Endoplasmic reticulum stress promotes epithelial-mesenchymal transition via the PERK signaling pathway in paraquat-induced pulmonary fibrosis

Abstract

Pulmonary fibrosis is the primary reason for mortality in patients with paraquat (PQ) poisoning. Our previous study demonstrated that epithelial-mesenchymal transition (EMT) had a role in PQ-induced pulmonary fibrosis. However, the role of endoplasmic reticulum (ER) stress in PQ-induced EMT remains clear. The present study aimed to determine the role of ER stress in EMT in PQ-induced pulmo- nary fibrosis. A549 and RLE-6TN cells were incubated with LY294002 (a PI3K inhibitor) or transfected with protein kinase RNA-like ER kinase (PERK) small interfering RNA (si) for 24 h prior to being exposed to PQ. Next, the expression levels of ER stress-related proteins, PI3K/AKT/GSK-3β signaling pathway-related proteins and EMT-related markers were analyzed by performing western blotting, reverse transcrip- tion-quantitative PCR and immunofluorescence assays. The results of the present study revealed that the protein expres- sion levels of PERK, phosphorylated (p)-PERK, p-eukaryotic initiation factor 2 (eIF2)α were significantly upregulated in the PQ group, whereas p-PI3K, p-AKT and p-GSK-3β were significantly upregulated in the sicontrol + PQ group compared with the sicontrol group. In vitro, following trans- fection with siPERK or treatment with the PI3K inhibitor, the protein expression levels of E-cadherin (an epithelial marker) were upregulated, whereas the protein expression levels of α-SMA (a mesenchymal marker) were downregu- lated. Immunofluorescence analysis revealed that the levels of E-cadherin were markedly upregulated, whereas the levels of α-SMA were notably downregulated following transfection with siPERK compared with the sicontrol group. The results of wound healing assay demonstrated that cell migration in the siPERK + PQ group was markedly decreased compared with the sicontrol + PQ group. These indicated that PQ-induced EMT was suppressed after silencing PERK. The expression levels of p-GSK-3β, p-AKT and p-PI3K were also markedly downregulated in the siPERK + PQ group compared with the sicontrol + PQ group. In conclusion, the findings of the present study suggested that ER stress may promote EMT through the PERK signaling pathway in PQ-induced pulmonary fibrosis. Thus, ER stress may represent a potential therapeutic target for PQ-induced pulmonary fibrosis.