Epigallocatechin-3-gallate has an anti-platelet effect in a cyclic AMP-dependent manner

Aim: in this study,we investigated the effect of (-)-epigallocatechin-3-gallate (egcg) on cyclic nucleotide production and vasodilator-stimulated phosphoprotein (vasp) phosphorylation in collagen (10 μg/ml)-stimulated platelet aggregation. methods: washed platelets (10 8/ml) from sprague-dawley rats (6-7 weeks old,male) were preincubated for 3 min at 37° in the presence of 2 mm exogenous cacl2 with or without egcg or other materials,stimulated with collagen (10 μg/ml) for 5 min,and then used for the determination of intracellular cytosolic ca 2+ ([ca 2+]i),thromboxane a 2 (txa 2),adenosine 3',5'-cyclic monophosphate (camp),guanosine 3',5'-cyclic monophosphate (cgmp),and vasp phosphorylation. results: egcg dose-dependently inhibited collagen-induced platelet aggregation by inhibiting both [ca 2+]i mobilization and txa 2 production. of two aggregation-inhibiting molecules,camp and cgmp,egcg significantly increased intracellular levels of camp,but not cgmp. egcg-elevated camp level was decreased by sq22536,an adenylate cyclase inhibitor,but not by etazolate,a campspecific phosphodiesterase inhibitor. in addition,egcg elevated the phosphorylation of vasp-ser 157,a camp-dependent protein kinase (a-kinase) substrate,but not the phosphorylation of vasp-ser 239,a cgmp-dependent protein kinase substrate,in intact platelets and collagen-induced platelets,and vaspser157 phosphorylation by egcg was inhibited by both an adenylate cyclase inhibitor sq22536 and an a-kinase inhibitor rp-8-br-camps. we have demonstrated that egcg increases camp via adenylate cyclase activation and subsequently phosphorylates vasp-ser 157 through a-kinase activation to inhibit [ca 2+]i mobilization and txa2 production on collagen-induced platelet aggregation. conclusions: these results strongly indicate that egcg is a beneficial compound elevating camp level in collagen-platelet interaction,which may result in the prevention of platelet aggregationmediated thrombotic diseases.