Supplementary Figure 7: Characterization of B cells in spleen and CNS of mice lacking functional Ly6G⁺ MDSCs during recovery from EAE.

a, Histologic analysis showing co-staining of CD3, CD19 and B220 in control mice (Ly6g^Cre/WT) and Stat3^ΔLy6G mice during early recovery (day 21). Scale bar, 20 μm. b, Flow cytometric analysis of live CD19⁺ B cell compartment in spleen and CNS of control mice (Ly6g^Cre/WT), Stat3^ΔLy6G mice and Ly6G-tdTomato⁺ MDSC-depleted mice (Ly6G Ab) during early recovery from EAE. CD1⁺CD5⁺ regulatory B cells (B_reg cells) are not increased upon MDSC loss of function; representative plots of n = 6 individuals per group; gate on CD19⁺B220⁺ cells. c, Analysis of cytokine production by intracellular cytokine staining in CD19⁺B220⁺ B cells purified from CNS of Ly6g^Cre/WT (n = 6), Stat3^ΔLy6G mice (n = 6) and Ly6g^Cre/WT mice treated with anti-Ly6G depleting antibody (Ly6G Ab, n = 6) at EAE recovery (day 21) and stimulated ex vivo with PMA/ionomycin in the presence of brefeldin A; symbols depict mean ± s.d. of biological replicates; Kruskal-Wallis test with Dunn’s post test; *P < 0.05, ***P < 0.001. d, Representative cytograms of live CD19⁺ B cells isolated from the CNS of Ly6g^Cre/WT control mice or Stat3^ΔLy6G mice (with dysfunctional MDSCs) and stimulated ex vivo with PMA/ionomycin in the presence of brefeldin A for intracellular cytokine staining (double staining for IL-6 and IL-10).