Fig. 4

IFN-α/β and TNF-α secreted by activated T cells cooperate to inhibit MDSC function. a, b The expression of M-CSFR (a) and ARG-1 (b) of MDSCs in the presence of ASN or denatured ASN, which was removed by heating, was determined using FACS (a) or western blotting (b). c–g MDSCs were treated with IFN-α (0.5 ng/mL), IFN-β (0.5 ng/mL), TNF-α (2 ng/mL), or their combination. The expression of M-CSFR (c), Ki-67 (d), and ARG-1 (e) was estimated at day 3; the cells were then cocultured with CFSE-labeled T cells for 6 days. The suppressive activity of MDSCs was monitored using FACS (f). The percentage of CD11b⁺CD49d⁻ cells was analyzed using FACS (g). h–j MDSCs were exposed to ASN with control IgG1 (10 μg/mL), TNF-blocking mAbs (2 μg/mL), IFNAR1 mAbs (2 μg/mL), or their combination for 3 days. The levels of M-CSFR and ARG-1 in MDSCs were determined using FACS (h) and western blotting (i); the suppressive capacity of T cells was measured using FACS (j). Immunoblots are representative of 3 subjects (b, e, i). The data were generated from three independent experiments (a, d, f, g, h, j) or six independent experiments (c). The data are shown as the mean ± SEM. *p < 0.05; **p < 0.01; ***p < 0.001