Cellular and Molecular Analysis of Body Wall Closure

体壁闭合的细胞和分子分析

• 批准号: 10133115
• 负责人: TREVOR J WILLIAMS
• 金额: $ 19.44万
• 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/10133115
• 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; 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.

哺乳动物腹体壁闭合缺陷，包括腹裂、胸腹裂和 脐膨出与腹体壁、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型之一， 大约每2000个活产婴儿中就有一个。虽然这些缺陷的发生相对 常见的是，令人惊讶的是缺乏有关腹体发育和闭合的知识 在小鼠或人中的壁。这一领域进一步复杂化的一系列理论的发病机制， 未经实验测试的人体壁缺陷。这一建议旨在产生一种范式， 我们对哺乳动物腹侧体闭合理解的转变，可以提供一个机械框架， 全面的资源，以便将来了解这一过程是如何因为遗传和/或 环境原因。由基因Tfap 2a编码的转录因子AP-2 β在以下方面具有重要作用： 小鼠体壁闭合和先前的研究表明，它调节外胚层，间充质， 外周神经系统和中胚层的相互作用，驱动腹体壁的发展。 Tfap 2a基因敲除小鼠具有严重形式的腹侧闭合缺陷，即胸腹裂，其中腹侧闭合缺陷的发生率为100%。 胸腹不能覆盖，心、肺、肝、肠外露。重要的是， 这是少数几个简单的小鼠模型之一，它给出了完全渗透和一致的体壁闭合缺陷。 可以用来理解这个重要的发育过程是如何失败的。在第一个目标中， 在特定的胚胎时间从对照和Tfap 2a缺失小鼠收集组织， 细胞RNA测序（scRNAseq）分析。这将确定小鼠体壁的正常基因表达模式 关闭以及受Tfap 2a损失影响的细胞类型和基因。几个额外的基因是 已知影响体壁闭合，我们将能够将这些基因归因于相关组织群体， 了解他们如何影响这一关键的发展过程。在第二个目标中，我们将 对所有对照时间点以及突变体的最相关时间点进行scATACseq分析。 然后，我们将整合所有的scRNAseq和scATACseq数据，以识别组织、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 体壁中的染色质可及性因Tfap 2a的缺失而改变。将进行验证分析， 确定与正常和异常相关的细胞群和基因表达谱的关键变化 体壁闭合随后，这些研究的信息将被综合成一部小说， 一个强大的正常体壁闭合模型，可以作为未来理解的重要框架 这类重要的人类出生缺陷的原因。