Figure 1: ILC2s localize to adventitial cuffs in multiple organs.

(A and B) 3D rendering with zoom of a 200μm thick lung slice with IL-5⁺ ILC2 (RFP), bronchioles (autofluor⁺ Lyve⁻), arteries (Lyve1^dim+, Autofluor⁺, bronchiole⁻adjacent), veins (Lyve1^dim+ Autofluor⁺, non-bronchiole adjacent) and lymphatics (Lyve1^bright+, Autofluor⁻) visualized. (C and D) IL-5 RFP⁺ ILC2 and CD31⁺ vessels visualized in (C) brain meninges and (D) perigonadal adipose tissue (GAT). (E and F) IL-5 RFP⁺ ILC2 visualized in the lung adventitial cuff, highlighted by a lack of (E) Aquaporin 5 and SMA, and positively by (F) Col1A1 rich cuff regions surrounding bronchioles and arteries. (G) 3D image quantification of IL-5 RFP⁺ ILC2s localized within the cuff volume, using automatic surfacing (lungs), or found within 40μm of SMA⁺ vasculature (meninges, GAT). (H) Percent of cells in lung cuffs: ILC2 (IL-5 RFP⁺), T-cells (CD3ε⁺), DCs (CD11c⁺ MHCII⁺ co-positive) and alveolar macrophages (AM, CD11c⁺ MHCII⁻). Analysis in (G) included lung cuff, parenchyma, and pleural regions, whereas (H) excluded pleura. (A-E) Images are representative of 3 or more mice or (F) 2 mice. (G and H) Data points represent averages of individual mice with n ≥5 animals with ≥55 cells of each type analyzed per mouse. See also Figures S1–3 and Movies S1 and S2.