Fig. 3: Measurement of the intracellular levels of ROS and lipid peroxidation and determination of cell death due to ferroptosis following cisplatin treatment.

A–D A549 and A549/DDP cells in the logarithmic growth phase were seeded in 6-well plates and treated with cisplatin for 48 h. Subsequently, 10 μM DCFH-DA (A, B) or 2 μM C11-BODIPY581/591 (C, D) fluorescent probes were added to the medium in each well for 30 min to ensure coverage of the cell layer by the probe. Then, green fluorescence was observed via an immunofluorescence microscope (A–C), or the cells were collected to analyze the levels and differences in intracellular ROS and lipid peroxidation via flow cytometry (B–D). The error bars represent the standard deviation from three replicates. Analysis was performed via an unpaired t-test, with ****p < 0.0001 relative to the control or differently treated groups. E A549 and A549/DDP cells were seeded in 6-well plates, followed by stimulation with cisplatin (20 μM) alone or in combination with the ferroptosis inducer erastin (10 μM) or the iron chelator DFO (50 μM) for 48 h. Immunofluorescence staining with propidium iodide (PI) was performed to compare the differences in cell death levels after different treatments. F Flow cytometry was used to analyze the degree of cell death after different cisplatin treatments. The error bars represent the standard deviation from three replicates. Analysis was performed via an unpaired t-test, with ****p < 0.0001 relative to the control or differently treated groups.