Collective Cell Behavior in Mechanosensing of Substrate Thickness.

Extracellular matrix stiffness has a profound effect on the behavior of many cell types. Adherent cells apply contractile forces to the material on which they adhere and sense the resistance of the material to deformation-its stiffness. This is dependent on both the elastic modulus and the thickness of the material, with the corollary that single cells are able to sense underlying stiff materials through soft hydrogel materials at low (<10 µm) thicknesses. Here, we hypothesized that cohesive colonies of cells exert more force and create more hydrogel deformation than single cells, therefore enabling them to mechanosense more deeply into underlying materials than single cells. To test this, we modulated the thickness of soft (1 kPa) elastic extracellular-matrix-functionalized polyacrylamide hydrogels adhered to glass substrates and allowed colonies of MG63 cells to form on their surfaces. Cell morphology and deformations of fluorescent fiducial-marker-labeled hydrogels were quantified by time-lapse fluorescence microscopy imaging. Single-cell spreading increased with respect to decreasing hydrogel thickness, with data fitting to an exponential model with half-maximal response at a thickness of 3.2 µm. By quantifying cell area within colonies of defined area, we similarly found that colony-cell spreading increased with decreasing hydrogel thickness but with a greater half-maximal response at 54 µm. Depth-sensing was dependent on Rho-associated protein kinase-mediated cellular contractility. Surface hydrogel deformations were significantly greater on thick hydrogels compared to thin hydrogels. In addition, deformations extended greater distances from the periphery of colonies on thick hydrogels compared to thin hydrogels. Our data suggest that by acting collectively, cells mechanosense rigid materials beneath elastic hydrogels at greater depths than individual cells. This raises the possibility that the collective action of cells in colonies or sheets may allow cells to sense structures of differing material properties at comparatively large distances.

Epithelial mechanobiology, skin wound healing, and the stem cell niche.

Skin wound healing is a vital process that is important for re-establishing the epithelial barrier following disease or injury. Aberrant or delayed skin wound healing increases the risk of infection, causes patient morbidity, and may lead to the formation of scar tissue. One of the most important events in wound healing is coverage of the wound with a new epithelial layer. This occurs when keratinocytes at the wound periphery divide and migrate to re-populate the wound bed. Many approaches are under investigation to promote and expedite this process, including the topical application of growth factors and the addition of autologous and allogeneic tissue or cell grafts. The mechanical environment of the wound site is also of fundamental importance for the rate and quality of wound healing. It is known that mechanical stress can influence wound healing by affecting the behaviour of cells within the dermis, but it remains unclear how mechanical forces affect the healing epidermis. Tensile forces are known to affect the behaviour of cells within epithelia, however, and the material properties of extracellular matrices, such as substrate stiffness, have been shown to affect the morphology, proliferation, differentiation and migration of many different cell types. In this review we will introduce the structure of the skin and the process of wound healing. We will then discuss the evidence for the effect of tissue mechanics in re-epithelialisation and, in particular, on stem cell behaviour in the wound microenvironment and in intact skin. We will discuss how the elasticity, mechanical heterogeneity and topography of the wound extracellular matrix impact the rate and quality of wound healing, and how we may exploit this knowledge to expedite wound healing and mitigate scarring.