Oxidation of Apolipoprotein B-100 in LDL

描述(申请人提供)：有大量证据 提示低密度脂蛋白的氧化修饰 对发起和/或发展 动脉硬化。尽管大量关于低密度脂蛋白氧化的研究已经 发表后，人们对细胞的氧化修饰知之甚少。 载脂蛋白。载脂蛋白的化学多样性大于低密度脂蛋白 脂类为提高生物标记物的特异性提供了机会 区分氧化的特定机制的贡献。第一 本提案的具体目的是检验氧化作用 通过作为氧化低密度脂蛋白候选机制的方法在体外对低密度脂蛋白进行研究 在体内产生载脂蛋白(apo B-100)的修饰 足以区分为氧化低密度脂蛋白的生物标志物 在体内由各自的机制决定。在我们到目前为止的研究中，我们观察到 低密度脂蛋白氧化形成的apoB-100氧化产物的明显差异 以Cu2+、HOCl、髓过氧化物酶(MPO)为底物。我们建议调查 同样地，低密度脂蛋白在体外通过Fe2+催化的氧化、硝化 (NO和ONOO供体、MPO或嗜酸性粒细胞过氧化物酶[PO]+NO2-)；以及 培养细胞介导的氧化，包括内皮细胞、血管 平滑肌细胞和单核细胞。这些研究将采用隔离方法 基于高效液相色谱和结构分析 基于质谱学的表征方法。第二个具体目标是 这项提议是为了检验循环中的低密度脂蛋白亚组分 从某些个体分离出的apoB-100具有特定的修饰 将反映氧化产物的有限子集的氧化 低密度脂蛋白在体外具有特异性1.本研究的第三个特异性目的 建议检验从动脉粥样硬化材料中分离出的低密度脂蛋白的假设 将展示载脂蛋白氧化产物的有限子集 具有特定目标的特点1.我们到目前为止的结果提供了非常强劲的 支持我们作为本应用程序基础的工作假设。这个 提出的研究的目标是开发一种方法来区分 低密度脂蛋白氧化的特定机制被怀疑有助于 体内低密度脂蛋白的氧化及其与动脉粥样硬化的关系。这样做的长期目标是 研究是为了创造更具机械性的特异性治疗的基础 努力延缓动脉粥样硬化的发生和发展，并 开发这些不同氧化机制的生物标记物来监测 这些治疗干预措施的效果。最后，我们提出的研究 不仅与动脉粥样硬化有关，甚至与低密度脂蛋白的氧化有关， 但在这些研究中提出的概念和方法很容易应用 与一系列重要的人类疾病有关，而未指明的氧化 建议通过流程来做出贡献。

DESCRIPTION (provided by the applicant): There is a large body of evidence suggesting that oxidative modifications of low-density lipoproteins (LDL) contribute significantly to the initiation and/or progression of atherosclerosis. Although numerous studies of the oxidation of LDL lipids have been published, far less is known about the oxidative modification of the apoprotein. The greater chemical diversity of the apoprotein than of the LDL lipids offers the opportunity for greater biomarker specificity for distinguishing contributions of specific mechanisms of oxidation. The first Specific Aim of the present proposal is to test the hypothesis that oxidations of LDL in vitro by methods that are candidate mechanisms for oxidation of LDL in vivo produce modifications of the apoprotein (apo B-100) that are sufficiently distinguishable to be empIoyed as biomarkers for oxidation of LDL in vivo by the respective mechanisms. In our studies to date we have observed clear differences in products of apoB-1 00 oxidation formed in oxidation of LDL in vitro by Cu2+, HOCl, and myeloperoxidase (MPO). We propose to investigate similarly the oxidation of LDL in vitro by Fe2+-catalyzed oxidation, nitration (NO and ONOOdonors, MPO or eosinophil peroxidase aboutPO] plus NO2-), and oxidation mediated by cultured cells, including endothelial cells, vascular smooth muscle cells and monocytes. These studies will employ isolation methods based on high performance liquid chromatography (HPLC) and structural characterization methods based on mass spectrometry. The second Specific Aim of this proposal is to test the hypothesis that subfractions of circulating LDL isolated from some individuals will exhibit specific modifications apoB-1 00 oxidation that will reflect a limited subset of the products of oxidation of LDL in vitro characterized in Specific Aim 1. The third Specific Aim of this proposal is to test the hypothesis that LDL isolated from atheromatous material will exhibit a limited subset of the products of apoprotein oxidation that were characterized in Specific Aim i. Our results to date provide very strong support for the working hypotheses upon which we base this application. The goal of the studies proposed is to develop a means of distinguishing the specific mechanisms of LDL oxidation that are suspected to contribute to the oxidation of LDL in vivo and to atherogenesis. The longer-term goals of this research are to create the basis for more mechanistically specific therapeutic efforts to retard the initiation and progression of atherosclerosis and to develop biomarkers of these distinct mechanisms of oxidation to monitor efficacies of these therapeutic interventions. Finally, the studies we propose are not limited in relevance to atherosclerosis or even to oxidation of LDL, but the concepts and methods developed in these studies are readily applicable to a wide range of important human diseases for which unspecified oxidative processes are proposed to contribute.