[摘要]：Background. Controversy exists about the conditions effecting the development of forkhead/winghead helix transcription factor P3 (FOXP3) expressing T cells and their relevance in transplant recipients.Methods. We generated carboxy-fluorescein diacetate succinimidyl ester-labeled CD4(+)CD25(high) FOXP3(+) cells in mixed lymphocyte reactions (MLRs) ("the Treg MLR"), with varying human leukocyte antigen (HLA) disparities and cell components. Five color flow cytometry and H-3-thymidine uptakes were the readouts.Results. (1) Despite lower stimulation indices (SIs) than two DR-mismatched MLRs, 2 DR-matched MLRs generated more than twofold higher percentages when gating on proliferating CD4(+)CD25(high) FOXP3(+) cells; (2) Even with low numbers of proliferating cells, autologous and HLA identical MLRs generated the highest FOXP3(+):FOXP3(-) cell ratios; (3) Elimination of either non-CD3(+) responding cells (resulting in "direct presentation" only) or responding CD25(+) (Treg generating) cells increased the SI but inhibited proliferating CD4(+)CD25(high) FOXP3(+) cell development; (4) MLR-generated CD4(+)CD25(high) FOXP3(+) cells added as third components specifically inhibited the same freshly set MLR SI and caused recruitment of new CD4(+)CD25(high) FOXP3(+) cells. As an example of the "Treg MLR" immune monitoring potential, addition of third component peripheral blood mononuclear cell containing high percentages of CD4(+)CD25(high) FOXP3(+) cells from an HLA identical kidney transplant recipient (in a tolerance protocol) caused donor-specific Treg MLR inhibition or recruitment. This was similar to the third component MLR Tregs generated entirely in vitro.Conclusion. In the Treg MLR, the generation of CD4(+)CD25(high) FOXP3(+) cells is more pronounced in the context of self-recognition (HLA matching, indirect presentation). These cells can be assayed for MLR inhibitory and Treg recruitment functions, so as to immunologically monitor the allospecific regulation after transplantation.
