FIGURE 3.
A-C, ELISPOT analysis of Th1 (IFN- γ (A) and Th2 (B and C) cytokine profile of splenocytes derived from transplant recipients treated with CTLA4Ig 2-3 wk after PDL1 blockade. In the CTLA4Ig treatment group, the frequency of alloreactive IFN-γ-producing splenocytes was significantly decreased as compared with the untreated control recipients (A). Anti-PDL1-mAb administration during the induction phase resulted in significant increase of the frequency of alloreactive IFN-γ-producing splenocytes (A). CTLA4Ig treatment produced a slight but nonsignificant decrease in the frequency of IL-4-producing cells as compared with controls, whereas administration of anti-PDL1-mAb significantly increased the IL-4-producing T cell frequency (B). Interestingly, CTLA4Ig treatment significantly increased in the frequency of IL-5-producing cells as compared with controls (C), and anti-PDL1 mAb further increased the IL-5-producing T cell frequency as compared with the CTLA4Ig-treatment group (C). Data are representative of three independent experiments and indicate the mean of triplicate results in each experiment. D and E, Frequency of alloreactive CD8 effector T cells in splenocytes derived from transplant recipients treated with CTLA4Ig 2-3 wk after PDL1 blockade. The frequency of effector CD8+ T cells (expressing a CD62LlowCD44high phenotype) was significantly decreased in the CTLA4Ig treatment group as compared with controls (3.6 ± 1.0% vs 23.7 ± 0.4%, p < 0.001). Anti-PDL1 mAb significantly increased the frequency of effector CD8+ T cells, as compared with the CTLA4Ig treatment group (12.1 ± 2.5% vs 3.6 ± 1.0%, p < 0.001). A representative experiment is shown (D).