Role of phospholipase A2 in spinal cord secondary injury

说明（由申请人提供）：磷脂酶A2在脊髓继发性损伤中的作用损伤后脊髓损伤有两种机制：原发性机械损伤和由多种损伤机制介导的继发性损伤。迄今为止，三种损伤机制，即，炎症、氧化和兴奋性神经毒性在脊髓损伤（SCI）后被广泛研究。由于涉及多种机制，因此阻断一种特定机制不太可能显著阻止继发性SCI的进程。然而，这些不同的机制可能共享一个中心或汇聚路径来发挥其有害影响。如果是这样的话，阻断这种会聚通路应该比阻断单一通路导致更大的解剖和功能恢复。一个候选分子，可以作为收敛介质是酶磷脂酶A2（PLA 2）。PLA 2是水解磷脂的sn-2位的酯键以产生游离脂肪酸如花生四烯酸（AA）和溶血磷脂的多种酶家族。这些产品是生物活性类花生酸和血小板活化因子（PAF）的前体，它们是众所周知的炎症、氧化和细胞毒性介质。此外，PLA 2可以直接攻击细胞膜，诱导神经元和神经胶质细胞死亡。虽然PLA 2的下游产物，如AA，已被广泛研究，但令人惊讶的是，关于PLA 2本身在创伤性SCI中的作用和机制知之甚少。最近，我们首次证明，在急性挫伤性SCI后，总PLA 2的活性和胞质PLA 2（cPLA 2; PLA 2的亚型）的表达均显著增加（Liu et al.，Ann Neurol 59：606-619，2006）。值得注意的是，在SCI后30分钟给予小鼠AACOCF 3（一种cPLA 2抑制剂）显著减少了组织损伤并改善了行为恢复。在这里，我们提出了一个中心假设，PLA 2是一个汇聚分子，介导与继发性SCI相关的多种损伤途径。如果我们的假设是正确的，那么阻断PLA 2的激活应该会抑制多种损伤途径，从而促进SCI后更大的神经保护和功能恢复。由于cPLA 2是参与受体介导的AA释放的最重要的PLA 2同工酶，因此本申请将集中于cPLA 2在介导SCI中的作用和机制。因此，提出了以下三个具体目标来确定1）cPLA 2是否作为会聚分子介导自由基、兴奋性氨基酸和炎性细胞因子的细胞毒性作用，2）cPLA 2活化是否是介导继发性SCI所必需的且充分的，以及3）cPLA 2介导继发性SCI的机制，重点是线粒体功能障碍。完成这一应用可能会导致新的和有效的战略，旨在促进更大的解剖和功能恢复后SCI的发展。 公共卫生相关性：本申请将检验磷脂酶A2（PLA 2）是一种会聚分子，介导与继发性脊髓损伤（SCI）相关的多种损伤机制的中心假设。为了验证这一假设，我们提出了三个具体的目标，以确定1）cPLA 2是否作为一个收敛分子，介导自由基，兴奋性氨基酸和炎症细胞因子的细胞毒性作用，2）cPLA 2激活是否是必要的和足够的介导继发性SCI，和3）的机制，cPLA 2介导继发性SCI，重点放在线粒体功能障碍。我们希望，通过完成这项申请，我们将确定一个新的治疗干预目标，旨在促进更大的解剖和功能恢复后SCI。

DESCRIPTION (provided by applicant): Role of phospholipase A2 in spinal cord secondary injury There are two mechanisms of damage to the spinal cord after injury: a primary mechanical injury and a secondary injury mediated by multiple injury mechanisms. To date, three injury mechanisms, i.e., inflammation, oxidation and excitatory neurotoxicity, are extensively studied following spinal cord injury (SCI). Since multiple mechanisms are involved, it is unlikely that blocking one particular mechanism would significantly prevent the course of secondary SCI. However, it is possible that these different mechanisms may share a central or convergence pathway to exert their detrimental effects. If so, blocking such a convergence pathway should result in greater anatomical and functional recovery than blocking a single pathway. A candidate molecule that could serve as a convergence mediator is the enzyme phospholipase A2 (PLA2). PLA2 is a diverse family of enzymes that hydrolyze the ester bond at the sn-2 position of phospholipids to produce a free fatty acid such as arachidonic acid (AA) and a lysophospholipid. These products are precursors of bioactive eicosanoids and platelet activating factor (PAF) that are well-known mediators of inflammation, oxidation and cytotoxicity. Additionally, PLA2 can attack cell membranes directly to induce neuronal and glial death. Although the downstream products of PLA2, such as AA, have been extensively studied, to our surprise, little is known concerning the role and mechanism of the PLA2 itself in traumatic SCI. Recently, we demonstrated, for the first time, that both the activity of total PLA2 and expression of cytosolic PLA2 (cPLA2; a subtype of PLA2) increased significantly following an acute contusive SCI (Liu et al., Ann Neurol 59:606-619, 2006). Remarkably, AACOCF3, a cPLA2 inhibitor, administered at 30 min post-SCI in mice significantly reduced tissue damage and improved behavioral recovery. Here, we propose a central hypothesis that PLA2 is a convergence molecule that mediates multiple injury pathways associated with the secondary SCI. If our hypothesis is correct, blocking PLA2 activation should induce inhibition of multiple injury pathways and, therefore, promotion of greater neuroprotection and functional recovery following SCI. Since cPLA2 is the most important PLA2 isozyme implicated in receptor-mediated release of AA, this application will focus on the role and mechanisms of cPLA2 action in mediating SCI. As such, the following three specific aims are proposed to determine 1) whether cPLA2 serves as a convergence molecule mediating the cytotoxic effects of free radicals, excitatory amino acids and inflammatory cytokines, 2) whether cPLA2 activation is both necessary and sufficient to mediate secondary SCI, and 3) the mechanism by which cPLA2 mediates secondary SCI with an emphasis being placed on the mitochondria dysfunction. Completion of this application may lead to the development of novel and effective strategies aimed at promoting greater anatomical and functional recoveries after SCI. PUBLIC HEALTH RELEVANCE: This application will test a central hypothesis that phospholipase A2 (PLA2) is a convergence molecule that mediates multiple injury mechanisms associated with secondary spinal cord injury (SCI). To test this hypothesis, we have proposed three specific aims to determine 1) whether cPLA2 serves as a convergence molecule that mediates the cytotoxic effects of free radicals, excitatory amino acids and inflammatory cytokines, 2) whether cPLA2 activation is both necessary and sufficient to mediate secondary SCI, and 3) the mechanism by which cPLA2 mediates secondary SCI with an emphasis being placed on the mitochondria dysfunction. We hope that, by completion of this application, we will identify a novel target for therapeutic intervention aimed at promoting greater anatomical and functional recoveries after SCI.