Fig. 6.

Th-polarizing ability of pDCs exposed to H₂O₂. Freshly isolated pDCs were treated with 0.01 µM H₂O₂ and R837, separately and in combination for 24 h, and then washed and cocultured for up to 6 days with naïve autologous CD4⁺CD45RA⁺ T cells. (A) Th17-polarizing ability of H₂O₂-treated pDCs. To assess the development of Th17 cells in the cocultures, levels of secreted IL-17 were determined in the culture supernatants by ELISA. (B) Th1- and Th2-polarizing ability of H₂O₂-exposed pDCs. After 6 days of cocultivation, T cells were restimulated with anti-CD3 mAb, phorbol 12-myristate 13-acetate, and ionomycin, and intracellular IFN-γ and IL-4 staining was performed. The percentages of IFN-γ- and IL4-positive cells were analyzed by means of flow cytometry. The dot-plot diagrams represent the results from one of four individual experiments. (C) Expression of OX40-L, a regulator of Th2 cell development, on the surface of pDCs treated with H₂O₂. Freshly isolated pDCs were stimulated with H₂O₂ only, R837 only, or both together for 24 h and then stained for OX40-L. Changes in the frequency of OX40-L-positive cells were analyzed using flow cytometry. Results are presented as means ± SE of three individual experiments. **P<0.01, ***P<0.001 vs T cells cocultured with untreated pDCs.