Figure 4. CD8⁺ ACAID Tregs need to respond to IFN-γ to express their regulatory function.

(A) CD8⁺ T cells were isolated from WT or C57BL/6 IFN-γR^−/− mice, which were injected with nonadherent BALB/c spleen cells in the AC. DTH-positive control mice received CD4⁺ effectors from immunized mice. C57BL/6 APCs pulsed with BALB/c alloantigens in vitro were added to all cell mixtures. This experiment was performed twice with similar results; n = 5 mice/group/experiment; *P < 0.001. (B) CD8⁺ T cells were isolated from WT, IFN-γR^−/−, or IFN-γ^−/− mice, which were primed in the AC with nonadherent BALB/c splenocytes on Day 0 and immunized s.c. with nonadherent BALB/c splenocytes 7 days later. Positive control mice were not primed in the AC but were immunized s.c. with nonadherent BALB/c splenocytes. CD4⁺ T cells and CD8⁺ T cells were isolated from naïve, negative control mice and immunized mice 7 days following s.c. immunizations and coinjected with APCs pulsed with BALB/c alloantigens into the ears of naïve C57BL/6 mice. Specific ear swelling was assessed 24 h later. Results are expressed as mean ± sem. This experiment was conducted twice with five mice/group/experiment; *P = 0.018. Differences in the means between the IFN-γ^−/− CD8 and SC CD4 groups were not significantly different (P>0.05).