Besides the fibrinolytic system, leukocyte recruitment and angiogenesis are two major determinants of thrombus resolution. Previous studies in a murine model of stagnant flow venous thrombosis have shown that splenectomy effectively delays thrombus resolution, and is associated with accumulation of anionic phospholipids. Here we propose that thrombus angiogenesis is inhibited in the presence of anionic, procoagulant phospholipids, e.g. phosphatidylserine (PS), as compared to neutral phospholipids such as phosphatidylcholine (PC) or phosphatidylethanolamine (PE).Methods
Phospholipid vesicles were prepared, containing (1) 100% PC, (2) 60% PC / 40% PS, (3) 60% PC / 40% PE, (4) 60% PC / 20% PS / 20% PE. Biological activity of the mixtures was confirmed by thrombin generation assay. For analysing effects on endothelial cells, a BrdU cell proliferation assay and a lactate dehydrogenase cytotoxicity assay were performed with human umbilical vein endothelial cells (HUVEC).Results
In the BrdU cell proliferation assay, exposure of HUVEC towards 50 μM or 100 μM phospholipids containing PS for 24 h and 48 h inhibited DNA synthesis rates significantly, whereas neutral phospholipids had hardly any effect. The lactate dehydrogenase cytotoxicity assay confirmed a lack of cytotoxicity in all cases.Conclusions
Together with their procoagulant function, anionic phospholipids such as phosphatidylserine attenuate proliferation of endothelial cells and may thus reduce or inhibit thrombus-associated angiogenesis, a factor relevant for thrombus resolution.