Fig. 7: Resistance of SK-Hep1 ρ⁰ cells against sorafenib-induced ferroptosis.

A SK-Hep1 ρ⁺ and ρ⁰ cells were treated with 10 μM sorafenib for the indicated periods, and cell death was determined using LDH release assay. B SK-Hep1 ρ⁺ and ρ⁰ cells were treated with 10 μM sorafenib in the presence or absence of 2.5 µM Fer-1, 0.1 µM Lip-1, 10 µM ZIL, 50 µM DFO, 50 µM Trolox or 50 µM z-VAD for 72 h. Cell death was determined using an LDH release assay. C, D In SK-Hep1 ρ⁺ and ρ⁰ cells treated with 10 μM sorafenib in the presence or absence of Fer-1 for 48 h, accumulation of cellular ROS (C) and lipid peroxides (D) was analyzed by flow cytometry after staining with CM-H2DCFDA and BODIPY-C11, respectively (right). Representative histograms are shown (left). E, F In SK-Hep1 ρ⁺ and ρ⁰ cells treated with 10 μM sorafenib in the presence or absence of 2.5 μM Fer-1 for 48 h, accumulation of mitochondrial ROS (E) and mitochondrial lipid peroxides (F) was determined by flow cytometry after staining with MitoSOX and MitoPerOx, respectively (right). Representative histograms are shown (left). G, H In SK-Hep1 ρ⁺ and ρ⁰ cells treated with 10 μM sorafenib in the presence or absence of 100 μM MitoTEMPO or 500 nM MitoQ for 48 h, accumulation of mitochondrial ROS (G) and mitochondrial lipid peroxides (H) was determined by flow cytometry after staining with MitoSOX and MitoPerOx, respectively (right). Representative histograms are shown (left). I After treatment of SK-Hep1 ρ⁺ cells with 10 μM sorafenib in the presence or absence of 100 μM MitoTEMPO or 500 nM MitoQ, cell death was evaluated using LDH release assay. J After treatment of SK-Hep1 ρ⁺ cells with 10 μM sorafenib in the presence or absence of varying doses of DecylQ or MitoQ, cell death was evaluated using LDH release assay. Data represent means ± SD from three independent experiments. Data were analyzed by one-way ANOVA with Tukey’s multiple comparison test (A–I except ρ⁺ and ρ⁰ cell comparisons) or two-tailed unpaired Student’s t test (ρ⁺ and ρ⁰ cell comparisons in A, J). (*p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001; ns, not significant).