Characteristics of 5-HT–induced apoptosis in group 1 BL cells.

(left) (A) L3055 cells transfected with either bcl-2,bcl-x_L, or control vector were treated with 125 μM 5-HT and DNA synthesis (expressed as mean % control ± SEM of 3 experiments) was assessed exactly as for Figure 1A. (B) After 24 hours of culture, with control medium [(i), (iii)] or 125 μM 5-HT [(ii), (iv)], L3055 bcl-2-transfectants [(i), (ii)], and L3055 vector control cells [(iii), (iv)] were stained with the fluorescent nuclear dye acridine orange and were visualized by microscopy. Results are representative of 3 experiments. (right) L3055 cells were treated with control medium or with 150 μM 5-HT as indicated. (C) Cultured for 24 hours, stained with the cationic dye JC-1, and visualized by confocal microscopy. (D, E) Cultured for 6 hours, then stained with a fluorescent probe for active caspases (FAM-VAD-FMK). (D) Results obtained by flow cytometry (% cells active caspase positive indicated in top right corner). (E) Confocal microscopy analysis of cells counterstained with the Hoechst nuclear dye (blue), with PI to detect dead and membrane-compromised cells (red); activated caspases are stained green. Results are representative of 4 experiments.