Role of Endothelin Receptor B signaling in the genesis of aganglionic megacolon

内皮素受体 B 信号在无神经节巨结肠发生中的作用

• 批准号: 8045356
• 负责人: MILES L EPSTEIN
• 金额: $ 31.33万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-04-01 至 2013-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8045356
• 关键词: Address; Affect; Alleles; Animal Model; Autologous; Biological Neural Networks; Cell Count; Cell Culture Techniques; Cell Transplants; Cell physiology; Cells; Colon; Congenital Megacolon; Data; Defect; Development; Disease; Disease model; Embryo; Endothelin; Endothelin B Receptor; Endothelin Receptor; Endothelin-3; Enteral; Enteric Nervous System; Environment; Excision; Exhibits; Failure; Fetus; Fluorescence-Activated Cell Sorting; Ganglia; Gastrointestinal Motility; Gastrointestinal tract structure; Gene Proteins; Genes; Genotype; Goals; Health; Hindgut; Image; Implant; In Vitro; Individual; Intercalated Cell; Intestines; Knock-out; Label; Learning; Left; Life; Ligands; Link; Mediating; Mesenchymal; Mesenchyme; Methods; Modeling; Molecular; Motor; Motor Activity; Movement; Mus; Mutation; Myenteric Plexus; Neural Crest; Neural Crest Cell; Neural tube; Neuroglia; Neurons; Operative Surgical Procedures; Partner in relationship; Patients; Pharyngeal structure; Phenotype; Preparation; Principal Investigator; Process; Proliferating; Proteins; Relative (related person); Reporting; Research Personnel; Role; Signal Transduction; Small Intestines; Smooth Muscle Myocytes; Stem cells; Testing; Time; Tissues; Transgenic Organisms; Transplantation; Two-Dimensional Gel Electrophoresis; cell motility; cohort; implantation; in vivo; migration; motility disorder; mutant; nerve supply; neuronal circuitry; novel; novel strategies; postnatal; programs; protein expression; public health relevance; receptor

DESCRIPTION (provided by applicant): The enteric nervous system (ENS) develops primarily from a cohort of neural crest- derived cells (ENCCs) that enter at the pharynx before advancing caudally through the developing gut. The advance of ENCCs through the developing gut requires their coordination of proliferation, differentiation, and migration. Defects in any of these cellular processes alone or combined are sufficient to stop ENCC advance. When ENCCs do not complete their advance through the colon, a region of the gut is left without intrinsic innervation and is representative of Hirschprung's disease (HD). One set of mutations commonly associated with HD is found in genes encoding the ligand (EDN3), or its receptor endothelin-B (EDNRB). The receptor is expressed in both ENCCs and gut mesenchyme. To date, the precise cellular defect and molecular mechanism responsible for EDNRB-mediated HD remain unknown. To address these issues we have utilized a novel conditional floxed allele that allows us to knockout ednrb expression in neural crest cells only. By mating our floxed ednrb mice with Wnt1-Cre transgenics we delete ednrb specifically in neural crest-derived ENCC. In addition, the ENCCs also express the yellow fluorescent protein, allowing their movement to be recorded. Our studies have revealed two previously unrecognized cellular defects related to defective EDNRB signaling. First, the ENCC advance in null ednrb embryos is delayed shortly after neural crest cells enter the gut. Second, as null ednrb ENCCs reach the colon, they display aberrant trajectories and defective migration. Our data indicate that these two defects are related; the initial delay results in ENCCs entering the colon a day later than normal. We hypothesize that the initial delay is caused by pre- enteric neural crest defect, and the subsequent failure to advance in the hindgut results from changes in the hindgut environment. The first specific aim is to determine the cellular defect responsible for the initial delay of ENCC advance in the null mutant. The second aim is to determine the mechanism for defective ENCC migration in the colon. The third specific aim is to determine whether ENCCs transplanted into the aganglionic postnatal hindgut can restore coordinated motor function to the hindgut. Together, these studies will identify cellular defects responsible for HD resulting from the absence of EDNRB signaling and determine the feasibility of establishin coordinated motor function with transplanted null ednrb ENCCs. PUBLIC HEALTH RELEVANCE: Our goals are to elucidate the mechanisms underlying Hirschsprung's disease and to devise a method to provide functional innervation to the region of aganglionic colon. These studies will benefit patients with Hirchsprung's disease and other gastrointestinal motility disorders because providing functional innervation is a superior alternative to surgical excision, which leaves patient with lifelonggastrointestinal problems. The information gained here will also be relevant to transplantation of other tissues including neurons.

描述（由申请人提供）：肠神经系统（ENS）主要由一组神经嵴衍生细胞（ENCC）发育而成，这些细胞在通过发育中的肠道向尾侧推进之前进入咽部。ENCC通过发育中的肠道的推进需要它们的增殖、分化和迁移的协调。这些细胞过程中的任何一个单独或组合的缺陷都足以阻止ENCC的进展。当ENCC没有完成其通过结肠的推进时，肠道的一个区域没有内在的神经支配，并且是Hirschprung病（HD）的代表。通常与HD相关的一组突变发现于编码配体（EDN 3）或其受体内皮素-B（EDNRB）的基因中。该受体在ENCC和肠间充质中表达。迄今为止，负责EDNRB介导的HD的精确细胞缺陷和分子机制仍然未知。为了解决这些问题，我们已经利用了一种新的条件floxed等位基因，使我们能够敲除ednrb表达的神经嵴细胞。通过将我们的floxed ednrb小鼠与Wnt 1-Cre转基因小鼠交配，我们在神经嵴来源的ENCC中特异性地删除ednrb。此外，ENCC还表达黄色荧光蛋白，允许记录它们的运动。我们的研究揭示了两个以前未被认识到的细胞缺陷有关的缺陷EDNRB信号。首先，在无效ednrb胚胎中的ENCC进展在神经嵴细胞进入肠道后不久被延迟。其次，当无效ednrb ENCC到达结肠时，它们显示出异常的轨迹和有缺陷的迁移。我们的数据表明，这两个缺陷是相关的;最初的延迟导致ENCC进入结肠比正常晚一天。我们假设最初的延迟是由肠前神经嵴缺陷引起的，随后的后肠推进失败是由后肠环境的变化引起的。第一个具体目标是确定在无效突变体中负责ENCC推进的初始延迟的细胞缺陷。第二个目的是确定有缺陷的ENCC在结肠中迁移的机制。第三个具体目标是确定ENCC移植到无神经节的出生后后肠是否可以恢复协调的后肠运动功能。总之，这些研究将确定因EDNRB信号传导缺失而导致HD的细胞缺陷，并确定移植无效ednrb ENCC建立协调运动功能的可行性。公共卫生关系：我们的目标是阐明先天性巨结肠的发病机制，并设计一种方法来提供功能性神经支配的区域无神经节结肠。这些研究将使患有先天性巨结肠和其他胃肠动力障碍的患者受益，因为提供功能性神经支配是外科切除的一种上级选择，外科切除会给患者留下终身的胃肠问题。这里获得的信息也将与包括神经元在内的其他组织的移植有关。