title: Epigallocatechin-3-gallate has an anti-platelet effect in a cyclic AMP-dependent manner.
abstract: OBJECTIVE
  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°C in the presence of 2 mM exogenous CaCl(2) 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 VASP-Ser(157) 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 TXA(2) 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 aggregation-mediated thrombotic diseases.
authors.affiliation: Department of Biomedical Laboratory Science, College of Biomedical Science and Engineering and Regional Research Center, Inje University, Gyungnam, Korea.
authors.forename: Woo-Jeong
authors.initials: WJ
authors.lastname: Ok
authors.forename: Hyun-Jeong
authors.initials: HJ
authors.lastname: Cho
authors.forename: Hyun-Hong
authors.initials: HH
authors.lastname: Kim
authors.forename: Dong-Ha
authors.initials: DH
authors.lastname: Lee
authors.forename: Hye-Yeon
authors.initials: HY
authors.lastname: Kang
authors.forename: Hyuk-Woo
authors.initials: HW
authors.lastname: Kwon
authors.forename: Man Hee
authors.initials: MH
authors.lastname: Rhee
authors.forename: Mujo
authors.initials: M
authors.lastname: Kim
authors.forename: Hwa-Jin
authors.initials: HJ
authors.lastname: Park
mesh_terms: D000225:Adenine / Q000031:analogs & derivatives / Q000494:pharmacology; D000067956:Adenylyl Cyclase Inhibitors; D000818:Animals; D002392:Catechin / Q000031:analogs & derivatives* / Q000494:pharmacology; D003094:Collagen / Q000494:pharmacology; D000242:Cyclic AMP / Q000096:biosynthesis / Q000378:metabolism*; D006152:Cyclic GMP / Q000494:pharmacology; D008297:Male; D010766:Phosphorylation; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D000067956:Adenylyl Cyclase Inhibitors; D010975:Platelet Aggregation Inhibitors; C017759:9-(tetrahydro-2-furyl)-adenine; D002392:Catechin; D003094:Collagen; C045651:epigallocatechin gallate; D000242:Cyclic AMP; D006152:Cyclic GMP; D000225:Adenine