Project 1 Heavy Metal Induced Airway Remodeling and COPD

肺是可吸入环境毒物的主要门户，包括重金属，如砷 (As)镉（Cd）和锰（Mn），所有这些都被认为是导致慢性阻塞性肺疾病的原因。 疾病（COPD）。COPD是美国第三大死亡原因。COPD的患病率是 与超级基金所在地亚拉巴马伯明翰的受影响地区相比， 控制区。肺巨噬细胞中的肽基精氨酸脱亚胺酶-2（PAD 2）被重金属激活 金属，并诱导波形蛋白脱亚胺化（瓜氨酸）为瓜氨酸化波形蛋白 通过精氨酸残基不可逆地改变为非编码瓜氨酸残基。我们的假设是 暴露于含有重金属（As、Cd和Mn）的颗粒物会导致 肺巨噬细胞中的肽基精氨酸脱亚胺酶2和波形蛋白的脱亚胺化。气道中TLR 4的活化 脱亚氨基（瓜氨酸化）波形蛋白对成纤维细胞的作用导致促侵袭、促纤维化表型， 随后的气道重塑和COPD。我们将在以下具体目标中检验这一假设： 目的1：我们将使用一种新的、选择性的PAD 2药理学抑制剂（AFM 30 a）以及泛PAD抑制剂 （BB-Cl-脒），以评估这是否导致波形蛋白的瓜氨酸酶抑制。我们还将评估， 瓜氨酸波形蛋白通过调节气道成纤维细胞进入3D肺球的侵袭性表型 TLR 4在体外的上调。目的2：确定重金属暴露是否导致气道重塑， 与PAD 2的活化、瓜氨酸酶和波形蛋白的分泌有关 和肺成纤维细胞的侵袭性促纤维化表型。对PAD 2的药理学或遗传学抑制将阻断 COPD的发展。我们将使用TLR 4-/-小鼠来评估瓜氨酸化波形蛋白是否直接导致气道 重塑和COPD以及使用3D肺的成纤维细胞的侵袭性促纤维化表型 肺球目的3：确定肺巨噬细胞，BAL， 来自受影响地区的一组受试者的血浆和EBC是COPD的生物标志物。现有生物标本 已在受试者的发现队列中进行了测试，并将在验证队列中进行前瞻性测试 COPD受试者与CT扫描、血浆和呼出气上的气道分形维数（AFD）平行 冷凝水（EBC）测量。呼出气冷凝物中COPD的早期生物标志物可能有助于我们 认识疾病易感性。重要的是，这些研究可能提供新的治疗策略， 阻断PAD 2在慢性肺病（如COPD）患者中的作用。

The lung is a major portal for respirable environmental toxicants including heavy metals such as arsenic (As), cadmium (Cd), and manganese (Mn), all of which are recognized to cause chronic obstructive pulmonary disease (COPD). COPD is the third largest cause of mortality in the US. The prevalence of COPD is twice as high in the Affected Area in Birmingham, Alabama where the Superfund site is located when compared to the Control Area. Peptidyl arginine deiminase-2 enzyme (PAD2) in lung macrophages is activated by heavy metals in a calcium dependent manner and induces deimination (citrullination) of vimentin to citrullinated vimentin by the irreversible alteration of the arginine residue to the non-coded citrulline residue. Our hypothesis is that exposure to particulate matter containing heavy metals (As, Cd and Mn) leads to induction and activation of peptidyl arginine deiminase 2 in lung macrophages and deimination of vimentin. Activation of TLR4 in airway fibroblasts by deiminated(citrullinated) vimentin leads to a pro-invasive, pro-fibrogenic phenotype, with subsequent airway remodeling and COPD. We will examine this hypothesis in the following specific aims: Aim 1: We will use a novel, selective pharmacologic inhibitor of PAD2 (AFM30a) as well as a pan PAD inhibitor (BB-Cl-amidine) to evaluate if this leads to inhibition of citrullination of vimentin. We will also evaluate if citrullinated vimentin modulates airway fibroblast into an invasive, phenotype in 3D lung pulmospheres through upregulation of TLR4 in vitro. Aim 2: Determine whether heavy metal exposure leads to airway remodeling in a murine model of COPD and is associated with the activation of PAD2, the citrullination and secretion of vimentin and an invasive profibrotic phenotype of lung fibroblast. Pharmacologic or genetic inhibition of PAD2 will block the development of COPD. We will use TLR4-/- mice to evaluate if citrullinated vimentin directly causes airway remodeling and COPD as well as an invasive pro-fibrogenic phenotype of fibroblasts using 3D lung pulmospheres. Aim 3: Determine whether PAD2 and citrullinated vimentin, present in lung macrophages, BAL, plasma and EBC of a cohort of subjects from the Affected Area are biomarkers for COPD. Existing biospecimens have been tested in a discovery cohort of subjects and prospective testing will be conducted in a validation cohort of COPD subjects in parallel with airway fractal dimension (AFD) on CT scans, plasma and exhaled breath condensate (EBC) measurements. Early biomarkers of COPD in exhaled breath condensate may help us recognize disease susceptibility. Importantly, these studies may provide novel therapeutic strategies to block the effects of PAD2 in patients with chronic lung disease such as COPD.