B7-H1 transmits an antiapoptotic signal to tumor cells. (A) B7-H1/P815 were cultured in the wells, which were precoated with either control IgG (Ctl Ig) or murine PD-1Ig (PD-1Ig) at 5 μg/mL for 18 hours. After extensive washing, cells were labeled with ⁵¹Cr and further incubated with activated PD-1⁰2C CTLs at indicated E/T ratios for 4 hours. CTL activity was determined in a ⁵¹Cr release assay. Each point is the mean of triplicates with SD. The data are representative of at least 3 experiments. (B) PD-1 stimulation of B7-H1⁺ tumor cells does not change the expression of Fas. The expression of Fas on mock/P815 or B7-H1/P815 cells before and after culture with immobilized murine PD-1Ig was determined by antimurine Fas mAb in flow cytometric analysis as described above. (C) PD-1 stimulation reduced the susceptibility of tumor cells to Fas mAb-mediated apoptosis. Mock/P815 or B7-H1/P815 cells were cultured in the presence of immobilized control IgG or murine PD-1Ig (5 μg/mL) for 18 hours. After extensive washing, cells were then transferred to the wells coated with anti-mouse Fas antibody (5 μg/mL). After 48 hours of culture, the cells were subjected to an annexin-V and PI binding assay. Mock/P815 without treatment was served as negative control (control). (D) PD-1 stimulation reduced the susceptibility of tumor cells to STP-induced apoptosis. B7-H1/P815 cells were cultured in the presence of immobilized control IgG or murine PD-1Ig (5 μg/mL) for 18 hours. After extensive washing, cells were treated with STP at 0.25 μM for 8 hours and subjected to an annexin-V binding assay. The numbers in the graph indicate the percentage of annexin-V⁺ or annexin-V⁺ and PI⁺ cells.