FIG. 4.

Overexpression of the active form of Akt overcomes the cytotoxic effect of VP-16. (A and B) HT1080 cells were transfected with the pcDNA3-EGFP plasmid together with the empty vector pUSEamp (Mock) or the pUSEamp vector encoding the active form of Akt (Myr-Akt). After transfection for 24 h, HT1080 cells were treated with 10 μg of VP-16 per ml for an additional 36 h. After fixation, cells were stained with DAPI. Nuclear fragmentation was visualized with a fluorescence microscope. (B) The number of surviving cells was determined by counting 100 EGFP-positive (transfected) cells that did not show nuclear condensation and fragmentation. Each vertical bar represents the mean ± the standard deviation of three independent experiments. (C) A549 cells were treated with the indicated concentrations of VP-16 in the presence (solid columns) or absence (open columns) of 20 μM LY294002 for 48 h. (D) 293T cells were treated with the indicated concentrations of ADR in the presence (solid columns) or absence (open columns) of 20 μM LY294002 for 48 h. The apoptotic sub-G₁ fraction of the population was determined by FACScan analysis as described in Materials and Methods.