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. Author manuscript; available in PMC: 2014 Jul 18.
Published in final edited form as: J Immunol. 2010 Nov 1;185(11):6599–6607. doi: 10.4049/jimmunol.1002041

FIGURE 6.

FIGURE 6

LPS, IFN-α, and IL-27 induction of IL-10 is STAT1 and STAT3 dependent. A, Total of 0.5 × 106 WT BMDMs were unstimulated or stimulated with LPS (50 ng/ml), IFN-α (250 U/ml), or IL-27 (10 ng/ml) for indicated time points. Cell lysate was collected and immune-blotted for STAT1 and STAT3 expression and STAT1 and STAT3 phosphorylation using specific Abs. B, Total of 0.5 × 106 WT and STAT1−/− BMDMs were unstimulated or stimulated with LPS (50 ng/ml), IFN-α (250 U/ml), or IL- 27 (10 ng/ml) for 3 h. RNA was harvested, and relative IL-10 transcript was detected via qPCR normalized to L32. Student t test was performed. *p < 1 × 103. C, Total of 0.5 × 106 macrophage-specific STAT3+/+ and STAT3−/− BMDMs were unstimulated or stimulated with LPS (50 ng/ml), IFN-α (250 U/ml), or IL-27 (10 ng/ml) for 3 h. RNA was harvested, and relative IL-10 transcript was detected via qPCR normalized to L32. Data represent two independent experiments. *p < 0.05. D, ChIP using Abs against STAT1, STAT2, STAT3, c-Maf, and Sp1 using primers specific to the IL-10 promoter. Transcription factor enrichment presented as percent of input representing two independent experiments. E, Model of LPS-mediated induction of IL-10 gene expression involves sequential induction and signaling type I IFN followed induction and signaling by IL-27 signaling in macrophages.