Cellular and Molecular Analysis of Body Wall Closure

体壁闭合的细胞和分子分析

• 批准号: 9978529
• 负责人: TREVOR J WILLIAMS
• 金额: $ 23.33万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-01 至 2022-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9978529
• 关键词: Abdomen; Address; Affect; Anterior eyeball segment structure; Bioinformatics; Biological Models; Birth; Categories; Cell Lineage; Cell Nucleus; Cells; Chest; Child; Chromatin; Communication; Complement; Congenital Abnormality; Congenital omphalocele; Data; Data Set; Defect; Development; Developmental Process; Ectoderm; Embryo; Embryonic Development; Enhancers; Expression Profiling; Face; Family; Future; Gastroschisis; Gene Expression; Gene Expression Profile; Gene Expression Profiling; Gene Expression Regulation; Genes; Genetic; Genetic Transcription; Heart; Human; Human body; In Situ; Infant; Knockout Mice; Knowledge; Life; Link; Live Birth; Liver; Lung; Manuscripts; Mesenchymal; Mesenchyme; Mesoderm; Modeling; Molecular Analysis; Morphogenesis; Mus; Mutant Strains Mice; Mutate; Nerve Tissue; Parents; Pathogenesis; Pathology; Patients; Peripheral Nervous System; Population; Process; Proxy; Quality of life; Regulator Genes; Regulatory Element; Resources; Role; Signal Transduction; Structure; TFAP2A gene; Techniques; Testing; Time; TimeLine; Tissues; Writing; amnion; base; cell type; conditional knockout; enhancer-binding protein AP-2; insight; malformation; migration; mouse model; mutant; novel; promoter; public repository; single-cell RNA sequencing; theories; transcription factor; transcriptome

Mammalian ventral body wall closure defects, including gastroschisis, thoracoabdominoschisis, and omphalocele are linked with the morphogenesis of the ventral body wall, the amnion, and the umbilical ring. Malformations of the ventral body wall comprise one of the leading categories of human birth defects and are present in about one out of every 2000 live births. Although the occurrence of these defects is relatively common, there is a surprising lack of knowldege concerning the development and closure of the ventral body wall in mouse or human. This field is further complicated by the array of theories on the pathogenesis of human body wall defects that have not been experimentally tested. This proposal aims to produce a paradigm shift in our understanding of mammalian ventral body closure that can provide a mechanistic framework and comprehensive resource for future understanding of how this process goes awry because of genetic and/or environmental causes. The transcription factor AP-2, encoded by the gene Tfap2a, has an essential role in mouse body wall closure and previous studies have shown that it regulates ectodermal, mesenchymal, peripheral nervous system, and mesodermal interactions that drive development of the ventral body wall. Tfap2a null mice have a severe form of ventral closure defect, a thoracoabdominoschisis, in which the ventral covering of the chest and abdomen fails to form so that the heart, lungs, liver, and gut are exposed. Critically, this is one of the few simple mouse models that gives a fully penetrant and consistent body wall closure defect that can be used to understand how this important developmental process can fail. In the first Aim, relevant tissue will be collected from control and Tfap2a null mice at specific embryonic times and subjected to single cell RNA seq (scRNAseq) analysis. This will identify the normal gene expression patterns of mouse body wall closure as well as cell types and genes that are impacted by loss of Tfap2a. Several additional genes are known to affect body wall closure and we will be able to attribute these genes to relevant tissue populations to understand how they may be influencing this critical developmental process. In the second Aim, we will perform scATACseq analysis on all control time points as well as the most relevant time points for the mutant. We will then integrate all the scRNAseq and scATACseq data to identify the tissues, cell types, and gene expression signatures that are normally associated with body wall closure. Further, we will identify how chromatin accessibility in the body wall is altered by loss of Tfap2a. Verification analyses will be performed to identify critical changes in cell populations and gene expression profiles associated with normal and abnormal body wall closure. Subsequently, the information from these studies will be synthesised into a novel and powerful model for normal body wall closure that can serve as an important framework for future understanding of this important class of human birth defect.

哺乳动物腹侧体壁闭合缺陷，包括腹裂、胸腹裂和 脐膨出与腹侧体壁、羊膜和脐环的形态发生有关。 腹侧体壁畸形是人类出生缺陷的主要类别之一， 尽管这些缺陷的发生率相对较高，但每 2000 名活产婴儿中就有 1 人存在这种缺陷。 常见的是，人们对腹侧体的发育和闭合知之甚少。 小鼠或人类的墙壁因一系列关于其发病机制的理论而变得更加复杂。 该提案旨在产生一个范例。 我们对哺乳动物腹侧身体闭合的理解发生了转变，它可以提供机械框架和 未来了解这一过程如何因遗传和/或因素而出错的综合资源 由 Tfap2a 基因编码的转录因子 AP-2 在环境因素中发挥着重要作用。 小鼠体壁闭合和先前的研究表明它调节外胚层、间质、 周围神经系统和中胚层相互作用驱动腹侧体壁的发育。 Tfap2a 缺失小鼠具有严重的腹侧闭合缺陷，即胸腹劈裂，其中腹侧闭合缺陷 胸部和腹部的覆盖物无法形成，因此心脏、肺、肝脏和肠道都暴露在外。 这是为数不多的能够提供完全渗透且一致的体壁闭合缺陷的简单小鼠模型之一 可以用来理解这个重要的发展过程如何会在第一个目标中失败。 将从特定胚胎时间的对照小鼠和 Tfap2a 缺失小鼠收集组织并进行单次处理。 细胞 RNA seq (scRNAseq) 分析这将识别小鼠体壁的正常基因表达模式。 关闭以及受 Tfap2a 丢失影响的细胞类型和基因。 已知会影响体壁闭合，我们将能够将这些基因归因于相关组织群体 在第二个目标中，我们将了解它们如何影响这一关键的发展过程。 对所有对照时间点以及突变体最相关的时间点进行 scATACseq 分析。 然后我们将整合所有 scRNAseq 和 scATACseq 数据来识别组织、细胞类型和基因 通常与体壁闭合相关的表达特征此外，我们将确定如何进行。 体壁中染色质的可及性因 Tfap2a 的缺失而改变，将进行验证分析。 识别与正常和异常相关的细胞群和基因表达谱的关键变化 随后，这些研究的信息将被综合成一种新颖的和 正常体壁闭合的强大模型可以作为未来理解的重要框架 这一类重要的人类出生缺陷。