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Endothelial cell PAF synthesis following thrombin stimulation utilizes Ca(2+)-independent phospholipase A(2)
J McHowat1, P J Kell, H B O'Neill
1Department of Pathology, St. Louis University School of Medicine, 1402 South Grand Boulevard, St. Louis, Missouri 63104, USA. Mchowatj@slucare1.sluh.edu
Biochemistry
|December 6, 2001
Summary
Platelet activating factor (PAF) synthesis in human umbilical artery endothelial cells (HUAEC) requires calcium-independent phospholipase A2 (iPLA2) activation. This study shows iPLA2 drives PAF production via plasmalogen hydrolysis, not direct PLA2 action.
Area of Science:
- Biochemistry
- Cell Biology
- Lipid Metabolism
Background:
- Platelet activating factor (PAF) is a potent lipid mediator synthesized by endothelial cells upon stimulation.
- PAF production pathways include de novo synthesis, direct phospholipase A2 (PLA2) action, and remodeling pathways.
Purpose of the Study:
- To investigate the role of PLA2 and plasmalogen phospholipid hydrolysis in PAF synthesis in thrombin-stimulated human umbilical artery endothelial cells (HUAEC).
Main Methods:
- Assessed PLA2 activity (Ca2+-dependent and independent) in HUAEC.
- Analyzed phospholipid hydrolysis and lipid mediator release (arachidonic acid, prostacyclin, PAF) following thrombin stimulation.
- Utilized bromoenol lactone to inhibit iPLA2 activity.
Main Results:
- Thrombin stimulation preferentially increased membrane-associated, Ca2+-independent PLA2 (iPLA2) activity, particularly on plasmenylcholine substrates.
- iPLA2 activation correlated with increased arachidonic acid and prostacyclin release, and elevated levels of lysoplasmenylcholine, lysophosphatidylcholine, and PAF.
- Inhibition of iPLA2 blocked plasmalogen hydrolysis and PAF synthesis.
Conclusions:
- PAF production in thrombin-stimulated HUAEC necessitates iPLA2 activation.
- The findings suggest PAF synthesis occurs via the CoA-independent transacylase remodeling pathway, rather than direct PLA2 hydrolysis of alkylacyl glycerophosphocholine.