Figure 4. CTLA-4 coinhibitory signals control CD8⁺ memory T cell/secondary effector cytokine secreting function.

Thy1.1⁺ OT-I T cells (1 × 10⁴) were adoptively transferred into naive B6 Thy1.2 hosts and infected with Listeria-OVA to generate recipients containing memory OT-I T cells. On day 30 after infection, mice received an OVA-expressing skin graft and were treated with 200 μg CTLA-4 Ig, 100 μg anti-CD28 dAb, or 100 μg anti-CD28 dAb + 250 μg anti–CTLA-4 on days 0, 2, and 4 after transplant. Animals were sacrificed on day 5 after transplant, and splenocytes were harvested and restimulated in vitro with 10 nM SIINFEKL as described in Methods. (A) Representative flow cytometry plots of IFN-γ and IL-2 secretion from unstimulated (top row) and stimulated (bottom row) CD8⁺Thy1.1⁺ T cells. (B) Representative flow cytometry plots of IFN-γ and TNF secretion from unstimulated (top row) and stimulated (bottom row) CD8⁺Thy1.1⁺ T cells. (C–E) Frequencies (left) and absolute numbers (right) of (C) IFN-γ–, (D) IFN-γ ⁺IL-2⁺–, and (E) IFN-γ⁺TNF⁺–producing CD8⁺ Thy1.1⁺ T cells. Data are representative of 3 independent experiments with a total of 10–15 mice per group. *P < 0.05 by 1-way ANOVA. dAb, domain antibody.