Fig. 6

The effect of CPP on Wnt/β-catenin pathway activity. a-b. CPP alone (A) and combination with TRAIL (B) decreased Wnt/β-catenin activity in the TOP-flash reporter gene assay. TOP-flash contains the wild-type LEF/TCF-binding sites and can be activated by LiCl (20 mM). TOP-flash or FOP-flash (containing mutant LEF/TCF-binding sites) plasmids was co-transfected with the Renilla plasmid into cells to determine the transcriptional activity of Wnt/β-catenin signaling. c. Western blot analysis of the change of β-catenin in the overall levels after drug treatment of YES-2 and KYSE-150 cells. d. Western blot analysis of the change of p-β-catenin (Ser675) in the nucleus and cytoplasm after drug treatment of YES-2 and KYSE-150 cells. p-β-catenin (Ser675) is the form of β-catenin that enters the nucleus. e-g. qRT-PCR analysis of the change of Wnt/β-catenin pathway target gene (Survivin, c-Myc, and cyclin D1) mRNA levels. LiCl (20 mM) was used to activate the Wnt/β-catenin pathway after treatment of YES-2 and KYSE-150 cells with various concentrations of CPP for 24 h. Data are shown as the mean ± SEM. *P < 0.05