(A) iNKT cells adoptively transferred into PR8-infected Jα18^–/– mice reduced the total number of lung MDSCs. PR8-infected WT, Jα18^–/–, and CD1d^–/– mice were injected with iNKT cells. Lung homogenates were stained with CD11b and Gr-1 Abs. (B) Injection of sublethal doses of PR8 (3 × 10² PFU) reduces the total numbers of lung MDSCs. (C) Adoptive transfer of iNKT cells reduces in vitro suppressive activity of PR8-induced MDSCs. MDSCs were purified from lungs of PR8-infected mice and cocultured with CFSE-labeled OT-I splenocytes. (D) The suppressive activity of MDSCs from PR8-infected mice is reduced by treatment with anti–CD40 agonist Ab and NOS2 and ARG1 inhibitors. Lung-purified MDSCs were left untreated (black bars) or treated in vitro with either anti–CD40 agonist Ab (white bars) or with N^G-monomethyl-l-arginine (L-NMMA) and NOHA (light gray bars) and then added to OT-I splenocytes. Proliferation of unpulsed OT-I splenocytes in the absence of MDSCs is shown (dark gray bars). (E) Adoptive transfer of MDSCs from infected mice suppresses the expansion of UTY_246–254–specific CD8⁺ T cell responses. UTY_246–254 CTL responses were assessed in vaccinated mice injected with MDSCs from PR8-infected mice (see Methods). H-2D^b/UTY_246–254 tetramer (UTY_246–254D^b tetramer) versus CD8 dot plots are shown for gated propidium iodide–negative lymphocytes, and the percentage of cells staining positively with the tetramers is indicated. In A–E, data represent the average ± SD of n = 5 mice/group, and results of statistical analyses, performed using Student’s t test, are shown.