Svep1 expression pattern and Svep1-/- hypoplastic lung lobes anomalies.
(a) In situ hybridization of lungs from E14.5 wild type embryos showed strong Svep1 mRNA expression in the mesenchyme at the lung periphery and around bifurcating distal airway tips (right inset outlined in red, arrowhead). (b) Arrowhead shows Mesenchymal expression of SVEP1 protein in normal E12.5 lung. White arrows indicate the location of the SVEP1 protein on the epithelial membrane stained with ECAD (purple) in a distal airway bud. (c) SVEP1 expression is strong in the mesenchyme towards the lobe tip of the E14.5 lungs and branching cleft sites (arrowheads). (d) SVEP1 protein localizes in the lung parenchyma including the primary septa (upper box) and adjacent to the proximal airway epithelium (lower box). (e) microCT scan of E18.5 embryos showing multiple defects including thinning of the diaphragm and lung hypoplasia (red arrows). (f) Ventral view of E18.5 of Svep1+/+ and Svep1-/- lungs. The arrowheads indicate the tips of the lung lobes, which are short and rounded (highlight in black) in Svep1-/- mice and have like a cauliflower morphology. (g) H&E-stained lung sections embryos demonstrate that E16.5 lungs from Svep1-/- embryos are smaller and have irregular lobe edges. Black arrowheads indicate left lobes with highlighted surface. (h) The red arrowheads show the dorsal side of the E18 lungs, whereas the black arrows highlight the ventral side. Svep1-/- embryos show defective saccular development mainly at the edge on the ventral side of the lung (inserts arrowheads). Co-localization of ECAD and SOX9 confirms high expression of SOX9 in distal airways (white arrowheads) in the lung of Svep1-/- embryos at E18.5. (i) Plots showing Sox9 relative mRNA expression at E14.5, E16.5, and E18.5. mRNA expression is significantly high at E18.5 in Svep1-/- embryos (average number of tips per area ± SEM; p < 0.05; n ≥ 5). Right cranial (RCr), Right Caudal (RCa), Right Medial (RMe), and Right Accessory (RAc) lobes. Scale bars: 50μm (a, b, c, d), 100 μm.