Figure 3.

Role of PROCR in Th17 differentiation. (A) Cells from spleen, peripheral LNs, and thymus were isolated from WT and EPCRδ/δ mice, and frequencies of CD8⁺, CD4⁺, memory CD62L⁻CD44⁺CD4⁺, naive CD62L⁺CD44⁻CD4⁺, and regulatory Foxp3⁺CD4⁺ T cells were determined by flow cytometry (four mice/group). (B–E) Naive CD4⁺ T cells from WT and EPCRδ/δ mice were differentiated into Th17 cells by anti-CD3/anti-CD28 stimulation in the presence of TGF-β1 + IL-6 (T1+6) or no cytokines (Th0) or IL-12 as control. Il17a, Ifng, and Procr mRNA levels were determined after 96 h using quantitative PCR (B), and IL-17A and IFN-γ protein secretion, as well as PROCR expression levels, were examined by flow cytometry (C). (D and E) Cells were differentiated by anti-CD3/anti-CD28 stimulation with or without 300 nM aPC and cytokine production was analyzed 96 h later using flow cytometry (D) and ELISA (E). (B–D) Bar graphs represent mean ± SD of three independent experiments; Student’s t test; *, P < 0.05; ns, not significant.