Fig. 2

E2A suppressed the tumor-initiating capacity of CRC cells in vitro. a Effect of shE2A on SW480 cell sphere-formation ability. The sphere-formation ability of SW480 cells was enhanced by shE2A. P < 0.05, Scale bars: 50 μm. b Transfection of plasmids pEZ-M29-E12 and pEZ-M29-E47 to Caco-2 cells decreased the number of spheres per field. Left panel: Representative images of sphere formation assay are shown with original magnification of 40×. Scale bars: 50 μm. Right panel: Data in the histograms is expressed as means ± SD from three separate experiments. c Knockdown of E2A induced Lgr5 expression. Left panel: Immunoblot analysis showed that cancer stem-cell marker Lgr5 was increased in the shE2A-expressing SW480 clone, indicating induction of tumor-initiating ability. GAPDH was used as loading control. Right panel: Densitometric analysis of left panel normalized to GAPDH. d E12 and E47 suppressed Lgr5 expression. Left panel: Immunoblot analysis of Lgr5 and GAPDH expression with or without transfection of E12 or E47. Right panel: Densitometric analysis of left normalized to GAPDH. Data in the histograms is expressed as the means ± SD from three separate experiments. e Immunofluorescence analysis with anti-Lgr5 antibodies as indicated. Nuclei were counterstained with 4′, 6-diamidino-2-phenylindole (DAPI). Magnification: 400×; Scale: 50 μm. *, P < 0.05