Project 4: Urinary Protein Biomarkers for Assessing the Potential Toxicity

肺部疾病是美国人群发病率和死亡率的重要原因。接触空气、水和食物中的化学物质会导致这些疾病。许多化学物质经过生物活化，在啮齿动物中产生选择性肺损伤。该项目的重点是萘（NA）和1-硝基萘（NN），这是目前在超级基金废物场和代表一类重要的环境化合物。这两种化学物质都经过代谢成为亲电子中间体，这些中间体与靶细胞中的蛋白质共价结合。蛋白质结合是细胞毒性的关键组成部分。我们已经证明，在啮齿动物和灵长类动物鼻上皮中靶向的蛋白质是相似的。这强调了开发与毒性密切相关的生物标志物的必要性。核心假设是：/在靶呼吸组织中与NA/NN的反应性代谢物形成关键蛋白加合物与细胞毒性有因果关系;尿液和鼻刷中加合肽/蛋白的测量提供了暴露和效应的高度灵敏的分子特征。/拟议的工作建立在最近的发现基础上，这些发现显示：1）NA处理的小鼠尿液中的高比活度加合物，以及2）小鼠、大鼠和猴的肺和鼻腔中大量鉴定的蛋白质加合物。拟议的研究将利用项目3的抗体捕获和浓缩尿液中的加合肽，并评价动物模型系统中的加合物水平。将通过加速器质谱法（核心A）验证这些方法评估暴露于少量14 C-NA后加合物水平的能力，将利用质量 核心A和B的质谱设施用于分析加合肽，并且将严重依赖于核心B提供的蛋白质组学服务。总体而言，这些研究预计将：1）获得关于呼吸组织中形成的加合蛋白如何在整个动物中处理的基本信息，2）探索环境相关浓度下的浓度-反应关系，3）提供可应用于暴露人群的方法，这将是有用的-与修改-为评估风险的其他代谢活化化学品和4）提供了可以阐明基因多态性的重要性的方法，这些基因负责NA和NN等化学物质的生物处置。

Lung diseases are a significant cause of morbidity and mortality in the US population. Exposure to chemicals in air, water and food contributes to these diseases. A number of chemicals undergo bioactivation to produce selective lung injury in rodents. This project focuses on naphthalene (NA) and 1-nitronaphthalene (NN), which are present in superfund waste sites and which represent a class of environmentally important compounds. Both chemicals undergo metabolism to electrophilic intermediates which become bound covalently to proteins in target cells. Protein binding is a key component of cytotoxicity. We have demonstrated that the proteins targeted are similar in rodents and primate nasal epithelium. This underscores the need to develop biomarkers which are tightly tied to toxicity. The central hypothesis is: /the formation of key protein adducts with reactive metabolites of NA/NN in target respiratory tissue is causally related to cytotoxicity; measurement of adducted peptides/proteins in urine and nasal brushings provides a highly sensitive molecular signature of exposure and effect./ The proposed work builds on recent findings showing 1) high specific activity adducts in urine of NA-treated mice, and 2) numerous identified protein adducts in lungs and nasal cavity of mice, rats and monkeys. The proposed studies will utilize antibodies from project 3 to trap and concentrate adducted peptides from the urine and to evaluate adduct levels in animal model systems. These approaches will be validated for their ability to assess adduct levels following exposures to small amounts of 14C-NA by accelerator mass spectrometry (Core A), will utilize the mass spectrometry facilities of cores A and B for analysis of adducted peptides and will depend heavily on the proteomics services offered by Core B. Overall, these studies are expected to: 1) yield fundamental information on how adducted proteins, formed in respiratory tissues, are handled in the whole animal, 2) explore concentration-response relationships at environmentally relevant concentrations, 3) provide methods which can be applied to exposed human populations, which will be useful-with modification-for assessing risk of other metabolically activated chemicals and 4) provide methods which can clarify the importance of genetic polymorphisms in genes responsible for the biodisposition of chemicals like NA and NN.