FIGURE 4.

Activation of Tim-1 on the surface of T cells directly delivers a positive signal that triggers Ca²⁺/calmodulin/NF-AT pathway. A, Naive CD4⁺CD25⁻ T were labeled with fluo-3-AM and IgG2a or agonist anti-Tim-1 mAb was added at the time indicated by (↓). Data are representative of three consecutive experiments yielding similar results. B, Naive CD4⁺CD25⁻ T were cultured with mature syngeneic BM-DC and anti-Tim-1 mAb in the presence or in the absence of cyclosporine (100 ng/ml). Tritiated thymidine incorporation (cpm) was measured after 48 h of culture. Bars represent the mean ± SEM of one of three consecutive experiments yielding similar results. C, Naive CD4⁺CD25⁻ T were cultured with mature syngeneic BM-DC and anti-Tim-1 mAb or isotype control mAb in the presence or absence of cyclosporine (100 ng/ml) for 21 h. NF-AT activity was measured from the nuclear extract of the various experimental settings. Bars represent the mean ± SEM of one of three consecutive experiments yielding similar results. *, p < 0.05 (Mann-Whitney U test). D, Naive CD4⁺CD25⁻ T were cultured with mature allogeneic BM-DC in the presence of IgG2a isotype control mAb or anti-Tim-1 mAb (5 μg/ml), and in the absence or the presence of cyclosporine (100 ng/ml), for 2 days. Expression of IL-2 transcripts was then quantified by quantitative RT-PCR. Bars represent the mean ± SEM of three consecutive experiments.