Phospholipids released from activated platelets improve platelet aggregation and endothelial cell migration

This study was undertaken to isolate phospholipids released from activated platelets and to investigate their biological activities. Freshly washed pl atelets were activated with freezing/thawing, thrombin, ionophore 23187, an d arachidonic acid. Thrombin was incubated with platelet-rich plasma to pro mote synthesis and release of phospholipids from platelets. Phospholipids i n supernatants of activated platelets were extracted with butanol and separ ated by thin-layer chromatography. Release of phosphatidylserine (PS) and p hosphatidic acid (PA) increased when platelets were treated with freezing/t hawing, ionophore, and thrombin. The lysophosphatidyl ethanolamine (LPE) ap peared not to be induced with freezing/thawing, but increased significantly by thrombin, ionophore, and arachidonic acid. The effects of platelet phos pholipids on hemostasis and angiogenesis were studied with platelet aggrega tion and endothelium chemotaxis. Phospholipids isolated from thrombin-stimu lated platelet-rich/platelet-poor plasmas were used as synergistic agonists in platelet aggregation and as chemotactic agents in endothelial cell migr ation. Several phospholipids increased chemotaxis and platelet aggregation; these were PS, PA, LPE, and sphingosine-1-phosphate. Also, chemotaxis of t hose phospholipids increased when combined with charcoal-stripped fetal bov ine serum, suggesting that cofactors in serum enhanced phospholipid-induced cell migration. These observations suggest that activated platelets releas e biologically active phospholipids into the blood stream, where they may p lay an important role in thrombosis and angiogenesis.