Macrophage Expression of APOAI and Atherosclerosis

性状（由申请方提供）：载脂蛋白（apo）Al是血浆HDL的主要结构组分。HDL和apoAl的血浆水平与CHD率呈负相关，并且它们的操纵是动脉粥样硬化的治疗干预。ApoAl是胆固醇逆向转运（RCT）的驱动因子，所述胆固醇逆向转运是将过量的外周胆固醇递送至肝细胞的途径。RCT促进血管健康的机制尚不清楚，存在apoAl和HDL在动脉壁中发挥直接保护作用的可能性。在这种情况下，高密度脂蛋白水平可能只是与动脉壁中高密度脂蛋白浓度有关，这可能是斑块稳定的真正效应。在此授权的前一个周期中，我们已经表明，动脉巨噬细胞的apoAl的独家生产导致高浓度的apoAl在斑块中，血浆中apoAl的量最小，血浆HDL没有变化。这种干预导致在具有不同血浆HDL水平的小鼠模型中完全预防动脉粥样硬化，范围从几乎没有到正常的两倍。我们还表明，动脉表达apoAl显着减少apoE缺乏诱导的动脉粥样硬化形成，表明巨噬细胞apoAl可以对抗沉重的脂质负担。我们已经通过转基因、巨噬细胞特异性apoAl表达和使用apoAl表达逆转录病毒载体的骨髓细胞转导的组合方法实现了这一点。我们现在提出这些研究路线的自然进展，旨在进行机械发现，技术创新和临床可翻译性。我们将开发表达由生理性ME 2启动子驱动的apoAl构建体的慢病毒载体，所述启动子通过生理性LXR环对胆固醇加载敏感。在该系统中，apoAl的巨噬细胞特异性表达将在巨噬细胞转化为泡沫细胞期间上调。我们还将产生表达由相同胆固醇敏感启动子驱动的apoAl的转基因小鼠。另一项技术进步将是使用纯化的造血干细胞（HSC）和亚致死剂量的辐射。使用这些工具，我们将评估人apoAl和单个氨基酸变体对体内和体外血管脂质积聚和炎症的影响。已知天然apoAl变体降低HDL水平，对动脉粥样硬化形成具有不同的作用。待检验的主要假设是：1）ApoAl的血管效应可以与血浆HDL水平分离; 2）ApoAl对巨噬细胞胆固醇稳态、炎症和氧化具有直接影响。这些研究有可能揭示HDL对动脉斑块的治疗作用的基础。

DESCRIPTION (provided by applicant): Apolipoprotein (apo) Al is the main structural component of plasma HDL. Plasma levels of HDL and apoAl are inversely related to CHD rates, and their manipulation is a therapeutic intervention for atherosclerosis. ApoAl is the driver of reverse cholesterol transport (RCT), the pathway through which excess peripheral cholesterol is delivered to the hepatocyte. The mechanism through which RCT promotes vascular health is not understood, and the possibility exists that apoAl and HDL exert direct protective effects in the artery wall. In this light, higher HDL levels might simply be related to higher concentrations of HDL in the artery wall, which may be the true effector of plaque stabilization. In the previous cycle of this grant we have shown that the exclusive production of apoAl from arterial macrophages leads to high concentrations of apoAl in the plaque, with minimal amounts of apoAl in plasma and no changes in plasma HDL. This intervention resulted in the complete prevention of atherosclerosis in mouse models with different levels of plasma HDL, ranging from almost none to twice normal. We have also shown that arterial expression of apoAl significantly reduces the atherogenesis induced by apoE deficiency, suggesting that macrophage apoAl can counter heavy lipid burden. We have accomplished this through a combined approach of transgenic, macrophage-specific apoAl expression, and bone marrow cell transduction using apoAI-expressing retroviral vectors. We are now proposing the natural progression of these investigative lines, aiming for mechanistic discoveries, technical innovation, and clinical translatability. We will develop lentiviral vectors expressing apoAl constructs driven by the physiologic ME2 promoter, which is sensitive to cholesterol loading through the physiologic LXR loop. In this system, macrophage-specific expression of apoAl will be up-regulated during the transformation of the macrophage into a foam cell. We will also produce transgenic mice that express apoAl driven by the same cholesterol-sensitive promoter. Another technological advance will be the use of purified hematopoietic stem cells (HSC) and sub-lethal doses of radiation. Using these tools, we will evaluate the effects of human apoAl and single amino-acid variants on vascular lipid accumulation and inflammation in vivo and in vitro. The natural apoAl variants are known to lower HDL levels with diverging effects on atherogenesis. The main hypotheses to be tested are: 1) The vascular effects of apoAl can be dissociated from plasma HDL levels; 2) ApoAl has direct effects on macrophage cholesterol homeostasis, inflammation, and oxidation. These studies have the potential to uncover the basis for the therapeutic effect of HDL on the arterial plaque.