Synergistic effect of SSZ and RT on ferroptosis induction in the DLD-1 cell line under hypoxic conditions. DLD-1 and HCT116 cells were treated with SSZ/Ferrostatin-1 (16 h) under hypoxic conditions (1%), and the induction of ferroptosis was determined. (A) The amount of lipid peroxidation after SSZ treatment was evaluated using the C11BODIPY dye in DLD-1 (top) and HCT116 (bottom) cell lines. (B) The amount of lipid peroxidation after SSZ treatment + RT (16Gy) 16 h after RT was determined in DLD-1 (top) and HCT116 (bottom) cell lines. (C) Mitochondrial membrane potential determination by a TMRE staining and flow cytometry was performed in DLD-1 (top) and HCT116 (bottom) cell lines. (D) The survival fraction after RT with 8Gy + SSZ and Ferrostatin-1 treatment was determined in DLD-1 (top) and HCT116 (bottom) cell lines. Data are represented as mean ± SEM, n ≥ 3. A two-way ANOVA followed by a Dunnett’s or Bonferroni multiple comparison test, or a one-way ANOVA followed by a Dunnett’s multiple comparison test, was performed for statistical analyses. * p < 0.05, ** p < 0.01.