How Patterned Mesenchymal-Epithelial interactions Shape Intestinal Crypts

项目摘要/摘要 结肠上皮是人类许多疾病的发源地，因此了解其发育和 人体机能对人体健康至关重要。结肠上皮在形态上和功能上分为 不同的区域，增殖干细胞被安置在Lieberkühn的隐窝底部，而 分化的细胞分布在最靠近管腔的上皮中。血管内皮细胞动态平衡的维持 地穴在很大程度上是由来自底层间充质的空间图案化信号决定的。这个 间充质细胞参与成人结肠隐窝的动态平衡最近引起了人们的兴趣。然而，几乎没有什么是 了解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体和信号如何与发育中的上皮细胞相互作用 推动结肠隐窝的形态发生，并确定干细胞在发育和再生过程中的动力学。 我们将通过三个目标来完成这项工作：第一，我们将绘制间充质-上皮相互作用图 用单细胞RNAseq和组织学方法研究结肠隐窝的发育过程。然后我们将使用 转基因小鼠模型研究间充质WNT信号在三个阶段的具体作用 结肠隐窝的发育：结肠隐窝的内陷、延长和分裂。第二，我们将调查 损伤后胚胎程序在上皮和间充质中重新激活的可能性 DSS处理诱导的。我们将测试胚胎或损伤相关间充质刺激的能力 成体的可塑性，定义为承诺的祖细胞去分化为更多能的状态 上皮组织。第三，我们将采用一种合成的方法来重建结肠周围的间质信号。 球体，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 在体外形成。总体而言，我们的项目结合了体内研究、体外模型、人工合成 工程学和生物信息学方法。这一全面的范围是通过高度的 加州大学旧金山分校的协作环境和两名首席调查员开展的富有成效的伙伴关系 这个项目。这项建议的发现将显著提高我们对结肠发育的基本知识 和再生，突出了间充质-上皮细胞的动态串扰。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.