Group V sPLA2 in Atherosclerosis

描述(由申请人提供)：早期动脉粥样硬化形成的一个关键事件是低密度脂蛋白(LDL)颗粒通过与内膜蛋白多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中的几种修饰酶，包括脂肪酶、氧化酶和蛋白酶。对这种修饰的低密度脂蛋白的一系列生物学反应，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了积极的作用，它不仅介导了低密度脂蛋白颗粒的滞留，而且还调节了各种酶对低密度脂蛋白的活性。因此，在低密度脂蛋白积聚的区域，低密度脂蛋白、细胞外基质和低密度脂蛋白修饰酶的共同定位导致了一系列自我延续的事件，最终导致动脉粥样硬化。分子的识别和表征以及它们相互作用促进这一过程的方式，对于我们理解病变形成的易感性机制是至关重要的。这项建议旨在研究一种新的分泌型磷脂酶A2，V组sPLA2，我们现在证明它存在于人类和小鼠动脉粥样硬化病变中，与巨噬细胞特异性相关。我们推测，巨噬细胞附近的V组sPLA2和低密度脂蛋白导致聚集和/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，提出了以下特定目标：特定目标1)：测试V组sPLA2在低密度脂蛋白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水解后的低密度脂蛋白颗粒来确定颗粒组成、密度以及聚集和/或融合的程度；量化V组sPLA2水解对向巨噬细胞输送低密度脂蛋白的影响，并确定动脉内皮下的其他因素，即鞘磷脂酶和细胞外基质是否促进V组sPLA2介导的效应。具体目的2)：验证V组sPLA2在血管壁巨噬细胞表达导致动脉粥样硬化增加的假说。这将通过将过表达野生型V组sPLA2或缺乏蛋白多糖结合的酶的突变形式的胎儿肝脏造血干细胞移植到低密度脂蛋白受体/低密度脂蛋白受体-/-小鼠中来实现。测量病变大小和胆固醇/胆固醇酯含量将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.