Iron metabolism regulation of epithelial-mesenchymal transition in idiopathic pulmonary fibrosis

Background: Iron overload has been found in the lungs of patients with idiopathic pulmonary fibrosis (IPF) and is thought to be involved in disease progression; however, the underlying mechanism is complex and not yet fully understood. We sought to assess the in vitro role of iron in the progression of fibrosis in lung epithelial cells, and examine the possible regulation of iron and IPF. Methods: Erastin was used to establish a cell model of iron accumulation in mouse lung epithelial cell line 12 (MLE-12). A Cell Counting Kit-8 assay and annexin V staining were applied to measure cell viability and apoptosis, quantitative polymerase chain reaction (qPCR) and quantitative immunoblot analysis of the protein was conducted to analyze the expression of E-cadherin, N-cadherin, alpha-smooth muscle actin (alpha-SMA), Vimentin and beta-Actin. The autophagy was visualized by microtubule-associated protein 1A/1B-light chain 3 (LC3) staining and western blot. Results: The results showed that cell proliferation was significantly inhibited and apoptotic and necrotic cells were significantly increased with 2 mu M of erastin treatment. Western blotting showed that reactive oxygen species (ROS) production and the level of heme oxygenase-1 were increased in the cells. Epithelialmesenchymal transition (EMT) represented by the suppression of E-cadherin and the upregulation of alpha-smooth muscle actin (alpha-SMA) and Vimentin was induced by erastin. Additionally, autophagy represented by activated LC3B and up-regulated Beclin-1 were also induced by erastin. To further ascertain the role of autophagy in erastin-induced EMT, chloroquine, which is an autophagy inhibitor, was employed, and was found to effectively reduce EMT in this process. Conclusions: These results support the role of the enhanced accumulation of iron as a mechanism for increasing the vulnerability of lung epithelial cells to iron-driven oxidant injury that triggers further autophagy during EMT.