Metals in exhaled breath condensate as COPD biomarkers

描述(由申请人提供)： 慢性阻塞性肺疾病(COPD)是世界性的重大健康问题，且发病率高、发病率高、费用高，且呈上升趋势。COPD是一个重要的医学领域，需要生物标志物来研究其发病机制、诊断和临床治疗。呼出的呼吸冷凝液(EBC)可以用非侵入性方法收集，是开发COPD生物标志物的一种有前途的媒介。本研究旨在将最灵敏、选择性和特异性最高的参考分析技术应用于慢性阻塞性肺疾病患者外周血红细胞成分的研究。采用电热原子吸收光谱分析(ETAAS)和电感耦合等离子体质谱(ICPMS)测定COPD患者外周血中有毒金属和必需微量元素的含量。这种新的方法(以前从未做过)将被用于识别和验证暴露和对有毒金属易感性的生物标记物，已知的有毒金属包含在烟草烟雾中，可能在导致COPD的致病途径中发挥基本的病因学作用。液-质联用(LC-MS/MS)将作为一种补充手段来开发适用于COPD患者长期监测的生物标志物(来自脂质过氧化的醛)。这一新的方法代表着对替代介质(BALF、血液、血清、尿液、头发)的分析的重大进步，替代介质(由于复杂基质中的干扰物质)不那么可靠，并且反映了全身而不是肺(目标组织)的有毒金属和必需微量元素的水平。烟草烟雾和污染的环境大大增加了肺部有毒化学品的负担，特别是重金属。暴露的生物标志物(铝、镉、铬、镍、铅的EBC水平)应提供目标组织剂量的定量估计，从而区分暴露与非暴露对照。肺对吸入有毒物质的反应表现出很高的个体间变异性，这是不同解毒能力的函数。考虑到某些微量元素(锰、铜、锌、硒和钼)是金属蛋白(超氧化物歧化酶、谷胱甘肽过氧化物酶、黄嘌呤氧化酶)的组成部分，可调节对有毒物质的反应，因此将开发易感生物标志物，这可能是吸烟者患COPD的有限比例(15-20%)的原因。易感性生物标志物可用于识别和咨询暴露在烟草烟雾或环境污染物中的疾病风险增加的人。作用的生物标志物将从细胞膜中脂质过氧化后释放到EBC中的醛开始开发。醛的模式可以区分由于氧化应激或其他机制造成的膜损伤，例如自由基的直接攻击。综上所述，我们提出了一种开发COPD生物标志物的新方法，依靠(I)非侵入性收集程序获得(Ii)简单、无干扰的基质(EBC实际上是水)，并通过(Iii)高度选择性和特异性(如果不是完全基于质谱分析)的参考分析方法来确定(Iv)稳定的分析物(重金属和微量元素)。通过EBC生物标记物，我们试图解决两个具体目标：a)确定EBC中金属和醛水平作为COPD患者暴露、易感性和疗效的生物标记物的有效性；b)确定EBC生物标记物作为预测COPD自然病史的预后工具的临床实用性。将EBC中的重金属和有毒金属水平确定为COPD患者暴露、影响和易感性的新生物标志物，可以帮助卫生保健提供者设计新的初级和二级干预措施，包括药物，以改善世界各地COPD患者的生活质量和预后。

DESCRIPTION (provided by applicant): Chronic obstructive pulmonary disease (COPD) is a major health problem worldwide and is increasing with a high prevalence, high morbidity and high cost. COPD is an important medical area for which biomarkers are needed for the study of the pathogenesis, the diagnosis and the clinical management. Exhaled breath condensate (EBC) can be collected with non-invasive methods and is a promising medium to develop biomarkers of COPD. The present research project is aimed at applying the most sensitive, selective and specific reference analytical techniques to the study of the composition of EBC in COPD patients. EBC levels of toxic metals and essential trace elements will be measured in COPD patients by electro-thermal atomic absorption spectroscopy (ETAAS) and inductively coupled plasma - mass spectrometry (ICP-MS). Such a novel approach (never done before) will be applied to identify and validate biomarkers of exposure and susceptibility to toxic metals, known to be contained in tobacco smoke, and probably playing a fundamental etiologic role in the pathogenetic path leading to COPD. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) will be used as a complementary approach to develop biomarkers of effect (aldehydes from lipid peroxidation) suitable for the long-term monitoring of COPD patients. This novel approach represents a significant advance over the analysis of alternative media (BALF, blood, serum, urine, hair), which are not as reliable (owing to interfering substances in the complex matrix) and reflect systemic rather than lung (target tissue) levels of both toxic metals and essential trace elements. Tobacco smoke and polluted environments substantially increase the lung burden of pneumotoxic chemicals, particularly heavy metals. Biomarkers of exposure (EBC levels of AI, Cd, Cr, Ni, Pb) should provide a quantitative estimate of the target tissue dose, thus distinguishing exposed from non-exposed controls. The lung response to inhaled pneumotoxic substances shows a high inter-individual variability as a function of different detoxifying capacity. Biomarkers of susceptibility will be developed relying on the consideration that some trace elements (Mn, Cu, Zn, Se, and Mo) are components of metalloproteins (superoxide dismutase, glutathione peroxidase, xanthine oxidase) known to modulate the response to toxic substances, possibly accounting for the limited proportion (15-20%) of smokers developing COPD. Biomarkers of susceptibility could be useful to identify and counsel people who are at increased risk of disease when exposed to tobacco smoke or environmental pollutants. Biomarkers of effect will be developed starting from aldehydes released into the EBC after lipid peroxidation in cell membranes. The pattern of aldehydes could distinguish membrane damage due to either oxidative stress or other mechanisms, such as a direct attack by free radicals. In summary, we propose a novel approach to the development of COPD biomarkers, relying on (i) noninvasive collection procedures to obtain (ii) a simple, interference-free matrix (EBC is practically water) to be analyzed by (iii) reference analytical methods, which are highly selective and specific (mainly, if not entirely, based on mass spectrometry), to determine (iv) stable analytes (heavy metals and trace elements). Through EBC biomarkers we sought to address two specific aims: a) to determine the validity of metals and aldehydes levels in EBC as biomarkers of exposure, of susceptibility and of effect in patients with COPD; b) to determine the clinical utility of EBC biomarkers as prognostic tools to predict the natural history of COPD. Establishing heavy and toxic metal levels in EBC as novel biomarkers of exposure, effect and susceptibility in patients with COPD could assist health care providers in devising new primary and secondary interventions, including drugs, to improve the quality of life and outcome of patients with COPD worldwide.