TGFβ2 and TGFβ3 Have Separate and Sequential Activities during Epithelial–Mesenchymal Cell Transformation in the Embryonic Heart

Abstract

Heart valve formation is initiated by an epithelial-mesenchymal cell transformation (EMT) of endothelial cells in the atrioventricular (AV) canal. Mesenchymal cells formed from cardiac EMTs are the initial cellular components of the cardiac cushions and progenitors of valvular and septal fibroblasts. It has been shown that transforming growth factor beta (TGFbeta) mediates EMT in the AV canal, and TGFbeta1 and 2 isoforms are expressed in the mouse heart while TGFbeta 2 and 3 are expressed in the avian heart. Depletion of TGFbeta3 in avian or TGFbeta2 in mouse leads to developmental defects of heart tissue. These observations raise questions as to whether multiple TGFbeta isoforms participate in valve formation. In this study, we examined the localization and function of TGFbeta2 and TGFbeta3 in the chick heart during EMT. TGFbeta2 was present in both endothelium and myocardium before and after EMT. TGFbeta2 antibody inhibited endothelial cell-cell separation. In contrast, TGFbeta3 was present only in the myocardium before EMT and was in the endothelium at the initiation of EMT. TGFbeta3 antibodies inhibited mesenchymal cell formation and migration into the underlying matrix. Both TGFbeta2 and 3 increased fibrillin 2 expression. However, only TGFbeta2 treatment increased cell surface beta-1,4-galactosyltransferase expression. These data suggest that TGFbeta2 and TGFbeta3 are sequentially and separately involved in the process of EMT. TGFbeta2 mediates initial endothelial cell-cell separation while TGFbeta3 is required for the cell morphological change that enables the migration of cells into the underlying ECM.

An improved protocol for the isolation and cultivation of embryonic mouse myocytes

“…5) in vitro. Both of these growth factors are known to be present within the myocardium of the developing heart and to be necessary for proper heart development (Dickson et al 1993;Boyer et al 1999;Tomanek et al 1999;Giordano et al 2001;Camenisch et al 2002). From these results, we conclude that our in vitro embryonic myocyte cultures recapitulate cardiac gene expression and growth factor production related to proliferative potential and myocyte maintenance.…”

Cilia and coordination of signaling networks during heart development

“…In line with this, mutations in the NODAL and LEFTY2 genes cause heterotaxy and/or isolated CHD in humans. 94,95 In vitro collagen gel experiments, using AV or OFT explants, suggest that TGFβ signaling is involved in EMT and EndoMT during the formation of endocardial cushions, [96][97][98] and targeted mutation of genes encoding TGFβ ligands causes defects of the OFT (double outlet right ventricle (DORV)), abnormal semilunar valves, and AV cushions, supporting an involvement in development of the endocardial cushions. 99,100 Null mutants for the TGFβ receptor genes, Tgfbr1 and Tgfbr2, are embryonic lethal in mice, but tissue specific deletion of TGFβ receptor genes support a role for TGFβ signaling in development of the OFT.…”

Co-ordinating Notch, BMP, and TGF-β signaling during heart valve development

“…In chick explants, addition of TGFβ1 and TGFβ3 induces EMT in the absence of the normally required AV myocardium [70,80]. Addition of neutralizing antibodies for TGFβ2 or TGFβ3 to chick AVC explants inhibits activation of endocardial cells (TGFβ2), and formation and migration of cushion mesenchymal cells (TGFβ3) [81]. This data indicates that TGFβ signalling plays a significant role in chick cardiac cushion development and therefore may play a similar role in mouse valve development.…”