ObjectiveTo investigate the role and molecular mechanism of dual-specificity phosphatase 26 (DUSP26) in the proliferation, migration and invasion of lung adenocarcinoma (LUAD) A549 cells.MethodsThe expression profile of DUSP26 was retrieved from the GEPIA2 tumor database, and its differential expression in LUAD tissues and normal human lung tissues were analyzed. Twelve pairs of LUAD tissue and paracancerous tissue surgically resected at Pingxiang People's Hospital between October 2022 and October 2023 were collected. The difference in DUSP26 expression between LUAD tissues and paracancerous tissues was analyzed using immunohistochemical (IHC) staining and Western blotting (WB). Additionally, the expression of DUSP26 in four LUAD cells (A549, SK-LU-1, Calu-3, H1299) and two normal bronchial epithelial cells (BEAS-2B, HBEC) was detected using WB method. A549 cells stably overexpressing DUSP26 (DUSP26-OE) or corresponding negative control (DUSP26-OENC) were constructed via lentiviral transfection. The effects of DUSP26 overexpression on cell proliferation, migration and invasion were detected using colony formation, scratch assay, and Transwell chamber assay, respectively. The expression levels of TGF-β1/SMAD2/3 pathway-and epithelial-mesenchymal transition (EMT)-related proteins were detected using WB method, and the expression levels of Ki-67 and cyclin D1 in cells were detected by immunofluorescence staining. Rescue experiments were conducted by adding 5 ng/mL recombinant TGF-β1. A nude mouse xenograft model was established using A549 cells to observe the effect of DUSP26 overexpression on the in vivo growth of transplanted tumors. The expression levels of TGF-β1/SMAD2/3 pathway-and EMT-related proteins in transplanted tumor tissues were assessed using WB method, and the expression levels of Ki-67 and cyclin D1 in transplanted tumor tissues were detected using immunofluorescence staining.ResultsDUSP26 expression was downregulated in both LUAD tissues and cells (P < 0.05, P < 0.01, P < 0.001 or P < 0.000 1). Compared with the DUSP26-OENC group, the DUSP26-OE group showed significantly reduced proliferation, migration and invasion of A549 cells (P < 0.01 or P < 0.001). Furthermore, the protein levels of TGF-β1, p-SMAD2/3, vimentin, N-cadherin, snail, Ki-67, and cyclin D1 were significantly reduced (P < 0.01, P < 0.001 or P < 0.000 1), while E-cadherin level was increased in the DUSP26-OE group (P < 0.000 1). The addition of 5 ng/mL TGF-β1 recombinant protein partially reversed the effects of DUSP26 overexpression in vitro experiments. The nude mice A549 cell xenograft model was successfully constructed. The growth rate of transplanted tumors was significantly slower in the DUSP26-OE group, with reduced volume and mass (all P < 0.001). The protein levels of TGF-β1, p-SMAD2/3, vimentin, N-cadherin, snail, Ki-67, and cyclin D1 in the transplanted tumor tissues were all reduced (P < 0.01 or P < 0.001), while E-cadherin level was increased (P < 0.000 1).ConclusionDUSP26 is downregulated in both LUAD tissues and cells. Upregulation of DUSP26 suppresses the proliferation, migration and invasion of A549 cells by inhibiting the TGF-β1/SMAD2/3 signaling pathway.