Ozone, lipid-protein adducts, and biological effects

描述（由申请人提供）：臭氧是最常见的环境污染物之一，人类暴露于这种活性分子水平的增加显然与促炎反应和呼吸系统疾病的加剧有关。不饱和脂质特别容易受到臭氧的影响，许多亲电醛和环氧化物是脂质臭氧暴露的主要产物。例如，胆固醇很容易与臭氧反应，并产生由甾醇的B环双键转化为活性羰基和环氧化物的产物。内源性形成的氧甾醇是众所周知的肝x受体（LXR）的配体，它调节参与胆固醇稳态、脂肪酸合成和逆向胆固醇运输的基因的表达。最近的研究也表明LXR具有强大的抗炎和免疫调节功能。与内源性形成的氧甾醇相反，一些臭氧衍生的氧甾醇抑制LXR功能，但这一机制在臭氧发病机制中的作用是已知的。众所周知，臭氧可以改变细胞功能并激活上皮细胞，但这些过程的生化机制尚未明确。臭氧衍生的氧甾醇是亲电试剂，与存在于蛋白质中的常见亲核残基反应，形成稳定的加合物。我们认为，已知的臭氧化学反应性，其在气道中的脂质暴露，这些暴露导致的氧化应激，以及人类疾病与环境损害的关联，要求进行协调研究，旨在发现将脂质、环境暴露和病理生理反应联系起来的基本化学和生物事件。臭氧衍生的氧化甾醇是拟议研究中的一个共同主题，我们在这里概述了旨在：1。提供化学纯臭氧衍生的氧甾醇，以评估这些化合物及其代谢物对上皮细胞功能的影响，特别关注LXR信号通路。2. 开发基于“点击化学”和合成炔基甾醇类似物的方法，允许分离和鉴定氧甾醇-蛋白加合物，并定义上皮细胞和巨噬细胞中的氧甾醇加合蛋白组。3. 利用合成甾醇和氧甾醇类似物来跟踪和鉴定各种上皮细胞区室中的脂质。4. 基于我们的细胞研究，开发脂质-蛋白内合的生物标志物的测定方法。范德比尔特大学和北卡罗来纳大学教堂山分校的实验室为研究一个重要的环境问题提供了独特的专业知识组合。这些专业知识包括化学合成、分离和表征方面的技能，以及研究暴露于空气污染物后引起的不良健康影响和介导这些反应的细胞机制的经验。

DESCRIPTION (provided by applicant): Ozone is one of the most commonly encountered environmental pollutants and human exposures to increased levels of this reactive molecule are clearly linked to proinflammatory responses and exacerbation of respiratory illness. Unsaturated lipids are particularly vulnerable to ozone and a number of electrophilic aldehydes and epoxides are known as primary products of lipid ozone exposure. Cholesterol, for example, reacts readily with ozone and gives oxysterol products that result from conversion of the ring- B double bond of the sterol to reactive carbonyl and epoxide functionality. Endogenously formed oxysterols are well-known ligands for the liver-X-receptor (LXR), which regulates the expression of genes involved in cholesterol homeostasis, fatty acid synthesis, and reverse cholesterol transport. More recent studies also implicate LXR in having potent anti-inflammatory and immune regulatory function. In contrast to endogenously formed oxysterols, some of the ozone-derived oxysterols inhibit LXR function, yet the role of this mechanism in ozone pathogenesis is known. Ozone is known to modify cellular function and activate epithelial cells but the biochemical mechanisms of these processes have yet to be defined. Ozone-derived oxysterols are electrophiles that react with common nucleophilic residues present in proteins, forming stable adducts. We contend that the known chemical reactivity of ozone, its presumed exposure to lipids in the airway, the oxidative stress that results from these exposures, and the association of human diseases with environmental insults calls for coordinated studies aimed at discovering the fundamental chemical and biological events linking lipids, environmental exposures and the pathophysiological response. Ozone-derived oxysterols are a common theme in the proposed research and we outline strategies here that are designed to: 1. Provide chemically pure ozone-derived oxysterols to assess the effect of these compounds and their metabolites on the function of epithelial cells, with specific focus on the LXR signaling pathway. 2. Develop methods based on "click chemistry" and synthetic alkynyl sterol analogs that permit the isolation and identification of oxysterol-protein adducts and define the oxysterol adduction proteome in epithelial cells and macrophages. 3. Utilize synthetic sterol and oxysterol analogs to track and identify lipids in various epithelial cellular compartments. 4. Develop assays for biomarkers of lipid-protein adduction based on our cellular studies. The laboratories at Vanderbilt University and University of North Carolina at Chapel Hill provide a unique combination of expertise for the study of an important environmental problem. This expertise includes skills in chemical synthesis, isolation and characterization coupled with experience studying adverse health effects induced upon exposure to air pollutants and the cellular mechanisms mediating these responses.