Selective hydrolysis of plasmalogens in endothelial cells following thrombin stimulation

The present study was performed to characterize thrombin-stimulated phospho lipase A(2) (PLA(2)) activity and the resultant release of lysophospholipid s from endothelial cells. The majority of PLA(2) activity in endothelial ce lls was membrane associated, Ca2+ independent, and arachidonate selective. Incubation with thrombin increased membrane-associated PLA(2) activity usin g both plasmenylcholine and alkylacyl glycerophosphocholine substrates in t he absence of Ca2+, with no increase in activity observed with phosphatidyl choline substrate. The increased PLA(2) activity was accompanied by arachid onic acid and lysoplasmenylcholine (LPlasC) release from endothelial cells into the surrounding medium. Thrombin-induced changes were duplicated by st imulation with the thrombin-receptor-directed peptide SFLLRNPNDKYEPF. Pretr eatment with the Ca2+-independent PLA(2) inhibitor bromoenol lactone blocke d thrombin-stimulated increases in PLA(2) activity, arachidonic acid, and L PlasC release. Stimulation of protein kinase C (PKC) increased basal PLA(2) activity and LPlasC production. Thrombin-stimulated PLA(2) activity and LP lasC production were enhanced with PKC activation and completely prevented with PKC downregulation. Thus thrombin treatment of endothelial cells activ ates a PKC-activated, membrane-associated, Ca2+-independent PLA(2) that sel ectively hydrolyzes arachidonylated, ether-linked phospholipid substrates, resulting in LPlasC and arachidonic acid release.