Pathways of Oxidation of HDL by Macrophages

巨噬细胞氧化 HDL 的途径

• 批准号: 6969285
• 负责人: JAY W HEINECKE
• 金额: $ 39.89万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-12-01 至 2009-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6969285
• 关键词: atherosclerosis; blood lipoprotein metabolism; blood lipoprotein transport; cholesterol; clinical research; familial hyperlipoproteinemia; high density lipoproteins; high performance liquid chromatography; human subject; lipoprotein disorder; macrophage; mass spectrometry; oxidation; phagocytes; protein structure function; tyrosine

The interaction of HDL apolipoproteins, particularly apoA-I, with the cell membrane transporter ABCA1 removes excess cellular cholesterol and protects against atherogenesis. This process is mediated by lipidpoor apolipoproteins generated by either de novo synthesis or dissociation from HDL particles. Thus, factors that impair the generation or lipid efflux activity of apoA-I could have profound atherogenic effects. Oxidative damage is implicated in the pathogenesis of atherosclerosis, a chronic inflammatory disease. 3- Chlorotyrosine and 3-nitrotyrosine, stable products of protein oxidation generated by phagocyte-derived hypochlorous acid (HOCI) and reactive nitrogen species (RNS), have been detected in human atherosclerotic lesions. However, the underlying factors that control tyrosine oxidation in proteins remain poorly understood. We found that oxidation of apoA-I by HOCI severely impairs its ability to remove cellular cholesterol by the ABCA1 pathway, consistent with the possibility that oxidation of HDL apolipoproteins in vivo would be atherogenic. We propose to test the hypothesis that site-specific oxidation of tyrosine residues by phagocyte-derived HOCI and RNS alters the biological and atheroprotective function of HDL. We will characterize the effects of site-specific oxidation of tyrosines in apoA-I on its ability to remove cellular cholesterol, identify protein motifs that direct site-specific tyrosine oxidation by HOCI and RNS, and determine if HDL is a physiological target for oxidative modification in the artery wall. This project will use HPLC and tandem mass spectrometry to locate and quantify the sites of tyrosine oxidation in apoA-I and model peptides, cell biology procedures to characterize the effects of apoA-I oxidation on lipid transport activity and interactions with ABCA1, tissue analysis to identify and characterize oxidized apoA-I in human atherosclerotic lesions, and mouse model approaches to test for the effects of macrophage-generated HOCI on apoA-I oxidation and atherogenesis in vivo. The proposed studies will provide insights into the structural features that direct oxidative modification of proteins, with important implications for the physiological significance of oxidative reactions in atherosclerosis and other inflammatory diseases.

HDL载脂蛋白，尤其是ApoA-I与细胞膜转运蛋白ABCA1的相互作用 去除过量的细胞胆固醇并预防动脉粥样硬化。此过程是由LipidPoor介导的 由从头合成或与HDL颗粒解离产生的载脂蛋白。因此，因素 这种损害ApoA-I的产生或脂质外排活性可能具有深远的动脉粥样硬化作用。氧化 损害与动脉粥样硬化（一种慢性炎症性疾病）的发病机理有关。 3- 氯丁嘧啶和3-硝基酪氨酸，由吞噬细胞衍生产生的蛋白质氧化产物的稳定产物 在人类中检测到次伐多酸（HOCI）和反应性氮种（RN） 动脉粥样硬化病变。但是，控制蛋白质中酪氨酸氧化的潜在因素仍然存在 理解不佳。我们发现，HOCI对ApoA-I的氧化严重损害了其去除细胞的能力 ABCA1途径的胆固醇与HDL载脂蛋白氧化的可能性一致 体内将是动脉粥样硬化的。我们建议检验以下假设：酪氨酸的位点特异性氧化 吞噬细胞衍生的HOCI和RN的残基改变了HDL的生物学和动脉保护功能。 我们将表征ApoA-I中酪氨酸的位点特异性氧化对去除细胞的能力的影响 胆固醇，鉴定蛋白质基序，这些蛋白质基序是通过HOCI和RN的直接位点特异性酪氨酸氧化的，并且 确定HDL是否是动脉壁中氧化修饰的生理靶标。这个项目将使用 HPLC和串联质谱法定位和量化ApoA-I中酪氨酸氧化的位点 模型肽，细胞生物学程序，以表征ApoA-I氧化对脂质转运的影响 活性和与ABCA1的相互作用，组织分析以识别和表征人类氧化的apoA-I 动脉粥样硬化病变和小鼠模型测试巨噬细胞生成的HOCI的影响 在体内的APOA-I氧化和动脉粥样硬化上。拟议的研究将提供有关结构的见解 直接氧化蛋白质修饰的特征，对生理具有重要意义 氧化反应在动脉粥样硬化和其他炎症性疾病中的重要性。