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. Author manuscript; available in PMC: 2014 Oct 1.
Published in final edited form as: Nat Immunol. 2014 Feb 23;15(4):354–364. doi: 10.1038/ni.2830

Figure 1. Human splenic ILCs express a mucosa-like group-3 phenotype and occupy MZ and perifollicular areas.

Figure 1

(a) Flow cytometry of CD4, CD25, CD56, CD69, CD96, CD161, NKp44, NKp46 and CCR6 on splenic Lin–CD117+CD127+ ILCs (red gate). Gray shading, negative control. (b) qRT-PCR of RORC (RORγt), AHR (AhR), IL22 (IL-22), TNF (TNF), LTA (LT-α), LTB (LT-β), PRF1 (Perforin-1) and IFNG (IFN-γ) mRNAs from splenic or tonsillar ILCs and NK cells and from splenic macrophages (Mϕ), B cells and T cells. Results are normalized to ACTB (β-actin) mRNA and presented as relative expression (RE) compared with that of fresh splenic NK cells. Error bars, s.e.m.; *P <0.05 (two-tailed unpaired Student's t test). (c) Flow cytometry of intracellular RORγt and T-bet in splenic ILCs (red lines) and NK cells (blue lines). (d) Flow cytometry of Lin–CD117+CD127+ ILCs in splenic and tonsillar lymphocytes. *P < 0.05 (Mann-Whitney U-test). (e) Flow cytometric analysis of frequency of viable Annexin-V–propidium iodide (PI)– ILCs after culture of splenic and tonsillar ILCs with medium alone (Ctrl), IL-1β, IL-7 or IL-23 for 72 h. (f) ELISA of IL-22 from splenic and tonsillar ILCs cultured as in (e). (g) IFA of spleen stained for MR (purple), RORγt (green) and DNA-binding 4’-6-diamidine-2’-phenylindole (DAPI; blue). RP, red pulp. Original magnification, ×10. FO: center of the follicle (h) Immunohistochemical quantification of CD117+Tryptase– ILCs from MZ-PFZ and red pulp (RP) areas in nine microscopic ×20 fields from two spleens. *P< 0.05 (Wilcoxon Matched-pairs signed Rank test). Data summarize three measurements from three pooled experiments with 1 donor in each (spleen and tonsil) (b), display values from thirty seven spleens and twelve tonsils (d), or show one of four experiments with similar results (a,c,e,f,g).