Gdnf and endothelin-3 regulate colorectal enteric nervous system development

Gdnf 和内皮素 3 调节结直肠肠神经系统发育

• 批准号: 8277360
• 负责人: ALLAN M GOLDSTEIN
• 金额: $ 37.73万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-06-01 至 2014-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8277360
• 关键词: Address; Affect; Animal Model; Award; Biological Assay; Biological Models; Birds; Cecum; Cells; Child; Chimera organism; Chronic; Cloaca Chamber; Colon; Colonic Aganglionosis; Colorectal; Complex; Congenital Disorders; Congenital Megacolon; Data; Defect; Development; Diagnosis; Disease; Distal; Embryo; Embryonic Development; Endothelin B Receptor; Endothelin-3; Enteral; Enteric Nervous System; Equilibrium; Etiology; Excision; Ganglia; Gastrointestinal tract structure; Gene Expression; Gene Silencing; Genes; Genetic; Goals; Health; Hindgut; Human; Immunohistochemistry; In Vitro; Intestinal Diseases; Intestinal Motility; Intestinal Obstruction; Intestines; Large Intestine; Ligands; Modeling; Molecular; Motor Activity; Mutation; Nerve; Nervous system structure; Neural Crest; Neural Crest Cell; Neural tube; Neuroglia; Neuronal Differentiation; Neurons; Neuropathy; Newborn Infant; Organ Culture Techniques; Pathway interactions; Patients; Phenotype; Population; Quail; RNA Interference; Retroviridae; Rodent; Role; Signal Pathway; Signal Transduction; Staging; Stem cells; Testing; Work; base; cost; improved; in vivo; inhibitor/antagonist; insight; loss of function; migration; mouse model; nervous system development; overexpression; prevent; progenitor; research study; vector

DESCRIPTION (provided by applicant): The enteric nervous system (ENS) is a complex network of neurons and glia present in the bowel wall and critical for regulating intestinal motility. Abnormal development of the ENS is the underlying cause of Hirschsprung's disease, a congenital intestinal disorder caused by the absence of enteric ganglia, usually in the distal colorectum, and leading to severe intestinal obstruction in newborns. Defects in the Ret and endothelin receptor B (EdnrB) signaling pathways are required for ENS development and are responsible for many cases of Hirschsprung's disease. However, how these key pathways function in vivo to regulate ENS development in the colorectum, and why mutations cause colorectal aganglionosis in humans, is poorly understood. The broad objective of this proposal is to determine the molecular mechanisms that regulate colorectal ENS development in order to identify potential targets for the treatment of Hirschsprung's disease and other congenital intestinal neuropathies. We hypothesize that Ret and EdnrB signaling act coordinately to influence the migration, proliferation, and differentiation of ENS progenitor cells as they cross the cecal and cloacal regions and that this function is critically important for ENS colonization of the colon. To test this hypothesis, we will use the avian embryo to activate and inhibit gene expression in vivo in order to study the role of these signaling pathways. The major advantages of the avian model system are the ease of performing experimental manipulations throughout embryogenesis and the ability to carry out genetic gain- and loss-of-function studies more quickly and at lower cost than in rodents. We propose to use replication-competent retrovirus for gene overexpression, vector-based RNAi for gene silencing, and organ culture assays. Specific Aim I will establish the role of EdnrB signaling on the migration, survival, proliferation, and differentiation of vagal and sacral ENS progenitor cells during formation of the distal intestinal ENS. Specific Aim II will examine the role of Ret signaling in the etiology of colorectal aganglionosis by focusing on the function of this pathway on migration and proliferation of ENS progenitors as they cross the cecal and cloacal regions. Specific Aim III will use an organ culture model of colorectal aganglionosis that we generate by inhibiting EdnrB signaling in the distal intestine. Activators and inhibitors of Ret activity will be added to EdnrB-deficient intestine in order to rescue the aganglionic phenotype by modulating the balance of activity between these two pathways. These experiments will establish the role of Ret and EdnrB signaling in the distal ENS, provide new insights into mechanisms underlying colorectal ENS development, and identify potential targets for the treatment of neurointestinal disorders. PUBLIC HEALTH RELEVANCE: Abnormalities in the development of the nerves that control intestinal function are a major cause of intestinal disease in children. Hirschsprung's disease, for example, is caused by the congenital absence of nerve cells in the large intestine, leading to severe bowel obstruction in newborns and requiring surgical removal of the affected intestine. The goal of this proposal is to understand the mechanisms that control development of the intestinal nervous system in order to improve our ability to diagnose and treat children born with these chronic and disabling conditions.

描述（由申请人提供）：肠神经系统（ENS）是肠壁中存在的神经元和神经胶质的复杂网络，对于调节肠道运动至关重要。 ENS的异常发育是Hirschsprung氏病的根本原因，这是由于缺乏肠神经节而引起的先天性肠道疾病，通常在远端的结肠菌中，并导致新生儿的严重肠道阻塞。 ENS发育需要RET和内皮素受体B（EDNRB）信号通路的缺陷，并负责许多Hirschsprung病例。但是，这些关键途径如何在体内发挥作用，以调节结直肠的发育，以及为什么突变引起人类大肠动症的人体病的原因很众所周知。该提案的广泛目的是确定调节结直肠发育的分子机制，以确定治疗赫希斯普伦病和其他先天性肠道神经病的潜在靶标。我们假设RET和EDNRB信号传导在跨盲肠和泄殖腔区域时会协同影响ENS祖细胞的迁移，增殖和分化，并且该功能对于结肠的ENS定植至关重要。为了检验这一假设，我们将使用鸟类胚胎激活和抑制体内基因表达，以研究这些信号传导途径的作用。鸟类模型系统的主要优点是在整个胚胎发生过程中易于执行实验操作，以及比啮齿动物更快，更低的成本进行遗传增益和功能丧失研究的能力。我们建议将竞争能力的逆转录病毒用于基因过表达，基于载体的RNAi用于基因沉默和器官培养试验。具体目的我将确定EDNRB信号传导在远端ENS的迁移，生存，增殖和分化的迁移，生存，增殖和分化。特定的目标II将通过关注该途径对ENS祖细胞的迁移和扩散的功能，在跨盲肠和泄殖腔区域的迁移和增殖方面，研究RET信号传导在结直肠剧院病的病因中的作用。特定的目标III将使用我们通过抑制远端肠中EDNRB信号传导而产生的结直肠动胞病的器官培养模型。 RET活性的激活剂和抑制剂将被添加到EDNRB缺陷式肠道中，以通过调节这两种途径之间的活性平衡来挽救Aganglionic表型。这些实验将确定RET和EDNRB信号在远端ENS中的作用，提供有关结直肠发育基础机制的新见解，并确定治疗神经局部疾病的潜在靶标。公共卫生相关性：控制肠道功能的神经发展的异常是儿童肠道疾病的主要原因。例如，Hirschsprung氏病是由大肠中神经细胞的先天性缺乏引起的，导致新生儿的严重肠梗阻，需要手术去除受影响的肠道。该提案的目的是了解控制肠道神经系统发展的机制，以提高我们诊断和治疗患有这些慢性和残疾状况的儿童的能力。