Ferroptosis inhibitors prevent cell death and ROS accumulation induced by HS in Arabidopsis roots. 6-d-old seedlings were preincubated with 1 µM Fer-1, 10 µM CPX, or DMSO (Control) as indicated. Cell death was induced by treating roots at 55°C for 10 min. (a) Kinetics of cell death induced by a 55°C HS. (b) Kinetics of ROS levels induced after a 55°C HS treatment. ROS accumulation in roots was detected by using the H2DCFDA probe. (c) ROS production is mediated by NOX activity and prevented by cotreatment with ferroptosis inhibitors or cotreatment with the NOX inhibitor DPI. Data are the mean ± SEM of three independent experiments. (d) Representative confocal images showing ROS detected by the H2DCFDA probe in A. thaliana roots. Bars, 50 µm. (e) Suppression of cell death after a 55°C HS by D4-linoleate (D4 Lin). 6-d-old seedlings were preincubated with 1 µM Fer-1, 10 µM CPX, 8 µM D4-linoleate, or DMSO (control) as indicated. Cell death was induced by treating roots at 55°C or 77°C for 10 min. (a and e) Root hairs were stained with Sytox green, and Sytox-positive (interpreted as dead cells) and Sytox-negative cells were quantified. Results are expressed as a percentage of dead cells. Bars with different letters denote statistical difference (one-way analysis of variance, P < 0.05).
