The ferret bleomycin-induced model of lung fibrosis shares features of human idiopathic pulmonary fibrosis

In the present study, we report a bleomycin-induced ferret PF model characterized by an irreversible decrease in pulmonary compliance and increase of opacification, accompanied by “honeycomb cyst-like” structures, “proximalization” of distal lung epithelium. Cellular and molecular analysis by single-nucleus RNA sequencing analysis revealed a significant shift in distal lung epithelium towards proximal epithelial phenotype. Importantly, a histopathological pattern of bronchiolization encompassing divergent atypical epithelial cells, and KRT17+/TP63+/KRT5low “basaloid-like” cells, were present in the distal fibrotic lung lesions. Trajectory analysis revealed AT2 cells transition through multiple cell-states in bleomycin injured ferret lungs, particularly AT2 to KRT8high/KRT7low/SOX4+ to eventual KRT8high/KRT7high/SFN+/TP63+/KRT5low “basaloid-like” cells. Further, immunofluorescence analyses demonstrated KRT7 and KRT8 populations reside in close proximity to the ACTA2 positive myofibroblasts in fibrotic foci thereby driving fibrogenic phenotype in bleomycin injured ferret lungs. Collectively, our results provide evidence that the bleomycin ferrets can reproduce pathophysiological, cellular, and molecular features of human IPF disease, suggesting that they may be a reliable model for understanding mechanisms of IPF pathogenesis and for testing therapeutic strategies for treatment of IPF. Overall design: Multiple (3x) low doses (2.5U/kg) of bleomycin were delivered to the lung of ferrets to induce a more rapidly progressive and irreversible pulmonary fibrosis. Lung structure/function studies (Pulmonary function tests and CT), cellular phenotyping by single nucleus RNA sequencing (snRNA-seq), and histopathologic cellular phenotyping were utilized to demonstrate the progressive and irreversible features of IPF that lead to honeycomb cysts and bronchiolization of the distal airspaces.