Fig. 4

Oxygen-dependent ferroptosis in cerebral reperfusion injury. a LPO of the hippocampus and cortex of mice were assayed following MCAO/R for 24 h. Data are means ± SEM, n = 5 animals per group. Two-way ANOVA with post hoc Sidak test was performed. b MDAs of the hippocampus and cortex of mice were assayed following MCAO/R for 24 h. Data are means ± SEM, n = 5 animals per group. Two-way ANOVA with post-hoc Sidak test was performed. c MDA was detected in the hippocampus of mice treated with RSL3 following MCAO/R for 24 h. Data are means ± SEM, n = 5 animals per group. One-way ANOVA with post-hoc Tukey test was performed. d Neurological score (higher numbers indicating more severe impairment) was performed at 0, 6, and 24 h after MCAO/R. Data are means ± SEM. Sham, n = 5; MCAO + DMSO, n = 7; MCAO + RSL3, n = 5. Two-way ANOVA with post-hoc Tukey test was performed. e Representative 2,3,5-triphenyl tetrazolium chloride (TTC)-stained serial brain sections of mice 24 h after MCAO/R, where viable tissue stains red. Quantification of infarction volume indicated by TTC staining using Image J. Data are means ± SEM. Sham, n = 5; MCAO + DMSO, n = 7; MCAO + RSL3, n = 5. One-way ANOVA with post-hoc Tukey test was performed. f Neurological score was performed at 1, 6, and 24 h after permanent-MCAO. Data are means ± SEM, n = 6 animals per group. Two-way ANOVA with post-hoc Tukey test was performed. g Representative TTC-stained serial brain sections of mice 24 h after permanent-MCAO, where viable tissue stains red. Quantification of infarction volume indicated by TTC staining using Image J. Data are means ± SEM, n = 6 animals per group. One-way ANOVA with post hoc Tukey test was performed. h Neuronal cell viability after OGD 2 h/reoxygenation 18 h. Data are means ± SEM, n = 10 wells from one representative of 4 independent experiments. t test was performed. i Detection of intracellular Fe²⁺ in N27 cells using FerroOrange at 2 h of OGD. Data are means ± SEM, n = 6 wells from one representative of 3 independent experiments. t test was performed. j Lipid ROS in N27 cells treated with OGD for 2 h (representative histogram plot for fluorescence of oxidized BODIPY-C11). k Relative lipid ROS is expressed as the ratio of oxidized to reduced BODIPY-C11 mean fluorescence intensity (MFI) in N27 cells treated with OGD for 2 h. Data are means ± SEM, n = 6 wells from one representative of 3 independent experiments. t test was performed. l Cell viability of N27 cells 24 h after OGD and Liproxstatin-1 (Lip-1) of concentration gradient co-treatment. Data are means ± SEM, n = 6 wells from one representative of 3 independent experiments