The role of mechanics in the growth and homeostasis of the intestinal crypt

We present a mechanical model of tissue homeostasis that is specialised to the intestinal crypt. Growth and deformation of the crypt, idealised as a line of cells on a substrate, are modelled using morphoelastic rod theory. Alternating between Lagrangian and Eulerian mechanical descriptions enables us precisely to characterise the dynamic nature of tissue homeostasis, whereby the proliferative structure and morphology are static in the Eulerian frame, but there is active migration of Lagrangian material points out of the crypt. Assuming mechanochemical growth, we identify the necessary conditions for homeostasis, reducing the full, time-dependent system to a static boundary value problem characterising a spatially heterogeneous "treadmilling" state. We extract essential features of crypt homeostasis, such as the morphology, the proliferative structure, the migration velocity, and the sloughing rate. We also derive closed-form solutions for growth and sloughing dynamics in homeostasis, and show that mechanochemical growth is sufficient to generate the observed proliferative structure of the crypt. Key to this is the concept of threshold-dependent mechanical feedback, that regulates an established Wnt signal for biochemical growth. Numerical solutions demonstrate the importance of crypt morphology on homeostatic growth, migration, and sloughing, and highlight the value of this framework as a foundation for studying the role of mechanics in homeostasis.Comment: 21 pages, 7 figure

Hydrogel crosslinking modulates macrophages, fibroblasts, and their communication, during wound healing

Biomaterial wound dressings, such as hydrogels, interact with host cells to regulate tissue repair. This study investigates how crosslinking of gelatin-based hydrogels influences immune and stromal cell behavior and wound healing in female mice. We observe that softer, lightly crosslinked hydrogels promote greater cellular infiltration and result in smaller scars compared to stiffer, heavily crosslinked hydrogels. Using single-cell RNA sequencing, we further show that heavily crosslinked hydrogels increase inflammation and lead to the formation of a distinct macrophage subpopulation exhibiting signs of oxidative activity and cell fusion. Conversely, lightly crosslinked hydrogels are more readily taken up by macrophages and integrated within the tissue. The physical properties differentially affect macrophage and fibroblast interactions, with heavily crosslinked hydrogels promoting pro-fibrotic fibroblast activity that drives macrophage fusion through RANKL signaling. These findings suggest that tuning the physical properties of hydrogels can guide cellular responses and improve healing, offering insights for designing better biomaterials for wound treatment

Chronic TNFα-driven injury delays cell migration to villi in the intestinal epithelium

The intestinal epithelium is a single layer of cells which provides the first line of defence of the intestinal mucosa to bacterial infection. Cohesion of this physical barrier is supported by renewal of epithelial stem cells, residing in invaginations called crypts, and by crypt cell migration onto protrusions called villi; dysregulation of such mechanisms may render the gut susceptible to chronic inflammation. The impact that excessive or misplaced epithelial cell death may have on villus cell migration is currently unknown. We integrated cell-tracking methods with computational models to determine how epithelial homeostasis is affected by acute and chronic TNFα-driven epithelial cell death. Parameter inference reveals that acute inflammatory cell death has a transient effect on epithelial cell dynamics, whereas cell death caused by chronic elevated TNFα causes a delay in the accumulation of labelled cells onto the villus compared to the control. Such a delay may be reproduced by using a cell-based model to simulate the dynamics of each cell in a crypt-villus geometry, showing that a prolonged increase in cell death slows the migration of cells from the crypt to the villus. This investigation highlights which injuries (acute or chronic) may be regenerated and which cause disruption of healthy epithelial homeostasis

A Multicellular Model of Intestinal Crypt Buckling and Fission

Crypt fission is an in vivo tissue deformation process that is involved in both intestinal homeostasis and colorectal tumourigenesis. Despite its importance, the mechanics underlying crypt fission are currently poorly understood. Recent experimental development of organoids, organ-like buds cultured from crypt stem cells in vitro, has shown promise in shedding light on crypt fission. Drawing inspiration from observations of organoid growth and fission in vivo, we develop a computational model of a deformable epithelial tissue layer. Results from in silico experiments show the stiffness of cells and the proportions of cell subpopulations affect the nature of deformation in the epithelial layer. In particular, we find that increasing the proportion of stiffer cells in the layer increases the likelihood of crypt fission occurring. This is in agreement with and helps explain recent experimental work

