Figure 3
TH17 amplification is independent of AhR stimulation and also observed after culture on coated CFA.
(a) TH17 polarization with and without cytokine coating in AhR ligand rich (IMDM) versus low (RPMI) cell culture media for 4 days (n = 8 from 4 indep. exp., Bonferroni after ANOVA). (b) TH17 polarization with soluble cytokine in the presence of coated AhR agonist FICZ at the indicated doses and antagonist CH-223191 on coated cytokine preparation (n = 6 from 3 indep. exp., Bonferroni of selected conditions after ANOVA). (c) Cell culture plate coating with TH17 polarizing cytokines was performed with or without trypsin digestion as described in methods. (4 days, n = 4 from 2 indep. exp.). (d–g) CFA (d,e) and Montanide (Mon, f,g) water-in-oil adjuvants were added in solution or coated to the plate during TH17 polarization and the proportion of TH17 cells (d,f) and the mean TCR expression on live T cells (e,g) determined (4 days, n = 6–8, 3–4 indep. exp., Bonferroni after ANOVA). For all TH17 polarizations cytokines were added at standard concentration (50 ng/ml IL-6, 1 ng/ml TGFβ, 20 ng/ml IL-23) to the media of all cells, coating with 5x of these concentrations was as indicated in panels A and B, all cells were restimulated with PMA/ionomycin.
