Project 1 Heavy Metal Induced Airway Remodeling and COPD

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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.