Figure 7. In vivo IL-12 neutralization during vaccination generates defective effector memory but not central memory CTLs.

All mice were treated with Abs against IL-12p40 or an isotype control during CPS vaccination. A–C) To confirm the effect of IL-12 neutralization, PECs and spleens were analyzed for the activation of tgd057-specific CTLs on day 8 post-vaccination in IL-12p40 treated (Rx) or isotype Rx mice. CD44^hi K^b/SVLAFRRL+ CD8α+ TCRβ+ cells were enumerated in A, and the subpopulation distributions of K^b/SVLAFRRL+ CD8α+ cells were given in B. Samples were restimulated with CPS parasites and the frequency of IFN-γ+ cells in total K^b/SVLAFRRL+ CD8α+ cells is shown in C. D and E) Resting numbers and subpopulation distributions of tgd057-specific CTLs were analyzed in IL-12p40 Rx or isotype Rx immune rested mice as in A and B. F) Immune rested mice were in vivo rechallenged with 2×10⁶ CPS parasites for 12 hrs. PECs from IL-12p40 Rx or isotype Rx rechallenged immune mice were harvested and ex vivo stained for IFN-γ and granzyme B expression in total CD8α+ TCRβ+ cells. FACS plots are from one of three mice per group. G–I) Immune rested mice were in vivo rechallenged with 10⁷ CPS parasites. On day 6 post-rechallenge, PECs and spleens were analyzed for the secondary activation of tgd057-specific CTLs. G and H) Resting numbers and subpopulation distributions of tgd057-specific CTLs were analyzed in IL-12p40 Rx or isotype Rx immune rested mice as in A and B. I) IFN-γ production was assessed in K^b/SVLAFRRL+ CD8α+ cells after CPS restimulation. PEC samples were pools of 3–4 mice. For unpooled samples, bar graphs are shown as mean ± SEM of 3–4 mice and pie charts are mean values of 3–4 mice. One of two representative experiments is shown.