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. 2020 Sep 6;236(4):2725–2739. doi: 10.1002/jcp.30039

Figure 3.

Figure 3

Cell death of annulus fibrosus cells (AFCs) and nucleus pulposus cells (NPCs) induced by tert‐butyl hydroperoxide (TBHP) was inhibited and was similar to RSL3‐induced AFCs and NPCs ferroptosis. AFCs and NPCs were treated with 100 μM TBHP with or without the specific inhibitors deferoxamine mesylate (DFO; 10 μM) or ferrostatin‐1 (Fer‐1; 5 μM) or were exposed to RSL3 at different concentration (0, 10, 15, 20 μM). (a) Western blotting assay of nuclear receptor coactivator 4 (NCOA4), glutathione peroxidase 4 (GPX4), ferritin heavy chain (FTH), and GAPDH in AFCs. (b,c) The difference in GPX4, NCOA4, and FTH expression in AFCs under different treatments. (d) Lipid peroxidation level under different treatment conditions of AFCs. (e) Immunofluorescence detection of FTH in AFCs. (f) Western blotting assay of NCOA4, GPX4, FTH, and GAPDH in NPCs. (g,h) The difference in GPX4, NCOA4, and FTH expression in NPCs under different treatments. (i) Lipid peroxidation level under different treatment conditions of NPCs. (j) Immunofluorescence detection of FTH in NPCs. ***p < .001, **p < .01, *p < .05 (n = 3)