Fig. 4

Pan-caspase inhibitor Z-VAD-FMK and the cathepsin B inhibitor CA-074-Me inhibit Methylbenzoprim-induced apoptosis in melanoma cells. The percentage of apoptotic cells was evaluated after 24, 48 and 72 h of MBP (0.8, 8 μg/ml), the positive control Pyr (8 μg/ml) and untreated cells in Mel501 (panel a) and MeWo (panel b) cell lines pretreated for 2 h with the pan-caspase inhibitor Z-VAD-FMK (50 μmol/L) or the cathepsin B inhibitor CA-074-Me (10 μmol/L) or both. Columns, mean values of three independent experiments; bars, SD. *, P < 0.05; **, P < 0.01, *** P < 0.001 significance compared with Pyr and MBP treated cells. Z-VAD-FMK induced a protective effect in a time dependent manner in Mel501 and MeWo cell lines (panels a and b, respectively). When cathepsin B inhibitor CA-074-Me was used, apoptosis was significantly abrogated after 24 h treatment; this effect persisted after 48 h, while it was less evident after 72 h. Z-VAD-FMK and CA-074-Me, administered simultaneously, increased protective effect in both cell lines at all time points analyzed (panels a and b)