Sulforaphane attenuates pulmonary fibrosis by inhibiting the epithelial-mesenchymal transition

Abstract

Background: Idiopathic pulmonary fibrosis (IPF) is a progressive and fatal disease with no effective treatment. The epithelial-mesenchymal transition (EMT) is a critical stage during the development of fibrosis. To assess the effect of sulforaphane (SFN) on the EMT and fibrosis using an in vitro transforming growth factor (TGF)-beta 1-induced model and an in vivo bleomycin (BLM)-induced model. Methods: In vitro studies, cell viability, and cytotoxicity were measured using a Cell Counting Kit-8. The functional TGF-beta 1-induced EMT and fibrosis were assessed using western blotting and a quantitative real-time polymerase chain reaction. The lungs were analyzed histopathologically in vivo using hematoxylin and eosin and Masson's trichrome staining. The BLM-induced fibrosis was characterized by western blotting and immunohistochemical analyses for fibronectin, TGF-beta 1, E-cadherin (E-cad), and alpha-smooth muscle actin (SMA) in lung tissues. Results: SFN reversed mesenchymal-like changes induced by TGF-beta 1 and restored cells to their epithelial-like morphology. The results confirmed that the expression of the epithelial marker, E-cadherin, increased after SFN treatment, while expression of the mesenchymal markers, N-cadherin, vimentin, and alpha-SMA decreased in A549 cells after SFN treatment. In addition, SFN inhibited TGF-beta 1-induced mRNA expression of the EMT-related transcription factors, Slug, Snail, and Twist. The SFN treatment attenuated TGF-beta 1-induced expression of fibrosis-related proteins, such as fibronection, collagen I, collagen IV, and alpha-SMA in MRC-5 cells. Furthermore, SFN reduced the TGF-beta 1-induced phosphorylation of SMAD2/3 protein in A549 cells and MRC-5 cells. BLM induced fibrosis in mouse lungs that was also attenuated by SFN treatment, and SFN treatment decreased BLM-induced fibronectin expression, TGF-beta 1 expression, and the levels of collagen I in the lungs of mice. Conclusions: SFN showed a significant anti-fibrotic effect in TGF-beta-treated cell lines and BLM-induced fibrosis in mice. These findings showed that SFN has anti-fibrotic activity that may be considered in the treatment of IPF.