Figure 4. DRE activates the death-receptor-mediated extrinsic pathway of apoptosis.

Following treatment with DRE, at indicated time points and concentrations, MV-4-11 cells were collected, washed and incubated with lysis buffer to obtain cell lysate. The cell lysate was incubated with caspase substrates, specific to each caspase (3,8 and 9) and incubated for an hour. Fluorescence readings were obtained using a spectrofluorometer. An average of 6 readings per well and a minimum of three wells were run per experiment. The results here are reported as activity per µg of protein (in fold) and the average of three experiments are shown (a,b). (c). Prior to DRE treatment, MV-4-11 cells were pre-treated with a pan-caspase inhibitor, Z-VAD-fmk for an hour and then treated with DRE at the indicated concentration for indicated time points. These cells were then incubated with caspase-8 substrate and fluorescence readings were obtained. These cells were also analyzed for the induction of apoptosis by Hoechst and annexin-V staining (d). Magnification: 400×.