Paneth cell - rich regions separated by a cluster of Lgr5+ cells initiate crypt fission in the intestinal stem cell niche

The crypts of the intestinal epithelium house the stem cells that ensure the continual renewal of the epithelial cells that line the intestinal tract. Crypt number increases by a process called crypt fission, the division of a single crypt into two daughter crypts. Fission drives normal tissue growth and maintenance. Correspondingly, it becomes less frequent in adulthood. Importantly, fission is reactivated to drive adenoma growth. The mechanisms governing fission are poorly understood. However, only by knowing how normal fission operates can cancer-associated changes be elucidated. We studied normal fission in tissue in three dimensions using high-resolution imaging and used intestinal organoids to identify underlying mechanisms. We discovered that both the number and relative position of Paneth cells and Lgr5+ cells are important for fission. Furthermore, the higher stiffness and increased adhesion of Paneth cells are involved in determining the site of fission. Formation of a cluster of Lgr5+ cells between at least two Paneth-cell-rich domains establishes the site for the upward invagination that initiates fission

Integrated single-cell RNA-sequencing data of unwounded and wounded mouse skin and fibroblasts.

<p>This repository contains the .h5ad files that store the integrated scRNA-seq data we generated for the work, Almet et al. (2023), "Fibroblasts evolve in single-cell state to drive extracellular matrix and signaling changes across wound healing", to be published in the Journal of Investigative Dermatology.</p><p>The integrated* files contain both raw counts, normalized counts, as well as unspliced and spliced count estimates that were obtained using kallisto|bustools and velocyto. We integrated the data from the following published datasets:</p><ol><li><a href=" https://doi.org/10.7554/eLife.60066">Phan et al. (2021)</a>: Unwounded P21 mice and small wound P21 + 7 mice</li><li><a href="https://doi.org/10.1016/j.celrep.2020.02.091">Haensel et al. (2020)</a>: Unwounded P49 mice and small wound P49 + 4 mice</li><li><a href="https://doi.org/10.1038/s41467-018-08247-x)">Guerrero-Juarez et al. (2019)</a>: Large wound day 12 mice</li><li><a href="https://doi.org/10.1016/j.stem.2020.07.008">Abbasi et al. (2020)</a>: Large wound day 14 mice</li><li><a href="https://doi.org/10.1126/sciadv.aay3704">Gay et al. (2020)</a>: Large wound fibrotic (hairless) and regenerative (hair follicle neogenesis) day 18 mice</li></ol><p>The unwounded_* files were used to briefly integrated unwounded skin scRNA-seq from mouse models of different ages that have been used to analyze wound healing in <a href="https://doi.org/10.1016/j.celrep.2020.02.091">Haensel et al. (2020),</a> <a href=" https://doi.org/10.7554/eLife.60066">Phan et al. (2021)</a>, and <a href="https://doi.org/10.1016/j.celrep.2022.111155">Vu et al. (2022)</a>, which generated scRNA-seq for unwounded skin from mice aged P21, P49, and P616, respectively. </p><p>The data can be loaded using the Python package Scanpy or AnnData, but you can also load it in R if you use zellkonverter. </p&gt

