Functional Hierarchy of Remnant Lipoprotein Receptors

描述（由申请人提供）：粥样斑块巨噬细胞生活在一个特殊的环境中，其中胆固醇过量触发主动吞噬和炎症爆发的发散信号。在该资助的前几个周期中，我们专注于两种蛋白质，它们在细胞胆固醇运输和炎症调节方面具有合作和独立的功能，即apoE和LRP 1。ApoE是一种LXR调节蛋白，在胆固醇负荷条件下在巨噬细胞中高度表达，具有驱动胆固醇从细胞流出的能力，并具有直接的抗动脉粥样硬化作用。然而，apoE也可以与致动脉粥样硬化的脂蛋白结合，然后与LDLR和LRP 1等内化受体结合，从而将胆固醇驱回细胞。LRP 1是多种配体的受体，在肝脏中作为LDLR清除残余脂蛋白的后备系统。肝脏LRP 1需要局部产生的apoE来清除已经富含血浆来源的apoE的进入残留物，而LDLR即使在缺乏局部产生的apoE的情况下也能清除含apoE的脂蛋白。与工作的假设，载脂蛋白E和LRP 1之间的相互作用比在脂蛋白配体和它的内化受体之间的关系更复杂，我们移动到实验阶段，LDLR是不是占主导地位的脂蛋白受体和相互作用可能有复杂的生物学后果，动脉粥样硬化巨噬细胞。我们证明了巨噬细胞中LRP 1的缺失增加了动脉粥样硬化形成，这是一种矛盾的效应，因为它也减少了残余脂蛋白的内化并增加了apoE的表达。这些数据表明LRP 1的作用与大量胆固醇转运无关，并且apoE的血管效应是通过与LRP 1的相互作用介导的。我们惊讶地发现后者是不正确的，因为apoE的缺失加重了LRP 1-/-巨噬细胞的致动脉粥样硬化表型，从而表明这些蛋白质具有调节血管稳态的独立和累加作用。apoE-/-/LRP 1-/-小鼠的斑块显示出独特的严重的巨噬细胞凋亡模式，凋亡小体被活吞噬细胞内化的缺陷。注意到LRP 1对诱导炎症反应的独特作用，因为携带LRP 1-/-巨噬细胞的小鼠显示出显著更高数量的循环和脾促炎Ly 6chi单核细胞以及动脉壁Ly 6chi和CCR 2阳性巨噬细胞。LRP 1的另一个独特作用是对鞘脂激活蛋白原运输的调节。鞘脂激活蛋白原导致鞘脂激活蛋白（鞘脂激活蛋白）的形成，这对溶酶体和膜鞘脂的加工和清除至关重要。该途径与自噬机制交织在一起，最近显示自噬机制调节泡沫细胞的胆固醇流出，并与炎症反应交织在一起。拟议的研究旨在表征负责由LRP 1缺失引起的夸大的细胞死亡，炎症和细胞碎片清除功能障碍的分子途径。我们努力了解调节大块斑块消退的因素。

DESCRIPTION (provided by applicant): The atheroma macrophage lives in a peculiar environment, where cholesterol excesses trigger diverging signals of active phagocytosis and inflammatory bursts. During the previous cycles of this grant, we have focused on two proteins that have both cooperative and independent functions on cellular cholesterol trafficking and regulation of inflammation, apoE and LRP1. ApoE is an LXR-regulated protein, highly expressed in macrophages under conditions of cholesterol loading, with the ability to drive cholesterol efflux from the cell and with direct anti-atherogenic effects. However, apoE can also associate with atherogenic lipoproteins and then bind to internalizing receptors such as LDLR and LRP1, thus driving cholesterol back into the cell. LRP1, a receptor for multiple ligands, acts in the liver as a back-up system for the LDLR to clear remnant lipoproteins. Hepatic LRP1 needs locally produced apoE to clear incoming remnants already enriched in plasma-derived apoE, whereas LDLR clears apoE-containing lipoproteins even in the absence of locally produced apoE. With the working hypothesis that apoE and LRP1 interact more intricately than in a relation between lipoprotein ligand and its internalizing receptor, we moved to an experimental stage where LDLR is not the dominant lipoprotein receptor and where the interaction may have complex biological consequences, the atheroma macrophage. We demonstrated that absence of LRP1 in macrophages increases atherogenesis, a paradoxical effect given that it also reduced internalization of remnant lipoproteins and increased expression of apoE. These data suggest both that the effect of LRP1 is not linked to bulk cholesterol transport and that the vascular effect of apoE is mediated by an interaction with LRP1. We were surprised to determine that the latter is not true, as the deletion of apoE aggravated the atherogenic phenotype of LRP1-/- macrophages, thus suggesting that these proteins have independent and additive effects that regulate vascular homeostasis. The plaques of apoE-/-/LRP1-/- mice showed a uniquely severe pattern of macrophage apoptosis with a deficit in the internalization of apoptotic bodies by viable phagocytes. An effect unique to LRP1 was noted on the induction of inflammatory responses, as mice carrying LRP1-/- macrophages showed significantly higher numbers of circulating and splenic pro-inflammatory Ly6chi monocytes and arterial wall Ly6chi and CCR2 positive macrophages. Another effect unique to LRP1 was noted on the regulation of prosaposin trafficking. Prosaposin leads to the formation of the saposins (sphingolipid activator proteins), critical for processing and clearance of lysosomal and membrane sphingolipids. This pathway is intertwined with the autophagic machinery, recently shown to regulate cholesterol efflux from foam cells, and with the inflammatory response. The proposed studies aim at characterizing the molecular pathways responsible for the exaggerated cell death, inflammation, and dysfunctional clearance of cell debris caused by the absence of LRP1. We strive to understand the factors that regulate bulk plaque regression.