Figure 3. Origin of RORγt⁺CCR6⁺ Treg cells.

(A) CD4⁺YFP⁺CCR6⁻ Treg cells isolated from Foxp3^YFP-Cre reporter mice (CD45.2⁺) were mixed with CD4⁺CD25⁻CD44^loCD62L⁺ naïve T cells isolated from B6.SJL mice (CD45.1⁺) in the 1:9 ratio and then were adoptively transferred to CD45.1⁺CD45.2⁺ recipient mice. One day after the transfer, recipient mice were subcutaneously immunized with MOG/CFA. Seven days later, CD4⁺ T cells in dLNs were analyzed for Foxp3, RORγt and CCR6 expressions after gating populations based on CD45.1 and CD45.2 expression. (B) Helios expression in CD4⁺Foxp3⁺ or CD4⁺Foxp3⁻ cells in dLNs was compared with RORγt or CCR6 expression. (C–D) OVA-specific CD4⁺KJ1-26⁺GFP⁺CCR6⁻ Treg cells isolated from DO11.10xFoxp3^GFP reporter mice were adoptively transferred to BALB/c mice. One day after the transfer, mice were s.c. immunized with OVA/CFA or KLH/CFA or left untreated. Seven days after immunization, KJ1-26⁺ or KJ1-26⁻ CD4⁺Foxp3⁺ Treg cells in dLNs were analyzed for RORγt and CCR6 expressions. Data are representatives of two independent experiments. *, p<0.05. **, p<0.005. See also Figure S2.