Biomechanics of intestinal crypt morphogenesis

The intestinal epithelium exhibits remarkable rates of self-renewal to protect the small intestine and colon from damage during digestion and facilitate nutrient absorption. This monolayer of epithelial cells is maintained by the crypts of Liehberkühn, test-tube-shaped glands that are robust in morphology and structure, undergoing significantly large deformations, despite comprising a heterogeneous composition of cells with varying proliferative capacities and mechanical properties. While the genetic and molecular processes governing crypt morphogenesis have been studied in detail, there is a lack of understanding regarding the evident contribution of biomechanical factors, leading to a poor understanding of crypt morphogenesis as a whole. Additionally, it is not known how the crypt's unique, yet incredibly robust, proliferation structure arises. However, morphoelastic rod theory allows one to consider the interplay between growth and tissue mechanics in a unified framework, where we can exploit the slenderness aspect of the crypt and model the tissue as a growing, elastic rod. In this thesis, we use the framework of morphoelastic rods, which extends the classical Kirchhoff rod theory to account for local tissue growth, to explore various aspects of morphogenesis, growth, and homeostasis that are motivated by the crypt. We restrict ourselves to a planar geometry to model the transverse deformations of the crypt epithelium. Morphogenesis is modelled by the buckling and subsequent deformation of an elastic rod tethered to an underlying foundation, representing the crypt and the supporting extracellular matrix and stroma. First, we consider an abstracted model of the crypt, a morphoelastic rod supported by an elastic foundation. We consider crypt morphogenesis in the context of buckling, exploring how growth and spatial heterogeneous properties-two key aspects of the crypt-impact mechanical pattern formation. We investigate the buckling and post-buckling behaviour of the simplified crypt model, extending previous linear stability analyses with a weakly nonlinear analysis and complementing the analysis with numerical continuation of the full nonlinear system. We analyse how incorporating spatial heterogeneity in growth, rod and foundation stiffness affects the underlying bifurcation structure. Then, we specialise the framework to simulate tissue morphogenesis more realistically. We develop different models for the processes that are believed to play a role in crypt morphogenesis, but were not previously included, such as tissue relaxation, chemical signalling, self-contact, and so on. We use simulation results to determine which of these models contribute most significantly to a realistic crypt morphology. By combining several of these processes, we show that a realistic crypt morphology, which is highly-invaginated but narrow in structure, can be generated. To close, we consider a simplified 1D geometry to analyse how the unique growth structure of the crypt rises in development and subsequently is maintained in homeostasis. We develop a minimal mechanochemical model for tissue growth that captures the proliferation structure observed in the crypt, where proliferation activity is maximal away from the crypt base and the crypt top and is thus bimodal in shape. We finish by identifying the necessary conditions for dynamic tissue homeostasis, in which the proliferation structure is fixed with respect to the observable reference frame, but there is a continuous flux of tissue material due to the balance between growth and cell death, modelled through the sloughing of material.</p

The promising application of cell-cell interaction analysis in cancer from single-cell and spatial transcriptomics.

Cell-cell interactions instruct cell fate and function. These interactions are hijacked to promote cancer development. Single-cell transcriptomics and spatial transcriptomics have become powerful new tools for researchers to profile the transcriptional landscape of cancer at unparalleled genetic depth. In this review, we discuss the rapidly growing array of computational tools to infer cell-cell interactions from non-spatial single-cell RNA-sequencing and the limited but growing number of methods for spatial transcriptomics data. Downstream analyses of these computational tools and applications to cancer studies are highlighted. We finish by suggesting several directions for further extensions that anticipate the increasing availability of multi-omics cancer data

Analyzing network diversity of cell–cell interactions in COVID-19 using single-cell transcriptomics

Cell-cell interactions (CCI) play significant roles in manipulating biological functions of cells. Analyzing the differences in CCI between healthy and diseased conditions of a biological system yields greater insight than analyzing either conditions alone. There has been a recent and rapid growth of methods to infer CCI from single-cell RNA-sequencing (scRNA-seq), revealing complex CCI networks at a previously inaccessible scale. However, the majority of current CCI analyses from scRNA-seq data focus on direct comparisons between individual CCI networks of individual samples from patients, rather than "group-level" comparisons between sample groups of patients comprising different conditions. To illustrate new biological features among different disease statuses, we investigated the diversity of key network features on groups of CCI networks, as defined by different disease statuses. We considered three levels of network features: node level, as defined by cell type; node-to-node level; and network level. By applying these analysis to a large-scale single-cell RNA-sequencing dataset of coronavirus disease 2019 (COVID-19), we observe biologically meaningful patterns aligned with the progression and subsequent convalescence of COVID-19