Colony formation assay, BrdU incorporation, and Ki67 staining in HCT-116 and Panc-1 cancer cells. (A) Colony formation assay in HCT116 cells in the absence and presence of siRNA directed against Bnip3Δex3 isoform subjected to hypoxia, transfected with HA-Bnip3, or treated with doxorubicin (10 µM DOX), respectively. (B) Histogram represents quantitative data for A. Data were obtained from at least n = 3–4 independent experiments for each condition tested. Data are expressed as mean ± SD (error bars). *, statistically different from each other. (C) Quantitative data for BrdU incorporation in Panc-1 cells in the absence and presence of siRNA directed against Bnip3Δex3 isoform subjected to hypoxia, transfected with HA-Bnip3, or treated with doxorubicin (10 µM DOX), respectively. Data were obtained from three independent cell cultures for each condition tested. *, statistically different from each other; **, statistically different from CTRL; †, not statistically different from each other. (D) Epifluorescence microscopy of Panc-1 cells stained for Ki67 protein (green) and Hoechst 33258 nuclear dye (blue). Cells were subjected to hypoxia, transfected with HA-Bnip3, or treated with doxorubicin (10 µM DOX) in the absence and presence of siRNA directed against Bnip3Δex3 isoform. Bars, 10 µm.