Figure 5.

Suppression by m-MDSCs is mediated by iNOS. (A–B) m-MDSCs purified from VV-infected mice were co-cultured with naïve CFSE-labeled CD4⁺ (CD4) or CD8⁺ (CD8) T cells at a 2:1 m-MDSC to T cell ratio (+m-MDSC), in the presence of 1 μg/ml of anti-CD3 and anti-CD28 antibodies (CD3/CD28). Some T cells were incubated with media only (Control). Where indicated, SOD, L-NMMA, or nor-NOHA was added to the culture. Proliferation was determined 72 h later. (A) FACS plots showing the percentage of CFSE^low cells representing proliferating T cells. (B) The mean percentages ± SD of CFSE^low cells among total CD4 or CD8 cells are shown (n = 4). (C–D) m-MDSCs were co-cultured with CFSE-labeled naïve HA-specific CD4⁺ (6.5) or CD8⁺ (Clone 4) T cells at a 1:2 m-MDSC to T cell ratio (m-MDSC:T cells) in the presence of HA-pulsed CD11c⁺ DCs (Pulsed). Some 6.5 and Clone 4 cells were incubated with control CD11c⁺ DCs (Unpulsed). Where indicated, SOD, L-NMMA, or nor-NOHA was added to the culture. Proliferation was assessed 72 h later. (C) FACS plots showing the percentage of CFSE^low cells. (D) The mean percentages ± SD of CFSE^low cells among total 6.5 or Clone 4 T cells are shown (n = 4). Asterisks indicate the p value (unpaired student t-test) compared to the CD3/CD28 (B) or the pulsed (D) group: * = p <0.05; ** = p <0.01; *** = p <0.001. Data is representative of three independent experiments.