CD56^bright and CD56^dim NK cell subsets exhibit memory-like properties after IL-12 + IL-18preactivation. (A-B) CD56^bright and CD56^dim NK subsets in purified NK cell (≥ 95% CD56⁺CD3⁻) cultures exhibit memory-like IFN-γ production. (A) Representative data from control and IL-12 + IL-18–preactivated NK cells demonstrating the NK subset analysis gating strategy and differential IFN-γ production after the 7-day rest and a 6-hour IL-12 + IL-15 restimulation. (B) Summary data demonstrating that IL-12 + IL-18–preactivated CD56^bright and CD56^dim NK cells exhibit a memory-like IFN-γ response after restimulation with IL-12 + IL-15, IL-12 + IL-18, or K562 leukemia cells. Purified (≥ 95% CD56⁺CD3⁻) NK cells were cultured as in Figure 1A for 7 days and restimulated with cytokines or K562 (4:1 effector: target ratio) as indicated for 6 hours and then assayed for IFN-γ. Data shown are the means ± SEM percentage of IFN-γ⁺ NK cells, gated on CD56^bright or CD56^dim subsets, with n = 12 donors (4 independent experiments) for IL-12 + IL-15 restimulation, n = 6 donors (3 independent experiments) for IL-12 + IL-18 restimulation, and n = 9 donors (4 independent experiments) for K562 restimulation. (C-D) Flow-sorted CD56^bright and CD56^dim NK cells cultures exhibit memory-like IFN-γ production. (C) NK cell subsets were flow sorted (≥ 99% purity) and then preactivated as per Figure 1A, rested for 7 days, restimulated (6 hours) with IL-12 + IL-15, and stained for intracellular IFN-γ. The NK cell subset phenotype was retained in each sorted population after 7 days of culture. (D) Summary data are shown as means ± SEM of the percentage of IFN-γ⁺ sorted CD56^bright and CD56^dim NK cells after 7 days of rest followed by 6 hours of restimulation with IL-12 + IL-15 (n = 6; 3 independent experiments). *P < .05; **P < .01; ***P < .001.