Platelet Lipidomic Profiling: Novel Insight into Cytosolic Phospholipase A2α Activity and Its Role in Human Platelet Activation
Platelet Lipidomic Profiling: Novel Insight into Cytosolic Phospholipase A2α Activity and Its Role in Human Platelet Activation
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DOI：
10.1021/acs.biochem.5b00549
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发表时间：
2015-09-15
期刊：
影响因子：
2.9
通讯作者：
Hamm, Heidi E.
中科院分区：
文献类型：
--
作者：
Duvernay, Matthew T.;Matafonov, Anton;Hamm, Heidi E.
随着一种新的、更有选择性和更有效的胞质磷脂酶A（2）α（cPLA（2）α）抑制剂的出现，我们重新审视了cPLA（2）α活性在血小板活化中的作用，发现血小板信号传导的一个组成部分，甚至比以前认识到的更大，依赖于这种酶。在一项基于全血剪切的流动室试验中，giripladib（一种cPLA（2）α抑制剂）可减少血小板在胶原蛋白上的粘附和积聚。此外，giripladib差异影响P-选择素的表达和GPIIbIIIa激活取决于所采用的激动剂。虽然蛋白酶激活受体1（PAR 1）介导的血小板活化不受giripladib的影响，但PAR 4和GPVI介导的血小板活化水平显著降低。同时，血栓素A（2）受体拮抗剂SQ 29548对PAR、GPVI或嘌呤能受体介导的血小板活化没有影响，这表明cPLA（2）α在花生四烯酸释放下游产生的另一种类花生酸是PAR 4和GPVI介导的血小板活化的主要成分。与此同时，我们分析了PAR介导的甘油磷脂（GPL）质量的变化，有和没有giripladib，以更好地了解cPLA（2）α介导的脂质代谢。磷脂酰胆碱和磷脂酰乙醇胺（PE）表现出最大的消费量在凝血酶刺激。此外，我们确认磷脂酰肌醇是cPLA（2）α的主要底物。PAR 1和PAR 4诱导的代谢的比较显示，PAR 1活化下游消耗更多假定的花生四烯酸-PE物质。这些结果表明，cPLA（2）α活性在血小板功能中起着重要作用，并表明血小板活化产生了一种新型类花生酸，而这种类花生酸是PAR 4和GPVI介导的反应的一个重要组成部分。
With a newer, more selective and efficacious cytosolic phospholipase A(2)alpha (cPLA(2)alpha) inhibitor available, we revisited the role of cPLA(2)alpha 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 cPLA(2)alpha inhibitor, reduced platelet adhesion and accumulation on collagen. Moreover, giripladib differentially affected P-selectin expression and GPIIbIlla 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 A(2) 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 cPLA(2)alpha 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 cPLA(2)alpha-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 cPLA(2)alpha. A comparison of PAR1- and PAR4-induced metabolism revealed the consumption of more putative arachidonyl-PE species downstream of PAR1 activation. Instead of enhanced cPLA(2)alpha activity and therefore more arachidonic acid liberation downstream of PAR4, these results indicate the major role that cPLA(2)alpha 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.