Effect of TZD18 on apoptosis of tested cell lines. (A) Apoptotic cells in BV173 after treatment with PGZ (20 μM) or TZD18 (20 μM) were measured by TUNEL assay, as described in “Materials and methods.” (B) SD1 cells were cultured in the presence of TZD18 (10 or 20 μM) for different hours, washed, and lysed. Apoptosis was measured by cell death ELISA, as described in “Materials and methods.” Results are expressed as the fold change of enrichment of nucleosomes in the cytoplasm of cells treated with drugs compared with controls (without treatment). Data represent the mean ± SD of triplicate experiments. (C) Cells were cultured in the presence of TZD18 (20 μM) for 4 days, washed, and lysed in lysis buffer. Caspase 8 and caspase 9 activities in the cell lysates were measured as described in “Materials and methods.” Results were expressed as the fold increase of OD values compared with control (without treatment). Figure is representative of the results of 3 independent experiments. (D) Cells were incubated in the presence of TZD18 (20 μM), Z-VAD-FMK (20 μM), or both for 4 days. Cell apoptosis was examined as described in “Materials and methods.” Apoptosis was expressed as fold change in enrichment of nucleosomes in the cytoplasm of cells treated with drugs compared with the sample with the lowest value (cells treated with Z-VAD-FMK). Results are the mean ± SD of 3 individual experiments.