Qualitative Rather than Quantitative Changes Are Hallmarks of Fibroblasts in Bleomycin-Induced Pulmonary Fibrosis

Abstract

Pulmonary fibrosis is characterized by accumulation of activated fibroblasts that produce excessive amounts of extracellular matrix components such as collagen type I. However, the dynamics and activation signatures of fibroblasts during fibrogenesis remain poorly understood, especially in vivo. We examined changes in lung tissue cell populations and in the phenotype of activated fibroblasts after acute injury in a model of bleomycin-induced pulmonary fibrosis. Despite clustering of collagen type I-producing fibroblasts in fibrotic regions, flow cytometry-based quantitative analysis of whole lungs revealed that the number of fibroblasts in the lungs remained constant. At the peak of inflammation, fibroblast proliferation and apoptosis were both increased, suggesting that the clustering was not merely a result of proliferation, but also of fibroblast migration from nearby alveolar walls. Parabiosis experiments demonstrated that fibroblasts were not supplied from the circulation. Comprehensive gene expression analysis of freshly isolated fibroblasts revealed a detailed activation signature associated with fibrogenesis, including changes in genes responsible for migration and extracellular matrix construction. The Spp1 gene, which encodes osteopontin, was highly up-regulated and was an identifying characteristic of activated fibroblasts present at the sites of remodeling. Osteopontin may serve as a useful marker of profibrotic fibroblasts. These results provide insights into the cellular and molecular mechanisms underlying pulmonary fibrosis and provide a foundation for development of specific antifibrotic therapies.

Integrative analyses of miRNA and proteomics identify potential biological pathways associated with onset of pulmonary fibrosis in the bleomycin rat model

“…Interestingly, a recent study has demonstrated that quite similar functions are affected in isolated activated fibroblasts in BLM-induced pulmonary fibrosis, suggesting that activated resident fibroblasts may migrate to damaged sites, differentiate into myofibroblasts, and generate fibrotic legions by secreting ECM components (Tsukui et al, 2013). Taking these results into consideration, the biofunctional changes detected on day 3 may reflect BLM-induced activation of resident fibroblasts in the lung, although no sign of fibrosis was observed in the histopathological analysis.…”

FOXM1 is a critical driver of lung fibroblast activation and fibrogenesis

“…We sought to evaluate the contribution of FOXM1 to in vivo fibrogenesis by selectively deleting it from Col1-expressing cells. Under baseline conditions, Col1a2 expression in the lung is almost entirely limited to resident fibroblasts (57). It is well known that during fibrotic responses, other cells, including epithelial cells, can undergo mesenchymal transition (39,58), thus expressing Col1 and being subject to Cre-mediated deletion under the control of the Col1 promoter.…”

Inflammasome activation in airway epithelial cells after multi-walled carbon nanotube exposure mediates a profibrotic response in lung fibroblasts

“…We demonstrate here a significant increase in expression of these markers after exposure to conditioned medium from MWCNT treated HBE cells. Increased levels of OPN, PC-1 and TIMP-1 are noted in rodent lungs in bleomycin induced pulmonary fibrosis [36-38]. TN-C is prototypic family member of Tenascin family of extracellular matrix glycoproteins.…”