Fig. 2

RPRD1B increases cell migration and tumor lymph node metastasis. A Upregulation of RPRD1B was detected in seven GC cell lines compared with an immortalized gastric cell line (GES1) using qRT–PCR and western blotting. B qRT–PCR and western blot analyses showing the ectopic expression of RPRD1B in RPRD1B-overexpressing cells and decreased expression of RPRD1B in shRNA-transfected cells. GAPDH expression was used as a loading control (C) Transwell assay showing that RPRD1B promoted cell migration and invasion. Scale bar, 200 μm. D Silencing RPRD1B expression effectively inhibited cell migration and invasion, as analyzed using the Transwell assay. Scale bar, 200 μm. E, F Wound-healing assay showing that RPRD1B overexpression promoted the migration of HGC27 cells and RPRD1B knockdown inhibited the migration of AGS cells at 0, 24, and 48 h after scratch wounding. G An in vivo lymph node metastasis assay was performed to evaluate the effect of LacZ- and RPRD1B-transfected cells and Scr- and shRPRD1B-transfected AGS cells on tumor metastasis (left panel). Representative images of H&E-stained lymph node metastases after the footpad injection of the indicated cells are shown (right panel). Scale bar, 200 μm. H Representative lymph node metastases formed in nude mice injected with the indicated cells. The number of metastatic lymph nodes is summarized (n = 7 mice/group). Data are presented as the means ± SD of three independent experiments. (***, P < 0.001)