Figure 2.

EPBS downregulates EGFR and the Wnt/β-catenin pathway and decreases the nuclear pool of β-catenin. (A) A549 cells and PC-9 cells (5 × 10⁵ cells/well) were treated with EPBS (0, 30, 50, and 100 µM) for 4 h. Western blot analysis was performed. For confirming equal protein loading, the same blot membrane was stripped and re-probed with a β-actin antibody. (B) A549 cells (5 × 10⁵ cells/well) were treated with the indicated concentration of EPBS for 4 h or with 100 µM of EPBS for indicated time intervals. EPBS-treated cells were collected and lysed, and then an equal amount of lysates were detected by Western blot analysis. (C) EPBS (0, 30, 50, and 100 µM for 4 h)-treated A549 cells (5 × 10⁵ cells/well) were collected, and then cytoplasmic extracts (CE) and nuclear extracts (NE) were prepared. As loading control of cytoplasmic extracts or nuclear extracts, α-tubulin or lamin B antibodies were used. (D) A549 cells were treated with EPBS (100 µM) for 4 h, and β-catenin translocation was analyzed by immunocytochemistry. (E) A549 cells were treated with the indicated concentration of LiCl (inhibitor of GSK-3β) for 4 h, and then Western blotting was carried out. (F,G) A549 cells were treated with 20 mM of LiCl and 100 µM of EPBS for 4 h. Then, Western blot analysis and immunocytochemistry were performed. The fold values at the bottom of the blots indicate the fold activation for the untreated control group. Representative data of at least three independent experiments are shown.