Fig. 3

Downregulation of pro-SFTPB expression stimulates NSCLC progression by activating the Akt pathway. a Gene Set Enrichment Analysis using the TCGA dataset showing that the SFTPB level is negatively correlated with the Akt pathway in early-stage LUAD. b Western blot analysis showing that downregulation of pro-SFTPB activated the Akt/mTOR signaling pathway in NSCLC cells. The cells were subjected to analysis 72 h after transfection with pro-SFTPB shRNA. c Immunohistochemistry analysis showing that downregulation of pro-SFTPB increased Akt phosphorylation (Ser 473) in NSCLC tissues in subcutaneous xenograft models and lung metastasis models. d The Akt inhibitor MK2206 significantly inhibited the silencing of the pro-SFTPB-induced upregulation of Akt phosphorylation in NSCLC cells. The indicated NSCLC cells were transfected with shRNA pro-SFTPB shRNA. Forty-eight hours after transfection, the cells were treated with 5 µM MK2206 for 12 h and then subjected to Western blotting. e–f Soft agar (e) and invasion (f) assays showing that inhibition of the Akt pathway by the Akt inhibitor MK2206 significantly suppresses the downregulation of pro-SFTPB-stimulated soft agar colony formation and invasion by NSCLC cells. Forty-eight hours after transfection with pro-SFTPB shRNA (sh pro-SFTPB), the cells were treated with 5 µM MK2206 for 12 h and then subjected to analysis. *, compared to the scrambled control group; #, compared to the pro-SFTPB downregulation group. *, p < 0.05; **, p < 0.01; ***, p < 0.001; ^#, p < 0.05; ^##, p < 0.01; ^###, p < 0.001