MoDCs, LCs, and DDC-IDCs are equally capable of activating NK cells via IL-2 secretion by DC-activated alloreactive CD4⁺ T cells. (A) Proliferation of PBMCs was assessed by ³H-thymidine incorporation (y-axis; note log scale) after culture with monocyte-derived dendritic cells (moDC; ▾) or CD34⁺ HPC-derived Langerhans cells (LC; •) or dermal-interstitial dendritic cells (DDC-IDC; ○) for 6 days at the responder-stimulator ratios (10:1 to 1000:1) indicated along the X-axis. Data points represent the average of duplicate microwell cultures and their standard deviation. The data are representative of 3 independent experiments. (B) Proliferation of purified peripheral blood NK cells was assessed by ³H-thymidine incorporation (Y-axis; note log scale) after culture with monocyte-derived dendritic cells (moDC) or CD34⁺ HPC-derived Langerhans cells (LC) or dermal-interstitial dendritic cells (DDC-IDC) for 6 days. Data points for the indicated NK-cell-dendritic cell ratio (X-axis; NK/DC from 3:1 to 1000:1) represent the average of duplicate microwell cultures and their standard deviation. NK-cell/DC cultures were supplemented with 80 IU/mL rIL-2 (DC + IL-2 ▾), irradiated allogeneic CD4⁺ T cells (DC + CD4; NK/T-cell ratio = 1:1 ○), or nothing (DC only •). The data are representative of 3 independent experiments. (C) LCs, DDC-IDCs, and moDCs were cocultured with peripheral blood NK cells for 7 days at an NK-cell-DC ratio of 10:1. These primary cultures were supplemented with 80 IU/mL rIL-2 (DC + IL-2 ▾), irradiated allogeneic CD4⁺ T cells (DC + CD4; NK/T-cell ratio = 1:1 ○), or nothing (DC only •). NK cytotoxicity of the cultures was determined by ⁵¹Cr release assay against the MHC class I-negative target LCL721.221. Radiolabeled LCL721.221 targets were exposed for 4 to 6 hours to the NK-cell effectors, which were the responder lymphocytes from the original or primary DC/NK cocultures. The effectors were added as total lymphocytes, without adjustment for specific numbers of NK-cell blasts, to ascertain differences between the different DCs in stimulating NK cells in the primary cultures. The effector-target ratio is indicated along the X-axis, and specific lysis is plotted against the Y-axis. The data are representative of 3 experiments.