Effect of mature (LPS-induced) TGF-M-DCs, in comparison with mature M-DCs, in regulation of T-cell proliferation and tolerance. (A) Mature TGF-M-DCs are similar to mature M-DCs in induction of allogeneic MLR, both at lower efficiency than mature GM-DCs in MLR. LPS-activated TGF-M-DCs, M-DCs, and GM-DCs were stimulated with allogeneic naive CD4⁺ T cells for 6 days. T-cell proliferation was determined by [methyl-³H]thymidine incorporation for the last 16 hours of cell culture. (B) TGF-M-DCs induce T-cell tolerance/regulatory T cells with similar efficiency to that of M-DCs. T1 and T2 are naive CD4⁺ T cells (donor A) prestimulated with mature TGF-M-DCs or M-DCs (donor B) for 7 days. T1 and T2 (2.5 × 10⁴) were restimulated with allogeneic GM-CSF/IL-4–induced DCs in secondary MLR, and both of T1 and T2 were hyporesponsive T cells (top panel). Also, T1 and T2 (2.5 × 10⁴) were added to another MLR containing naive CD4⁺ T cells (5 × 10⁴; donor A) and GM-CSF/IL-4–induced DCs (5 × 10³; donor B) (bottom panel). Proliferation of T cells was determined by [³H]thymidine incorporation after 6 days of culture. Results are representative of 3 independent experiments. (C) CB CD4⁺ T cells were cocultured with LPS-activated GM-DCs, M-DCs, and TGF-M-DCs derived from both CB and AB in MLR for 7 days. Cells were harvested and stained with anti–CD4-FITC, anti–CD25-PE, and intracellular anti–Foxp3-APC. The top row is CD4⁺ T cells isolated from CB stained with anti-CD4, anti-CD25, and isotype control IgG for APC. The percentages of gated cell population are shown on the upper-right corner of dot plots. Results are one representative of 3 CB and 6 AB.