Therapeutic ECM Resorption in Cellular Systems and Precision Cut Lung Slices.

Project Summary Idiopathic Pulmonary Fibrosis (IPF) is characterized by progressive replacement of functional alveolar gas exchange tissue with collagen rich scar. The accumulation, crosslinking and stiffening of this matrix are defining features of the disease, and definitive barriers to effective repair or regeneration. Emerging evidence indicates that fibrotic scar remains highly resorbable under appropriate conditions], a process that appears to be impaired or absent in humans with IPF. The conditions and pathways that promote fibroblasts (and other cell types) to resorb collagen rich scar are largely unknown. Development of methods and approaches to address this critical gap in understanding is the focus of this U01 proposal. We propose to develop model systems in which ECM deposition and resorption can be efficiently studied in both primary cultured lung fibroblasts as well as precision cut lung slices (PCLS). Our preliminary data show that under appropriate stimuli, IPF-derived human lung fibroblasts can be prompted to degrade and resorb fibrillar collagen. We hypothesize that under appropriate stimuli fibroblasts can be stimulated to not only resorb collagen rich ECM in vitro, but also resorb scar-associated ECM in the lungs from patients with IPF. We propose to develop and leverage novel in vitro tools to test this hypothesis, with the goal of identifying biological pathways and therapeutic interventions that mediate physiologic collagen resorption. We propose to pursue these goals through two aims. In the first aim we will develop culture systems allowing us to identify the signals that promote ECM resorption by lung fibroblasts and delineate the molecular mechanisms of collagen resorption. We will validate these assays for high-throughput discovery and perform a focused screen as proof of concept of the value of this approach. We will also compare the innate ECM deposition and degradation characteristics of IPF and control fibroblasts and fibroblast subsets. In the second aim we will develop ex vivo lung tissue assays to test therapeutic modulation and mechanisms of clearance of scar-associated IPF ECM. We will characterize baseline and stimulus evoked collagenolytic activity in control and IPF lung tissue, and define the association of this activity with lung cell types and histopathological appearance of the tissue. We will also test candidate hits identified in aim 1 for their capacity to increase targeted fibrillar collagen degradation in the native IPF ECM environment. Together the proposed studies will establish robust models of ECM deposition and resorption in primary human IPF fibroblasts and ex vivo lung slices. This platform will open new avenues for identifying signals and mechanisms that shift fibroblasts in IPF toward a matrix resorbing state, generating new opportunities to develop advanced therapeutics for IPF.