Abstract 5299: Release of Sphingosine-1-Phosphate From Platelets Requires Thromboxane Synthesis and Thromboxane Receptor Activation
The sphingosine-derived lipid signaling molecule sphingosine-1-phosphate (S1P) is an important mediator of vascular homeostasis. S1P signaling has recently been shown to be a downstream signaling component of the receptors for thrombin, the protease-activated receptors (PARs). The present study investigates the requirement of thromboxane formation for the release of S1P induced by the prototypic thrombin receptor PAR1 from human platelets and the consequences for human monocyte and endothelial cell migration. S1P was detected by thin layer chromatography in [3H]sphingosine-labeled platelets and by mass spectrometry. Release of S1P was stimulated about 5 fold (0.3±0.1 vs. 1.6±0.5 nmol/1×106 platelets) by the selective PAR1-activating-peptide (PAR1-AP, 100 μM). Acetylsalicylic acid (ASA) or the reversible cyclooxygenase inhibitors diclofenac and ibuprofen suppressed S1P release. Oral ASA (500 mg single dose) attenuated S1P release from platelets in healthy volunteers ex vivo which was paralleled by inhibition of TX formation. S1P release was increased by the thromboxane receptor (TXR) agonist U46619, and inhibited by the TXR antagonist ramatroban. In platelets from TXR-deficient mice, thrombin-induced release of S1P was attenuated and not affected by ASA. Supernatants from PAR1-AP-stimulated platelets increased human umbilical vein endothelial cell (HUVECs) migration and the chemotactic capacity of human peripheral blood monocytes (2- and 3-fold, respectively). These effects were inhibited when platelets were pretreated with ASA. S1P receptor-specific antagonists revealed that the chemotactic responses in HUVECs were mediated by the S1P receptor-1 (S1P1) and by S1P1 and S1P3 in monocytes. These data suggest that S1P release from platelets after thrombin receptor activation requires TX synthesis and TXR activation. This novel pathway likely contributes to the regulation of vascular homeostasis during platelet activation and inflammatory processes. Inhibition of S1P may be involved in the anti-inflammatory actions of ASA in vivo, for example by affecting recruitment of endothelial cells and monocytes to sites of vascular injury.