Differential effects of CD4⁺ and CD8⁺ cells on lymphocyte development from human cord blood cells in murine fetal thymus explants

The possibility that mature lymphocytes play a role in the regulation of human T cell development was studied in the experimental model of fetal thymus organ cultures (FTOC), by reconstituting lymphocyte-depleted murine fetal thymus (FT) lobe with cells isolated from human umbilical cord blood (CB). Cultures were incubated with human cytokines (IL-7, FLT-3 ligand and Steel Factor), or remained untreated. When CD4⁺, or CD8⁺ CB cells, were co- cultured with FT explants, they expanded and maintained their original phenotypic markers, with no significant effect of the cytokines. Cultures of human hematopoietic stem cells (CD34⁺) gave rise to CD4⁺CD8^- cells, which were mainly CD3^-, with no indication of further intermediate developmental stages. However, a limited number of CD4⁺CD8⁺ (double positive [DP]) cells were detected when the CD34⁺ cells were co-cultured with CD4⁺ cells from the same CB samples. In contrast, FT with unseparated CB cells resulted in the different CD4/CD8 subsets, and their numbers increased in the presence of cytokines. The appearance of DP cells depended on the presence of either CD4⁺ or CD8⁺ cells in the cultured CB samples. Hence, DP cells were not detected when the CB was depleted of CD4⁺ and CD8⁺ cells ('depCB') before culture, and they appeared when depCB were co-cultured with either CD4⁺ or CD8⁺ cells. In contrast, CD4⁺ cells inhibited the development of CD8⁺CD3⁺ cells, and this was most pronounced in the absence of the cytokines. There was no symmetrical down-regulatory effect of CD8⁺ cells on the development of CD4⁺CD3⁺ cells. Addition of IL-15 to the cytokine mixture led to an increased proportion of CD56⁺ cells in cultures of CD34⁺ cells. The presence of CD4⁺, and not CD8⁺ cells, interfered with this process. Our results thus imply differential effects of CD4⁺ and CD8⁺ cells on thymocytopoiesis.