyP.


CONCLUSIONS
GSE causes endothelium-dependent NO-mediated relaxations of coronary arteries. This effect involves the intracellular formation of ROS in endothelial cells leading to the Src kinase/phosphoinositide 3-kinase/Akt-dependent phosphorylation of eNOS.
Madeira
Socorro Vanesca Frota
SV
UMR CNRS 7175, Pharmacologie et Physico-Chimie, Faculté de Pharmacie, Université Louis Pasteur, Illkirch, France.
Auger
Cyril
C
Anselm
Eric
E
Chataigneau
Marta
M
Chataigneau
Thierry
T
Soares de Moura
Roberto
R
Schini-Kerth
Valérie B
VB
D000818:Animals; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003331:Coronary Vessels / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology; D004789:Enzyme Activation / Q000187:drug effects; D005419:Flavonoids / Q000494:pharmacology*; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000502:physiology; D009569:Nitric Oxide / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism*; D009924:Organ Culture Techniques; D010084:Oxidation-Reduction; D010636:Phenols / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects / Q000502:physiology; D010936:Plant Extracts / Q000494:pharmacology*; D059808:Polyphenols; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D013552:Swine; D014664:Vasodilation / Q000187:drug effects / Q000502:physiology; D027843:Vitis; D019061:src-Family Kinases / Q000378:metabolism
D005419:Flavonoids; D010636:Phenols; D010936:Plant Extracts; D059808:Polyphenols; D017382:Reactive Oxygen Species; D009569:Nitric Oxide; D052250:Nitric Oxide Synthase Type III; D019061:src-Family Kinases; D051057:Proto-Oncogene Proteins c-akt