Ingression of the cardiac outflow tract wall is disrupted in mice with altered β-catenin, Shh and FGF8 signaling

Previous studies from us and others suggest that the second heart field (SHF)-derived outflow tract (OFT) wall can undergo a cell fate transition to mesenchymal cells within the intercalated valve swelling (ICVS). However, the trajectory of this transition and its intrinsic regulation remain unknown. Here, we investigate cell fate determination in mouse OFT development and identify specific NKX2-5 expression in the inferior midline OFT (imOFT) wall and adjacent ICVS. scRNA-seq of E10.5 OFT revealed a transcriptional signature specific to the imICVS within the heterogeneous cell populations. We observed that cells of the imOFT wall ingress while maintaining myocyte properties, whereas the leading cells initiate a transition to a mesenchymal fate. Furthermore, these ingressing cells exhibit 'lily-bulb' collective cell migration. In addition, we found that β-catenin, Shh and FGF8 signaling are associated with imICVS formation. These results identify a previously unreported type of cell ingression and demonstrate that the imICVS appears to spatially coordinate the distribution of the OFT main cushion derived from neural crest. This study provides new insights into the role of the SHF in OFT septation and the pathogenesis of congenital heart diseases.