Figure 4
From: High mobility group box 1-induced epithelial mesenchymal transition in human airway epithelial cells
Inhibition of the AKT/GSK3β/β-catenin signaling pathway decreased HMGB1-induced EMT.
(a) BEAS-2B cells were treated with 10 μM PI3K inhibitor (LY294002) and GSK-3β inhibitor (SB415286) for 10 min and then treated with HMGB1 (300 ng/mL) for 24 h. Protein expression was detected by western blot analysis. Quantification of E-cadherin, ZO-1 and vimentin was performed using ImageJ software. Data are expressed as mean ± SD (n = 5). #p < 0.05, as compared with the control group. *p < 0.05, as compared with the HMGB1-treated group. (b) BEAS-2B cells were transduced with lentiviral-expressed β-catenin shRNA (1 MOI) for 72 h and selected by puromycin. The mRNA from stable clones expressing β-catenin-targeting shRNAs was analyzed by quantitative real-time PCR. Data are expressed as mean ± SD (n = 5). ##p < 0.01, ###p < 0.001, as compared with the control group. **p < 0.01, ***p < 0.001, as compared with the vector group. (c) β-catenin protein expression was detected by western blot analysis in a stable β-catenin shRNA BEAS-2B cell clone. Quantification of protein expression was performed using ImageJ software. Data are expressed as mean ± SD (n = 3). ##p < 0.01, ###p < 0.001, as compared with the control group. **p < 0.01, ***p < 0.001, as compared with the vector group. (d) A stable β-catenin shRNA BEAS-2B cell clone was treated with HMGB1 (300 ng/mL) for 24 h and protein expression of E-cadherin, ZO-1 and vimentin was detected by western blot analysis. Quantification of protein expression was performed using ImageJ software. Data are expressed as mean ± SD (n = 4). #p < 0.05, as compared with the control group. *p < 0.05, as compared with the HMGB1 group. In immunoblotting assay, gels have been run under the same experimental conditions. Then cropped blots were incubated with different primary antibodies for analysis of signaling pathway.
