Induction of apoptosis and DEVDase activation by the liposomal ether-lipid ELL-12 in Jurkat, H9, and U-937 cells. (A) Jurkat T cells were treated in serum-free medium for 5 hours (solid squares and solid bars) or in the presence of serum for 24 hours (open squares and open bars) with the indicated concentrations of ELL-12 or with non–ET-18-OCH₃ liposomes at a concentration equivalent to the highest concentration of ELL-12 (lp) or 50 ng/mL anti-Fas antibody. The percentage of apoptotic cells was evaluated by the DNA-specific fluorochrome Hoechst. Nuclei were visualized by fluorescence microscopy and a minimum of 1,000 cells were scored. Mean values ± SD from at least 3 different experiments are shown. (B) Jurkat T cells were treated with 10 μg/mL ELL-12 for the indicated times and the percentage of apoptotic nuclei (solid squares) was determined as described above. DEVDase activity (open squares) in extracts from duplicate cultures was measured with the fluorogenic substrate Ac-DEVD-AMC. Results are the means ± SD of at least 3 independent experiments. (C) H9 cells (solid squares) and U-937 cells (open squares) were treated in serum-free medium for 16 hours with the indicated doses of ELL-12 or with non–ET-18-OCH₃ liposomes at a concentration equivalent to the highest dose of ELL-12. Cells were then stained with the DNA-specific fluorochrome Hoechst and nuclei were visualized by fluorescence microscopy. Apoptosis of cells incubated with non–ET-18-OCH₃ liposomes was 11.5% in H9 cells and 5.6% in U-937 cells. Data shown are representative of 3 independent experiments. DEVDase activity in extracts from H9 (D) and U-937 cells (E) treated for the indicated times with 10 μg/mL (open squares) or 50 μg/mL ELL-12 (solid squares) was measured with the fluorogenic substrate Ac-DEVD-AMC. Results are the means ± SD of at least 3 independent experiments.