The EGCg-induced redox-sensitive activation of endothelial nitric oxide synthase and relaxation are critically dependent on hydroxyl moieties.
Several rich sources of polyphenols stimulate the endothelial formation of nitric oxide (NO), a potent vasoprotecting factor, via the redox-sensitive activation of the PI3-kinase/Akt pathway leading to the phosphorylation of endothelial NO synthase (eNOS). The present study examined the molecular mechanism underlying the stimulatory effect of epicatechins on eNOS. NO-mediated relaxation was assessed using porcine coronary artery rings in the presence of indomethacin, and charybdotoxin plus apamin, inhibitors of cyclooxygenases and EDHF-mediated responses, respectively. The phosphorylation level of Akt and eNOS was assessed in cultured coronary artery endothelial cells by Western blot, and ROS formation using dihydroethidine. (-)-Epigallocatechin-3-O-gallate (EGCg) caused endothelium-dependent relaxations in coronary artery rings and the phosphorylation of Akt and eNOS in endothelial cells. These responses were inhibited by membrane-permeant analogues of superoxide dismutase and catalase, whereas native superoxide dismutase, catalase and inhibitors of major enzymatic sources of reactive oxygen species including NADPH oxidase, xanthine oxidase, cytochrome P450 and the mitochondrial respiration chain were without effect. The EGCg derivative with all hydroxyl functions methylated induced neither relaxations nor the intracellular formation of ROS, whereas both responses were observed when the hydroxyl functions on the gallate moiety were present. In conclusion, EGCg causes endothelium-dependent NO-mediated relaxations of coronary artery rings through the Akt-dependent activation of eNOS in endothelial cells. This response is initiated by the intracellular formation of superoxide anions and hydrogen peroxide, and is critically dependent on the gallate moiety and on the presence of hydroxyl functions possibly through intracellular auto-oxidation.
Auger
Cyril
C
UMR CNRS 7213, Laboratoire de Biophotonique et Pharmacologie, Faculté de Pharmacie, Université de Strasbourg, Illkirch, France.
Kim
Jong-Hun
JH
Chabert
Philippe
P
Chaabi
Mehdi
M
Anselm
Eric
E
Lanciaux
Xavier
X
Lobstein
Annelise
A
Schini-Kerth
Valérie B
VB
D000818:Animals; D002392:Catechin / Q000031:analogs & derivatives* / Q000737:chemistry / Q000494:pharmacology; D003331:Coronary Vessels / Q000187:drug effects* / Q000201:enzymology / Q000502:physiology; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000201:enzymology / Q000502:physiology; D004789:Enzyme Activation; D006861:Hydrogen Peroxide / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000096:biosynthesis*; D010084:Oxidation-Reduction; D017382:Reactive Oxygen Species / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism; D013552:Swine; D014664:Vasodilation
D017382:Reactive Oxygen Species; D013481:Superoxides; D002392:Catechin; D006861:Hydrogen Peroxide; C045651:epigallocatechin gallate; D052250:Nitric Oxide Synthase Type III