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. 2008 Jul 7;105(28):9721–9726. doi: 10.1073/pnas.0804231105

Fig. 5.

Fig. 5.

Ahr regulates the activation of Stat1 in Th17 cell development. (A) MACS-sorted naïve T cells were cultured with anti-CD3/CD28 beads and stimulated with IL-6 or TGF-β, either alone or combined, for 2 days. Whole cell lysates were immunoprecipitated with anti-Ahr antibody, after which Stat1, Stat3, Stat5, Stat6, and Ahr were detected with Western blotting. IP, immunoprecipitation; IB, immunoblot. (B) Naïve T cells isolated from Ahr WT and He mice were stimulated with anti-CD3/CD28 beads and TGF-β plus IL-6 in the presence or absence of IFN-γ for 3 days, followed by re-stimulation with PMA and ionomycin for 5 h and with GolgiStop (final 2 h), and then staining for intracellular cytokines. Dot plots show intracellular staining for IFN-γ and IL-17. (C and D) Naïve T cells isolated from Ahr WT and KO splenocytes were stimulated with anti-CD3/CD28 beads and TGF-β plus IL-6 for 30 min or 24 h, fixed and permeabilized in 90% methanol, and finally stained with Alexa Fluor 488-conjugated phospho-Stat1 and PE-conjugated phospho-Stat3. Intracellular levels of phospho-Stat1 (C) and Stat3 (D) were measured by means of flow cytometry. These results are representative of three independent experiments.