title: (-)-epicatechin activation of endothelial cell endothelial nitric oxide synthase, nitric oxide, and related signaling pathways.
abstract: Recent reports indicate that (-)-epicatechin can exert cardioprotective actions, which may involve endothelial nitric oxide synthase (eNOS)-mediated nitric oxide production in endothelial cells. However, the mechanism by which (-)-epicatechin activates eNOS remains unclear. In this study, we proposed to identify the intracellular pathways involved in (-)-epicatechin-induced effects on eNOS, using human coronary artery endothelial cells in culture. Treatment of cells with (-)-epicatechin led to time- and dose-dependent effects that peaked at 10 minutes at 1 mumol/L. (-)-Epicatechin treatment activates eNOS via serine 633 and serine 1177 phosphorylation and threonine 495 dephosphorylation. Using specific inhibitors, we have established the participation of the phosphatidylinositol 3-kinase pathway in eNOS activation. (-)-Epicatechin induces eNOS uncoupling from caveolin-1 and its association with calmodulin-1, suggesting the involvement of intracellular calcium. These results allowed us to propose that (-)-epicatechin effects may be dependent on actions exerted at the cell membrane level. To test this hypothesis, cells were treated with the phospholipase C inhibitor U73122, which blocked (-)-epicatechin-induced eNOS activation. We also demonstrated inositol phosphate accumulation in (-)-epicatechin-treated cells. The inhibitory effects of the preincubation of cells with the calmodulin-dependent kinase II (CaMKII) inhibitor KN-93 indicate that (-)-epicatechin-induced eNOS activation is at least partially mediated via the Ca(2+)/CaMKII pathway. The (-)-epicatechin stereoisomer catechin was only partially able to stimulate nitric oxide production in cells. Together, these results strongly suggest the presence of a cell surface acceptor-effector for the cacao flavanol (-)-epicatechin, which may mediate its cardiovascular effects.
authors.affiliation: Department of Medicine, University of California, San Diego, Calif, USA.
authors.forename: Israel
authors.initials: I
authors.lastname: Ramirez-Sanchez
authors.forename: Lisandro
authors.initials: L
authors.lastname: Maya
authors.forename: Guillermo
authors.initials: G
authors.lastname: Ceballos
authors.forename: Francisco
authors.initials: F
authors.lastname: Villarreal
mesh_terms: D000328:Adult; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D002392:Catechin / Q000378:metabolism / Q000494:pharmacology*; D022461:Caveolins / Q000378:metabolism; D002478:Cells, Cultured; D003331:Coronary Vessels / Q000166:cytology; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D008297:Male; D008875:Middle Aged; D009569:Nitric Oxide / Q000378:metabolism*; D052250:Nitric Oxide Synthase Type III / Q000187:drug effects / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D011336:Probability; D012016:Reference Values; D012494:Sampling Studies; D015398:Signal Transduction / Q000187:drug effects
chemical_list: D022461:Caveolins; D009569:Nitric Oxide; D002392:Catechin; D052250:Nitric Oxide Synthase Type III; D019869:Phosphatidylinositol 3-Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases
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