title: Calcium-independent phospholipase A2 is regulated by a novel protein kinase C in human coronary artery endothelial cells.
abstract: We demonstrated previously that thrombin stimulation of endothelial cells activates a membrane-associated, Ca(2+)-independent phospholipase A2 (iPLA2) that selectively hydrolyzes arachidonylated plasmalogen phospholipids. We report that incubation of human coronary artery endothelial cells (HCAEC) with phorbol 12-myristate 13-acetate (PMA) to activate protein kinase C (PKC) resulted in hydrolysis of cellular phospholipids similar to that observed with thrombin stimulation (0.05 IU/ml; 10 min). Thrombin stimulation resulted in a decrease in arachidonylated plasmenylcholine (2.7 +/- 0.1 vs. 5.3 +/- 0.4 nmol PO4/mg of protein) and plasmenylethanolamine (7.5 +/- 1.0 vs. 12.0 +/- 0.9 nmol PO4/mg of protein). Incubation with PMA resulted in decreases in arachidonylated plasmenylcholine (3.2 +/- 0.3 nmol PO4/mg of protein) and plasmenylethanolamine (6.0 +/- 1.0 nmol PO4/mg of protein). Incubation of HCAEC with the selective iPLA2 inhibitor bromoenol lactone (5 mM; 10 min) inhibited accelerated plasmalogen phospholipid hydrolysis in response to both PMA and thrombin stimulation. Incubation of HCAEC with PMA (100 nM; 5 min) resulted in increased arachidonic acid release (7.1 +/- 0.3 vs. 1.1 +/- 0.1%) and increased production of lysoplasmenylcholine (1.4 +/- 0.2 vs. 0.6 +/- 0.1 nmol PO4/mg of protein), similar to the responses observed with thrombin stimulation. Downregulation of PKC by prolonged exposure to PMA (100 nM; 24 h) completely inhibited thrombin-stimulated increases in arachidonic acid release (7.1 +/- 0.6 to 0.5 +/- 0.1%) and lysoplasmenylcholine production (2.0 +/- 0.1 to 0.2 +/- 0.1 nmol PO4/mg of protein). These data suggest that PKC activates iPLA2 in HCAEC, leading to accelerated plasmalogen phospholipid hydrolysis and increased phospholipid metabolite production.
authors.affiliation: Department of Pathology, St. Louis University School of Medicine, St. Louis, Missouri 63104, USA.
authors.forename: Maureen C
authors.initials: MC
authors.lastname: Meyer
authors.forename: Pamela J
authors.initials: PJ
authors.lastname: Kell
authors.forename: Michael H
authors.initials: MH
authors.lastname: Creer
authors.forename: Jane
authors.initials: J
authors.lastname: McHowat
mesh_terms: D016718:Arachidonic Acid / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured; D003331:Coronary Vessels / Q000187:drug effects / Q000378:metabolism*; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D054522:Group VI Phospholipases A2; D006801:Humans; D015151:Immunoblotting; D010741:Phospholipases A / Q000187:drug effects / Q000378:metabolism*; D054467:Phospholipases A2; D010743:Phospholipids / Q000378:metabolism; D010972:Platelet Activating Factor / Q000187:drug effects / Q000378:metabolism; D016133:Polymerase Chain Reaction; D020033:Protein Isoforms / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013917:Thrombin / Q000494:pharmacology
chemical_list: D010743:Phospholipids; D010972:Platelet Activating Factor; D020033:Protein Isoforms; D016718:Arachidonic Acid; D011493:Protein Kinase C; D010741:Phospholipases A; D054522:Group VI Phospholipases A2; C516475:PLA2G6 protein, human; D054467:Phospholipases A2; D013917:Thrombin; D013755:Tetradecanoylphorbol Acetate