Atherogenic Effects of Ambient PM In Susceptible Animals

描述(申请人提供)：本申请的目标是邀请一个由领先的环境、心血管和蛋白质组学研究人员组成的多学科团队，研究环境颗粒(PM)通过哪些途径促进载脂蛋白(ApoE)和低密度脂蛋白受体(LDL-R)缺陷小鼠的动脉粥样硬化形成。主要假设是PM诱导的氧化应激与氧化脂质成分协同作用，增强动脉粥样硬化病变中的炎症和细胞凋亡。有三个具体目标。目标1将确定环境PM暴露是否会加剧这些遗传易感菌株的动脉粥样硬化。动物将暴露在洛杉矶盆地粒子浓缩器收集的环境超细(空气动力学直径&lt；0.1微米)和细(&lt；2.5微米)颗粒物中。终点将是对早期和晚期动脉粥样硬化病变发展的定量评估。这些研究将得到体内和体外研究的补充，这些研究着眼于浓缩环境颗粒物(CAPS)对血管内皮细胞、平滑肌细胞和吞噬细胞的促氧化、促炎和促凋亡作用。目的2将确定PM诱导的氧化应激在炎症和细胞凋亡中的作用。暴露于CAPS的动物的主动脉血管组织将被检查脂质过氧化和血红素加氧酶1(HO-1)的表达、MAP激酶激活和细胞凋亡。血液将用于研究CAPS对急性时相蛋白、纤维蛋白原和氧化修饰的保护性高密度脂蛋白活性的影响。转基因HO-1启动子-荧光素酶小鼠将用于体内成像氧化应激产生的动力学和血管部位。体外研究将包括对内皮细胞氧化应激反应的蛋白质组分析。所有的氧化应激效应都将与CAPS的多环芳烃和苯二酚含量以及它们的体外氧化还原循环能力相关，由DTT分析确定。目的3通过与HO-1转基因和对氧磷酶基因敲除动物杂交，确定修饰的抗氧化防御机制是否影响CAPS对apoE基因缺陷小鼠动脉粥样硬化病变的诱导作用。研究人员还将确定调节抗氧化酶表达的NRF-2转录因子的缺失是否会影响载脂蛋白E t/-动物的病变发展。这些小鼠将暴露于AIMS 1和AIMS 2中讨论的CAPS，并选择用于展示其对动脉粥样硬化性炎症和细胞凋亡的影响的终点。

DESCRIPTION (provided by applicant): The goal of this application is to engage a multidisciplinary team of leading environmental, cardiovascular and proteomics researchers in the study of the pro-oxidative and pro-inflammatory pathways by which ambient pariculate matter (PM) enhance atherogenesis in apolipoprotein (apoE) and LDL receptor (LDL-R) deficient mice. The principal hypothesis is that PM-induced oxidative stress synergizes with oxidized lipid components to enhance inflammation and apoptosis in atherosclerotic lesions. There are three specific aims. Aim 1 will determine whether ambient PM exposures exacerbate atherosclerosis in these genetically susceptible strains. Animals will be exposed to ambient ultrafine (aerodynamic diameter < 0.1 mu m) and fine (< 2.5 mu m) particulates collected by particle concentrators in the Los Angeles basin. The endpoints will be a quantitative assessment of early and late atherosclerotic lesion development. These studies will be supplemented by in vivo and in vitro studies looking at the pro-oxidative, pro-inflammatory, and pro-apoptotic effects of concentrated ambient particulates (CAPs) on vascular endothelial, smooth muscle and phagocytic cells. Aim 2 will determine the role of PM-induced oxidative stress in inflammation and apoptosis. Aortic vascular tissue from CAPs exposed animals will be examined for lipid peroxidation and for heme oxygenase 1 (HO-1) expression, MAP kinase activation, and apoptosis. Blood will be used to study CAPs effects on acute phase proteins, fibrinogen and oxidative modification of protective HDL activity. Transgenic HO-1 promoter-luciferase mice will be used for in vivo imaging of the kinetics and vascular sites of oxidative stress generation. In vitro studies will include proteome analysis of the oxidative stress response in endothelial cells. All oxidative stress effects will be correlated with the PAH and quinone content of the CAPs, as well as their in vitro redox cycle capacity, determined by the DTT assay. Aim 3 will determine whether modified antioxidant defense mechanisms affect the induction of atherosclerotic lesions by CAPs in apoE deficient mice, which have been crossed with HO-1 transgenic as well as paraoxonase knockout animals. The investigators will also determine whether a deficiency of the Nrf-2 transcription factor, which regulates antioxidant enzyme expression, will affect lesion development in apoE t/- animals. These mice will be exposed to CAPs as discussed in Aims 1 and 2, and select endpoints used to demonstrate the effect on atherosclerotic inflammation and apoptosis.