Abstract 1740 Characterizing the role of matrix metalloproteinase-3 (MMP-3) in the pathogenesis of acute respiratory distress syndrome (ARDS)

Background: The acute respiratory distress syndrome (ARDS) is a major U. S. healthcare burden affecting over 200, 000 people annually with a mortality rate of up to 50% and healthcare costs exceeding 10 billion. ARDS disrupts alveolar-capillary units and epithelial barrier function, leading to protein-rich pulmonary edema and hypoxemic respiratory failure. Current treatments primarily rely on endogenous mechanisms of lung injury repair and supportive care, creating an urgent need for faster therapeutic strategies. Matrix metalloproteinases (MMPs) represent a group of >20 matrix-degrading proteolytic enzymes that have been strongly implicated in the pathogenesis of ARDS because of their direct role in extracellular matrix and intercellular junction degradation, their ability to regulate inflammation by cleaving cytokines and chemokines, and their expression by a variety of cells that are key to the pathobiology of ARDS. Objective: Previous research supports an association between upregulated matrix metalloproteinase-3 (MMP-3) expression in the lung parenchyma and the development of ARDS in patients and mouse models of acute lung injury (ALI). However, the mechanism by which MMP-3 expression influences ARDS pathogenesis remains unclear. The goal of this study is to understand the cellular mechanisms of MMP-3 impact on alveolar epithelial repair, investigate the consequences of MMP-3 expression in different lung epithelial cell types, and assess the therapeutic potential of a selective MMP-3 chemical inhibitor. Methods: To evaluate the role of MMP-3 in ARDS development, we induced ALI/ARDS through acid aspiration or viral pneumonia in wild type (WT) C57BL/6 or MMP-3 (-/-) knockout mice, which are representative of clinical ARDS in humans. MMP-3 expression and fibroproliferation were assessed up to 21 days post-ALI induction. Furthermore, CALU-3 and NuLi human lung epithelial cell lines were treated with recombinant active MMP-3 to assess cell adhesion junction integrity and epithelial barrier function. To further address human relevance, we performed single-cell RNA sequencing on human ARDS and healthy organ donor lungs. Results: From our single cell RNA sequencing data, we found the colocalization of upregulated MMP-3 and Keratin 5+ (KRT5+) transcript expression in lung basal cells of human ARDS samples. In our animal models of ALI, we found increased KRT5 protein expression that colocalized with MMP-3 expression in the lung parenchyma and increased fibroproliferation associated with the upregulation of MMP-3 and KRT5. Furthermore, we found downregulation of KRT5 protein expression in the lung parenchyma of MMP-3 (-/-) knockout mice. Lastly, our cell culture models showed that MMP-3 induced proteolytic cleavage of E-cadherin and MMP-3 influenced transcriptional alterations in junctional and adhesion molecules. Conclusion: Our data suggests that KRT5+ lung epithelial cell type-specific expression of MMP-3 may contribute to ARDS pathogenesis in part through the degradation of E-cadherin and other components of cell junctions. We acknowledge the funding from NIH R01 HL157424 to GPD, DCR, and ESR.