Figure 4

Tagln-Cre:Tgfbr2 ^flx/flx shows failure of anterior diaphragm formation. (a,c) micro-CT cross sections in mutant and WT littermates. (a) the liver herniates into the chest, the dilated SVC (arrow) is seen compressing the right lung upper lobe. (b) transverse section in the thoracic region at E13.5 showing the liver herniating to the thoracic cavity. The lateral body wall skeletal muscles labelled by (MyHC) and rib primordia by (Sex-Determining Region Y; SOX9) are normally developed. Of note, the lung lobar configuration is maintained, left, right and posterior caval lobes are seen. (c) in the WT the diaphragm reaches the sternum (arrow). (d) transverse section in WT embryo in the thoracic region at E14.5 showing developed anterior diaphragm (arrow) made of skeletal muscle cells (MyHC+). (e,f) coronal sections to compare the posterior (e) and anterior (f) diaphragm. (e) the posterior diaphragm is present, extends to the lateral body wall and multiple layers of skeletal muscle cells (MyHC+) are present (e’). (f) failure of formation of a muscular anterior diaphragm and a thin sac (Laminin+) separate the abdominal and thoracic cavities (f’). (g) the leading edge of the anterior diaphragm at E14.5 fails to progress ventrally in the mutant. Condensation of Tcf4 + fibroblasts is seen, however no myogenic progenitor cells can be detected (inset). (h–k) sagittal volume rendering with segmentation of the lungs and liver in mutant and WT littermates. (h) 3D reconstruction of the liver and lungs showing the herniating liver (yellow) compressing on the lower lobes of the lungs (blue). (i) hypoplastic lungs in Tagln-Cre:Tgfbr2 ^flx/flx. The lung has normal lobar structure, however the inferior (arrowheads) and superior (arrows) lobes are smaller. (j) WT littermate showing position of liver and lung, the SVC (arrow) is positioned distant to the right lung (pink). (k) WT littermate lung. R: right. Scale bars:1,000 µm in (a,c,h,j) 500 µm in (b,d–g) & inset 100 µm.