Administration of Ang II systematically enhances ILC2 responses. (A) Images of lung tissue from IL-5–tdTomato (red) reporter mice stained for AGT (green) and DAPI (blue). White arrows show IL-5⁺ ILC2s, and red arrows show AGT⁺ cells; scale bar, 20 µm. Images are from three independent experiments. n = 4/group. (B) Purified lung ILC2s from naive mice were stimulated with Ang II or PBS for 6 h. mRNA expression of Il5, Il13, and Mki67 was determined by qRT-PCR. n = 3/group, and data are repeated from two independent experiments. (C) Lung ILC2s were cultured with Ang II or PBS in the presence of IL-2, IL-7, and IL-33 cytokines for 3 d. The amounts of IL-5 and IL-13 in supernatants were measured by ELISA. N = 4/group, and data are from two independent experiments. (D) mLN lymphocytes were cultured with the indicated treatments for 6 h, and the intracellular cytokines IL-5 and IL-13 from ILC2s were determined by flow cytometry. IL-33: 100 ng/ml; Ang II: 0.5 µM; PMA: 50 ng/ml; ionomycin: 1 µg/ml. n = 3/group, and data are from two independent experiments. (E) Lung ILC2s were stimulated with IL-33 alone or IL-33 plus Ang II for 6 h, and the production of intracellular cytokines was evaluated by flow cytometry. n = 4, and data are from two independent experiments. (F–K) Naive mice were injected i.p. with Ang II (30 µg/mouse, daily) or PBS and sacrificed on day 6. (F–H) Flow-cytometric analysis of ILC2s (CD45⁺Lin⁻CD90⁺) (F), effector cytokines (G), and proliferation (H) from lungs. (I and J) Absolute cell counts of ILC2s (I) and effector cytokines and cell proliferation (J) in mLNs from naive mice treated with Ang II or PBS. (K) Abundance of ILC2s in visceral adipose tissue from naive mice treated with Ang II or PBS. (F–K) Data are from three independent experiments; n = 3 or 4/group. In all graphs, mean ± SEM is shown. Unpaired t test was used. *, P < 0.05; **, P < 0.01. Lin, lineage.