Modifying the intestinal extracellular matrix to optimize enteric neuronal stem cell therapy for Hirschsprung disease

项目概要： 先天性或后天性肠神经系统（ENS）的破坏导致神经肠道疾病， 包括先天性巨结肠、食管失弛缓症、胃轻瘫、慢性假性肠梗阻， 等人功能性ENS的正常发育依赖于肠上皮细胞之间的协调相互作用。 神经嵴来源的细胞和它们的微环境，包括细胞外基质（ECM），如肠 神经嵴来源的细胞沿着胚胎肠间充质沿着吻尾方向迁移。1这些相互作用 确保适当的细胞迁移、增殖和分化成肠神经元和神经胶质细胞， 调节许多胃肠道过程，包括运动和感觉功能。许多ECM和 ECM结合蛋白已显示促进肠神经嵴来源的细胞迁移，而其他ECM结合蛋白则促进肠神经嵴来源的细胞迁移。 （包括聚集蛋白和硫酸软骨素蛋白聚糖）已显示出抑制迁移，证明了 ECM在ENS形成中的关键作用。尽管对正常ENS发展的理解不断提高， 目前缺乏针对神经肠道疾病的治疗方法，因为它们针对的是症状而不是潜在的疾病。 我们的实验室和其他实验室正在研究肠神经元的潜力， 干细胞移植作为一种新的方法来恢复这些患者的神经支配和肠道功能，但 移植细胞的移植、迁移和增殖有限，阻碍了研究。 由于ENS的发展对当地环境的依赖，我们认为肠溶系统的成功发展 神经元干细胞移植依赖于允许的局部ECM来支持移植的细胞。我们 假设抑制性ECM蛋白（聚集蛋白和硫酸软骨素蛋白聚糖）过表达 在肠神经元干细胞移植后，阻碍干细胞的迁移。在这份协议中，我们 因此目的是鉴定无神经节细胞和正常结肠之间的ECM基因表达差异， 表征肠神经元干细胞移植后ECM蛋白表达，并利用我们的 了解正常ENS发育以改变肠神经元干细胞移植后的微环境 通过敲低抑制细胞迁移的ECM蛋白以增强细胞的迁移， 移植细胞

PROJECT SUMMARY: Congenital or acquired disruption of the enteric nervous system (ENS) leads to neurointestinal diseases, including Hirschsprung disease, esophageal achalasia, gastroparesis, chronic intestinal pseudo-obstruction, and others. Normal development of a functional ENS relies on the coordinated interaction between enteric neural crest-derived cells and their microenvironment, including the extracellular matrix (ECM), as the enteric neural crest-derived cells migrate rostrocaudally along the embryonic gut mesenchyme.1 These interactions ensure proper cell migration, proliferation, and differentiation into the enteric neurons and glial cells that regulate numerous gastrointestinal processes, including motor and sensory function. Numerous ECM and ECM binding proteins have been shown to promote enteric neural crest-derived cell migration, while others (including agrin and chondroitin sulfate proteoglycans) have been shown to inhibit migration, demonstrating a critical role of the ECM in ENS formation. Despite an improving understanding of normal ENS development, current therapies for neurointestinal diseases are lacking, as they target symptoms rather than the underlying pathology of an absent or abnormal ENS. Our lab and others are investigating the potential of enteric neuronal stem cell transplantation as a novel approach to restoring innervation and gut function in these patients, but studies have been hampered by limited engraftment, migration, and proliferation of transplanted cells.1 Given the reliance of ENS development on the local environment, we believe that successful development of enteric neuronal stem cell transplantation relies on a permissive local ECM to support the transplanted cells. We hypothesize that inhibitory ECM proteins (agrin and chondroitin sulfate proteoglycans) are over-expressed following enteric neuronal stem cell transplantation, thwarting the migration of stem cells. In this grant we therefore aim to identify ECM gene expression differences between the aganglionic and normal colon, characterize ECM protein expression following enteric neuronal stem cell transplantation, and to leverage our understanding of normal ENS development to modify the microenvironment after enteric neuronal stem cell transplantation by knocking down ECM proteins that inhibit cell migration in order to enhance the migration of transplanted cells.