Fig. 7: USP24 promotes autophagy-dependent ferroptosis in HCC cells.

A USP24 overexpressed SMMC-7721 and HCCLM3 cells were treated with Fer-1 (2 μM), Nec-1 (1 μM) and z-VAD-fmk (10 μM) separately. The cell viability was determined by CCK8 assay (n = 3). B Erastin (20 μM) was used to induce ferroptosis for 24 h, simultaneously treated with Fer-1 (2 μM), Nec-1 (1 μM) and z-VAD-fmk (10 μM). Cell viability was determined by CCK8 assay (n = 3). C Intracellular Fe²⁺ was detected with the FerroOrange probe (1 μM) under an inverted fluorescence microscope. Scale bar, 100 μm. D The Fe²⁺ fluorescence intensity was quantified by ImageJ (n = 3). E The intracellular Fe²⁺ was measured by colorimetric method (n = 3). F–G The mRNA and protein expression of NCOA4, TfR, FPN, ferritin and GPX4 after overexpressing USP24 was measured by qRT-PCR and WB (n = 3). H The level of GSH in HCC cells transfected with LV-CON or LV-USP24 vectors (n = 3). I The level of MDA in HCC cells transfected with LV-CON or LV-USP24 vectors (n = 3). J Control and USP24 overexpressed SMMC-7721 cells were treated with Sorafenib (0, 5, 10, 15, 20 and 25 μM) for 48 h, the cell viabilities were determined by CCK8 assay (n = 3). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.