Selective hydrolysis of plasmalogens in endothelial cells following thrombin stimulation

The present study was performed to characterize thrombin-stimulated phospholipase A(2) (PLA(2)) activity and the resultant release of lysophospholipids from endothelial cells. The majority of PLA(2) activity in endothelial cells was membrane associated, Ca2+ independent, and arachidonate selective. Incubation with thrombin increased membrane-associated PLA(2) activity using both plasmenylcholine and alkylacyl glycerophosphocholine substrates in the absence of Ca2+, with no increase in activity observed with phosphatidylcholine substrate. The increased PLA(2) activity was accompanied by arachidonic acid and lysoplasmenylcholine (LPlasC) release from endothelial cells into the surrounding medium. Thrombin-induced changes were duplicated by stimulation with the thrombin-receptor-directed peptide SFLLRNPNDKYEPF. Pretreatment with the Ca2+-independent PLA(2) inhibitor bromoenol lactone blocked thrombin-stimulated increases in PLA(2) activity, arachidonic acid, and LPlasC release. Stimulation of protein kinase C (PKC) increased basal PLA(2) activity and LPlasC production. Thrombin-stimulated PLA(2) activity and LPlasC production were enhanced with PKC activation and completely prevented with PKC downregulation. Thus thrombin treatment of endothelial cells activates a PKC-activated, membrane-associated, Ca2+-independent PLA(2) that selectively hydrolyzes arachidonylated, ether-linked phospholipid substrates, resulting in LPlasC and arachidonic acid release.