Uncovering the role of LRH-1 in enteroendocrine cell development and ‘gut-brain’ communication

项目摘要/摘要 背景： 肠上皮是身体最大的暴露表面积之一，必须 与消化、感染监测和新陈代谢相关的多种功能。帮助协调这些 活动的是肠内分泌细胞(EECS)，表达细胞机械的特化肠上皮细胞 用于化学和机械传感，以及通过“肠道-脑轴”进行主机通信。肝脏受体 Homolog-1(LRH-1，NR5A2)是一种表达于肠上皮细胞的核受体。我们已经向LRH展示了- 1对这些独特细胞的产生至关重要，LRH-1的缺失会导致显著的内脏 感觉迟钝。在这一应用中，我们建议阐明LRH-1促进LRH-1的机制(S) EECs的分化及其缺失对肠脑通讯的功能影响。 方法： 我们将结合使用遗传小鼠模型和肠道器官技术来研究LRH-1- 依赖于EEC分化。使用单细胞测序方法，我们将识别EEC前体细胞 在失去LRH-1之后，人口转移。利用EEC前身荧光标记，我们将在 实时检测LRH-1基因敲除对EEC细胞分化的影响及LRH-1与DNA的结合 目标。我们的分子研究将得到LRH功能后果评估的补充- 1-依赖EEC损失。最后，在我们的急性LRH-1基因敲除模型中使用详细的解剖学研究将 使我们能够探索快速的EEC破坏对粘膜神经纤维的影响，粘膜神经纤维是肠道的关键组成部分- 脑轴。 目标： 这项建议旨在扩展我在K08调查中发现的一个关键发现，并充分 建立我的独立研究计划。这些实验将为提交 成功的R01方案建立在肠上皮生理学、核的非凡融合上 受体生物学和神经元信号传递。

Project Summary/Abstract Background: The intestinal epithelium represents one of the largest exposed surface areas of the body and must perform multiple functions related to digestion, infectious surveillance, and metabolism. Helping to coordinate these activities are enteroendocrine cells (EECs), specialized intestinal epithelial cells expressing cellular machinery for chemical and mechanical sensing and for host communication via the ‘Gut-Brain Axis.’ Liver receptor homolog-1 (LRH-1, NR5A2) is a nuclear receptor expressed in the intestinal epithelium. We have shown LRH- 1 is critical for the production of these unique cells and that loss of LRH-1 leads to a remarkable visceral hyposensitivity. In this application, we propose to elucidate both the mechanism(s) by which LRH-1 promotes the differentiation of EECs and the functional consequences of their loss on gut-brain communication. Approach: We will use a combination of genetic mouse models and intestinal organoid technology to investigate LRH-1- dependent EEC differentiation. Using a single cell sequencing approach, we will identify EEC progenitor population shifts following loss of LRH-1. Leveraging an EEC progenitor fluorescent marker, we will observe in real time the effects of LRH-1 knockout on EEC cell differentiation and to delineate the LRH-1 DNA binding targets. Our molecular studies will be complemented by an evaluation of the functional consequences of LRH- 1-dependent EEC loss. Finally, use of detailed anatomic studies in our acute LRH-1 knockout models will enable us to explore the effects of rapid EEC disruption on mucosal nerve fibers, a key component of the gut- brain axis. Goals: This proposal is designed to expand upon a key finding uncovered during my K08 investigation and to fully establish my independent research program. These experiments will provide important data for submission of a successful R01 proposal built at the extraordinary confluence of intestinal epithelial physiology, nuclear receptor biology, and neuronal signaling.