Purpose: Malformations affecting the outflow of the heart are a major cause of morbidity and mortality in childhood. While many of these malformations occur sporadically, studies of families with congenital heart defects, alongside animal studies, have revealed that phenotypically discrete heart malformations can have diverse causes, resulting from disruption of a number of different genes, embryonic lineages or developmental processes. Clarifying the fundamental processes that underpin outflow development is essential to understand this complexity. Planar cell polarity (PCP) is the mechanism by which cells orient themselves in the plane of an epithelium or during directed cell migration, and is regulated by a highly conserved signalling pathway. Mutations in the PCP gene Vangl2, as well as in other key components of the pathway, cause a spectrum of cardiac outflow tract defects. Our objective was to establish which of the several cell lineages that form the developing heart require Vangl2-regulated PCP signalling and why disruption leads to outflow tract defects.
Methods and Results: Using a new conditionally floxed allele we show that Vangl2 is required solely within the second heart field (SHF) to direct normal outflow tract lengthening, a process that is required for septation and normal alignment of the aorta and pulmonary trunk with the ventricular chambers. Thus, in the absence of Vangl2 in the SHF the mice develop double outlet right ventricle and ventricular septal defects. Analysis of a range of markers of polarised epithelial tissues showed that in the normal heart, undifferentiated SHF cells move from the dorsal pericardial wall into the distal outflow tract where they acquire an epithelial phenotype, before moving proximally where they differentiate into cardiomyocytes. Thus there is a transition zone in the distal outflow tract where SHF cells become more polarised, turn off progenitor markers and start to differentiate to cardiomyocytes. Membrane-associated Vangl2 marks the proximal extent of this transition zone and, moreover, in the absence of Vangl2, the SHF-derived cells are abnormally polarised cells and do not acquire an epithelial phenotype. As a consequence they prematurely down-regulate the SHF-progenitor marker Isl1 and differentiate to cardiomyocytes. This leads to a shortened outflow tract.
Conclusion: Vangl2-regulated polarisation and consequent epithelialisation of the distal outflow tract is essential to lengthen the tubular outflow vessel. This leads to the double outlet right ventricle and ventricular septal defects.