FIG 3.

Myeloid CITED2 promotes anti-inflammatory gene expression and function. (A and B) Wild-type murine BMDMs were stimulated with 10 ng/ml of IL-4 or IL-13 separately for 6 h. CITED2 mRNA and protein expression were evaluated by quantitative-PCR and Western blot analyses, respectively (n = 4). (C to F) Wild-type murine BMDMs were pretreated with the STAT6-specific inhibitor AS1517499 (C and F) or transfected with siStat6-specific siRNA (D to F). The cells were stimulated with IL-4 or IL-13 separately and evaluated for CITED2 mRNA (C and E) and protein (F) expression (n = 3). (G) Lyz2^cre and Cited2^fl/fl:Lyz2^cre mouse BMDMs were stimulated with IL-4 or IL-13 separately and analyzed for Stat6 mRNA expression by quantitative PCR. (H) Lyz2^cre and Cited2^fl/fl:Lyz2^cre mouse BMDMs were stimulated with IL-4 and analyzed for STAT6 protein expression and phosphorylation (STAT6-Tyr641) by Western blotting. (I) BMDMs from Lyz2^cre and Cited2^fl/fl:Lyz2^cre were stimulated with 10 ng/ml of IL-4 for 18 h. Total RNA was isolated, and mRNA expression of Arg1, Retnla, Mgl2, Chil3, and Mrc1 were analyzed by quantitative PCR (n = 4). (J) RAW 264.7 cells transfected with either pCMV6 or pCMV6-Cited2 were stimulated with IL-4, and expression of Arg1, Retnla, Mgl2, Chil3, and Mrc1 was analyzed by quantitative PCR (n = 4). The beta-actin and 36B4 genes were used as housekeeping genes for Western blot and quantitative-PCR analyses, respectively. Each experiment consisted of three replicates. Data were analyzed by Student's t test. N.S., not significant; *, P < 0.01; **, P < 0.0001. All values are reported as means and SD.