Two Distinct Pools of Mesenchyme Contribute to the Development of the Atrial Septum

Abstract

Closure of the primary atrial foramen is achieved by fusion of the atrioventricular cushions with the mesenchymal cap on the leading edge of the muscular primary atrial septum. A fourth component involved is the vestibular spine, originally described by His in 1880 as an intra-cardiac continuation of the extra-cardiac mesenchyme of the dorsal mesocardium. The morphogenesis of this area is of great clinical interest, because of the high incidence of atrial and atrioventricular septal defects. Nonetheless, the origin of the participating components is largely unknown. Here we report that the primary atrial foramen is surrounded in its entirety by mesenchyme derived from endocardium. A second population of mesenchyme not derived from endocardium was observed at the caudal margin of the mesenchymal atrial cap, entirely embedded within the mesenchyme derived from endocardium and contiguous with the mesenchyme of the dorsal mesocardium. Our reconstructions show this second population does indeed take the form of a short spine, albeit that it is the right pulmonary ridge, rather than this spine, that protrudes into the atrial lumen. From the stance of morphological description, therefore, there is little thus far to substantiate the existence of an atrial spine. It is currently thought that the atrioventricular (AV) cushions, along with the primary atrial septum and the mesenchymal cap carried on its leading edge, are the main contributors to the process of atrial septation. A growing body of evidence points to the involvement of a fourth component, namely the vestibular spine ("spina vestibuli"). [1][2][3][4][5][6][7][8] The vestibular spine was initially nominated as playing such a role in 1880, by Wilhelm His the elder. 9 He described the spine as a triangular mesenchymal wedge, which protruded into the lumen of the atrium from a non-muscular area, which he called the "area interposita," in the dorsal wall of the common atrium ( Figure 1, stippled circle). The spine then disappeared from view for more than a century, eventually being retrieved by several workers, 1,2,4 -6,10,11 albeit with disagreements concerning the form and origin of this tissue. To clarify this, we have performed a lineage study using Tie2-Cre mice 12 to label endocardium and endocardium-derived mesenchyme, in combination with threedimensional reconstructions 13 to permit independent evaluation of the structures involved in atrial septation. Materials and MethodsThe Tie2-Cre and R26R transgenic mouse lines have been described previously. 12,14 Detection of ␤-galactosidase activity and immunohistochemistry were performed on 20-m thick cryostat sections. 15 Non-radioactive in situ hybridization analysis was performed on 12-m thick paraffin sections. 16 Three-dimensional visualization and geometry reconstruction of patterns of gene expression was then achieved. 13,17 Movie clips of these reconstructions are available on request. ResultsAt embryonic day (E) 9.5, it is possible to recognize two ridges, of equal size, at the connection of the dorsal m...

Development of the Pulmonary Vein

“…This fact had also not escaped us when we supported, in a series of investigations, the notion that the pulmonary vein did not originate from the systemic venous sinus (Webb et al, 1998a(Webb et al, ,b, 2000(Webb et al, , 2001Soufan et al, 2004;Christoffels et al, 2006). Throughout our studies, the key point of our own argumentation has been that, in terms of morphology, the systemic venous sinus cannot be distinguished as a morphologic entity separate from the remainder of the atrial chambers until after the formation of the so-called venous valves (Anderson et al, 2006). In our opinion, Manner and Merkel (2007) fail to give sufficient emphasis to the investigations, already mentioned above, in which we showed that, at the stages before the systemic venous sinus could be distinguished, and before the appearance of the pulmonary vein, the systemic venous tributaries still are embedded in the mesenchyme of the septum transversum, and not yet incorporated into the pericardial cavity (Soufan et al, 2004;Christoffels et al, 2006).…”

“…Moreover, that structures within the right atrium are marked positively for Pitx2 cannot be used as evidence that the developing pulmonary vein was initially in contact with one of the components of the definitive right atrium, namely the systemic venous sinus, as part of normal embryological development. As is shown by molecular studies, the formation of the primary atrial septum, derived from the right pulmonary ridge, composed of mediastinal myocardium, and marked by Pitx2, walls the orifice of the developing pulmonary vein into the morphologically left atrium (Soufan et al, 2004;Mommersteeg et al, 2006). It is not possible that the atrial septum is derived from the morphologically left side, because this would wall the pulmonary veins, surrounded by the mediastinal or mesocardial mesenchyme, into the right atrium (Fig.…”

Probing chromatin landscape reveals roles of endocardial TBX20 in septation

“…AP1 proteins regulate a variety of processes including cell proliferation and survival, differentiation, growth, migration, and transformation. Intriguingly, conditional deletion of the AP1 family member Jun using Tie2-Cre leads to DORV, VSDs, and valve defects (54), suggesting that perhaps JUN cooperates with TBX20 in endocardial lineages to effect ture required for proper atrioventricular septation (13,53). Notably, this defect was not observed in our analysis of Nfatc1-Cre Tbx20 fl/fl mutants.…”

“…Notably, this defect was not observed in our analysis of Nfatc1-Cre Tbx20 fl/fl mutants. Unlike the atrioventricular cushions and the mesenchymal cap of the atrial septum, the DMP is not derived from Tie2-Cre lineages (13). Therefore, defects in DMP development in Tie2-Cre Tbx20 fl/null mutants suggest a crosstalk between Tbx20-expressing cells of endocardial lineage and myocardial progenitors of the DMP.…”

“…Mutant hearts had ventricular septal defects (VSDs) and ostium primum type atrial septal defects (ASDs) ( Figure 1C), which are considered a type of atrioventricular septal defect (AVSD) (12). Additionally, the dorsal mesenchymal protrusion (DMP), a second heart fieldderived structure that normally appears as a protrusion at the right pulmonary ridge (13), appeared to be lacking in the mutant hearts ( Figure 1C). At E10.5 and E13.5, no defects in vascular patterning were observed (Supplemental Figure 1; supplemental material available online with this article; doi:10.1172/JCI85350DS1).…”