How Patterned Mesenchymal-Epithelial interactions Shape Intestinal Crypts

项目概要/摘要 结肠上皮是许多人类疾病的发生部位，因此了解其发育和 功能对人类健康至关重要。结肠上皮在形态上和功能上分为 不同的区域，增殖性干细胞被安置在Lieberkühn的隐窝底部，而 分化的细胞聚集在最靠近管腔的上皮中。维持上皮细胞的稳态， 隐窝在很大程度上由来自下层间充质的空间图案化信号决定。的 最近，参与成人结肠隐窝内环境稳定的间充质细胞引起了人们的兴趣。然而， 关于发育过程中结肠隐窝形成的分子机制，我们的建议 将决定间充质细胞群和信号如何与发育中的上皮细胞相互作用， 驱动结肠隐窝形态发生，并在发育和再生过程中定义干细胞动力学。 我们将通过我们的三个目标来完成这项工作：首先，我们将绘制间充质-上皮相互作用， 使用单细胞RNAseq和组织学方法的结肠隐窝发育过程。然后我们将使用 转基因小鼠模型，以研究间充质WNT信号在三个阶段中的具体作用 结肠隐窝的发育：结肠隐窝的内陷、延长和分裂。第二，我们会调查 损伤后上皮和间充质中胚胎程序的潜在再激活 由DSS治疗引起。我们将测试胚胎或损伤相关的间充质刺激 可塑性，定义为定向祖细胞的去分化到一个更多能的状态，在成年 上皮第三，我们将采取一种合成的方法来重建结肠周围的模式间充质信号 我们将使用这个新模型来测试极化WNT信号驱动结肠隐窝的充分性 体外形成。总的来说，我们的项目利用了体内研究，体外模型，合成 工程和生物信息学方法。这种全面的范围是通过一个高度 在UCSF的合作环境和两个主要研究人员进行富有成效的伙伴关系， 这个项目这项提议的发现将大大推进我们对结肠发育的基础知识 和再生，突出了间充质-上皮串扰的动力学。这将产生影响 超越了我们对结肠生物学的基本理解，并将有助于为发现治疗方法铺平道路。 治疗和调节结肠疾病的策略。

Project Summary/Abstract The colonic epithelium is the site of a host of human diseases, and therefore understanding its development and function is essential for human health. The colon epithelium is separated into morphologically and functionally distinct domains, with proliferative stem cells being housed at the bottom of the crypts of Lieberkühn, whereas differentiated cells populate the epithelium closest to the lumen. The maintenance of epithelial homeostasis in the crypt is dictated in large part by spatially patterned signals from the underlying mesenchyme. The mesenchymal cells involved in adult colonic crypt homeostasis have been of recent interest. However, little is known about the molecular mechanisms underlying colonic crypt formation during development. Our proposal will determine how mesenchymal cell populations and signals interact with the developing epithelium to drive colonic crypt morphogenesis and define stem cell dynamics during development and regeneration. We will accomplish this work through our three aims: First, we will map mesenchymal-epithelial interactions over the course of colonic crypt development using single cell RNAseq and histological approaches. We will then use genetically modified mouse models to investigate the specific role of mesenchymal WNT signals in three stages of colonic crypt development: invagination, elongation, and fission of colonic crypts. Second, we will investigate the potential re-activation of an embryonic program in both the epithelium and mesenchyme following injury induced by DSS treatment. We will test the ability of embryonic or injury-associated mesenchyme to stimulate plasticity, defined as de-differentiation of committed progenitors to a more multipotent state, in the adult epithelium. Third, we will take a synthetic approach to reconstruct patterned mesenchymal signals around colon spheroids, and we will use this new model to test the sufficiency of polarized WNT signals to drive colonic crypt formation in vitro. Overall, our project leverages a combination of in vivo studies, in vitro models, synthetic engineering and bioinformatic approaches. This comprehensive scope is made possible through a highly collaborative environment at UCSF and the productive partnership of the two principal investigators carrying out this project. The findings from this proposal will significantly advance our basic knowledge of colon development and regeneration, highlighting the dynamics of mesenchymal-epithelial crosstalk. This will have implications beyond our fundamental understanding of colon biology and will help pave the way for discovery of therapeutic strategies to treat and modulate colonic disease.