Cell-Cell Interactions during intestinal development

描述(申请人提供)：肠道巨大的吸收表面积主要是由其长度以及数百万指状绒毛突起提供的广泛的表面放大作用造成的。导致肠道表面积显著丧失的先天性或获得性病变严重损害了肠道吸收营养的能力，并可能危及生命。在过去的项目期间，工作已经开始在小鼠模型中识别控制子宫内绒毛形成的一些细胞和分子机制。胚胎14.5天到胚胎15.5天(小鼠)之间的时间特别重要，因为在这段时间里，上皮和底层间充质中受控的形态发生重塑导致绒毛的出现。支持这些研究的工作假设是：在E14.5之前，BMP信号积极地抑制绒毛发育。在胚胎14.5天，上皮HH信号通过促进间充质簇的形成启动绒毛发育。簇模式通过图灵系统进行，与这些簇的血管连接先于这些簇，并且是绒毛出现所必需的。重要的是，由于所有这些事件的形态特征也存在于人类肠道中，很可能在小鼠模型中发现的大多数信号范例将适用于人类肠道。这项建议利用遗传小鼠模型以及一种新的肠道外植体培养系统来机械地剖析这些表面生成过程中涉及的相互关联的信号和图案化事件。此外，一种新的软件工具可以高效地识别基因组DNA中的HH反应信号增强子，这将有助于识别HH靶基因。其具体目的是1)确定BMP信号如何控制绒毛形成的能力，并确定哪个组织(上皮或间充质)发挥这种控制作用；2)识别间充质簇形成过程中的HH靶基因；3)确定血管要素之间的关系，形成簇和簇模式。通过对这些相互关联的过程的详细分析，这些研究的目标是对肠道吸收表面的形成有新的认识。从器官的细胞成分中进行生物工程的能力不仅需要我们阐明对形态发生和细胞命运决定至关重要的分子信号，而且我们还需要了解管理组成器官的功能单元模式的规则。这项关于新生绒毛单位(上皮、间充质、血管系统)信号串扰的独特研究将对我们理解肠绒毛最初是如何在胚胎中形成的产生重大影响。 与公共健康相关：小肠表面被称为绒毛的指状突起高度缠绕；这种扩大的表面积对有效吸收营养至关重要。由于先天性肠道缺陷或因病理或外科事件造成的肠道表面积的丧失，可能危及生命。从肠道的细胞成分进行生物工程的能力不仅需要我们阐明对形态发生和细胞命运决定至关重要的分子信号，而且我们还需要了解管理组成器官的功能单元模式的规则。目前，关于绒毛是如何在子宫中生成的知之甚少。通过对扰乱绒毛形成的小鼠模型的研究和对一种新的肠道外植体培养系统的分析，我们建议剖析导致绒毛形成的分子过程。这项关于新生绒毛单位(上皮、间充质、血管系统)信号串扰的独特研究将对我们理解肠绒毛最初是如何在胚胎中形成的产生重大影响。

DESCRIPTION (provided by applicant): The impressively large absorptive surface area of the intestine is largely contributed by its length as well as the extensive surface amplification provided by millions of fingerlike villus projections. Congenital or acquired pathologies that result in significant loss of this intestinal surface area seriously compromise the ability of the intestine to absorb nutrients and can be life threatening. Work in the past project period has begun to identify some of the cellular and molecular mechanisms that control the formation of villi in utero in the murine model. The period between embryonic day (E) 14.5 and E15.5 (in the mouse) is particularly important since during this time, controlled morphogenic remodeling in both the epithelium and the underlying mesenchyme results in the emergence of villi. The Working Hypothesis underlying these studies is that: Villus development is actively inhibited by Bmp signaling prior to E14.5. At E14.5, epithelial Hh signals initiate villus development by promoting the formation of mesenchymal clusters. Cluster patterning proceeds via a Turing system and vascular coupling to these clusters precedes and is required for villus emergence. Importantly, since the morphological hallmarks of all of these events are also present in the human intestine, it is likely that the majority of the signaling paradigms uncovered in the mouse model will be applicable to the human intestine. This proposal makes use of genetic mouse models as well as a novel intestinal explant culture system to mechanistically dissect the interconnected signaling and patterning events involved in these surface- generating processes. Additionally, a new software tool that can recognize Hh-responsive signaling enhancers in genomic DNA with high efficiency will aid in the recognition of Hh target genes. The Specific Aims are designed to 1) Determine how Bmp signaling controls competence to form villi and establish which tissue (epithelium or mesenchyme) exerts this control; 2) Identify Hh target genes during the formation of mesenchymal clusters; and 3) Determine the relationship between vascular elements, forming clusters and cluster pattern. Through detailed analysis of these linked processes, the goal of these studies is to gain new insight into the formation of the intestinal absorptive surface. The ability to bioengineer organs from their cellular components will require not only that we elucidate the molecular signals that are important for morphogenesis and cell fate determination, but also that we understand the rules that govern the patterning of the functional units that comprise the organ. The unique focus of this investigation on signaling crosstalk in the nascent villus unit (epithelium, mesenchyme, vasculature) will have a major impact on our understanding of how intestinal villi are first formed in the embryo. PUBLIC HEALTH RELEVANCE: The surface of the small intestine is highly convoluted by finger-like projections called villi; this extended surface area is critical for efficient nutrient absorption. Loss of intestinal surface area either by congenital intestinal defects or by pathological or surgical events, can be life threatening. The ability to bioengineer the intestine from its cellular components will require not only that we elucidate the molecular signals that are important for morphogenesis and cell fate determination, but also that we understand the rules that govern the patterning of the functional units that comprise the organ. Currently, little is known about how villi are generated in utero. Through the study of mouse models with perturbed villus formation and through the analysis of a novel intestinal explant culture system, we propose to dissect the molecular processes responsible for villus emergence. The unique focus of this investigation on signaling crosstalk in the nascent villus unit (epithelium, mesenchyme, vasculature) will have a major impact on our understanding of how intestinal villi are first formed in the embryo.