Cytokine-induced NK-cell proliferation is suppressed by allogeneic Treg cells. (A) Healthy donor purified NK cells were CSFE labeled and cultured alone or with UCB-derived Tregs at a ratio of 1:1. Suppression of proliferation of NK cells incubated with IL-2 (0.5 ng/mL), IL-15 (0.5 ng/mL), or a combination of IL-2 + IL-15 (0.25 ng/mL) was measured. Shown is 1 representative donor of 6 experiments. (B) Healthy donor NK cells and UCB-derived Tregs were coincubated with 0.5 ng/mL of IL-2 or IL-15 and cocultured at various Treg:NK cell ratios. Percent suppression of NK proliferation by Tregs was evaluated by CFSE dilution. Treg suppression of CD3 bead-stimulated PBMNC effector T-cell proliferation was measured as a control. Data are an aggregate of 5 separate experiments. NK cells and Tregs were coincubated with IL-2 or IL-15 for 4 days, and (C) IL-2 or (D) IL-15 in the supernatant was measured by enzyme-linked immunosorbent assay. Various cytokine concentrations (0.2, 0.5, and 10 ng/mL) and Treg:NK cell ratios (1:1, 2:1, 4:1, 8:1) were compared. Results from 5 NK cells donors are shown (mean and SEM). *P = .05 compared with NK cells alone.