Opposing effects of IL-2 and TGF-β1 on NKG2D cell-surface expression. (A) NKG2D cell-surface expression in primary human NK cells. Cells were left untreated (medium) or were stimulated with IL-2, TGF-β1, or IL-2, and TGF-β1 for either 1 or 3 days and the surface expression of NKG2D was analyzed by flow cytometry. Gray-filled histograms represent isotype control staining while black line histograms show staining with anti-NKG2D antibody from a representative experiment (left panel). Summary of the data from 22 donors is shown on the right (note: primary NK cells are not viable if maintained for 3 days in medium plus TGF-β1 alone). (B) NKG2D cell-surface expression by NKL cells. NKL cells were cultured without IL-2 for 3 days, then stimulated as described in panel A and NKG2D expression was analyzed by flow cytometry. The left panel shows a representative experiment; gray-filled histograms represent isotype control staining, black line histograms show staining with anti-NKG2D antibody. Summary of the cell-surface expression of NKG2D on day 1 and day 3, after treatment with IL-2, TGF-β1, or IL-2 plus TGF-β1, is shown on the right; the data are from 5 independent experiments. The numeric values in the histograms indicate the median fluorescence intensity of NKG2D cell surface expression. Graphs in panels A and B represent the average values of median fluorescence intensity ± SEM. Asterisks indicate statistical significance (paired Student t test; *P < .05; **P < .01; ***P < .005).