Nicole Urtz1, Florian Gaertner1, Marie-Luise von Bruehl1, Sue Chandraratne1, Faridun Rahimi1, Lin Zhang1, Mathias Orban1, Verena Barocke1, Johannes Beil1, Irene Schubert1, Michael Lorenz1, Kyle R Legate1, Andrea Huwiler1, Josef M Pfeilschifter1, Christian Beerli1, David Ledieu1, Elke Persohn1, Andreas Billich1, Thomas Baumruker1, Michael Mederos y Schnitzler1, Steffen Massberg2. 1. From the Medizinische Klinik und Poliklinik I, Klinikum der Universität München (N.U., F.G., M.-L.v.B., S.C., F.R., M.O., V.B., J.B., I.S., M.L., K.R.L., S.M.), Department of Applied Physics, Center for NanoSciences (K.R.L.), and Walther-Straub-Institute of Pharmacology and Toxicology (M.M.y.S.), Ludwig-Maximilians-Universität, Munich, Germany; DZHK (German Centre for Cardiovascular Research), partner site Munich Heart Alliance, Munich, Germany (N.U., F.G., M.-L.v.B., S.C., F.R., M.O., J.B., I.S., M.L., M.M.y.S., S.M.); Heart Failure Institute, Research Center for Translational Medicine and Department of Cardiovascular Medicine, East Hospital, Tongji University School of Medicine, Shanghai, China (L.Z.); Institute of Pharmacology, University of Bern, Bern, Switzerland (A.H.); Pharmazentrum Frankfurt/ZAFES, Goethe University Hospital, Frankfurt am Main, Germany (J.M.P.); and Preclinical Safety (D.L., E.P.), and Autoimmunity, Transplantation and Inflammation (C.B., A.B., T.B.), Novartis Institutes for BioMedical Research, Basel, Switzerland. 2. From the Medizinische Klinik und Poliklinik I, Klinikum der Universität München (N.U., F.G., M.-L.v.B., S.C., F.R., M.O., V.B., J.B., I.S., M.L., K.R.L., S.M.), Department of Applied Physics, Center for NanoSciences (K.R.L.), and Walther-Straub-Institute of Pharmacology and Toxicology (M.M.y.S.), Ludwig-Maximilians-Universität, Munich, Germany; DZHK (German Centre for Cardiovascular Research), partner site Munich Heart Alliance, Munich, Germany (N.U., F.G., M.-L.v.B., S.C., F.R., M.O., J.B., I.S., M.L., M.M.y.S., S.M.); Heart Failure Institute, Research Center for Translational Medicine and Department of Cardiovascular Medicine, East Hospital, Tongji University School of Medicine, Shanghai, China (L.Z.); Institute of Pharmacology, University of Bern, Bern, Switzerland (A.H.); Pharmazentrum Frankfurt/ZAFES, Goethe University Hospital, Frankfurt am Main, Germany (J.M.P.); and Preclinical Safety (D.L., E.P.), and Autoimmunity, Transplantation and Inflammation (C.B., A.B., T.B.), Novartis Institutes for BioMedical Research, Basel, Switzerland. steffen.massberg@med.uni-muenchen.de.
Abstract
RATIONALE: Platelets are known to play a crucial role in hemostasis. Sphingosine kinases (Sphk) 1 and 2 catalyze the conversion of sphingosine to the bioactive metabolite sphingosine 1-phosphate (S1P). Although platelets are able to secrete S1P on 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 with Sphk1(-/-) or wild-type mice, as analyzed by mass spectrometry. Sphk2(-/-) platelets also failed to secrete S1P on stimulation. Blood from Sphk2-deficient mice showed decreased aggregation after protease-activated receptor 4-peptide and adenosine diphosphate stimulation in vitro, as assessed by whole blood impedance aggregometry. We revealed that S1P controls platelet aggregation via the sphingosine 1-phosphate receptor 1 through modulation of protease-activated receptor 4-peptide and adenosine diphosphate-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.
RATIONALE: Platelets are known to play a crucial role in hemostasis. Sphingosine kinases (Sphk) 1 and 2 catalyze the conversion of sphingosine to the bioactive metabolite sphingosine 1-phosphate (S1P). Although platelets are able to secrete S1P on 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 with Sphk1(-/-) or wild-type mice, as analyzed by mass spectrometry. Sphk2(-/-) platelets also failed to secrete S1P on stimulation. Blood from Sphk2-deficientmice showed decreased aggregation after protease-activated receptor 4-peptide and adenosine diphosphate stimulation in vitro, as assessed by whole blood impedance aggregometry. We revealed that S1P controls platelet aggregation via the sphingosine 1-phosphate receptor 1 through modulation of protease-activated receptor 4-peptide and adenosine diphosphate-induced platelet activation. Finally, we show by intravital microscopy that defective platelet aggregation in Sphk2-deficientmice 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-deficientmice 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.
Authors: Thiet M Vu; Ayako-Nakamura Ishizu; Juat Chin Foo; Xiu Ru Toh; Fangyu Zhang; Ding Ming Whee; Federico Torta; Amaury Cazenave-Gassiot; Takayoshi Matsumura; Sangho Kim; Sue-Anne E S Toh; Toshio Suda; David L Silver; Markus R Wenk; Long N Nguyen Journal: Nature Date: 2017-10-18 Impact factor: 49.962
Authors: Konstantin Stark; Vanessa Philippi; Sven Stockhausen; Johanna Busse; Antonella Antonelli; Meike Miller; Irene Schubert; Parandis Hoseinpour; Sue Chandraratne; Marie-Luise von Brühl; Florian Gaertner; Michael Lorenz; Alessandra Agresti; Raffaele Coletti; Daniel J Antoine; Ralf Heermann; Kirsten Jung; Sven Reese; Iina Laitinen; Markus Schwaiger; Axel Walch; Markus Sperandio; Peter P Nawroth; Christoph Reinhardt; Sven Jäckel; Marco E Bianchi; Steffen Massberg Journal: Blood Date: 2016-08-29 Impact factor: 22.113