Figure 4.

Recruitment of aryl hydrocarbon receptor (AhR) to the dioxin response elements (DREs) in the interleukin-33 (IL-33) promoter region. THP-1 macrophages were treated with 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin (TCDD) (10 nM) or kynurenine (KYN) (50 μM) for 1 h. A, Nuclear extracts from untreated or AhR ligand-treated cells were used for an electrophoretic mobility shift assay (EMSA), which was performed using double-stranded, 32P-labeled oligonucleotides containing the DRE binding sequence of the rat CYP1A1 promoter (left panel) or human IL-33 promoter (right panel). To confirm specificity, a 100-fold excess of unlabeled DRE oligonucleotide was added (cold). B, Chromatin immunoprecipitation (ChIP) was performed to assess the binding of AhR to DREs within the human IL-33 promoter region. Human genomic DNA (input for precipitation) was used as a positive control, and immunoprecipitation with a nonspecific antibody (IgG) was used as a negative control. The resulting precipitants were analyzed by real-time PCR, and the percent input was calculated. The values represent the mean ± SE (n = 3). Data were analyzed using a one-way analysis of variance (ANOVA), followed by Dunnett's test. **p < .01 versus the untreated group.