Inhibition of platelet-activating factor (PAF) acetylhydrolase by methyl arachidonyl fluorophosphonate potentiates PAF synthesis in thrombin-stimulated human coronary artery endothelial cells.
We have previously demonstrated that thrombin stimulation of endothelial cells results in increased membrane-associated, Ca(2+)-independent phospholipase A2 (iPLA2) activity, accelerated hydrolysis of membrane plasmalogen phospholipids, and production of several biologically active phospholipid metabolites, including prostacyclin and platelet-activating factor (PAF) that is abolished by pretreatment with the iPLA2-selective inhibitor bromoenol lactone. This study was designed to further investigate the role of alternative PLA2 inhibitors, including methyl arachidonyl fluorophosphonate (MAFP, an inhibitor of cytosolic PLA2 isoforms), on phospholipid turnover and PAF production from thrombin-stimulated human coronary artery endothelial cells (HCAECs). Paradoxically, pretreatment of HCAEC with MAFP (5-25 microM) resulted in a significant increase in PAF production in both unstimulated and thrombin-stimulated cells that was found to be a direct result of inhibition of PAF acetylhydrolase (PAF-AH) activity. Pretreatment with MAFP did not significantly inhibit HCAEC PLA2 activity, possibly due to the localization of PLA2 activity in the membrane fraction rather than the cytosol. Bromoenol lactone did not inhibit PAF-AH activity, even at concentrations as high as 20 microM. We conclude that MAFP augments thrombin-stimulated PAF production by inhibition of PAF catabolism without affecting membrane-associated iPLA2 activity.
Kell
Pamela J
PJ
Department of Pathology, St. Louis University School of Medicine, 1402 S. Grand Blvd., St. Louis, MO 63104, USA.
Creer
Michael H
MH
Crown
Kimberley N
KN
Wirsig
Karin
K
McHowat
Jane
J
D043203:1-Alkyl-2-acetylglycerophosphocholine Esterase / Q000037:antagonists & inhibitors*; D001095:Arachidonic Acids / Q000494:pharmacology*; D002478:Cells, Cultured; D002794:Choline / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D003331:Coronary Vessels / Q000187:drug effects / Q000378:metabolism*; D042783:Endothelial Cells / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004983:Ethanolamines / Q000378:metabolism; D020404:Glycerophospholipids / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D063065:Organophosphonates / Q000494:pharmacology*; D010741:Phospholipases A / Q000037:antagonists & inhibitors; D054467:Phospholipases A2; D010972:Platelet Activating Factor / Q000096:biosynthesis*; D013268:Stimulation, Chemical; D013917:Thrombin / Q000494:pharmacology*
D001095:Arachidonic Acids; D004791:Enzyme Inhibitors; D004983:Ethanolamines; D020404:Glycerophospholipids; D063065:Organophosphonates; D010972:Platelet Activating Factor; C104810:methyl arachidonylfluorophosphonate; D010741:Phospholipases A; D054467:Phospholipases A2; D043203:1-Alkyl-2-acetylglycerophosphocholine Esterase; D013917:Thrombin; D002794:Choline