Bleomycin initiates apoptosis of lung epithelial cells by ROS but not by Fas/FasL pathway.

Epithelial cells are considered to be a main target of bleomycin-induced lung injury, which leads to fibrosis in vivo. We studied the characteristics of in vitro bleomycin-induced apoptosis in a mouse lung epithelial (MLE) cell line. Bleomycin caused an increase of reactive oxygen species (ROS) resulting in oxidative stress, mitochondrial leakage, and apoptosis. These were associated with elevated caspase-8 and resultant caspase-9 activity and with upregulation of Fas expression. Glutathione and inhibitors of caspase-8 or caspase-9, but not of FasL, inhibited these effects, suggesting their dependence on ROS, caspase-8 and -9, in a Fas/FasL-independent pathway. However, postbleomycin-exposed MLE cells were more sensitive to Fas-mediated apoptosis. These results demonstrate that the initial bleomycin-induced oxidative stress causes a direct apoptotic effect in lung epithelial cells involving a regulatory role of caspase-8 on caspase-9. Fas represents an amplification mechanism, and not a direct trigger of bleomycin-induced epithelial cell apoptosis.