CD34⁺ HPC-derived LCs provide a more potent costimulatory cytokine environment through endogenous IL-15 for CTL activation than do moDCs. (A) Cytokine-generated CD34⁺ HPC-derived LCs or moDCs were pulsed with fluMP and recultured with purified autologous T cells already primed against fluMP (responder/stimulator ratio = 10:1). Cytokine or receptor blocking conditions are shown above each panel. Stimulation was stopped after 30 minutes at 37°C, and the proportion of T cells that had phosphorylated STAT5 was measured by flow cytometry. One representative experiment of 3 is shown. (B) Using the gating for pSTAT5 in panel A, the means from 3 independent experiments ± SEM are shown after stimulation by LCs or moDCs alone (P = .03; unpaired Student t test). (C) Again, using the pSTAT5 gating in panel A, the change in percentage of pSTAT5⁺ fluMP-reactive T cells, stimulated by either LCs or moDCs in the presence of IL-15 or anti–IL-15R-α, relative to stimulation by LCs or moDCs alone, is shown (n = 3 independent experiments each, mean ± SEM, P = .034 for the supplemental IL-15 condition, and P = .023 for the anti–IL-15R-α condition, comparing LCs vs moDCs by the unpaired Student t test). (D) The mean percentage ± SEM of antigen-nonspecific, rested conA T lymphoblasts that expressed pSTAT5 after restimulation by autologous LCs (n = 4 independent experiments) or 50% or 100% volume/volume LC-free supernatants (n = 5 independent experiments) was determined by cytofluororaphy using the same gating strategy as in panel A. The paired t test yielded P < .02 for the intact LC stimulation versus either concentration of LC-free supernatants, which were not significantly different from the negative control, T cells alone. ns indicates not significant.