GLP-1R functions in ICC by regulating FoxO1 signaling. (A) The mRNA levels of FoxO1 in RBE and HCCC-9810 cells with GLP-1R knockdown were determined by reverse transcription-quantitative polymerase chain reaction analysis. (B) The protein level and the phosphorylation state of FoxO1 in RBE cells were determined by western blot analysis. The p-FoxO1/FoxO1 ratio was calculated as (p-protein/control)/(total protein/control). (C) Expression of genes targeted by FoxO1 were measured in RBE cells with GLP-1R knockdown. Correlations between GLP-1R levels, FoxO1 levels, and FoxO1 phosphorylation states were evaluated in 20 tumor tissues by (D) immunohistochemical and (E) western blot analyses. Transwell assays were used to determine the effects of FoxO1 and FoxO1S256D in the (F) migration and (G) invasion of RBE cells with GLP-1R knockdown. n.s., not significant; GLP-1R, glucagon-like peptide-1; FoxO1, forkhead box O1; p, phosphorylated; si, small interfering RNA. Representative images for Transwell assays were obtained at ×20 magnification. n.s., not significant, *P<0.05 vs. siGLP-1R, **P<0.01 vs. siGLP-1R or siGLP-1R+S256D, ***P<0.001 vs. siGLP-1R.