Conditioned medium from actinomycin D-treated apoptotic cells induces mitochondria-dependent apoptosis in bystander cells

Abstract

Chemical-induced bystander effects have been known for several years, but the underlying mechanism is still seldom investigated. Previous researchers have found that mitomycin C and phleomycin induced micronuclei in bystander cells the same as in exposed cells. We previously demonstrated the ability of actinomycin D (ACTD) to induce bystander effects in normal Chinese hamster fibroblast V79 cells and found that conditioned medium (CM) obtained from ACTD-exposed apoptotic cells induced apoptosis in bystander cells. The present study further explores the probable mechanism of apoptosis in bystander cells. The main findings of this study are: (1) ACTD-treated CM induced apoptosis in bystander cells in a time-dependent manner, which was confirmed with morphological changes. (2) ACTD-treated CM increased the mRNA and protein levels of pro-apoptotic p53 and Bax, whereas it decreased those of anti-apoptotic Bcl-2 in bystander cells; these were all time-dependent effects. Reactive oxygen species (ROS) were also involved in apoptosis of bystander cells. (3) ACTD-treated CM reduced mitochondria membrane potential and induced cytochrome c release. (4) ACTD-treated CM induced G1 cell phase arrest, which may be another response in bystander cells when cultured with CM. These results suggest that chemical-treated CM induces p53-Bcl-2/Bax-cytochrome c signaling (i.e., mitochondria pathway)-dependent apoptosis in bystander cells, which is a kinetic response.