Time requirements of T cell help and the role of T<sub>regs</sub> cells.

<p>(<b>A</b>) 2×10<sup>4</sup> CD8<sup>+</sup> T cells were transferred into CD3ε<sup>−/−</sup> hosts. At 8 or 29 days after groups of hosts received CD4 T cells (2×10<sup>4</sup> cells) and the mice were sacrificed 7 weeks after the first cell transfer. The mean CD8<sup>+</sup> T cell recovery is shown for the three groups of hosts (***<i>p</i>≤0.001). It should be pointed out that the late transfer of CD4 T cells still induced the preferential accumulation of CD62L<sup>low</sup>CD8<sup>+</sup> T<sub>EM</sub> cells (not shown) (<b>B</b>) 2×10<sup>4</sup> CD8<sup>+</sup> T cells were transferred alone or with CD4<sup>+</sup> T cells from Lat-DTR mice into a CD3ε<sup>−/−</sup> host. A fraction of the mice were treated with diphtheria toxin (DT) for 10 days at either one or 4 weeks after cell transfer, and the mice were analyzed 7 weeks after cell transfer. The number of CD8<sup>+</sup> T cells recovered 7 weeks after cell transfer is shown. CD4<sup>+</sup> T cell depletion at week 1 or at week 4 reduced the number of CD8<sup>+</sup> T cells recovered (**<i>p</i>≤0.01). Similar findings were obtained in two independent experiments. Dot plots show representative examples of the CD62L and CD44 expression by the CD8<sup>+</sup> T cells recovered in the different groups of mice. (<b>C</b>) 10<sup>4</sup> CD8<sup>+</sup> T cells were transferred into CD3ε<sup>−/−</sup> hosts alone or with 5×10<sup>4</sup> CD4 T<sub>reg</sub> cells. The number of CD8<sup>+</sup> T cells recovered in individual mice is given as the mean value for each group. (<b>D</b>) 10<sup>4</sup> CD8<sup>+</sup> T cells were transferred into CD3ε<sup>−/−</sup> immune-deficient hosts alone, co-transferred with 10<sup>4</sup> CD4<sup>naive</sup> T cells or co-transferred with 10<sup>4</sup> CD4<sup>naive</sup> T cells and 5×10<sup>4</sup> CD4 T<sub>reg</sub> cells in three independent groups of hosts. The number of CD8<sup>+</sup> T cells recovered in individual mice is given as the mean value for each group. The increased expansion of CD8<sup>+</sup> T cells seen upon co-transfer with CD4<sup>naive</sup> T cells was abolished when the T<sub>reg</sub> cells were added in the transferred mix. (**<i>p</i>≤0.01) (<b>E</b>) 10<sup>5</sup> total CD8<sup>+</sup> T cells were transferred into CD3ε<sup>−/−</sup> hosts alone, co-transferred with 10<sup>4</sup> CD4<sup>naive</sup> T cells or co-transferred with 10<sup>4</sup> CD4 T cells and then given 5×10<sup>4</sup> T<sub>reg</sub> cells four weeks latter. Mice were culled 8 weeks after the initial CD8<sup>+</sup> T cell transfer. The data indicate the number of CD8<sup>+</sup> T cells recovered in each host. Statistically significant differences are shown (**<i>p</i><0.01). Similar findings were obtained in two independent experiments.</p>

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