Fig. 4
Cbfβ-deficient ILC2s show an exhausted-like phenotype after cytokine stimulation. ILC2s (CD45+ CD3– CD19– CD127+ CD25+ ST2+ KLRG1+) were sorted from the lungs of Cbfb+/f PLZF-Cre or Cbfbf/f PLZF-Cre mice and cultured with IL-2 and IL-33 (a, b, d–k) or IL-2 and IL-7 (c). The number of days is indicated below. a Flow cytometry analysis of IL-5 and IL-13 expression by ILC2s from the indicated mice cultured for 3 days. b, c The IL-5 (b, left; c, left) and IL-13 (b, middle) concentrations in the culture supernatants from the indicated mouse ILC2s on day 4 and the absolute number of cultured ILC2s (b, right; c, right) from the indicated mice on day 6. d MA plots of genes expressed in Cbfbf/f PLZF-Cre ILC2s versus Cbfb+/f PLZF-Cre ILC2s cultured with IL-5 and IL-13 for 4 days. Red dots indicate genes that were significantly differentially expressed (false discovery rate < 0.05). e Heat map of selected genes expressed in the cultured Cbfbf/f PLZF-Cre ILC2s versus Cbfb+/f PLZF-Cre ILC2s (e). f, g, h Mean fold change of RPKM values of the indicated genes in Cbfbf/f PLZF-Cre ILC2s versus Cbfb+/f PLZF-Cre ILC2s. i Gene set enrichment analysis of signature genes observed in exhausted CD8+ T cells performed on a gene set that was differentially expressed in Cbfbf/f PLZF-Cre ILC2s stimulated with IL-33 in vitro compared to Cbfb+/f PLZF-Cre ILC2s. j Flow cytometry analysis of TIGIT and IL-10 expression by cultured ILC2s from the indicated mice on day 6. k The frequency of TIGIT+ IL-10+ cells in the indicated ILC2s was determined by flow cytometry as in j. Numbers indicate the percentages of cells in each quadrant (a, j). In b, c, k, *p < 0.05 and **p < 0.01 by Student’s t-test. Data are representative of at least two independent experiments (mean ± s.d. of four mice in b, c and of three mice in k)
