Figure 2

Different γ-irradiation regimes induce different modalities of cell death and senescence in breast cancer cell lines. (A) Induction of apoptosis, necrosis, and senescence. Breast cancer cell lines were left untreated or γ-irradiated at single doses of 2 Gy, 20 Gy, or daily fractions of 2 Gy, respectively. Induction of apoptosis and necrosis was determined 1–4 days after irradiation by annexin V-FITC/PI staining and FACS analysis. Annexin V-FITC positive, PI negative cells were considered apoptotic, double positive cells were considered necrotic. Senescence induction was measured by flow cytometric SA-β-gal staining with the fluorogenic substrate C12-FDG-FITC. Cells with high C12-FDG-FITC and high SSC signal were considered senescent. Means ± s.d. of triplicates are depicted. (B) Induction of apoptosis and necrosis in the presence of zVAD-fmk. PS externalization and plasma membrane integrity were measured as in (A) in the presence of 50 μM of the poly-caspase inhibitor zVAD-fmk. Means ± s.d. of triplicates are shown. (C) Induction of apoptosis and necrosis in the presence of necrostatin-1. PS externalization and plasma membrane integrity were measured as in (A) in the presence of 50 μM of the necroptosis inhibitor necrostatin-1. Means ± s.d. of triplicates are shown. (D) Representative dot plots of HCC1937 cells stained for SA-β-gal activity with the fluorogenic substrate C12-FDG-FITC.