Naïve CD8+ T cells (1 × 105 cells/well) labeled with CFSE (Invitrogen; labeled according to the manufacturer’s protocol) were stimulated with CD3/CD28 Human T cell-Activator Dynabeads (5 μl/ml; Invitrogen) in the presence or absence of MDSCs (0.25 × 105 cells/well) and/or IL18/IFNα-activated NK cells (0.25 × 105 cells/well) in 96-well plates. As an alternative method for CD8+ T cell expansion, CFSE-labeled naïve CD8+ T cells (1 × 105 cells/well) were stimulated with staphylococcal enterotoxin B-pulsed mature monocyte-derived DCs (1 × 104 cells/well; matured for 48 h with 50 ng/ml TNFα), as previously described (18 (link)). When indicated, cells were co-cultured in the additional presence of small-molecule inhibitors or blocking antibodies against suppressive factors. On day 4–6, expanded CD8+ T cells were analyzed for proliferation via CFSE dilution and intracellular granzyme B expression. The following inhibitors/blocking antibodies were used in this study: celecoxib (20 μM; BioVision), 1-methyl-DL-tryptophan (1 mM; Sigma-Aldrich), L-NMMA (200 μM; Cayman Chemical), IL10 mAb (clone 25209; 1 μg/ml; R&D Systems), and nor-NOHA (200 μM; Cayman Chemical). The concentrations used did not affect viability in cell cultures, as confirmed by live cell counts.