Sphingosine 1-Phosphate Produced by Sphingosine Kinase 2 Intrinsically Controls Platelet Aggregation In Vitro And In Vivo
Rationale: Platelets are known to play a crucial role in hemostasis. Sphingosine kinases 1 and 2 (Sphk) catalyze the conversion of sphingosine (Sph) to the bioactive metabolite sphingosine 1-phosphate (S1P). Although platelets are able to secrete S1P upon activation, little is known about a potential intrinsic effect of S1P on platelet function.
Objective: To investigate the role of Sphk1- and Sphk2-derived S1P in the regulation of platelet function.
Methods and Results: We found a 100-fold reduction in intracellular S1P levels in platelets derived from Sphk2-/- mutants compared to Sphk1-/- or wildtype (WT) mice, as analyzed by mass spectrometry. Sphk2-/- platelets also failed to secrete S1P upon stimulation. Blood from Sphk2-deficient mice showed decreased aggregation after protease-activated receptor 4-peptide (PAR4-P) and adenosine-diphosphate (ADP) stimulation in vitro, as assessed by whole blood impedance aggregometry. We revealed that S1P controls platelet aggregation via the sphingosine 1-phosphate receptor 1 (S1PR1) through modulation of PAR4-P and ADP induced platelet activation. Finally, we show by intravital microscopy that defective platelet aggregation in Sphk2 deficient mice translates into reduced arterial thrombus stability in vivo.
Conclusions: We demonstrate that Sphk2 is the major Sphk isoform responsible for the generation of S1P in platelets and plays a pivotal intrinsic role in the control of platelet activation. Correspondingly, Sphk2-deficient mice are protected from arterial thrombosis after vascular injury, but have normal bleeding times. Targeting this pathway could therefore present a new therapeutic strategy to prevent thrombosis.
- Received May 22, 2015.
- Revision received June 26, 2015.
- Accepted June 30, 2015.