Figure 9

Effect of a pan-caspase inhibitor (Z-VAD(OMe)-FMK) in the apoptosis induced by ruthenium(II) complexes with 6-methyl-2-thiouracil in HL-60 cells determined by flow cytometry using annexin V-FITC/PI staining. (A) Representative flow cytometric dot plots showing the percentage of cells in viable (annexin V-FITC negative and PI negative cells), early apoptotic (annexin V-FITC positive, but PI negative cells), late apoptotic (annexin V-FITC positive and PI positive cells) and necrotic stages (PI positive, but annexin V-FITC negative cells). (B) Quantification of apoptotic HL-60 cells (annexin V-FITC positive cells). Cells were pre-treated for 2 h with 50 µM Z-VAD(OMe)-FMK, then incubated with the complexes at 2 μM for 24 h. Negative control (CTL) was treated with vehicle (0.2% DMSO) used for diluting the complexes, and doxorubicin (DOX, 1 µM) was used as positive control. Data are presented as mean ± S.E.M. of at least three independent experiments performed in duplicate. Ten thousand events were evaluated per experiment, and cellular debris was omitted from analysis. *p < 0.05 compared with negative control by ANOVA followed by Student Newman-Keuls test. ^#p < 0.05 compared with respective treatment without inhibitor by ANOVA followed by Student Newman-Keuls test.