Fig. 2

RRM2 suppresses ferroptosis in liver cancer cells. a, b RRM2 protein (a) and mRNA levels (b) in HepG2 and SMMC-7721 cells with or without RRM2 overexpression or knockdown, as analyzed by immunoblotting and qPCR, respectively. c–e Cell viability (c), cell death (d) and 4-HNE levels (e) were measured in HepG2 and SMMC-7721 cells with ectopically expressed or knocked down RRM2 before further treatment with Fer-1, ZVAD-FMK, Nec-1 or ectopically expressed RRM2. Cell viability was measured using a CellTiter-Glo luminescent cell viability assay, cell death was measured by staining with SYTOX Green followed by flow cytometry, and 4-HNE was measured by a kit from Abcam. f, g Cell death (f) and 4-HNE levels (g) were measured in HepG2 and SMMC-7721 cells treated with erastin (10 μM, 24 h) in the presence or absence of Fer-1 (1 μM, 24 h). The cells were also cultured with or without RRM2^T33E transfection, as indicated. h–j The levels of GSH (h), labile iron (i) and membrane-anchored phospholipids (j) in HepG2 and SMMC-7721 cells with or without RRM2 overexpression or knockdown were analyzed. The data are shown as the mean ± SD from three biological replicates (including IB). *P < 0.05, **P < 0.01 indicates statistical significance. Data from b−j were analyzed using a one-way ANOVA test