Fig. 8. ROS production resulting from STAT1 activation contributes to necroptosis.

(A) BMDMs were pretreated with BHA (100 µM) and/or zVAD (20 µM) for 30 min prior to LPS (1 µg/ml) or poly I:C (20 µg/ml) treatment. After incubation for 24 h, the cell viability was assessed by MTT assay. (B) Cells were pretreated with zVAD, LPS (1 µg/ml) and/or poly I:C for different periods, and then intracellular ROS was measured by DCFH₂-DA fluorescence. (C) Similar experiments as (B) were conducted in cells treated with Nec-1 (10 µM), AZD1480 (1 µM) or IFNR-Ab (1 µg/ml). veh, vehicle. (D) BMDMs from WT and STAT1 KO mice were treated with zVAD and/or LPS, and ROS levels at 8 h and 18 h were measured. Data were the mean ± SEM from at least three independent experiments. *P < 0.05, indicating enhancement effects of zVAD on cytotoxicity and ROS production. ^#P < 0.05, indicating reversal effects of BHA, Nec-1, AZD1480, IFNR-Ab, and STAT1 knockout on zVAD-induced cytotoxicity and ROS production.