The physical and molecular mechanisms of intestinal villus morphogenesis and repair

肠绒毛形态发生和修复的物理和分子机制

• 批准号: 10438924
• 负责人: Zev Jordan Gartner
• 金额: $ 56.96万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-15 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10438924
• 关键词: Adult; Architecture; Area; Artificial tissue; Atomic Force Microscopy; Atrophic; Automobile Driving; Calcium Signaling; Celiac Disease; Cells; Clinical; Computer Models; Connexin 43; Data; Development; Embryo; Endothelin; Endothelin Receptor; Engineering; Epithelial; Erinaceidae; Fibroblasts; Fingers; Future; Gap Junctions; Genes; Genetic Transcription; Goals; Human; Impairment; Infection; Inflammatory; Injury; Intestines; Investigation; Malabsorption Syndromes; Maps; Measurement; Measures; Mechanics; Mediating; Mesenchymal; Mesenchyme; Modeling; Molecular; Morphogenesis; Morphology; Mus; Myosin Type II; Natural regeneration; Nonmuscle Myosin Type IIA; Nutrient; Outcome; Pathway interactions; Patients; Pattern; Phase; Physical condensation; Play; Population; Process; Proteins; Radiation induced damage; Regenerative research; Resolution; Role; Scientist; Shapes; Signal Transduction; Site; Small Intestines; Smooth Muscle Actin Staining Method; Smooth Muscle Myocytes; Source; Stains; Surface; Testing; Tissue Engineering; Tissues; Villus; blebbistatin; cell motility; chemotherapy; design; driving force; experimental study; gastrointestinal; improved; in vivo regeneration; inhibitor; interfacial; intestinal epithelium; intestinal villi; live cell microscopy; monolayer; non-muscle myosin; novel therapeutics; programs; radiation effect; reconstitution; regenerative; repaired; replacement tissue; side effect; single-cell RNA sequencing; uptake

ABSTRACT Villi are finger-like projection that line the lumen of the small intestine. Villi play a critical role in nutrient uptake by increasing the intestinal absorptive surface area by several orders of magnitude. Loss of this absorptive surface through villus atrophy causes major digestive complications and nutrient malabsorption. Abnormalities in villi are found in many gastrointestinal maladies, such as inflammatory bowel and celiac diseases, and are also side effects of radiation, chemotherapy, and infection. Degenerated villi can sometimes fully reform, yet in other situations regeneration is impaired, resulting in persistent villus atrophy and patient suffering. Villi emerge during development from an initially flat intestinal surface. The mechanisms underlying villus formation and repair remain poorly described, and an understanding of these processes is essential to develop new therapies. The long term goal of this proposal is to build an understanding of the molecules and forces that sculpt the villus during development and regeneration so as to improve strategies for growing and regenerating the intestine for human patients. Recent data from our labs suggest that the mesenchyme plays a central role in sculpting the architecture of the villus. Specifically, our preliminary data implicate a specialized population of self- organizing sub-epithelial mesenchymal cells that condense immediately below the forming villus as the source of the physical forces necessary to pattern and fold the overlying epithelium into villi. We investigated the molecular mechanisms leading to condensation of these cells using single cell RNAseq and found they also express a unique transcriptional program. This program has an unusual overlap with genes regulating Ca2+ mediated contractility in smooth muscle cells but without expressing smooth muscle actin. We provide evidence that inhibition of key proteins in this program results in a loss of mesenchymal condensation and villus evagination. Guided by these preliminary data, we propose to test two hypotheses related to the complementary physical and molecular aspects of villus formation. We combine quantitative measurements and computational modeling to test the hypothesis that the formation of villus condensates occurs analogously to phase separation phenomena studied extensively by physicists and material scientists, and that condensates exert physical forces on the overlying epithelium initiating its folding. We then test the hypothesis that Endothelin released from the epithelium triggers increased calcium signaling in the subepithelial mesenchyme, which are synchronized through gap junctions to drive cell contractility leading to phase separation and condensation. This project has major implications for our fundamental understanding of the developmental of the gut and for tissue engineering of intestinal tissue, as we do not know how mammalian intestinal villi are built. Thus, these findings will be impactful because they provide a new mechanistic blueprint for this process that incorporates both signals and forces. These findings will also lay the groundwork for future studies of regeneration: we find that in adults, the sub-epithelial mesenchyme retains expression of many of these molecular features. Further, these features are upregulated following injury.

摘要 绒毛是指状突起，排列在小肠腔中。绒毛在营养吸收中起着关键作用 通过将肠吸收表面积增加几个数量级。这种吸收损失 表面通过绒毛萎缩引起主要消化并发症和营养吸收不良。异常 在许多胃肠道疾病，如炎症性肠病和乳糜泻中发现， 还有放疗化疗和感染的副作用退化的绒毛有时可以完全改革，但在 其他情况下，再生受损，导致持续的绒毛萎缩和患者痛苦。 绒毛在发育过程中从最初平坦的肠表面出现。绒毛的潜在机制 形成和修复的描述仍然很少，对这些过程的理解对于开发 新疗法这项提案的长期目标是建立对分子和力的理解， 在发育和再生过程中塑造绒毛，以改善生长和再生策略 人类患者的肠道。我们实验室的最新数据表明，间充质在 塑造绒毛的结构。具体来说，我们的初步数据表明，一个特殊的自我- 组织化的上皮下间充质细胞凝聚在形成绒毛的正下方， 将覆盖的上皮细胞折叠成绒毛所需的物理力。我们调查的 使用单细胞RNA测序导致这些细胞凝聚的分子机制，并发现它们也 表达独特的转录程序。这个程序与调节Ca 2+的基因有不寻常的重叠 介导平滑肌细胞的收缩，但不表达平滑肌肌动蛋白。我们提供的证据 在这个程序中，抑制关键蛋白质会导致间充质凝聚和绒毛的损失， 外翻在这些初步数据的指导下，我们提出了两个与互补性相关的假设。 绒毛形成的物理和分子方面。我们将联合收割机定量测量和计算 模拟以检验绒毛冷凝物的形成类似于相分离发生的假设 物理学家和材料科学家广泛研究的现象，以及冷凝物产生的物理力， 在上面的上皮上开始折叠。然后，我们测试了内皮素从血管中释放的假设， 上皮触发上皮下间充质中钙信号的增加， 通过缝隙连接驱动细胞收缩，导致相分离和凝聚。 该项目对我们对肠道发育的基本理解以及对 肠组织的组织工程，因为我们不知道哺乳动物的肠绒毛是如何建立的。因此这些 研究结果将产生影响，因为它们为这一过程提供了一个新的机制蓝图， 信号和力量。这些发现也将为未来的再生研究奠定基础：我们发现， 在成人中，上皮下间充质保留了许多这些分子特征的表达。此外，本发明还 这些特征在损伤后被上调。