Induction and suppression of Th17 cell development. (A) TAMs induced Th17 cells. Normal blood T cells (5 × 105/mL) were stimulated with blood macrophages or TAMs (2.5 × 105/mL) from 3 ovarian cancer patients (OC17, OC20, and OC38). Th17 cells were analyzed by FACS. Results are expressed as the percentage of Th17 cells in CD4+ T cells. Similar results were observed in 8 ovarian cancer patients (P < .01, compared with control). (B) TAMs induced T-cell IL-17 production. Normal T cells (5 × 105/mL) were stimulated with different concentrations of normal macrophages (Mφ) or TAMs from donor OC20. IL-17 was detected by ELISA in the culture supernatants. Results are expressed as mean ± SEM; n = 5. P < .01. (C) The cytokine expression in TAMs. TAMs were isolated from ovarian cancer. Expression of IL-1β and IL-23p19 was detected by real-time PCR. Results are expressed as mean ± SEM; n = 5. P < .01. (D) The importance of IL-1β in TAM-mediated Th17 cell induction. T cells were stimulated for 5 days with TAMs with or without the indicated neutralizing antibodies. IL-23 was blocked by specific IL-23 siRNA as we reported.20 Th17 cells were detected by FACS. Results are expressed as the mean of Th17 cells in CD4+ T cells ± SEM; n = 5. *P < .05 compared with control. (E-F) Treg cells suppressed Th17 and T-cell IL-17 production induced by TAMs. T cells (5 × 105/mL) were stimulated with TAMs (2.5 × 105/mL) in the presence or absence different concentrations of tumor-associated Treg cells. Th17 cells were analyzed by FACS (E). Results are expressed as the percentage of Th17 cells in CD4+ T cells. IL-17 was detected by ELISA in the culture supernatants (F). n = 6. *P < .05 compared with control. (G) The relevance of the adenosinergic pathway in Treg cell-mediated Th17 suppression. In the culture system described (E-F), ARL67156 was added. IL-17 was detected by ELISA in the culture supernatants; n = 6. *P < .05 compared with control.
