Fig 2: GITR is expressed on ILC2s and induces Th2 cytokine secretion in activated ILC2s. a Gating strategy of Lin-CD45+IL-7R+ST2+ ILC2 cells. b Expression of GITR in naïve (white) and IL-33-activated (green) murine white adipose tissue-derived ILC2s compared to the isotype control (gray). Corresponding quantitation of GITR expression shown as MFI+/- SEM, n = 6. c Visceral adipose tissue (VAT) resident ILC2s were isolated from a cohort of naïve C57BL/6 mice and a cohort of activated C57BL/6 mice intraperitoneally challenged with rmIL-33 on 3 consecutive days. Naïve and activated VAT ILC2s were cultured with recombinant mouse (rm) IL-2 and rmIL-7 with DTA-1 (5 µg/mL) or isotype control for 48 h. The levels of IL-5, IL-13, GM-CSF, IL-6, and IL-9 were measured by Luminex on the culture supernatants, n = 6. Error bars are the mean ± SEM. Student’s t-test, ***p < 0.001, ns, non-significant

Fig 3: Lung CD4+ T cells and CD4+ T cell subsets at day 28 postinfection. C57BL/6 mice were subcutaneously given 30 or 150 µg of anti-CD20 or isotype control antibody weekly. Three days after the first injection, all mice were inoculated with approximately 2 × 105 asci of P. murina inoculum. Twenty-eight days postinfection, the right upper and lower lung lobes were harvested, and single cells were prepared and stimulated with PC antigen for 6 h followed by staining for cell surface markers and intracellular cytokine staining. For CD4+ cells and subsets, 100,000 cells were acquired. One-way ANOVA and Tukey’s multiple-comparison tests indicated that there were significant differences for IL-4+ CD4+ and IL-5+ CD4+ subsets between anti-CD20- and ISO-treated groups at higher treatment dosages. There were significantly fewer CD4+ T cells in the higher-treatment-dosage group (P < 0.01) in panel A. Statistical significance is indicated as follows: *, P = 0.05; **, P = 0.01; ns, not significant.

Fig 4: Lung CD4+ T cells and CD4+ T cell subsets at day 14 postinfection. C57BL/6 mice were subcutaneously given 30 or 150 µg of anti-CD20 or isotype control antibody weekly. Three days after the first injection, all mice were inoculated with approximately 2 × 105 asci of P. murina inoculum. Fourteen days postinfection, the right upper and lower lung lobes were harvested, and single cells were prepared and stimulated with PC antigen for 6 h followed by staining for cell surface markers and intracellular cytokine staining. For CD4+ cells and subsets, 100,000 cells were acquired. One-way ANOVA and Tukey’s multiple-comparison tests indicated that there were significant differences for IL-4+ CD4+ and IL-5+ CD4+ subsets between anti-CD20- and ISO-treated groups at higher treatment dosages. Low (30 µg) and high (150 µg) doses of anti-CD20 treatment did not impact the CD4+ T cell number in panel A. Statistical significance is indicated as follows: *, P = 0.05; **, P = 0.01; ***, P = 0.001; ns, not significant.

Fig 5: HpBARI suppresses Alternaria-induced innate immune responses.Alternaria allergen (10 µg) and HpBARI (10 µg) were intranasally administered and mice culled 24 hr later. Eosinophils were enumerated in the BAL (A) and lung (B), and intracellular cytokine staining carried out for IL-13 in ICOS+lineage–CD45+ lung ILC2s (C). IL-5 was measured in cell-free BAL supernatants (D). Data representative of 2 repeat experiments, each with 3–4 mice per group. Error bars show SEM.