Figure 8.

ADV activated the intrinsic apoptosis pathway in freshly stimulated, and pre-activated T cells. Resting human T cells were freshly stimulated (A–C) or pre-activated for 48 h (D–F) with anti-CD3/CD28 antibodies and cultured with increasing concentrations of ADV. Cells were stained after 72 h with active caspase 9-binding FITC-LEHD-FMK reagent for flow cytometric analysis. Representative histograms (B, E) and mean ± SEM percentages of caspase 9 positive cells of n = 4 independent experiments are presented (A, D). Cells were stained after 72 h with CellEvent™ Caspase-3/7 Green detection reagent for flow cytometric analysis. Mean ± SEM percentage of Caspase-3/7–positive cells are shown (C, F). (G) For kinetic analyses human T cells were stimulated with anti-CD3/CD28 antibodies and cultured with increasing concentrations of ADV in the presence of Caspase-3/7 reagent and CytoTox Red. Kinetic measures of the number of caspase 3/7 positive cells (left) or CytoTox Red positive cells (right) were recorded by the IncuCyte S3 imaging system at 3 h intervals for 70 h. Data are presented as mean ± SEM from n = 3 independent experiments. Statistical analysis was performed with One-Way ANOVA and Dunnett’s Multiple Comparison Analysis Test as post hoc test. (****P ≤ 0.0001, ***P ≤ 0.001, **P ≤ 0.01, *P ≤ 0.05).