HEMOSTASIS, THROMBOSIS, AND VASCULAR BIOLOGY Sequential activation of p38 and ERK pathways by cGMP-dependent protein kinase leading to activation of the platelet integrin IIb 3

Integrin activation (inside-out signaling) in platelets can be initiated by agonists such as von Willebrand factor (VWF) and thrombin. Here we show that a mitogenactivated protein kinase (MAPK), p38, plays an important role in the activation of integrin IIb 3 induced by VWF and thrombin. A dominant-negative mutant of p38, p38AF, inhibits IIb 3 activation induced by VWF binding to its receptor, the platelet glycoprotein Ib-IX (GPIb-IX), and p38 inhibitors diminish platelet aggregation induced by VWF or low-dose thrombin. The inhibitory effect of p38 inhibitor is unlikely to be caused by the previous suggested effect on cyclo-oxygenase, as inhibition also was observed in the presence of high concentrations of cyclooxygenase inhibitor, aspirin. VWF or thrombin induces p38 activation, which is inhibited in cGMP-dependent protein kinase (PKG)–knockout mouse platelets and PKG inhibitor–treated human platelets, indicating that activation of p38 is downstream from PKG in the signaling pathway. p38AF or p38 inhibitors diminish PKG-induced phosphorylation of extracellular stimuli-responsive kinase (ERK), which also is important in integrin activation. Thus, p38 plays an important role in mediating PKG-dependent activation of ERK. These data delineate a novel signaling pathway in which platelet agonists sequentially activate PKG, p38, and ERK pathways leading to integrin activation. (Blood. 2006;107:965-972)