Immunological characteristics of Tr17 cells after CII immunization
We analyzed the expression of cell surface markers related to Treg cell stability and function in Tr17 cells or RORγt-negative Treg cells. Treg-specific molecules such as CD25, CTLA-4, and GITR were significantly increased in Tr17 cells compared to RORγt\sout--negative Treg cells both of which were isolated from draining LNs on 10 days after 1st-CII-immunization (Figure 3a, b, c). Compared to RORγt-negative Treg cells, ICOS expression was also upregulated in Tr17 cells, which was previously reported as a characteristic marker molecule of Tr17 cells15(Figure 3d). Moreover, most of the Tr17 cells expressed Helios, indicating that Tr17 cells in CIA might be differentiated from thymus-derived Treg cells, as in a previous report on the EAE model15(Figure 3e). In Foxp3creRORγtfl/flcKO mice, although CD25 expression on Treg cells was comparable to Foxp3wtRORγtfl/fl control mice, CTLA-4 and GITR expression on Treg cells was significantly decreased compared to Foxp3wtRORγtfl/flcontrol mice at 10 days after 1st CII-immunization (Supplemental Figure 2); thus, deletion of RORγt in Treg cells results in down-regulation of characteristic Treg cell molecules. These results suggested that Tr17 cells detected in the course of CIA retained a Treg cell nature with characteristic features of Tr17 cells as reported in the EAE model, and that deletion of RORγt might attenuate Treg cell function.