Myocardin in the pathogenesis of pleural remodeling

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Pleural injury often leads to extensive remodeling of the pleural surfaces, which in severe cases leads to pleural fibrosis (PF). PF can occur as a result of bacterial pneumonia, trauma, radiation exposure and asbestos-related pleural injury. When severe, PF can cause restrictive lung disease or fibrothorax. PF with lung restriction is often seen in medical practice but current treatment is unsatisfactory and effective pharmacotherapy is not available. PF is characterized by proliferation of alpha-smooth muscle actin (α-SMA) expressing myofibroblasts. These myofibroblasts contribute to the thickening of the pleura via increased extracellular matrix deposition. The expansion of myofibroblasts is largely due to mesenchymal transition (MT) of resident pleural mesothelial cells, termed MesoMT. Our preliminary data show that TGF-β induces myocardin expression and activity. Further, myocardin and its effector proteins, smooth muscle myosin, calponin, and KIF5A are upregulated in our mouse models of pleural injury. Conversely, myocardin down- regulation attenuates TGF-β mediated induction of MesoMT marker, α-SMA, secretion of collagen and fibronectin and pleural fibrosis. We also reported that KIF5A is upregulated in TGF-β induced PF and critical for collagen secretion. Based on these observations and strong preliminary data, we strongly infer that myocardin expression and activation substantively contribute to the progression of PF. In this project, we will test the central hypothesis that the activation and upregulation of myocardin and its effector proteins are critical determinants in the acquisition of the activated PMC phenotypes and the progression of PF. Our objective is to test this postulate using state of the art cellular, biochemical, molecular, physiologic and imaging techniques. Primary human (H) and mouse (M) PMCs will be used to define mechanisms by which MesoMT is regulated. Three murine models will be used to assess the role of myocardin and KIF5 signaling in PF: carbon black/bleomycin (CBB) induced PF, Streptococcus pneumoniae-induced empyema/PF and TGF-β induced PF. Our specific aims are: 1) To define the mechanism of myocardin-induced MesoMT of HPMCs, 2) to define the mechanism of KIF5 activation in MesoMT and ECM deposition by HPMCs and 3) to define the contribution of myocardin and related effector proteins to neo-matrix deposition in the progression of PF in vivo. We will use new murine models of fibrosing pleural injury, including mice with mesothelial labelling to enable fate-mapping analyses, molecular, biochemical and immunohistochemical techniques with which we have expertise and state of the art CT imaging and pulmonary function analyses to accomplish these aims. The mechanism(s) by which TGF-β and myocardin regulate MesoMT and PF are currently unclear, representing potentially important gaps in our understanding of the pathogenesis of pleural organization and our ability to identify new therapeutic targets. This proposal addresses each of these gaps and predictably will identify novel targets that could be developed to improve outcomes of patients with PF/fibrothorax.