Characterizing neurogenic progenitors in the adult intestine

成人肠道神经源性祖细胞的特征

• 批准号: 10545000
• 负责人: ALLAN M GOLDSTEIN
• 金额: $ 36.33万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-01-01 至 2024-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10545000
• 关键词: Ablation; Adult; Agonist; Birth; Cell Therapy; Cells; Chronic; Clinical; Colitis; Colon; Colonic Diseases; Congenital Megacolon; Constipation; Coupled; Data; Digestive System Disorders; Disease; Distal; Dominant-Negative Mutation; Enteral; Enteric Nervous System; Epigenetic Process; Esophageal achalasia; Esthesia; Excision; Experimental Models; Functional Gastrointestinal Disorders; Functional disorder; Gastroparesis; Gene Expression; Genetic Transcription; Glial Differentiation; Goals; Health; Human; Hyperplasia; Immunity; In Vitro; Injury; Intestinal Pseudo-Obstruction; Intestines; Irritable Bowel Syndrome; Knowledge; Lead; Life; Mediating; Methodology; Modeling; Morbidity - disease rate; Mucositis; Mus; Mutation; Nervous System Physiology; Neuroglia; Neurons; Pathologic; Pathway interactions; Population Heterogeneity; Process; Regulation; Reporter; Rodent; Role; Serotonin; Signal Pathway; Signal Transduction; Source; Structure; Subgroup; Surface; System; Testing; Transgenic Organisms; absorption; cell motility; density; design; experimental study; gastrointestinal; gastrointestinal function; gastrointestinal infection; genetic analysis; gut inflammation; improved; in vitro Assay; in vivo; inhibitor; insight; live cell imaging; motility disorder; mouse model; nerve stem cell; nervous system development; neural; neurogenesis; novel strategies; pharmacologic; postnatal; progenitor; response; serotonin receptor; single-cell RNA sequencing; stem cell biology; stem cells; transcriptome; transdifferentiation

Project Summary The enteric nervous system (ENS) is of fundamental importance to human health through its regulation of all aspects of gastrointestinal (GI) function, most notably gut motility. Congenital or acquired abnormalities of the ENS consequently can lead to serious functional GI disorders, including esophageal achalasia, gastroparesis, intestinal pseudo-obstruction, irritable bowel syndrome, Hirschsprung disease, and slow transit constipation. The adult intestine is known to possess neuronal progenitors, but their role and the mechanisms that activate them are unknown. We and others have observed the birth of new neurons in specific experimental injury models, including intestinal inflammation, GI infection, and following focal neuronal ablation. This neurogenic response can serve to replace neurons lost to injury, but it can also produce neuronal hyperplasia which can have significant pathologic consequences. Enteric neurogenesis is thus a double-edged sword that can be leveraged for a beneficial effect but needs to be modulated to limit its consequences. Our preliminary results suggest that experimental colitis in rodents promotes enteric glial cells to undergo a neurogenic transition via a 5-HT4-dependent pathway. However, this process is poorly understood. The overall objective of this proposal is to understand the mechanisms underlying postnatal enteric neurogenesis and its role in GI health and disease. To achieve this goal, we propose the following specific aims: (1) identify the downstream pathways that are activated by 5-HT4 signaling and lead to glial differentiation into neurons; (2) characterize the subpopulations of glial cells present in the intestine and determine the genetic and epigenetic changes that occur during the glia-to-neuron fate switch; and (3) leverage the intestine’s capacity for neurogenesis to treat the hypoganglionic transition zone associated with Hirschsprung disease. A variety of methodologies will be used to achieve these aims, including isolation and culture of enteric glia from reporter mice; in vitro assays using dominant-negative mutants and pharmacologic inhibitors to determine the signaling pathways involved in glial neurogenesis; single cell RNA seq to identify glial cell subpopulations; a dual reporter transgenic system for live cell imaging of glia-to-neuron cell fate transition; analysis of the genetic and epigenetic changes occurring during that transition; and induction of enteric neurogenesis in HSCR bowel to treat transition zone hypoganglionosis. Successful completion of these experiments will significantly enhance our understanding of the mechanisms underlying neurogenesis in the adult intestine, provide insights into the pathophysiology of neurointestinal diseases, identify new targets to modulate neurogenesis in vivo, offer novel approaches for expanding enteric neurons in the hypoganglionic transition zone of HSCR and in other neurointestinal diseases, and improve the in vitro expansion of enteric neuronal progenitors for cell therapy applications.

项目摘要 肠神经系统（ENS）通过其调节所有神经元的活动而对人类健康具有根本的重要性。 胃肠道（GI）功能方面，最显著的是肠道运动。先天性或后天性畸形 ENS因此可导致严重的功能性GI疾病，包括食管失弛缓症、胃轻瘫， 假性肠梗阻、肠易激综合征、先天性巨结肠和慢传输型便秘。 成年人的肠道具有神经元祖细胞，但它们的作用和激活机制尚不清楚。 他们是未知的。我们和其他人已经观察到在特定的实验性损伤中新神经元的诞生 模型，包括肠道炎症、GI感染和局灶性神经元消融后。这种神经性的 反应可以用来取代因损伤而失去的神经元，但它也可以产生神经元增生， 有严重的病理后果。因此，肠道神经发生是一把双刃剑， 但需要加以调整以限制其后果。我们的初步结果 表明啮齿类动物实验性结肠炎促进肠神经胶质细胞通过神经元转化， 5-HT 4依赖性通路。然而，人们对这一过程知之甚少。本提案的总体目标是 了解出生后肠道神经发生的机制及其在GI健康中的作用， 疾病为了实现这一目标，我们提出了以下具体目标：（1）确定下游通路 由5-HT 4信号激活并导致胶质细胞分化为神经元;（2）表征 神经胶质细胞的亚群存在于肠中，并决定遗传和表观遗传变化， 发生在神经胶质到神经元的命运转换过程中;（3）利用肠道的神经发生能力来治疗 与先天性巨结肠症相关的神经节下移行区。将采用各种方法 用于实现这些目标，包括从报告小鼠分离和培养肠神经胶质细胞;体外测定 使用显性阴性突变体和药理学抑制剂来确定参与的信号通路， 胶质神经发生;鉴定胶质细胞亚群的单细胞RNA测序;双报告基因转基因系统 用于胶质细胞向神经元细胞命运转变的活细胞成像;分析遗传和表观遗传变化 发生在过渡期间;并诱导HSCR肠道中的肠道神经发生以治疗过渡区 神经节机能减退成功完成这些实验将大大提高我们对 成人肠神经发生的机制，提供了对肠神经发生的病理生理学的见解。 神经肠道疾病，确定新的目标，以调节体内神经发生，提供新的方法， 在HSCR的下神经节过渡区和其他神经肠细胞中扩张肠神经元 疾病，并改善用于细胞治疗应用的肠神经元祖细胞的体外扩增。