Extended Data Figure 1: Characterization of lung development in Piezo2^−/− mice.

a, b, Whole-mount staining for αSMA in the left lobe of lungs from E16.5 (a) and E18.5 (b) wild-type and Piezo2^−/− mice. L1–L6, lateral branches of the left lobe. E16.5: wild-type n = 4, Piezo2^−/− n = 3; E18.5: wild-type n = 4, Piezo2^−/− n = 5. A normal conducting airway and vessel pattern was observed in Piezo2^−/− lungs at late embryonic stages. c, d, Whole-mount immunostaining for E-cadherin and Muc-1 (c) or E-cadherin, T1α, and SpC (d) in left lobes from E18.5 wild-type and Piezo2^−/− mice. Arrows indicate proximal (P) to distal (D) direction. Left panels, lower magnification; right panels, higher magnification of the distal region. Solid circles in right panels indicate alveolar progenitors in the distal region. Solid hexagons in left panels indicate nascent type I pneumocytes. Yellow dashed circles in left panels indicate nascent type II pneumocytes. A normal epithelial morphology and no defects in alveolar epithelial patterning and differentiation were observed Piezo2^−/− lungs. e, Representative ultrastructures of wild-type and Piezo2^−/− newborn lungs. Black arrows mark type I pneumocyte extensions. White dotted circles mark type II pneumocytes. White arrowheads within white dotted circles mark lamellar bodies. RBC, red blood cells. Samples from four mice per genotype were analysed. Normal morphology and similar abundances of type I and II pneumocytes, endothelial cells, red blood cells and surfactant proteins were observed in Piezo2^−/− lungs compared to wild-type lungs. f, Wet-to-dry lung ratio 6 h after delivery at E18.5 to assess clearance of fetal pulmonary fluid. NS, statistically not significant. Unpaired Student’s t-test. Bars represent mean ± s.d. g, Per cent weight of postnatal day (P)0 lung, heart, and liver normalized to whole body weight. Bars represent mean ± s.e.m. NS, statistically not significant. Kruskal–Wallis nonparametric test (lung and liver) or one-way ANOVA (heart). Scale bars, 500 μm (a, b); 33 μm (c, d, left); 22 μm (c, d, right); 5 μm (e).