Exploring the Role of the anterior SHF in AVSD Pathogenesis

本申请是对我们的有效R 01-HL 122906的行政补充申请， 根据NOT-OD-22-137（“管理补充材料的可用性， 包括-研究整个生命周期中的共现疾病以了解唐氏综合征- 项目”）。 与唐氏综合征（DS）相关的应用意义：房室间隔缺损（AVSD）是 复杂的先天性心脏缺陷，但在一般人群中相对罕见。约0.05%的婴儿 天生就有AVSD。然而，它们在DS患者中非常常见（35-40%的DS婴儿被诊断为患有 AVSD）。在这个项目中，我们将解决国家心脏，肺和血液的优先事项之一。 研究所（NHLBI），因为我们将使用动物模型，专注于参与的形态学事件， AVSD的发病机制，如果不治疗，可能是生命第一年内死亡的原因。 翻译后摘要：心脏发育的研究，因为它涉及到先天性心脏病一直是重点， 多年来一直在实验室工作。这一提议是对发现后开始的研究的逻辑延伸。 主要研究者进行的背侧间充质干细胞（PMM）。我们认为， 房室（AV）间充质复合体的发育，并确定房室间充质复合体是由 第二心脏领域（SHF）。我们提出了一个假设，即大脑皮层可能参与了 AVSD的发病机制。事实上，在我们对16三体（Ts 16）小鼠的研究中，我们发现， AVSD与小脑发育不全有关。此外，我们证明，实验删除 候选基因被认为是必要的适当的SHF发展（Alk 3和Smo）导致扰动 与AVSD发病机制的关系。对SHF;Smo小鼠的分析显示， 包括Sox 9基因。基于我们早期的研究，我们预测Sox 9将在细胞凋亡中发挥关键作用。 发展和AV分隔。令人惊讶的是，尽管SHF特异性Sox 9缺失导致50%的人发生AVSD， SHF; Sox 9后代，它不是胚胎发育不全的结果，而是与发育失败有关。 间充质细胞帽重要的是，几乎所有检查的样本都出现了室间隔缺损 （VSD）。这一（已发表的）观察结果与我们对AVSD发病机制的研究结果相结合（见： 研究计划）导致了关于前SHF和后SHF（aSHF和pSHF）在 DS背景下心房和心室缺陷的发病机制。我们提出了不同的实验 小鼠模型，包括DS的TcMAC 21模型，以检验这些假设。此外，我们还将启动一个 细胞RNAseq实验以深入了解AVSD发病机制中涉及的分子机制， 特别关注胚胎流出道在VSD发病机制中的作用。

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.