Figure 3.
Effects of acitretin on caspase activation. (A) Time course of caspases-8, -9, -3 and PARP cleavage in acitretin-treated SCL-1 cells. Cells were cultured with 10−5 M acitretin for the indicated times, and then the cells were harvested. Twenty micrograms cell extracts were resolved by 7.5% SDS-polyacrylamide denaturing gels and analysed by Western blotting with specific antibodies for activation of caspases-9 and -3 and cleavage of the caspase-3 substrate PARP. Immunoblotting of the same membranes with antibodies to β-actin was used to control for loading. The results represent one of three independent experiments, which yielded similar results. (B) Cells were treated for 48 hrs with 10−5 M acitretin alone or the caspases-8 and -9 inhibitors (see ‘Materials and methods’) respectively, or the combinations of acitretin with each of the above caspase inhibitors. The SCL-1 cells were harvested and analysed for apoptosis by the Cell Death Detection ELISA. Cells treated with DMSO were used as control. Data are shown as median of three independent cultures in one representative experiment. The experiment shown is representative of three independent experiments showing similar results. *P < 0.01 versus untreated cells. **P > 0.01 versus untreated cells. #P < 0.01 versus acitretin-treated cells. ##P > 0.01 versus acitretin-treated cells.