Group V sPLA2 in Atherosclerosis

描述（由申请人提供）：早期动脉粥样硬化的一个关键事件是低密度脂蛋白（LDL）颗粒通过与内膜蛋白聚糖结合而滞留在内皮下层。这些滞留的脂蛋白颗粒暴露于动脉壁上的几种修饰酶，包括脂肪酶、氧化酶和蛋白酶。对这种修饰LDL的多种生物学反应，包括巨噬细胞的募集和脂质负荷，导致动脉粥样硬化的发生和发展。细胞外基质（ECM）似乎在这一过程中发挥积极作用，不仅通过介导LDL颗粒的滞留，而且通过调节各种酶对LDL的活性。因此，在LDL积聚的区域，LDL、ECM和LDL修饰酶的共定位导致一系列自我延续的事件，最终导致动脉粥样硬化。分子的鉴定和表征及其相互作用促进这一过程的方式，是我们理解病变形成易感性机制的核心。本研究旨在研究一种新型的分泌型磷脂酶A2， V组sPLA2，我们现在发现它存在于人类和小鼠与巨噬细胞特异性相关的动脉粥样硬化病变中。我们假设，在巨噬细胞附近的V组spla2和LDL导致聚集和/或融合的脂蛋白颗粒的局部产生，从而导致泡沫细胞的形成。为了验证这一假设，提出了以下具体目的：具体目的1)：验证V组sPLA2在LDL颗粒中产生修饰导致巨噬细胞摄取增加的假设。这将通过分析V组sPLA2水解后的LDL颗粒来完成，以确定颗粒组成、密度以及聚集和/或融合的程度；量化V组sPLA2水解对LDL脂质向巨噬细胞传递的影响，并确定动脉内皮下的其他因素，即鞘磷脂酶和ECM是否促进了V组sPLA2介导的作用。特异性目的2)：验证巨噬细胞在血管壁表达V组sPLA2导致动脉粥样硬化增加的假设。这将通过将过度表达野生型V组sPLA2或缺乏蛋白聚糖结合的酶的突变形式的胎肝造血干细胞移植到LDL受体-/-小鼠中来实现。测量病变大小和胆固醇/胆固醇酯含量可以评估动脉粥样硬化的程度。

DESCRIPTION (provided by applicant): A critical event in early atherogenesis is the retention of low-density lipoprotein (LDL) particles in the subendothelium through their binding to intimal proteoglycans. These retained lipoprotein particles are exposed to several modifying enzymes in the arterial wall, including lipases, oxidizing enzymes, and proteases. A multitude of biological responses to such modified LDL, including the recruitment and lipid loading of macrophages, leads to the initiation and progression of atherosclerosis. The extracellular matrix (ECM) appears to play an active role in this process by not only mediating the retention of LDL particles, but by also modulating the activity of various enzymes towards LDL. Thus, in regions where LDL is being accumulated, the co-localization of LDL, ECM, and LDL modifying enzymes leads to a self-perpetuating cascade of events that culminates in atherosclerosis. The identification and characterization of molecules and the manner in which they interact to contribute to this process, is central to our understanding the mechanisms that underlie susceptibility to lesion formation. This proposal aims to study a novel form of secretory phospholipase A2, Group V sPLA2, which we now show is present in human and mouse atherosclerotic lesions specifically associated with macrophages. We hypothesize that Group V sPLA2and LDL in the proximity of macrophages leads to the localized production of aggregated and/or fused lipoprotein particles, which consequently leads to foam cell formation. To test this hypothesis, the following specific aims are proposed: Specific Aim 1): To test the hypothesis that Group V sPLA2 produces modifications in LDL particles that lead to increased uptake by macrophages. This will be accomplished by analyzing LDL particles after hydrolysis by Group V sPLA2 to determine particle composition, density, and extent of aggregation and/or fusion; quantifying the effect of Group V sPLA2 hydrolysis on the delivery of LDL lipid to macrophages, and determining whether other factors in the arterial subendothelium, namely sphingomyelinase and ECM, promote Group V sPLA2-mediated effects. Specific Aim 2): To test the hypothesis that macrophage expression of Group V sPLA2 in the vessel wall results in increased atherosclerosis. This will be achieved by transplanting fetal liver hematopoetic stem cells over-expressing wild-type Group V sPLA2, or a mutant form of the enzyme that is deficient in proteglycan binding, into LDL receptor-/- mice. Measuring lesion size and cholesterol/cholesterol ester content will assess the extent of atherosclerosis.