Jmjd3 induction in IL-4–treated macrophages. (A) BMDMs were incubated with IL-4 for 6 hours. The mRNA levels of JmjC family proteins were measured by quantitative RT-PCR. N.D. indicates not detected. (B-D) Jmjd3 mRNA levels were measured by quantitative RT-PCR after the following treatments. BMDMs were incubated with IL-4 (B) and indicated cytokines (C) for 6 hours. BMDMs were incubated with indicated cytokines for 2 days (D left). On day 2, cells were washed with phosphate-buffered saline and incubated with or without fresh IL-4, or with or without a cocktail of IL-4 and IL-13 for an additional 2 days as indicated (D right). #P < .05. (E) BMDMs were incubated with IL-4 as indicated. Jmjd3 expression was measured by immunoblotting. Gapdh was used as a loading control. Representative data were shown from 3 independent experiments. (F-H) Jmjd3 mRNA levels were measured by quantitative RT-PCR after the following treatments. Peritoneal macrophages (PMΦs) were incubated with IL-4 for 6 hours (F). BMDMs were incubated with LPS, a cocktail of LPS and IFN-γ, or CpG DNA (in indicated concentration) for 6 hours (G). BMDMs from WT (C57BL/6), Myd88^−/−, and Trif^−/− mice were incubated with or without IL-4 for 6 hours (H). All data of quantitative RT-PCR shown are “fold induction” relative to that in untreated WT cells. Data are representative of 3 independent experiments and are expressed as mean ± SEM. *P < .05; **P < .01; ***P < .001.