2.7 Preparation of bone marrow-derived MDSCs in vitro
Bone marrow cells were isolated from mice by flushing their femurs and
tibiae, centrifuging the cell-containing elution (1800 rpm, 5 min), and
resuspending them in complete RPMI 1640 (Thermo Fisher Scientific)
medium supplemented with IL-6 (40 ng/mL, Peprotech, Suzhou, Jiangsu,
China) and GM-CSF (40 ng/mL, Peprotech). The cells were maintained at 37
°C in a 5 % CO2 humidified atmosphere for 4 d. To
investigate the effect of IL-35 on the expansion of MDSCs, IL-35, IL-6,
and GM-CSF were added to the medium at the same time.