Figure 5: The inhibitory effect of type I IFN overrides IFN-γ-mediated priming for IL-12 secretion in CD16^neg monocytes.

(A) Purified human CD16^neg monocytes (10⁶/ml) were cultured overnight in the presence of medium alone (blue circles), IFN-γ (10 ng/ml) alone (white filled circles), or in the presence of both IFN-γ and IFN-α (10 ng/ml each) (gray filled circles) before stimulation with untreated (left) or MycB-treated T. gondii (middle), or R848 (right). Culture supernatants were collected 18 hours later and IL-12p40 was measured by ELISA. Connected symbols indicate the values obtained for monocytes from the same donor (n=6) treated in parallel under the 3 different conditions. (B) CD16^neg monocytes (10⁶/ml) were cultured with either IFN-γ (left panel) or IFN-α (right panel), or only in medium for 18 hours when supernatants were removed. The IFN-treated cultures were split into two wells, and medium added to one while the second was treated with the opposite IFN type from that used in the first culture. The monocytes cultured first in medium alone were treated with the same type of IFN used during second incubation. All cultures were then stimulated with R848 (300 ng/ml) in parallel for an additional 18 hours. IL-12p40 was measured by ELISA and connected symbols indicate values obtained for monocytes from the same donor (n=8) cultured in parallel under the 3 different conditions. **P <0.01.