Mechanochemical mechanisms of intestinal villus development and regeneration

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PROJECT SUMMARY Intestinal villi comprise a mesenchymal core ensheathed by an epithelial layer that together increase the intestinal absorptive surface area nearly 100 fold. Damage to villi incurred by irradiation, chemotherapeutics, and many gastrointestinal maladies cause villus atrophy, resulting in nutrient malabsorption and digestive complications. Remarkably, in many cases atrophied villi fully regenerate to restore their finger-like morphology, yet in other situations the regenerative process is impaired, resulting in persistent villus atrophy. How villi regenerate their morphology and why this process can fail are unknown. Moreover, we lack an understanding of how mammalian villi are initially built during development. The long-term goal of this project is to understand how signals and forces are integrated to sculpt mammalian villi during development and regeneration so as to improve strategies for growing and regenerating human intestinal tissue. Villus formation initiates in the embryo with the aggregation of mesenchymal cells adjacent to the overlying epithelium that form condensations, termed villus clusters. While villus clusters seem to be required for villus morphogenesis, the mechanisms underlying their formation have not been identified. Here, I explore a link between these events that first build villi in the mammalian embryo during development and those that rebuild them in the adult during regeneration. First, I quantify the physical properties of villus clusters and test whether mesenchymal condensation into clusters, and the subsequent buckling of the epithelium into villi, is initiated through mesenchymal cellular contractility and reinforced by differential interfacial tension. Next, I examine the genetic mechanisms underlying the mechanics of these mesenchymal condensation events and villus formation. Specifically, I put forth and test a model generated from single-cell RNA sequencing data wherein the contractile activities of the subepithelial mesenchyme are triggered by Endothelin-dependent Ca2+ signaling, and the contractions become coupled locally between neighboring cells through gap junctions to drive cluster formation and epithelial buckling. Finally, I test whether subepithelial mesenchymal cells utilize similar mechanochemical mechanisms to regenerate villus architecture in response to damage in the adult intestine, and additionally, characterize villus regeneration at single cell resolution to identify new genetic mediators of adult villus architecture. Together, these studies will define how signals and forces interact to form and reform mammalian intestinal villi, identify new therapeutic targets to stimulate villus regeneration, and, more broadly, will address the question of how tissue shape and size are regenerated.