Figure 4.

Donor CD8⁺ T cells are rapidly recruited to the T cell areas of recipient spleens, in direct proximity to host CD11c⁺ DCs. CFSE-labeled CD8⁺ T cells (2 × 10⁶) derived from C3H.SW mice (CD45.2⁺) were intravenously injected into lethally irradiated B6/SJL mice (CD45.1⁺). (a) Aggregation of CD11c⁺ DCs in the T cell areas of the spleens of irradiated mice. Cryosections of spleen harvested from lethally irradiated B6 mice 6 hours after TBI were immunostained with anti-CD11c Ab (brown) and anti-CD4 Ab (gray-black). Magnification: top row, ×100; bottom row, ×400. Data are representative of three experiments. (b) Spleens were taken from B6 mice 6 hours after PBS injection and from irradiated B6 mice 6 hours after infusion of CFSE-labeled C3H.SW CD8⁺ T cells (green) and cryosectioned for immunofluorescent staining with anti-CD11c Ab (red). Magnification, ×100. (c) Splenocytes were prepared and counted at the indicated timepoints. Cells were sequentially labeled with Cychrome-conjugated anti-CD8 Ab coupled with biotinylated anti-CD45.2 and streptavidin-conjugated PE. The percentage of CD8⁺CD45.2⁺ cells was obtained by flow cytometry analysis. The recruitment rate (percentage) of donor CD8⁺ T cells in spleen was calculated by dividing the absolute number of donor CD8⁺ T cells recovered from the spleens of irradiated recipient mice by the total number of intravenously injected donor CD8⁺ T cells.