Phospholipase D1 is a regulator of inflammatory platelet-endothelial interactions under high shear conditions
M. Klier1, N. Gowert1, S. Jäckel2, C. Reinhardt2, M. Elvers1 (1Düsseldorf, Germany, 2Mainz, Germany)
Time: 11:00 - 12:00
Objective: GPIb-dependent integrin activation is essential for stable thrombus formation under conditions of high shear. Recent evidence suggests that GPIb is not only required for stable thrombus formation but for platelet-mediated inflammatory responses. Because PLD1 is essential for GPIb-mediated integrin activation we here analysed if PLD1 is important for platelet-mediated inflammatory responses and further studied the relevance of PLD1 for platelet mediated endothelial activation and leukocyte recruitment.
Methods: In vitro and in vivo analysis of PLD1-/- mice to analyse the interaction of platelets, leukocytes and the (inflammatory) endothel under static and under flow conditions.
Results: Pld1-/- platelets showed strongly reduced adhesion to TNF-alpha stimulated endothelial cells (ECs) under high but not under intermediate shear conditions ex vivo. This together with significantly reduced CD40L expression and integrin activation of Pld1-/- platelets led to reduced platelet-mediated up-regulation of adhesion molecules and reduced IL-6 release of ECs. Additionally, we were able to show that the adhesion of leukocytes on inflammatory stimulated platelets was reduced upon PLD1 deficiency in vitro. Under inflammatory conditions in vivo, loss of PLD1 resulted in reduced platelet and leukocyte recruitment and arrest to the injured carotid artery. In a venous thrombosis model of the vena cava, platelet and leukocyte recruitment was significantly reduced as well. To analyse the signalling pathway that is modulated by PLD1 and responsible for impaired inflammatory responses, we performed immunoprecipitation studies and found a direct interaction of PLD1 and Src kinases suggesting that PLD1 is essential for the activation of Src. Altered phosphorylation of Src and PLCgamma2 in PLD1 deficient platelets confirmed that PLD1 plays an important role in these processes.
Conclusion: This study identifies PLD1 as a promising target to reduce platelet-mediated inflammatory processes.