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von Willebrand factor activates endothelial nitric oxide synthase in blood platelets by a glycoprotein Ib‐dependent mechanism
Author(s) -
RIBA R.,
OBERPRIELER N. G.,
ROBERTS W.,
NASEEM K. M.
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.1538-7836.2006.02195.x
Subject(s) - platelet activation , chemistry , thromboxane a2 , platelet , signal transduction , microbiology and biotechnology , cyclic guanosine monophosphate , enos , phosphorylation , nitric oxide , medicine , biochemistry , biology , receptor , nitric oxide synthase , organic chemistry
Summary. Background: The molecular regulation of endothelial nitric oxide synthase (eNOS) in blood platelets and the signalling events induced by platelet‐derived NO are poorly defined. In particular, the ability of von Willebrand factor (VWF) to stimulate cyclic guanosine monophosphate (cGMP) formation in platelets has produced conflicting data. Objectives: To determine the mechanisms leading to eNOS activation and clarify the downstream signaling pathways activated by platelet‐derived NO in response to VWF. Methods: We used three independent markers of NO signaling, [ 3 H] l ‐citrulline production, cGMP accrual and immunoblotting of vasodilator–stimulated phosphoprotein (VASP) to examine the NO signaling cascade in response to VWF. Results: VWF increased NO synthesis and bioavailability, as evidenced by increased [ 3 H] l ‐citrulline production and cGMP accrual, respectively. VWF‐induced eNOS activation was GPIb‐IX‐dependent and independent of integrin α IIb β 3 . cGMP formation in response to VWF required Ca 2+ mobilization, Src family kinases, phosphatidylinositol 3‐kinase and phospholipase C, but not protein kinase C. This suggests that a cross‐talk between the signaling mechanisms regulates platelet activation and NO synthesis. VWF‐induced cGMP accrual was completely blocked by apyrase and indomethacin, demonstrating an essential role for platelet‐derived ADP and thromboxane A 2 (TxA 2 ). Elevated cGMP levels led to increased VASP phosphorylation at serine 239 that was both protein kinase G (PKG)‐ and protein kinase A (PKA)‐dependent. Conclusions: We demonstrate that VWF activates eNOS through a specific Ca 2+ ‐dependent GPIb receptor‐signaling cascade that relies on the generation of platelet‐derived ADP and TxA 2 . Furthermore, we provide the first evidence to suggest that platelet derived‐NO/cGMP activates PKA in addition to PKG.