IL-22 regulation is most closely related to IFN-γ rather than IL-17. Naive T cells were cultured under Th0 (no polarizing cytokines), Th1 (1 ng/mL IL-12), Th2 (25 ng/mL IL-4), Th17 (10 ng/mL IL-1β, 20 ng/mL IL-6, 10 ng/mL TNF, 1 ng/mL TGF-β, 100 ng/mL IL-23), suboptimal Th17 (absence of individual Th17-promoting cytokines) in the presence of anti-CD3 + anti-CD28. Protein and transcript analyses were performed after 24 hours of restimulation with anti-CD3 + anti-CD28. Supplemental Table 1 contains more details. (A) Clustering analysis of Th cytokines produced in all experimental conditions using a Pearson correlation–based distance. Cytokines were separated in clusters by comparing their linkage distance. The agglomerative coefficient reflects the structure of the data (values close to 1 indicate well-separated clusters), and resampling similarity index (rsi) evaluates the robustness of the clustering. (B) Graphs of amounts of IL-17 or IL-22 protein were correlated to IFN-γ or TNF levels, using Pearson correlation. R indicates correlation coefficient. (C) RT-PCR for expression of RORC, RORA, AHR, TBx21 in Th0, Th1, Th2, and Th17 conditions. Threshold cycle values were normalized to mRNA of ribosomal protein L34 gene. Data were normalized to the maximal value obtained for each donor. Data are mean ± SEM of 9 donors. (D-E) Graphs of IL17A and IL22 transcript levels, obtained from 9 independent experiments with cells cultured as previously described, were correlated to RORC, RORA, AHR, and TBx21 transcript levels, using Pearson correlation. R indicates correlation coefficient.