Figure 4.

ADV induced DNA double-strand breaks in resting, freshly stimulated, and pre-activated T cells. Resting human T cells were incubated with increasing concentrations of ADV (A, B) or Cladribine (C, D) for 8 h. The levels of DNA double-strand breaks were assessed by γH2AX immunofluorescence staining and automated foci quantification. Representative microscopic images of γH2AX foci (green), DAPI nuclear staining (blue) (A, C) and γH2AX foci quantification (B, D) are shown. Each bar represents the mean ± SEM of n = 3 independent experiments. Resting human T cells were freshly stimulated (E, G) or pre-activated for 48 h (F, H) with anti-CD3/CD28 antibodies and cultured with increasing concentrations of ADV for additional 24 h, respectively. (E, F) Representative dot plots of flow cytometric analysis of γ-H2AX and propidium iodide staining of DNA content are shown. (G, H) Percentages of γ-H2AX positive cells (mean + SEM) are depicted for 3 independent experiments. Statistical analysis was performed by 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).