Tolerogenic potential of human thymic pDCs. (A) Ex vivo-isolated immature thymic pDCs (IpDCs) or (B) pDCs activated in response to CD40L plus IL-3 (MpDCs) were cocultured with CFSE-labeled autologous (Auto) or allogeneic (Allo) CD4⁺ SP thymocytes (CD4SP) or allogeneic PB CD4⁺ T lymphocytes (Allo CD4PB). CD25 expression of total cells (left black histograms) and CFSE expression of either total or electronically gated CD25⁺ and CD25⁻ cells (middle empty histograms) was analyzed by flow cytometry after 7 days. The CFSE profile of total cells cultured in the absence of DCs is shown in black. Numbers correspond to total percentages of CD25⁺ cells. Analysis of CD25 versus Foxp3 expression of total cultured cells is shown on the right. Numbers correspond to percentages of CD25⁺Foxp3⁺ cells. Background staining was defined with isotype-matched Abs. Results of 1 of 3 experiments are shown. (C) Relative numbers of CD25⁺Foxp3⁺ cells recovered from cocultures of either allo CD4SP thymocytes or allo CD4PB lymphocytes with MpDCs. Data are mean ± SEM for 3 independent experiments. *P < .05. (D) Suppressive function of CD25^hi cells derived from CD4PB lymphocytes primed with MpDCs. Naive peripheral CD4⁺ T lymphocytes stimulated with anti-CD3 and anti-CD28 mAbs plus IL-2 (20 ng/mL) were cultured in the absence (control) or presence of magnetically sorted CD25^hi or CD25^lo/− cells (upper right histograms). Data are mean ± SEM of [³H]-thymidine incorporation at day 5 of triplicate cultures (cpm) of 1 of 3 independent experiments. **P < .01.