In patients with acute respiratory distress syndrome (ARDS), disruption of the alveolar-capillary barrier leads to edema accumulation in the lungs which reduces lung compliance and impairs gas exchange. Hypercapnia (elevated CO2 levels) is often an inevitable consequence of lung protective strategies utilizing low tidal volume mechanical ventilation in ARDS patients. The repair of the alveolar epithelium following acute lung injury is critical for lung homeostasis, which is accomplished by alveolar type 2 (AT2) cell self-renewal, and differentiation into AT1 cells. We and others have reported that exposure to hypercapnia is associated with decreased AT2 cell proliferation and differentiation and worse clinical outcomes in patients with ARDS. Mitochondrial dysfunction and metabolic changes are commonly observed in patients with severe ARDS but whether this dysfunction is causally related to impaired epithelial repair is not known. Three tricarboxylic acid (TCA) cycle enzymes, including isocitrate dehydrogenase (IDH) produce CO2 making them susceptible to inhibition by mass action during hypercapnia. We have reported that elevated CO2 levels inhibit IDH2 in alveolar epithelial cells to activate downstream signaling pathways to impair alveolar epithelial barrier function and increase mortality in a murine model of ARDS induced by influenza A pneumonia. Our new preliminary data suggest inhibition of the TCA cycle by hypercapnia activates the integrated stress response (ISR) via OMA1, DELE1 and HRI, altering the alveolar epithelial cell state and increasing alveolar-capillary permeability. Further preliminary data suggest that hypercapnia-induced pathologic ISR activation causes AT2 cells to stall in a transitional state, precluding normal lung repair after injury via upregulation of ATF4. Collectively, we hypothesize that hypercapnia inhibits the TCA cycle and activates the ISR via the OMA1/DELE1 and HRI axis to induce an ATF4-mediated increase in alveolar epithelial permeability and stall the differentiation of AT2 into AT1 cells. We will test this hypothesis via three interrelated Specific Aims: Aim 1. To determine whether hypercapnia-induced TCA cycle inhibition activates the ISR and ATF4 to impair alveolar epithelial permeability. Aim 2. To determine whether hypercapnia inhibits the TCA cycle to activate the ISR via OMA1, DELE1 and HRI. Aim 3. To determine whether hypercapnia-induced pathologic ISR activation activates ATF4 to inhibit alveolar epithelial differentiation and delay lung repair after injury. We propose causal experiments using sophisticated genetic murine models to link hypercapnia to mitochondrial metabolism, ISR activation and failed epithelial differentiation to worsened lung injury and inhibition of lung repair. As hypercapnia is often an inevitable clinical consequence of lung injury and lung protective ventilation strategies, mitigating its effects by ISRIB or other small molecules may improve patient survival and improve reco...