Fig. 2.

PRDX6 knockdown through transient knockdown system enhances ferroptotic cell death. a Western blot analysis of PRDX6 expression in H1299 after infection of lentivirus targeting to PRDX6 for 48 h. b After infection for 48 h, H1299 cells was treated with Erastin (2 μM) for 12 h, and LOOH production was assayed by flow cytometry using C11-BODIPY. Representative data are shown and the statistical analysis is from nine independent experiments and the bar indicates the mean values. c After infection for 48 h, H1299 cells were treated with RSL-3 (50 nM) or sorafenib (20 μM) for 12 h, and LOOH production was assayed by flow cytometry using C11-BODIPY. The statistical analysis is from three independent experiments and the bar indicates the mean values. d After infection for 48 h, H1299 cells were treated for 24 h with Erastin (2 μM) with or without ferrostatin-1(Fer-1) (10 μM), and cell viability was assayed by SRB assay. The statistical analysis is from three independent experiments and the bar indicates the mean values. e After infection for 48 h, H1299 cells were treated with Erastin (2 μM) with or without Fer-1 (10 μM), necrostatin-1 (30 μM) and Z-VAD-FMK (10 μM) for 12 h, and LOOH production was assayed. The statistical analysis is from three independent experiments and the bar indicates the mean values. f Western blot analysis of PRDX6 overexpression in H1299 cells with transfection of PRDX6 cDNA. g After gene transfection for 24 h, H1299 cells are treated with Erastin (0, 1, 2, 4, and 8 μM) for 24 h, and cell viability was assayed by SRB assay. The statistical analysis is from three independent experiments and the bar indicates the mean values. h, i After gene transfection for 24 h, H1299 cells are treated with Erastin (8 μM) or RSL-3 (100 nM) for 12 h, and LOOH production was assayed. Representative data are shown and the statistical analysis is from three independent experiments and the bar indicates the mean values. (*P < 0.05; **P < 0.01; ***P < 0.001)