Lipid Peroxidation, GSH Depletion, and SLC7A11 Inhibition Are Common Causes of EMT and Ferroptosis in A549 Cells, but Different in Specific Mechanisms

Abstract

Epithelial-mesenchymal transition (EMT) induced by transforming growth factor-β1 (TGF-β1) is thought to be involved in the pathogenesis of pulmonary fibrosis. Emerging evidence suggested that there are some common causes between ferroptosis and pulmonary fibrosis. The interaction of EMT and ferroptosis and its mechanism were investigated by detecting the expression levels of α-smooth muscle actin (α-SMA), E-cadherin, solute carrier family 7 member 11 (SLC7A11), and glutathione peroxidase 4 (GPX4) and measuring the contents of reactive oxygen species (ROS), malondialdehyde (MDA), and glutathione (GSH). The cellular morphology and ultrastructure of mitochondria were studied by microscope and transmission electron microscope (TEM), respectively. The results showed that ferroptosis in A549 cells was induced by Erastin, which decreased the expression levels of E-cadherin (E-Ca), α-SMA, and SLC7A11, accompanied with ROS and MDA increase, as well as GSH decrease. TGF-β1 promoted ultrastructure variation of mitochondria similar to ferroptosis and mesenchymal changes in morphology during EMT of A549 cells, accompanied with reduced GSH content and expression of SLC7A11, as well as ROS and MDA increase. Ferrostatin-1 (Fer-1) recovered ferroptosis induced by Erastin, but had no effect on the morphological change caused by TGF-β1. Furthermore, Fer-1 reduced ROS and MDA production, and increased SLC7A11 expression in the early subsequently increased GSH. However, the effects of Fer-1 on above indicators were different in time. The expression of GPX4 had no significant change during EMT induced by TGF-β1 and ferroptosis induced by Erastin in A549 cells. It is indicating that Erastin promoted the de-epithelialization of lung epithelial cells, but inhibited the process of myofibroblast differentiation; Fer-1 could partially inhibit EMT induced by TGF-β1, but reverse ferroptosis induced by Erastin. TGF-β1 could delay the ferroptosis, but could not prevent it. Lipid peroxidation, GSH depletion, and SLC7A11 inhibition are common causes of EMT and ferroptosis in A549 cells, but different in specific mechanisms. The exact effects of GPX4 involved in EMT and ferroptosis in A549 cells need further study.

Keywords: GPX4; GSH; SLC7A11; epithelial-mesenchymal transition (EMT); ferroptosis.