Article Figures & Data
Figures
- FIGURE 1.
LLT1 is a ligand for the human CD161 receptor. A, 293T cells either untransfected or stably transfected with pIRES2-EGFP-CD161 or -LLT1 were stained with control hIgG- and LLT1-multimers. B, The binding of LLT1-multimer to 293T-CD161 transfectants was blocked specifically by anti-CD161 mAb DX12 (10 μg/ml). C, 293T-CD161 (squares) or 293T-LLT1 (circles) transfectants were stained with LLT1-multimer in the presence of increasing concentrations of anti-CD161 DX12 (filled symbols) mAb or anti-CD94 (open symbols) mAb as a control. Results are shown as mean ± SEM of three experiments.
- FIGURE 2.
LLT1 interaction with CD161 results in a down-regulation of CD161. A, Polyclonal NK cells fixed or not before incubation with C1R and C1R-LLT1 cells were monitored for cell surface expression of CD161. B, Polyclonal NK cells were incubated for 4 h with 293T or 293T-LLT1 cells and CD161 cell surface expression was monitored. The mean fluorescence intensities obtained after exposure to control 293T cells were arbitrarily set as 100%. Results are shown as mean ± SEM of three experiments.
- FIGURE 3.
LLT1 interaction with CD161 inhibits NK cell-mediated cytotoxicity. A, Monitoring of CD107a expression on NK cells unstimulated or stimulated with C1R and C1R-LLT1 for 5 h. The percentage of NK cells expressing CD107a is indicated. B, C1R and C1R-LLT1 were used as targets in a 4-h 51Cr release assay with polyclonal CD161+ NK cells. Blocking mAb and isotype control mAb were added at 10 μg/ml. C, Redirected killing assay using polyclonal NK cells and P815 cells transfected or not with LLT1. Suboptimal concentration of anti-CD16 mAb (0.5 μg/ml) was added to anti-CD161 (191B8) or isotype control (10 μg/ml) mAbs. Data are representative of four to eight experiments.
- FIGURE 4.
LLT1 binding to CD161 inhibits IFN-γ production by NK cells. Polyclonal NK cells were stimulated for 4 h with C1R and C1R-LLT1 in the presence of brefeldin A (10 μg/ml) and rIL-2 (200 U/ml). The percentage of NK cells secreting IFN-γ is indicated. A, Incubation with target cells only or with PMA (5 ng/ml) and ionomycin (0.5 μg/ml). B, Incubation in the presence of increasing concentrations of blocking anti-CD161 (191B8) or isotype control mAbs. Data are representative of eight independent experiments.
- FIGURE 5.
Simultaneous ligation of CD161 and CD3 enhances IFN-γ production by T cells. A, Polyclonal T cells were incubated for 4 h with plate-bound anti-CD3 (500 ng/ml) alone or together with isotype control or anti-CD161 (191B8) (5 μg/ml) mAbs. B, Polyclonal T cells were stimulated with P815 transfected or not with LLT1 in the presence of anti-CD3 mAb (5 μg/ml). Dot plots represent the percentage of T cells secreting IFN-γ. Data are representative of two to four independent experiments.
