Impaired cytokine production of NK cells after interaction with MSCs. After culture for 6 days with IL-2 in the presence or in the absence of MSCs, NK cells were cocultured with FO-1 melanoma cell line (as stimulator) at an E/T ratio of 8:1. After 5 hours of culture in the presence of 100 U/mL IL-2 and GolgiStop, cells were collected. Both surface and intracytoplasmic stainings were performed with anti–CD56-PC5 and anti–IFN-γ-PE mAbs, respectively. Then, NK cells were analyzed by flow cytometry by acquiring 5 × 10⁴ events in the lymphocyte region. (A) Unstimulated NK cells represent negative control. (B,C) NK cells, cultured alone or in the presence of MSCs, respectively, stimulated with FO-1 cells. Numbers indicate percentages of double (CD56 and IFN-γ) positive cells. (D) Means plus or minus SD of percentages of IFN-γ–positive NK cells (upon interaction with the NK-susceptible FO-1 cells) obtained in 6 independent experiments performed using different NK-cell populations after culture in IL-2 either in the presence or in the absence of MSCs. **P < .01.