Figure 5.

CD56^brightCD16^- NK_reg cell induction of CD4⁺ T-cell apoptosis. Apoptotic CD4⁺ T cells after the 96-hour suppression assay co-culture were identified with use of Annexin V and 7-AAD flow cytometric staining. Conditions tested included (A) activated CD4⁺ T cells in the absence of suppressor cells (control), (B) CD56^brightCD16^- regulatory NK (NK_reg) cells, and (C) CD56^dimCD16⁺ NK cells co-cultured with allogeneic CD4⁺ T cells at the 1:1 ratio. With use of FSC-A and SSC-A, lymphocytes were gated and using CD4 and 7-AAD staining live and dead CD4⁺ T cells were gated (plots not shown). After, Annexin V and 7-AAD were plotted. For the histogram, Annexin V-negative and 7-AAD-negative (live) CD4⁺ T cells are shown in blue, and Annexin V-positive, 7-AAD-negative (early apoptotic) and annexin V-positive, 7-AAD-positive (late apoptotic) CD4⁺ T cells are shown in purple. (D) The percentage of apoptotic CD4⁺ T cells in the CD4⁺ T-cell control as compared to the NK_reg and NK^dim conditions are shown (mean +/- standard deviation). NS: not significant. Statistical analyses were performed using Microsoft Excel version 2110 and a two-tailed t test – two-sample assuming unequal variance. The data is from a single experiment representative of 3 experiments.