MODELING ENTERIC NERVOUS SYSTEM DEVELOPMENT AND HIRSCHSPRUNG'S DISASE IN HUMAN PLURIPOTENT STEM CELLS

总结 人类肠神经系统（ENS）是一个复杂的网络，约有5亿个神经元，来自于 肠神经嵴（ENC）和胃肠（GI）功能所必需的。ENS缺陷是导致许多 常见的人类疾病包括胃轻瘫、肠易激综合征（IBS）、肥厚性幽门梗阻、肠易激综合征（IBS）、肠易激综合征（IBS）、肠易激综合征（肠易激综合征）、肠易激综合征（肠易激综合征）、肠易激综合征（肠易激综合征）、 狭窄先天性巨结肠是一种严重的先天性缺陷，其特征是缺乏ENS 前体在肠道的远端部分称为无神经节细胞症。无神经节细胞症导致缺乏 适当的蠕动导致功能性结肠阻塞（“巨结肠”） 由于缺乏可接近的组织，对人类ENS的发展知之甚少。使用 人多能干细胞（hPSC）代表了一种获得人ENS的新颖而独特的策略 开发和建模并可能治疗HD。我们小组最近的工作表明， 从人类PSC产生ENC的可行性，并为使用该技术开发ENC奠定了基础。 治疗HD和潜在的其他ENS疾病的药物和细胞为基础的策略。当前应用程序 在这些令人兴奋的初步工作的基础上，实现三个具体目标： 在目标1中，我们将建立条件来表征和操作神经元亚型的广泛库 和肠神经胶质细胞，并在与已知靶细胞的创新共培养测定中测试它们的功能。在 目的二，我们将应用ENC分化技术来模拟hPSC中的HD，并验证电流和识别 新的候选HD药物。在目标3中，我们将在小鼠模型中优化基于hPSC的细胞治疗方法 并进行机制研究，确定HD动物体内挽救的关键参数。 我们的研究是独一无二的，因为它代表了可靠地重建和研究早期人类ENS谱系的第一次努力， 体外为该提案而聚集的合作者在hPSC，ENS生物学， 化学筛选和细胞移植。ENS是一种没有被其他人充分研究的结构 尽管它对人类健康有重要贡献。我们的研究有可能直接产生新药， HD的细胞治疗。这项工作为更广泛的应用奠定了基础，并开发了必要的技术。 社区研究ENS的发展和ENS在人类健康和疾病中的功能。

SUMMARY The human enteric nervous system (ENS) is a complex network of ~500 million neurons derived from the enteric neural crest (ENC) and essential for gastro-intestinal (GI) function. ENS defects are the cause of many common human disorders including gastroparesis, irritable bowel syndrome (IBS), hypertrophic pyloric stenosis. Hirschsprung's disease (HD) is a severe congenital defect characterized by the lack of ENS precursors in the distal portions of the gut referred to as aganglionosis. Aganglionosis results in the lack of proper peristalsis causing functional colonic obstruction ("megacolon") Development of the human ENS is poorly understood given the lack of accessible tissue. The use of human pluripotent stem cells (hPSCs) represent an novel and unique strategy to access human ENS development and to model and potentially treat HD. Very recent work from our group demonstrates the feasibility of generating ENC from human PSCs and sets the stage for using the technology for developing drug and cell based strategies in treating HD and potentially other ENS disorders. The current application builds on such exciting preliminary work to pursue three specific aims: In Aim 1, we will establish conditions to characterize and manipulate the broad repertory of neuronal subtypes and enteric glia from hPSCs and to test their function in innovative co-culture assays with known target cells. In Aim 2, we will apply ENC differentiation technology to model HD in hPSCs and to validate current and identify novel candidate HD drugs. In Aim 3, we will optimize hPSC-based cell therapy approaches in a mouse model of HD and perform mechanistic studies that define the critical parameters for in vivo rescue of HD animals. Our study is unique as it represents the first effort to reliably recreate and study early human ENS lineages in vitro. The collaborators assembled for this proposal have complementary expertise in hPSCs, ENS biology, chemical screening and cell transplantation. The ENS is a structure that is not sufficiently studied by others despite its important contribution to human health. Our study has the potential to directly yield novel drug and cell based treatments for HD. The work sets the stage and develops the technology necessary for the broader community to study ENS development and ENS function in human health and disease.