Involvement of epithelial-mesenchymal transition in methotrexate-induced pulmonary fibrosis

• Published in: Journal of toxicological sciences 2014/04/01, Vol.39(2), pp.319-330
• Main Authors: Ohbayashi, Masayuki, Kubota, Satoshi, Kawase, Aya, Kohyama, Noriko, Kobayashi, Yasuna, Yamamoto, Toshinori
• Format: Article
• Language: English
• Subjects: Actins - metabolism; Alveolar epithelial cell; Animals; Cadherins - metabolism; Cell Proliferation - drug effects; Down-Regulation; Epithelial Cells - drug effects; Epithelial Cells - metabolism; Epithelial Cells - pathology; Epithelial mesenchymal transition; Epithelial-Mesenchymal Transition - drug effects; Epithelial-Mesenchymal Transition - genetics; Humans; Interleukin-6 - metabolism; Methotrexate; Methotrexate - adverse effects; Mice; Myofibroblasts - cytology; Myofibroblasts - drug effects; Myofibroblasts - pathology; Pulmonary Alveoli - cytology; Pulmonary Fibrosis - genetics; Pulmonary Fibrosis - physiopathology; Pulmonary toxicity; Transforming Growth Factor beta1 - metabolism; Tumor Cells, Cultured; Up-Regulation; Vimentin - metabolism
• ISSN: 0388-1350; 1880-3989
• DOI: 10.2131/jts.39.319

Epithelial-mesenchymal transition (EMT) plays a pivotal event in the development of pulmonary fibrosis. We have previously reported that methotrexate (MTX)-induced alveolar epithelial cell injury followed by pulmonary fibrosis as a result of the recruitment and proliferation of myofibroblasts. However, there is no data concerning whether EMT occurs in MTX-induced pulmonary fibrosis. In the present study, therefore, we investigated the expression of EMT markers such as E-cadherin, α-SMA, and vimentin by immunofluorescence analysis in mouse lung tissues after administration of MTX. We found that vimentin and α-SMA-positive cells of the MTX-induced pulmonary fibrosis were increased; on the other hand, E-cadherin was decreased, indicating that epithelial cells act as the main source of mesenchymal expansion. These results exhibited the down-regulation of E-cadherin expression and the up-regulation of α-smooth muscle actin (α-SMA) in primary mouse alveolar epithelial cells (MAECs) and A549 cell lines. Additionally, MTX-induced A549 cells exhibited an EMT-like phenotype accompanied by the elevation of the expression of interleukin-6 (IL-6) and transforming growth factor (TGF)-β1, as well as an enhancement of migration. All of these findings suggest that MTX-induced pulmonary fibrosis occurs via EMT.