This application for an Administrative Supplement to our active R01-HL122906 is being submitted in response to PA-20-272 in accordance with NOT-OD-22-137 (“Availability of Administrative Supplements for the INCLUDE - INvestigation of Co-occurring conditions across the Lifespan to Understand Down syndrome - Project”). Significance of application in relation to Down syndrome (DS): Atrioventricular septal defects (AVSDs) are complex congenital heart defects but relatively rare in the general population. Approximately 0.05% of babies born have an AVSD. They are, however, very common in DS patients (35-40% of DS babies are diagnosed with an AVSD). In this project we will address one of the priorities of the National Heart, Lung, and Blood Institute (NHLBI) as we will, using animal models, focus on the morphological events involved in the pathogenesis of AVSDs which, if untreated, can be the cause of death during the first year of life. Abstract: The study of heart development as it relates to congenital heart disease has been the focus of the Wessels lab for years. This proposal is a logical extension of studies that were initiated after the discovery of the Dorsal Mesenchymal Protrusion (DMP) by the PI. We identified the DMP as a structure critically important for the development of the atrioventricular (AV) mesenchymal complex and determined that the DMP was derived from the Second Heart Field (SHF). We developed the hypothesis that the DMP could be involved in the pathogenesis of AVSDs. Indeed, in our study of the Trisomy 16 (Ts16) mouse, a model for DS, we found that AVSDs were associated with hypoplasia of the DMP. Furthermore, we demonstrated that experimentally deleting candidate genes believed to be essential for proper SHF development (Alk3 and Smo) resulted in perturbation of DMP formation and AVSD pathogenesis. Analysis of the SHF;Smo mouse revealed downregulation of several genes including Sox9. Based on our earlier studies, we predicted that Sox9 would play a critical role in DMP development and AV septation. Surprisingly, while SHF-specific Sox9 deletion resulted in AVSD in 50% of SHF;Sox9 offspring, it was not the result of DMP hypoplasia, but rather associated with failed development of the mesenchymal cap. Importantly, virtually all specimens inspected had developed a ventricular septal defect (VSD). This (published) observation in combination with results of our studies on AVSD pathogenesis (see: Research Plan) have led to new theories about the role of the anterior and posterior SHF (aSHF and pSHF) in the pathogenesis of the atrial and ventricular defects in the context of DS. We propose experiments with different mouse models, including the TcMAC21 model for DS, to test these hypotheses. In addition, we will initiate single cell RNAseq experiments to gain insight into the molecular mechanisms involved in the pathogenesis of AVSDs, specifically focusing on the role of the embryonic outflow tract in VSD pathogenesis.

Atrioventricular defects (AVSDs) are complex heart malformations found in 35-40% of individuals with Down syndrome. The research proposed in this Supplement is aimed at determining the role of the Second Heart Field in the pathogenesis of the respective morphological components that form part of the AVSD spectrum. Elucidating the molecular and cellular mechanisms underlying the pathogenesis of AVSDs could lead to strategies to prevent and/or treat this severe congenital defect.