Project 1 Heavy Metal Induced Airway Remodeling and COPD

肺是呼吸性环境毒物的主要入口，包括砷等重金属。 (As)、镉(Cd)和锰(Mn)，所有这些都被认为会导致慢性阻塞性肺 疾病(COPD)。慢性阻塞性肺病是美国第三大致死原因。慢性阻塞性肺病的患病率是现在的两倍 与超级基金网站所在的阿拉巴马州伯明翰受影响地区相比， 控制区。肺巨噬细胞内肽精氨酸脱氨酶-2酶(PAD2)的激活 并诱导波形蛋白脱亚胺(瓜氨酸化)为瓜氨酸波形蛋白 通过将精氨酸残基不可逆地转变为非编码瓜氨酸残基。我们的假设是 接触含有重金属(As、Cd和Mn)的颗粒物会导致诱发和激活 肺巨噬细胞内肽基精氨酸脱亚氨酶2与波形蛋白的表达TLR4在呼吸道中的激活 脱亚胺(瓜氨酸)波形蛋白诱导的成纤维细胞具有侵袭性、促纤维化表型。 随后的气道重塑和COPD。我们将从以下具体目标来检验这一假设： 目的1：我们将使用一种新型的、选择性的PAD2抑制剂(AFM30a)和一种PAN PAD抑制剂 (BB-氯-嘧啶)，以评估这是否导致抑制波形蛋白的瓜氨酸化。我们还将评估是否 瓜氨酸波形蛋白通过调节肺成纤维细胞的侵袭性表型在3D肺组织中的作用 体外上调TLR4的表达。目的2：确定重金属暴露是否导致大鼠气道重塑 COPD小鼠模型，与PAD2激活、瓜氨酸化和波形蛋白分泌有关 和侵袭性肺成纤维细胞纤维化表型。药物或遗传抑制PAD2将阻断 慢性阻塞性肺疾病的发展。我们将使用TLR4-/-小鼠来评估瓜氨酸波形蛋白是否直接导致呼吸道 应用3D肺研究成纤维细胞的重塑和COPD以及侵袭性促纤维化表型 大气球。目的3：确定PAD2和瓜氨酸波形蛋白是否存在于肺巨噬细胞，BAL， 来自受影响地区的一群受试者的血浆和EBC是COPD的生物标志物。现有的生物群落 已经在发现的受试者队列中进行了测试，并将在验证队列中进行前瞻性测试 慢性阻塞性肺疾病患者的CT、血浆和呼气分维(AFD)平行研究 凝析油(EBC)测量。呼气冷凝液中COPD的早期生物标志物可能有助于我们 认识到疾病的易感性。重要的是，这些研究可能提供新的治疗策略 阻断PAD2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.