Fig. 4.

Memory cells exposed to tolerance signals respond to further reactivation but fail to accumulate. B6 mice immunized with 3K with/without LPS i.v. as indicated were injected with 3K-OVA and alum i.p. 40 d after Imm2. After an additional 4, 6, or 9 d, the numbers of splenic IA^b/3K tetramer⁺ cells were determined (A). Each point shows the mean of a total of 7–27 mice from seven separate experiments; error bars are SEM. The x axis is set at the limit of detection as determined by staining spleen cells from naive animals. On days 3–6 after Imm3, these mice were given drinking water containing BrdU and the percentages of CD44^hi IA^b/3K tetramer⁺ cells that were BrdU⁺ in each mouse were determined 6 d later (B). Each point represents one mouse and the line shows the mean of the group. Alternatively, the levels of expression of PD1 (C), ICOS (D), active caspase 3 and 7 (E), and Bcl2 (F) were determined in the antigen-specific CD4 T cells. In all experiments, data are representative of one to three independent experiments with three to four mice per group per time point.