Autologous cell transplantation for the treatment of colorectal aganglionosis

自体细胞移植治疗结直肠无神经节细胞病

基本信息

批准号：
10308113
负责人：
Ryo Hotta
金额：
$ 7.75万
依托单位：
MASSACHUSETTS GENERAL HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-12-01 至 2023-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10308113
关键词：
Action Potentials Address Affect Age Area Autologous Autologous Transplantation Cell Separation Cell Survival Cell Therapy Cell Transplantation Cells Child Childhood Chronic Clinical Colon Colorectal Congenital Disorders Congenital Megacolon Constipation Data Development Diphtheria Toxin Disease Distal Drug Screening Electrophysiology (science)Embryo Enteral Enteric Nervous System Enterocolitis Environment Esophageal achalasia Excision Exhibits Failure Fecal Incontinence Functional Gastrointestinal Disorders Functional disorder Gastrointestinal Diseases Gastrointestinal tract structure Gastroparesis Goals Growth Factor Healthcare Human Immunosuppression In Vitro Injections Intestinal Pseudo-Obstruction Intestines Irritable Bowel Syndrome Laboratories Length Life Mediating Methods Modeling Morbidity - disease rate Mus Nerve Neural Crest Neuroglia Neuronal Differentiation Neuronal Injury Neurons Only Child Operative Surgical Procedures Proliferating Rodent Savings Stem cell transplant Synapses Testing Time Transgenic Mice Transplantation base cell motility clinical application clinically significant combinatorial cost diphtheria toxin receptor effective therapy enteric neuropathy experimental study ganglion cell gastrointestinal gastrointestinal function in vivo induced pluripotent stem cell innovation migration mouse model nerve stem cell novel novel strategies postnatal progenitor reduce symptoms single-cell RNA sequencing stem stem cells transcriptome transcriptomics transplantation therapy

项目摘要

PROJECT SUMMARY Hirschsprung disease is a potentially lethal congenital disorder characterized by the absence of enteric nervous system (ENS) along variable lengths of distal intestine due to the failure of neural crest-derived cells to colonize the entire intestine during development. The surgical treatment of Hirschsprung disease involves removing the aganglionic segment. While this is life- saving, significant complications commonly occur after surgery, including constipation, fecal incontinence, and enterocolitis. Transplantation of enteric neural crest-derived cells (ENCCs) offers a promising new approach to replacing missing or abnormal enteric neurons in Hirschsprung disease and other neurointestinal diseases. Recent studies, including from our laboratory, demonstrate that ENCCs can be isolated from the postnatal intestine, propagated in culture, and transplanted into the gut wall. However, major challenges remain: (1) the numbers of neurons generated have been limited, (2) their ability to integrate into neuroglial networks capable of restoring gut function has yet to be demonstrated, and (3) mice with Hirschsprung disease die at 4-6 weeks of age, limiting the time window available for analysis after cell transplantation. To overcome these challenges, we propose the following aims: (1) to incorporate an innovative approach to isolating and expanding postnatal gut-derived ENCCs that generates greater numbers of progenitor ENCCs than prior methods and (2) to transplant autologously-derived ENCCs into a novel non-lethal model of colorectal aganglionosis generated by local injection of human diphtheria toxin (DT) into transgenic mice whose neural crest-derived cells express DT receptor. To optimize ENCC expansion, a combinatorial drug screen approach will be used. ENCCs will be cultured with candidate molecules, together with known growth factors, to identify the optimal “cocktail” for progenitor cell expansion. Single cell RNAseq will be performed to characterize the transcriptome profile of cells prior to transplantation. Autologously-derived donor cells will be delivered into the experimentally- generated aganglionic segment of colon. Analyses will include quantitative determination of ENCC survival, proliferation, and neuronal differentiation, and functional characterization of neuronal activity. The results obtained will establish an optimized approach to ENCC isolation, expansion, and transplantation, and demonstrate the potential of cell-based therapy to restore GI function in Hirschsprung disease and other neurointestinal diseases.

项目概要先天性巨结肠症是一种潜在致命的先天性疾病，其特征是缺乏由于肠神经系统（ENS）的故障，沿远端肠的不同长度的肠神经系统（ENS）神经嵴衍生细胞在发育过程中定植于整个肠道。先天性巨结肠症的治疗涉及去除无神经节段，而这是生命的一部分。手术后常会出现明显的并发症，包括便秘、粪便失禁和小肠结肠炎。肠神经嵴衍生细胞（ENCC）移植。提供了一种有前途的新方法来替代缺失或异常的肠神经元先天性巨结肠症和其他神经肠道疾病的最新研究，包括我们的研究。实验室证明 ENCC 可以从产后肠道中分离出来，并在然而，主要挑战仍然存在：（1）数量。生成的神经元数量有限，（2）它们整合到神经胶质网络的能力能够恢复肠道功能尚未得到证实，并且（3）患有先天性巨结肠的小鼠疾病在 4-6 周龄时死亡，限制了细胞后可用于分析的时间窗口为了克服这些挑战，我们提出以下目标：（1）采用创新方法分离和扩增产后肠道来源的 ENCC 比以前的方法产生更多数量的祖细胞 ENCC，并且 (2) 进行移植自体衍生的 ENCC 构建新型非致死结直肠无神经节细胞模型通过将人白喉毒素（DT）局部注射到转基因小鼠体内而产生，该小鼠的神经嵴衍生细胞表达 DT 受体，以优化 ENCC 扩增，这是一种组合药物。将使用筛选方法与候选分子一起培养 ENCC。已知的生长因子，以确定祖细胞扩增的最佳“鸡尾酒”。在进行 RNA 测序之前，将进行 RNAseq 来表征细胞的转录组谱自体来源的供体细胞将被输送到实验中。生成的结肠无神经节段的分析将包括定量测定。 ENCC 存活、增殖、神经元分化以及功能特征所获得的结果将建立一种优化的 ENCC 分离方法，扩张和移植，并证明基于细胞的治疗恢复的潜力先天性巨结肠症和其他神经肠道疾病中的胃肠道功能。