Induction of lymphocyte proliferation in primary and secondary allogeneic cultures. (A) The allostimulatory activity of DCs obtained in the different cytokine combinations was tested in primary cultures, using monocyte-depleted PBMCs. Proliferating lymphocytes were identified in density plots as CD4⁺/BrdU⁺ double-positive cells. (B) In primary cultures, IL-16-DCs and TPO-DCs showed higher allostimulatory activity than control SCF-DCs, whereas IL-16/TPO-DCs were unable to stimulate the proliferation of allogeneic T lymphocytes. The mean ± SEM from 9 independent experiments is shown. IL-16-DCs, TPO-DCs, and IL-16/TPO-DCs compared with SCF-DCs with the paired Wilcoxon signed rank test. (C) Tested in a 2-step culture system, IL-16/TPO-DCs induced tolerance of cocultured T cells. Purified naive T cells were cultured alone (left column) or cocultured with either SCF-DCs (center column) or IL-16/TPO-DCs (right column). After 5 days, the T cells were harvested, washed, and rested for 2 days. The rested T cells from primary cultures were subsequently rechallenged with SCF-DCs. Proliferation of T lymphocytes was assessed as BrdU incorporation after 4 days. One representative of 3 independent experiments is shown.