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Contribution of the sacral neural crest to the peripheral nervous system of the post-umbilical gastrointestinal tract

骶神经嵴对脐后胃肠道周围神经系统的贡献

基本信息

批准号：
10644256
负责人：
Marianne Bronner
金额：
$ 55.64万
依托单位：
CALIFORNIA INSTITUTE OF TECHNOLOGY
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-07-01 至 2027-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10644256
关键词：
Antibodies Birds Brain Burn injury CRISPR/Cas technology Cell Differentiation process Cells Central Nervous System Chick Colon Compensation Congenital Abnormality Congenital Megacolon Data Defect Development Disease Embryo Enteral Enteric Nervous System Environment Gastrointestinal Hormones Gastrointestinal Motility Gastrointestinal tract structure Gene Expression Profile Genetic Transcription Genome Genomic approach Genus Alpharetrovirus Hindgut Hormone secretion Human In Situ Individual Interspecific Recombination Intestines Knowledge Label Length Life Mediating Modernization Motor Neurons Neck Nervous System control Neural Crest Neural Crest Cell Neural tube Neuroglia Neuronal Differentiation Neurons Patients Peripheral Nervous System Plasmids Population Primitive foregut structure Process Proliferating Publishing Quail Regulator Genes Retroviridae Role SOX21 gene SOX4 gene Sampling Somites Sorting Tail Techniques Testing Time Tyrosine 3-Monooxygenase Umbilicus cell motility cholinergic enteric neuropathy experimental study fluorophore gut colonization hatching healing hindbrain insight loss of function migration neural novel programs public health relevance single-cell RNA sequencing time use transcription factor

项目摘要

The enteric nervous system (ENS), the largest portion of the peripheral nervous system, is derived from neural crest populations referred to as “vagal” and “sacral”, arising from the neck and tail regions, respectively. However, much less has been published about sacral than vagal neural crest. To rectify this knowledge gap, we propose to: 1. characterize the temporal sequence of migration and differentiation of sacral neural crest cells into neuronal subtypes in the hindgut; 2. transcriptionally profile the sacral neural crest as at multiple time points and compare with that of vagal neural crest; 3. test function of transcription factors that may drive sacral neural crest cell fate choice. The results will enable us to test whether vagal and sacral neural crest cells give rise to similar or distinct types of enteric neurons and elucidate the influence of the intestinal environment on their differentiation. We will characterize the chick sacral neural crest using a novel lineage labeling technique of Replication Incompetent Avian (RIA) retroviruses that enables us to specifically target and isolate by FACS the sacral neural crest-derived population to perform the following aims: Specific Aim 1: Retrovirus-mediated lineage analysis of the chick sacral compared with vagal neural crest: We will label the neural tube caudal to somite 27 with RIA retroviruses that permanently label sacral neural crest cells in order to follow their long term fate. Preliminary results suggest that sacral neural crest-derived cells populate the post-umbilical gut and differentiate into cholinergic motor neurons as well as tyrosine hydroxylase positive cells. Sacral neural crest will be compared with vagal neural crest and interactions between the two populations examined. Finally, clonal relationships between sacral crest cells will be characterized. Specific Aim 2: Single cell RNA-seq of sacral neural crest-derived cells in the post-umbilical gut. To elucidate gene regulatory programs controlling progressive differentiation of sacral neural crest cells into neurons and glia in the hindgut, we propose to characterize the transcriptional profile of sacral crest-derived cells FACS sorted from the post-umbilical gut and processed by single cell (sc) RNA-seq at embryonic days (E) 2.5, E6, E8, 10, 15, 21 (prehatching); these will be compared with vagal crest-derived cells at comparable stages. Preliminary scRNA-seq data on E10 suggest that there are differences in neuronal subtypes produced by sacral vs vagal crest. scRNA-seq will enable us to sample different neuronal subtypes and infer developmental trajectories. Specific Aim 3: Role of transcriptional regulators into differentiation of sacral neural crest into neuronal and glial subtypes. Focusing on transcription factors that are present in the enteric precursor cluster (e.g. Nfatc1, Foxn2, Sox4, Sox21, Elf2, Znf536), we will test whether sacral neural crest-enriched transcription factors are critical for mediating proliferation, migration within the gut, and/or cell fate decisions. To this end, we will perform targeted loss of function in the sacral neural crest using a single-plasmid CRISPR-Cas9 strategy and examine subsequent effects on distribution and/or differentiation of sacral neural crest-derived cells.

肠神经系统（enteric nervous system， ENS）是外周神经系统的最大部分，来源于