N-cadherin adhesion signals assembly of VE-cadherin junctions. (A) Representative confocal images of mouse lung sections from Cre⁻, Cdh2 iEC-KO, and Cdh2 iPC-KO mice stained for VE-cadherin (green on merged image), PECAM1 (red), and nuclei (DAPI, blue). Bar, 10 µm; insets, 5 µm. VE-cadherin at PECAM1-positive junctions is reduced in Cdh2 iEC-KO and Cdh2 iPC-KO mice as compared with controls. (B and C) Quantification of PECAM1 area (B) and VE-cadherin adhesion area normalized to PECAM1 area (C) in lungs of Cre⁻, Cdh2 iEC-KO, and Cdh2 iPC-KO mice; n = 15–26 images from three mice per group. ****, P < 0.0001, ANOVA with Tukey’s post hoc test. (D) Representative confocal images of human pulmonary arterial ECs (HPAECs) grown on either gelatin-coated glass or N-cad-BioS and stained for VE-cadherin (green), N-cadherin (red), and DAPI (blue). Bar, 10 µm. An anti–N-cadherin antibody targeting the cytosolic domain of N-cadherin was used to avoid staining of N-cad-BioS surface. (E) Quantification of VE-cadherin adhesion area from images in D; additional groups included denatured (Den) N-cad-BioS or N-cadherin depletion. n = 10–14 images per group. ****, P < 0.0001, ANOVA with Tukey’s post hoc test. (B, C, and E) Data are shown as mean ± SD. See also Fig. S2.