Figure 2

Cell death induced by MiTMABs is blocked by the pan-caspase inhibitor, ZVAD. A-D, G₂/M synchronized HeLa cells were treated for 20 h with MiTMABs and controls (untreated, 0.1% DMSO and 2-(DiMA)EM) in the presence and absence of the pan-caspase inhibitor, ZVAD. DNA content in these cells was analysed by flow cytometry. Representative flow cytometry histograms of propidium iodide stained control treated (untreated and 0.1% DMSO) HeLa cells and HeLa cells treated with MiTMABs in the presence (filled histograms) and absence (empty histograms) of ZVAD are shown (A). The percentage of cells (mean ± S.D. from two independent experiments) with < 2N (DNA fragmentation, which is indicative of cells undergoing apoptosis; (B), 4N (C), and >4N (D) DNA content are shown. Treatment with MiTMABs + ZVAD caused a decline in cells containing <2N DNA and a corresponding increase in cells containing ≥ 4N DNA content compared to MiTMABs alone. * p < 0.05, ** p < 0.01 (Student's t tests). E, HeLa cells were treated as described in A except that after 8 h cells were fixed and stained for α-tubulin, and the percentage of cells that were multinucleated was scored using immunofluorescence microscopy. Graph shows mean ± S.D. from two independent experiments. F, Representative microscopy images of (E) illustrating multinucleated HeLa cells treated with either MiTMAB or OcTMAB in combination with ZVAD. Red, α-tubulin. DNA, blue.