TGF-β mediates Clec-2-Pdpn-dependent lung development. (A) Neonatal lethality in Clec1b^fl/fl and Clec1b^ΔPLT neonates treated with mouse IgG (control) or anti-TGF-β. *P = .0049, Fisher’s exact test. (B) Immunostaining of α-SMA (green) and DAPI (blue) in E17.5 distal lung. AL, alveolar sac; M, lung mesothelium. (C) Quantification of α-SMA expression in E17.5 distal lung. The asterisk indicates the group in which α-SMA expression was significantly lower than that in the other 3 groups; mean ± SD, n = 3 each. *P < .0001, Holm-Sidak test. (D) Whole-mount immunohistochemistry of Wt1 (magenta) and Pdpn (green) in E17.5 luMCs. (E) Quantification of Wt1-negative rate in luMCs; mean ± SD, n = 3 each. *P = .0084, Tukey’s test. (F) Quantification of luMC density; mean ± SD, n = 3 each. *P = .0001, Tukey’s test. (G) Clec-2 expression on fetal platelets in fetuses treated with anti-TGF-β antibody; mean ± SD, n = 4, 4, 5, and 4, respectively, from the left of the histogram; Student t test. N.S., not significant. (H) Total TGF-β1 relative concentration in Clec1b^+/+ and Clec1b^–/– lungs; mean ± SD, n = 7 (Clec1b^+/+), n = 6 (Clec1b^–/–). *P = .0187, Student t test. Scale bars: 25 μm (B,D).