Human Mesenchymal Cells Regulates Naive and Memory T Regulatory Cells.

CD4(+)CD25(+)FoxP3(+) T regulatory (T reg) cells constitute 1–2% of peripheral blood cells in adults and function prevalently as immunosuppressors. 70% of T reg are memory/effector cells with a CD45RO+ phenotype. The other 30% have a naive T cell phenotype (CD45RA+). Both sub-populations exert a regulatory function and when sorted and/or purified each inhibits a mixed lymphocyte culture. In the present study, we investigated the effects of human recombinant IL-7 (rIL-7), human mesenchymal cells (hMSC) and hMSC engineered with the IL-7 gene (hMSC/IL-7) on regulatory T cells expressing a memory (CD4/CD25/CD45RO) or naive (CD4/CD25/CD45RA) phenotype. T cells from healthy donors were enriched by immnuselection to provide populations of CD45RA+ cells (95 % ± 2.9) and CD45RO+ cells (97 % ± 0.25). Enriched naive and memory cells were cultured in presence rIL-7 (100 ng/ml), hMSC (ratio 5:1) or hMSC/IL-7 (ratio 5:1). After 7 days’ culture we examined the CD4/CD25+ cells within the naive and memory populations. In the naive T cell population the T reg starting fraction of 0.05 % ± 0.01 of CD4/CD25 positive cells, did not change in the presence of rIL-7 while it rose to 0.2 % ± 0.14 in presence of human MSC and more interestingly reached 1.67 % ± 0.6 in presence of IL-7 engineered human MSC. In the memory T cell population the T reg starting fraction of 0.3 % ± 0.05 of CD4/CD25 positive cells, did not change in presence of rIL-7 while it rose 1.5 % ± 0.9 in the presence of human MSC and more interestingly reached 11.55 % ± 7.5 in the presence of human IL-7 engineered-MSC. We analyzed CD127 expression within different groups. In the naive T reg starting fraction 3 % ± 1.2 expressed CD127 which was down-regulated to 0.96 % ± 0.5 in the presence of rIL-7, to 0.29 % ± 0.2 with human MSC and to 0.37 % ± 0.02 with human IL-7engineered-MSC. Memory T reg cells expressed CD127 in 15% ± 1.2 of the starting fraction which was down-regulated to 1.2 % ± 0.45 in the presence of rIL-7, to 1.32 % ± 0.34 with hMSC and to 4.01% ± 0.74 in presence of hMSC/IL-7. FoxP3 expression was measured by real time quantitative PCR in sort-purified subsets of peripheral blood, identified by staining with a combination of CD4, CD25, CD45RA or CD45RO. In naive T reg FoxP3 expression was increased 1.15 fold in the presence of hMSC and 2.7 fold in presence of hMSC/IL-7. In memory T reg FoxP3 expression was increased 1.14 fold in the presence of hMSC and 2.67 fold in presence of hMSC/IL-7. In conclusion naive T reg cells are IL-7 independent and up-regulated by human MSC. Engineering human MSC with the IL-7 gene enhanced up-regulation. Memory T reg cells are also IL-7 independent and are up-regulated by human MSC. Engineering human MSC with the IL-7 gene markedly increased up-regulation. The different regulatory systems may underlie different functions within the T reg sub-populations.