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, Immune cell subsets were gated on live CD45 + cells, then further identified as follows: T cells, MHCII ? CD3 + ; dendritic cells, CD3 ? MHCII + CD11c + ; NK1.1 + innate cells, CD3 ? MHCII ? NK1.1 + ; B cells, CD3 ? MHCII + B220 + ; inflammatory monocytes, CD3 ? CD11b + Ly6C hi Ly6G ? ; neutrophils, CD3 ? CD11b + Ly6C int Ly6G +. (d) Quantitation by flow cytometry of T-bet-expressing CD45 + immune cells, T cells, dendritic cells or NK1.1 + innate cells from the CNS of T-bet ZsGreen reporter mice immunized with MOG 35-55 /CFA and pertussis toxin. CNS-infiltrating cells were harvested at the onset (day 7) or peak of EAE disease (day 13-14 post-immunization) and stained for lineage-specific markers as in (c). Data in (b-d) of stimulation, Stimulated cells were stained with viability dye, antibodies to lineage markers (TCR?, TCR??, B220, Gr1) and antiCD45, anti-NKp46, anti-IL-7R?, anti-CD49b (DX5), anti-ROR?t, anti-IL-17A and antiIFN-?. Production of IL-17A and IFN-? by meningeal NK cells, p.1
, ? CD45 + NKp46 + ROR?t ? DX5 ?/lo IL-7R? + ) and NKp46 + ILC3
, The box-and-whiskers plot depict the frequency of NKp46 + ILCs within Lin ? CD45 + population in the meninges of Tbx21 f/f or Tbx21 f/f NKp46-Cre + mice. (c) Absolute numbers of NK cells, ILC1 and NKp46 + ILC3 and IFN-?-producing NK cells, ILC1 and NKp46 + ILC3 (gated as in a) in the meninges of Tbx21 f/f or Tbx21 f/f NKp46-Cre + mice. (d-e) Mean clinical scores, percent disease incidence and maximum clinical score of Eomes f/f or Eomes f/f NKp46-Cre + mice following adoptive, ? CD45 + NKp46 + ROR?t + IL-7R? + ) was measured by intracellular cytokine staining. (b)
, 001 (two-tailed Student's t-test (b,c,e) or two-way ANOVA (d)). Data are combined from two independent experiments (b,c; n = 9, 10 mice per group) or three independent experiments