Figure 1. Inflammatory exFOXP3 T cells are enriched in the inflamed CNS in vivo.

EAE was induced in female FOXP3^Cre-GFP Rosa^RFP fate-mapping mice. Tissues were collected at onset, peak, or chronic phase of disease (represented by the arrows in Supplemental Figure 1), and immune cells were isolated from spleen, lymph nodes (LN), or pooled brain and spinal cord (CNS) for flow cytometry. (A) Percentage of FOXP3⁺ Tregs and exFOXP3 T cells in the CNS along EAE course. (B) Percentage of FOXP3⁺ Tregs and exFOXP3 T cells at the chronic phase in different tissues. (C–L) Isolated cells at peak or chronic phase were stimulated for 4 hours with PMA, ionomycin, and Golgiplug. Percentage of CD25⁺ (C and H), GITR⁺ (D and I), IFN-γ⁺ (E and J), GM-CSF⁺ (F and K), and IL-17⁺ (G and L) within either FOXP3⁺ Tregs or exFOXP3 T cells in CNS. Representative dot plot of RFP versus GFP in splenocytes and gating strategy in Supplemental Figure 1. n = 3–5; 2-way ANOVA with Bonferroni’s multiple-comparison test. Data are plotted as mean ± SEM. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001.