Involvement of mitochondria and ROS generation in edelfosine-induced cell death in Leishmania parasites and cancer cells.

(A) L. panamensis promastigotes were untreated (Control) or treated with 10 μM edelfosine at the indicated times, and cells with disrupted ΔΨm (DiOC6(3)low) and ROS production (HE→Eth) were measured by flow cytometry. The numbers in each quadrant refer to the percentages of cells in each population. (B) L. panamensis promastigotes untreated (Control) and treated with edelfosine (EDLF) for 3 h were incubated with 2 μM HE and 10 μg/ml Hoechst 33342, and then analyzed by fluorescence microscopy. (C) L. panamensis promastigotes and (D) Jurkat cells were preincubated with 10 μg/ml CsA for 1 h, or with 10 mM NAC or 10 mM GSH for 2 h, and then incubated in the absence or presence of 10 μM edelfosine for 9 h. After treatment, the percentage of hypodiploid cells was analyzed by flow cytometry. Untreated control cells were run in parallel. (E) L. panamensis promastigotes and (F) Jurkat cells were preincubated with 10 μM rotenone, 5 mM malonate, 10 μM antimycin A, 1.5 mM azide, 50 μM CCCP or with 1 and 10 μM oligomycin (L. panamensis and Jurkat cells, respectively) for 1 h, and then incubated in the absence or presence of 10 μM edelfosine for 9 h. After treatment, cells producing ROS were quantified by flow cytometry. Untreated control cells were run in parallel. Data shown are means ± SD or representative of three independent experiments performed. Asterisks denote that the differences between the indicated groups (C and D) and with control cells (E and F) are statistically significant. (*) P<0.05. (**) P<0.01.