Group V sPLA2 in Atherosclerosis

描述（由申请方提供）：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聚糖结合而滞留在内皮下。这些保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LDL的多种生物反应，包括巨噬细胞的募集和脂质负载，导致动脉粥样硬化的开始和进展。细胞外基质（ECM）似乎在这一过程中发挥了积极的作用，不仅介导LDL颗粒的保留，但也通过调节各种酶对LDL的活性。因此，在LDL正在积聚的区域中，LDL、ECM和LDL修饰酶的共定位导致在动脉粥样硬化中达到高潮的事件的自我延续级联。分子的识别和表征以及它们相互作用以促进这一过程的方式，是我们理解病变形成易感性的机制的核心。该提案旨在研究一种新形式的分泌型磷脂酶A2，V组sPLA 2，我们现在表明，这是目前在人类和小鼠动脉粥样硬化病变，特别是与巨噬细胞。我们推测V组sPLA 2和LDL在巨噬细胞附近导致聚集和/或融合的脂蛋白颗粒的局部产生，从而导致泡沫细胞形成。具体目的1）：检验V组sPLA 2在LDL颗粒中产生修饰，导致巨噬细胞摄取增加的假设。这将通过以下方式实现：分析V组sPLA 2水解后的LDL颗粒，以确定颗粒组成、密度和聚集和/或融合程度;定量V组sPLA 2水解对LDL脂质递送至巨噬细胞的影响，并确定动脉内皮下的其他因子（即鞘磷脂酶和ECM）是否促进V组sPLA 2介导的作用。具体目的2）：检验血管壁中V组sPLA 2的巨噬细胞表达导致动脉粥样硬化增加的假设。这将通过将过表达野生型V组sPLA 2或蛋白聚糖结合缺陷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.