Introduction: We have previously reported bleomycin (BLM)-induced pulmonary fibrosis and therapeutic studies in a translational sheep model. The present study uses this model to focus on the changing profile of alveolar macrophages (AMs) and lung fibroblasts during disease progression.
Methods: Lung fibrosis was induced in sheep (n=4) via staggered bronchoscopic instillations of BLM into separate lung lobes. One lobe was given BLM on weeks 1 and 3 of the experimental period, with a separate lobe given BLM on weeks 3 and 5. Saline (control) instillations were given to a healthy control lobe on weeks 1 and 3. Sheep were euthanized at week 7, allowing lung tissue and bronchoalveolar lavage (BAL) collections from a healthy lung lobe, as well as from BLM-exposed lobes showing ‘early’ and ‘later’ stages of fibrotic disease. AMs were characterized by flow cytometry and cytokine gene expression. Histopathology was assessed in lung tissues, while lung fibroblasts were grown in culture and functional changes investigated in vitro. RT-PCR, Western blot and ELISA were employed to assess changes in gene and protein expression.
Results: Lung histopathology, inflammation and fibrosis scores increased significantly with disease progression. This was accompanied by a shift towards an M2 alveolar macrophage (CD163, low MHC CLII) profile, together with elevated BAL and tissue TGF-β levels over time. Lung fibroblasts showed significant phenotypic changes, with elevated TGF-β, MMP-9 and αSMA expression in ‘later fibrotic’ lung fibroblasts.
Conclusion: This work provides a foundation for a better understanding of the cellular basis of disease progression in the sheep model of bleomycin-induced pulmonary fibrosis. Our data highlight alveolar macrophages and lung fibroblasts as key players in pulmonary fibrosis, with a role in transitional change from a proinflammatory to a more profibrotic lung environment. This work also provides a translational platform for therapeutic studies moving forward.