Project Summary/Abstract Pulmonary fibrosis (PF) describes a group of disorders in which the alveolar space is progressively replaced by fibrotic tissue which eventually leads to death from respiratory failure. Available therapies modestly slow disease progression but are not curative, difficult to tolerate and most patients die unless they receive a lung transplant. The prevalence of PF has tripled in the US Veterans Population between 2010 and 2019 and likely to continue increasing in light of the COVID-19 pandemic. We were the first to show that some patients with severe COVID- 19 develop severe lung fibrosis that is indistinguishable from IPF even at the level of the single cell transcriptome. These findings prompted us to perform the first of many lung transplants for patients with severe COVID-19 in the US. Even more concerning, however, is the prevalence of respiratory symptoms in more than 30% of patients with post-acute sequelae of COVID-19 some of whom have PF. Thus, if even a small fraction of the 66 million US survivors of COVID-19 to date develop fibrotic lung disease, the public health impact will be enormous. Our laboratory has made important contributions to a model that hypothesizes that lung fibrosis begins with damage to the alveolar epithelium which leads to the recruitment circulating monocytes to the alveolar space where they differentiate into profibrotic alveolar macrophages to create a temporary seal over the injured alveolar epithelium and in direct contact with alveolar fibroblasts. Trophic factors for fibroblasts are released from alveolar macrophages and fibroblasts secrete G-CSF in response, which is necessary to sustain alveolar macrophages and creates a profibrotic circuit that persists until the epithelial barrier is restored. Metformin is an oral drug used for Type 2 diabetes that also ameliorates fibrosis in animal models but has not been associated with improved outcomes in PF patients. Our preliminary data suggest these disparate findings might reflect the pharmacokinetics of the drug—high levels of drug in the lung are only achieved with intraperitoneal but not gut administration. Thus, while metformin may not be useful as a therapy for lung fibrosis, small molecules that mimic its mechanism of action might be. However, this requires a mechanistic understanding of metformin’s efficacy in lung fibrosis which is currently lacking. Metformin is concentrated in mitochondria where it functions to inhibit Complex I of the mitochondrial electron transport chain. We have identified a yeast protein, NDI1, that restores mitochondrial electron transport in the absence of complex I activity and is insensitive to metformin. Using this construct, we have shown that many of metformin’s biologic effects are attributable to its ability to inhibit complex I. We have also generated preliminary data suggesting a direct activator of AMPK, an indirect target of metformin, prevents fibrosis after the intratracheal administration of...