Fig. 4. DNA breakdown in L. donovani promastigotes after H₂O₂ exposure and effects of a caspase inhibitor on DNA breakage. (A) Number of cells positive for TUNEL staining at various hours after H₂O₂ treatment. Symbols: d, 4 mM H₂O₂ treatment; j, control with no treatment. Data are mean ± s.e.m. of four experiments. (B) DNA profile in agarose gels from treated and untreated promastigotes. (a) Without H₂O₂ exposure. (b) After 2 hours of 4 mM H₂O₂ treatment. (c) After 4 hours of 4 mM H₂O₂ exposure. (d) After 6 hours of 4 mM H₂O₂ treatment. (C) DNA breakage and the number of TdT labelled cells after inhibition of caspase activity by Z-DEVD-FMK. Symbols: j, 4 mM H₂O₂; m, 4 mM H₂O₂ + 1 µM inhibitor; ., 4 mM H₂O₂ + 10 µM inhibitor; d, control without treatment. Inset: DNA gel showing (a) marker, (b) DNA of cells pretreated with 1 µM inhibitor prior to exposure to 4 mM H₂O₂ and (c) DNA of cells treated with 4 mM H₂O₂. Data are mean ± s.e.m. of four experiments. (D) TdT labelled cells at 6 hours under different treatments. (a) Phase contrast of the same field as (b). (b) TdT labelled cells in 4 mM H₂O₂-treated group. (c) Phase contrast of same field as (d). (d). TdT labelled cells in group pretreated with Z-DEVD-FMK (1 µM) prior to exposure to H₂O₂. (e) Phase contrast of (f). (f) TdT labelled cells in the control group. Results are representative of three experiments. Bar, 100 µm.