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. 2022 Feb 24;119(9):e2108686119. doi: 10.1073/pnas.2108686119

Fig. 4.

Fig. 4.

Priming with ascr#7 attenuates both innate and adaptive type 2 immunity in the lung. Mice were sensitized with synthetic ascr#7 plus OVA in alum on days 0 and 14, and indicated assays were performed on day 15 as described in SI Appendix, Fig. S2A. (A) Representative staining profiles of CD44 and ST2 expression on CD4+CD44hi cells from spleen (Left) and percentages of CD44hiST2+CD4+ T cells in the spleen (Right). Data from one experiment are representative of two independent experiments (Left) and pooled from two independent experiments (Right). Mean values from eight mice per group are shown with SD. *P < 0.05, **P < 0.01, ***P < 0.001. Mice were sensitized with synthetic ascr#7 plus OVA in alum on days 0 and 14, and indicated assays were performed on day 24 as described in SI Appendix, Fig. S2B. (B) Representative staining profiles of CD44 and ST2 expression on CD3e+CD4+ cells from lungs (Left) and absolute cell numbers of CD44hiST2+CD3e+CD4+ T cells in lungs (Right). (C) Lung CD3e+CD4+ T cells from mice sensitized with ascr#7 or with control (solvent vehicle) were prepared and stimulated subsequently in vitro with OVA and APCs. An ELISA measured the indicated cytokines in the culture supernatants collected after 72 h. Mean values with SD are shown. (D) Lung mononuclear cells were isolated from mice sensitized with ascr#7 or control (solvent vehicle) and stimulated with anti-TCRb mAb for 6 h. Representative intracellular staining profiles of IL-5 and IL-13 gated on CD4+CD3e+ST2+ cells are shown (Left). Percentages of IL-13– or IL-5–producing cells in CD4+CD3e+ST2+ cells are shown (Right). (E) Representative staining profiles of CD25 and CD127 expression on lineage− (Lin−) cells of lung mononuclear cells are shown (Left). Absolute cell numbers of Lin− CD25+CD127+ cells in the lung are shown (Right). (F) Representative staining profiles of CD127 and intracellular GATA3 expression in Lin−CD45+ cells of lung mononuclear cells are shown. (G) CD45−PECAM−EpCAM+ cells were sorted from the lungs of immunized mice, and qRT-PCR of Il33 and Tslp was performed. Mean values from three (G) per group are shown with SD. *P < 0.05, **P < 0.01.