title: Platelet Lipidomic Profiling: Novel Insight into Cytosolic Phospholipase A2α Activity and Its Role in Human Platelet Activation.
abstract: With a newer, more selective and efficacious cytosolic phospholipase A2α (cPLA2α) inhibitor available, we revisited the role of cPLA2α activity in platelet activation and discovered that a component of platelet signaling, even larger than previously appreciated, relies on this enzyme. In a whole blood shear-based flow chamber assay, giripladib, a cPLA2α inhibitor, reduced platelet adhesion and accumulation on collagen. Moreover, giripladib differentially affected P-selectin expression and GPIIbIIIa activation depending on the agonist employed. While protease-activated receptor 1 (PAR1)-mediated platelet activation was unaffected by giripladib, the levels of PAR4- and GPVI-mediated platelet activation were significantly reduced. Meanwhile, the thromboxane A2 receptor antagonist SQ29548 had no effect on PAR-, GPVI-, or puriniergic receptor-mediated platelet activation, suggesting that another eicosanoid produced downstream of arachidonic acid liberation by cPLA2α was responsible for this large component of PAR4- and GPVI-mediated platelet activation. In parallel, we profiled PAR-mediated changes in glycerophospholipid (GPL) mass with and without giripladib to better understand cPLA2α-mediated lipid metabolism. Phosphatidylcholine and phosphatidylethanolamine (PE) demonstrated the largest consumption of mass during thrombin stimulation. Additionally, we confirm phosphatidylinositol as a major substrate of cPLA2α. A comparison of PAR1- and PAR4-induced metabolism revealed the consumption of more putative arachidonyl-PE species downstream of PAR1 activation. Instead of enhanced cPLA2α activity and therefore more arachidonic acid liberation downstream of PAR4, these results indicate the major role that cPLA2α activity plays in platelet function and suggest that a novel eicosanoid is produced in response to platelet activation that represents a large component of PAR4- and GPVI-mediated responses.
authors: Department of Pharmacology, Vanderbilt University , Nashville, Tennessee 37232, United States.
authors: Matthew T
authors: MT
authors: Duvernay
authors: Hematology/Oncology, Vanderbilt University , Nashville, Tennessee 37232, United States.
authors: Anton
authors: A
authors: Matafonov
authors: Center for Neuroscience Drug Discovery, Vanderbilt University , Nashville, Tennessee 37232, United States.
authors: Craig W
authors: CW
authors: Lindsley
authors: Department of Pharmacology, Vanderbilt University , Nashville, Tennessee 37232, United States.
authors: Heidi E
authors: HE
authors: Hamm
mesh_terms: D001565:Benzoates / Q000494:pharmacology; D001792:Blood Platelets / Q000737:chemistry / Q000187:drug effects / Q000201:enzymology*; D020404:Glycerophospholipids / Q000097:blood; D054513:Group IV Phospholipases A2 / Q000037:antagonists & inhibitors / Q000097:blood*; D006801:Humans; D008055:Lipids / Q000097:blood*; D009842:Oligopeptides / Q000494:pharmacology; D010446:Peptide Fragments / Q000494:pharmacology; D015539:Platelet Activation; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism; D044463:Receptor, PAR-1 / Q000378:metabolism; D018179:Receptors, Thrombin / Q000097:blood; D021241:Spectrometry, Mass, Electrospray Ionization; D013314:Stress, Mechanical; D013449:Sulfonamides / Q000494:pharmacology; D013917:Thrombin / Q000494:pharmacology
chemical_list: D001565:Benzoates; D020404:Glycerophospholipids; D008055:Lipids; D009842:Oligopeptides; C586663:PLA-695; D010446:Peptide Fragments; D010980:Platelet Membrane Glycoproteins; D044463:Receptor, PAR-1; D018179:Receptors, Thrombin; D013449:Sulfonamides; C455901:alanyl-tyrosyl-prolyl-glycyl-lysyl-phenylalanine; C098341:platelet membrane glycoprotein VI; C082835:thrombin receptor peptide (42-47); D054513:Group IV Phospholipases A2; D013917:Thrombin; C112908:protease-activated receptor 4