Association of CD133 with NRF2 upregulation in HCT116 spheres. (A) Transcript levels of NRF2, GCLC, AKR1C1, and NQO1 were measured in monolayer and sphere HCT116 cells using a relative qRT-PCR analysis. aP < 0.05 compared with the monolayer group. (B) Protein levels of NRF2, GCLC, AKR1C1 and NQO1 were assessed in monolayer and sphere HCT116 cells. Bar graph represents quantified protein levels from at least three experiments. aP < 0.05 compared with the monolayer group. (C) HCT116 with either the non-transfection (NT), non-specific siRNA (siCtrl) or CD133-specific siRNA (siCD133) transfection were grown in the sphere culture system, and transcript levels of CD133, GCLC, AKR1C1, NQO1, and NRF2 were measured. aP < 0.05 compared with the monolayer. bP < 0.05 compared with the siCtrl group. (D) Protein levels for NRF2, GCLC, AKR1C1, and NQO1 were monitored in sphere cultured HCT116 cells with either the non-specific siRNA (siCtrl) or CD133-specific siRNA (siCD133) transfection. Bar graph represents quantified protein levels from at least three experiments. Quantification results of western blotting were relative values to the loading control GAPDH. aP < 0.05 compared with the monolayer. bP < 0.05 compared with the siCtrl group. NT, non-transfection group.