Dunlop, I. E. | bioRxiv Science

bioRxiv2021

Mapping the mechanical microenvironment in the ovary

Follicle development in the human ovary must be tightly regulated to ensure cyclical release of oocytes (ovulation), and disruption of this process is a common cause of infertility. Recent ex vivo studies suggest that follicle growth may be mechanically regulated, however the actual mechanical properties of the follicle microenvironment have remained unknown. Here we map and quantify the mechanical microenvironment in mouse ovaries using colloidal probe atomic force microscope (AFM) indentation, finding an overall mean Youngs Modulus 3.3 {+/-} 2.5 kPa. Spatially, stiffness is low at the ovarian edge and centre, which are dominated by extra-follicular ECM, and highest in an intermediate zone dominated by large follicles. This suggests that large follicles should be considered as mechanically dominant structures in the ovary, in contrast to previous expectations. Our results provide a new, physiologically accurate framework for investigating how mechanics impacts follicle development and will underpin future tissue engineering of the ovary.

biophysics↗

bioRxiv2020

Epithelial-stromal cell interactions and ECM mechanics drive the formation of airway-mimetic tubular morphology in lung organoids

The complex cellular organisation of the human airway tract where interaction between epithelial and stromal lineages and the extracellular matrix (ECM) make it a difficult organ to study in vitro. Current in vitro lung models focus on modelling the lung epithelium such as air-liquid interface (ALI) cultures and bronchospheres, do not model the complex morphology and the cell-ECM interaction seen in vivo. Models that include stromal populations often separate them via a semipermeable barrier, which precludes the effect of cell-cell interaction or do not include the ECM or the effect of ECM mechanics such as viscoelasticity and stiffness. Here we investigated the effect of stromal cells on basal epithelial cell-derived bronchosphere structure and function through a triple culture of bronchial epithelial, lung fibroblast and airway smooth muscle cells. Epithelial-stromal cross talk enabled formation of epithelial cell-driven branching tubules consisting of luminal epithelial cells surrounded by stromal cells termed bronchotubules. Addition of agarose to the Matrigel scaffold (Agrigel) created a mechanically tunable ECM, where viscoelasticity and stiffness could be altered to enable long term tubule survival. Bronchotubule models enable the investigation of how epithelial-stromal cell and cell-ECM communication drive tissue patterning, repair and development of disease. Significance StatementCurrent models of airways diseases such as asthma and COPD do not reflect the physical characteristics of the diseased airway which may impact upon our understanding of disease pathophysiology. We have utilised the physical properties of agarose to modify the 3D stiffness of Matrigel to resemble the human airway. Using a primary airway epithelial cell-derived organoid model we demonstrate that a combined Matrigel/agrigel matrix allows sustained 3D organoid structure and the creation of tubules that can contract in response to a clinically relevant bronchoconstrictor. A complex 3D organoid composed of functioning epithelial cells, smooth muscle cells and fibroblasts may provide opportunities for refined drug discovery programmes. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=141 SRC="FIGDIR/small/408815v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@906212org.highwire.dtl.DTLVardef@1168e15org.highwire.dtl.DTLVardef@1ca16b2org.highwire.dtl.DTLVardef@980f65_HPS_FORMAT_FIGEXP M_FIG C_FIG O_LIMixture of healthy lung basal epithelial cells and healthy lung fibroblast cultured in matrigel result in tubules that fail in 4 days. C_LIO_LIAddition of healthy airway smooth muscle allows for a contractile phenotype. C_LIO_LITriple culture of cells in a stiffer scaffold agrigel allows maintenance of tubular organoids for a minimum of 20 days. C_LI

bioengineering↗