Synovial IFN-γ⁺ Th cells isolated ex vivo from RA patients are induced by cytokines. (A) Expression of CD45RA, CD45RO, CCR5, and IL-18Rα on SF-derived Th cells ex vivo; 1 representative patient of 6. (B,F) Live SF- or SM-derived Th cells were stained ex vivo for secreted IFN-γ and 4-1BB by the cytokine secretion assay technology. Numbers indicate frequencies among Th cells. (C,G) Magnetically enriched live IFN-γ⁺ Th cells were analyzed for 4-1BB expression. Numbers indicate frequencies within the IFN-γ⁺ population. (D,H) SF-MNCs or SM-MNCs were stimulated for 14 hours with αCD3 + αCD28. CD4⁺ Th cells were analyzed for 4-1BB and IFN-γ expression. Numbers indicate frequencies within the IFN-γ⁺ population. (B-D,F-H) Results from 4 representative SF and all analyzed SM samples. (E) Frequencies of live 4-1BB⁺IFN-γ⁺ and 4-1BB⁻IFN-γ⁺ SF-derived Th cells after magnetic enrichment according to secreted IFN-γ (all patients). P value was determined by Wilcoxon rank test.