S1P-receptor 1 signalling reduces arterial thrombosis via upregulation of endothelial thrombomodulin expression
M Benkhoff, P Mourikis, T Huckenbeck, B Knoop, M Barcik, G Al-Kassis, K Trojovsky, P Wollnitzke, L Dannenberg, M Kelm, B Levkau, A PolzinAbstract
Background
Sphingosine 1-phosphate (S1P) is an important lipid mediator in the cardiovascular system. Numerous links between S1P and blood coagulation have been described. However, results regarding the role of S1P in coagulation are inconclusive. A physiological antithrombotic regulator expressed by endothelial cells (EC) is thrombomodulin (TM). To date, the impact of S1P on TM-expression is unknown.
Methods
We evaluated S1P effects on TM-expression, platelet adhesion, activation in vitro and ex vivo as well as thrombus formation in vivo. In addition, we analysed the association of plasma S1P levels and circulating thrombin in 74 patients with cardiovascular disease.
Results
We revealed that S1P significantly increased TM-expression on HUVECs by 25 %. This effect was abolished by S1P receptor 1 (S1PR1)- inhibition with W146. Further analyses on the downstream signalling revealed that phosphoinositide 3-kinase (PI3K) inhibition blunted S1P-dependent TM-expression. Furthermore, S1P reduced platelet adhesion on ECs in flow-chamber experiments. In absence of ECs, S1P did not affect platelet activation. To assess the in vivo effects of our findings, we first investigated TM-expression in mice that were treated with S1P. We found increased TM-expression in aortic endothelium after S1P treatment. The increase of aortic TM-expression led to reduced in vivo thrombus formation as no occlusion and reduced maximal thrombus size could be observed in S1P treated animals. Importantly, S1P treatment did not enhance bleeding in these animals. Vice versa, SphK1-deficient mice with low S1P concentrations had reduced endothelial TM-expression and enhanced in vivo arterial thrombus formation. This could be rescued by TM-treatment of these mice. In line, in an all-comer cohort of 74 patients with cardiovascular disease, higher S1P concentrations were associated with lower circulating thrombin concentrations.
Conclusion
In conclusion, S1P inhibited thrombus formation in an endothelium- and TM-dependent manner. Mechanistically, activation of S1PR1 led to PI3K-mediated TM-upregulation. This might be a therapeutic target in prevention of thrombus formation without enhancing bleeding risk.