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P2Y 12 receptor‐mediated potentiation of thrombin‐induced thromboxane A 2 generation in platelets occurs through regulation of Erk1/2 activation
Author(s) -
SHANKAR H.,
GARCIA A.,
PRABHAKAR J.,
KIM S.,
KUNAPULI S. P.
Publication year - 2006
Publication title -
journal of thrombosis and haemostasis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.947
H-Index - 178
eISSN - 1538-7836
pISSN - 1538-7933
DOI - 10.1111/j.1538-7836.2006.01789.x
Subject(s) - thromboxane a2 , platelet , p2y12 , chemistry , thrombin , platelet activation , purinergic receptor , thrombin receptor , phospholipase c , receptor , p2y receptor , protease activated receptor , microbiology and biotechnology , endocrinology , medicine , biochemistry , biology , platelet aggregation
Summary. Background: Thromboxane A 2 (TXA 2 ) is a positive feedback lipid mediator that is generated upon stimulation of platelets with various agonists. Aspirin works as an antithrombotic drug by blocking the generation of TXA 2 . The aim of this study was to evaluate the role of the purinergic P2Y receptors in thrombin‐induced TXA 2 generation. Results: PAR1‐activating peptide (SFLLRN), PAR4‐activating peptide (AYPGKF), and thrombin, induced the activation of cytosolic phospholipase A 2 (cPLA 2 ), release of arachidonic acid (AA) from membrane‐bound phospholipids, and subsequent TXA 2 generation in human platelets. The actions of these agonists were significantly inhibited in the presence of the P2Y 12 receptor antagonist, AR‐C69931MX, but not the P2Y 1 receptor antagonist, MRS2179. In addition, AYPGKF‐ and thrombin‐induced TXA 2 generation was significantly reduced in platelets from mice dosed with clopidogrel, confirming the results obtained with the human platelets. Also, Pearl mouse platelets that lack releasable nucleotides generated significantly less TXA 2 when compared with the wild‐type littermates in response to PAR stimulation. Inhibition of extracellular signal‐regulated protein kinase 1/2 (Erk 1/2) activation using U0126, an inhibitor of MAP kinase kinase (MEK), suppressed PAR‐mediated cPLA 2 phosphorylation and TXA 2 generation. Further, platelets that were pretreated with AR‐C69931MX, as well as Pearl mouse platelets, displayed the reduced levels of Erk1/2 phosphorylation upon stimulation with the PAR agonists. Conclusions: Based on these findings, we conclude that thrombin‐induced Erk1/2 activation is essential for PAR‐mediated TXA 2 generation, which is potentiated by the P2Y 12 receptor‐mediated signaling pathway but not the P2Y 1 receptor‐mediated signaling pathway. Finally, using selective inhibitors of Src kinases, we show that PAR‐mediated Src activation precedes Erk1/2 activation.