Objectives : It has been reported that the trichothecene mycotoxin 4-acetyl-12,13-epoxyl-9-trichothecene-3,15-diol (AETD) from the fruiting bodies of Isaria japonica Yasuda is a potent inducer of apoptosis in human promyelocytic HL-60 cells. Methods : The present study aims to characterize the molecular events leading to AETD-induced apoptosis in HL-60 cells. Results : The percentage of apoptotic cells (annexin V-positive cell population) increased dose- and time-dependently after AETD exposure. Apoptosis of HL-60 cells by AETD was associated with the formation of intracellular reactive oxygen species (ROS), the depletion of intracellular glutathione (GSH), and the activation of caspase-3. Pre-treating the cells with the antioxidant N-acetyl-L- cystein (NAC) and the caspase-3 inhibitor Z-DEVD-fmk abrogated AETD-induced apoptosis and caspase-3 activation. NAC blocked intracellular ROS formation and GSH depletion, but Z-DEVD-fmk did not. Conclusions : These results indicate that AETD induces apoptosis in HL-60 cells by causing intracellular ROS formation and GSH depletion followed by the downstream event of caspase-3 activation.