The interplay of cell–cell and cell–substrate adhesion in collective cell migration

Collective cell migration often involves notable cell–cell and cell–substrate adhesions and highly coordinated motion of touching cells. We focus on the interplay between cell–substrate adhesion and cell–cell adhesion. We show that the loss of cell-surface contact does not significantly alter the dynamic pattern of protrusions and retractions of fast migrating amoeboid cells (Dictyostelium discoideum), but significantly changes their ability to adhere to other cells. Analysis of the dynamics of cell shapes reveals that cells that are adherent to a surface may coordinate their motion with neighbouring cells through protrusion waves that travel across cell–cell contacts. However, while shape waves exist if cells are detached from surfaces, they do not couple cell to cell. In addition, our investigation of actin polymerization indicates that loss of cell-surface adhesion changes actin polymerization at cell–cell contacts. To further investigate cell–cell/cell–substrate interactions, we used optical micromanipulation to form cell–substrate contact at controlled locations. We find that both cell-shape dynamics and cytoskeletal activity respond rapidly to the formation of cell–substrate contact.     

Modelling planar polarity of epithelia: the role of signal relay in collective cell polarization

Collective cell polarization is an important characteristic of tissues. Epithelia commonly display cellular structures that are polarized within the plane of the tissue. Establishment of this planar cell polarity requires mechanisms that locally align polarized structures between neighbouring cells, as well as cues that provide global information about alignment relative to an axis of a tissue. In the Drosophila ovary, the cadherin Fat2 is required to orient actin filaments located at the basal side of follicle cells perpendicular to the long axis of the egg chamber. The mechanisms directing this orientation of actin filaments, however, remain unknown. Here we show, using genetic mosaic analysis, that fat2 is not essential for the local alignment of actin filaments between neighbouring cells. Moreover, we provide evidence that Fat2 is involved in the propagation of a cue specifying the orientation of actin filaments relative to the tissue axis. Monte Carlo simulations of actin filament orientation resemble the results of the genetic mosaic analysis, if it is assumed that a polarity signal can propagate from a signal source only through a connected chain of wild-type cells. Our results suggest that Fat2 is required for propagating global polarity information within the follicle epithelium through direct cell–cell contact. Our computational model might be more generally applicable to study collective cell polarization in tissues.     

How the tooth got its stripes: patterning via strain-cued motility

We hypothesize that a population of migrating cells can form patterns when changes in local strains owing to relative cell motions induce changes in cell motility. That the mechanism originates in competing rates of motion distinguishes it from mechanisms involving strain energy gradients, e.g. those generated by surface energy effects or eigenstrains among cells, and diffusion–reaction mechanisms involving chemical signalling factors. The theory is tested by its ability to reproduce the morphological characteristics of enamel in the mouse incisor. Dental enamel is formed during amelogenesis by a population of ameloblasts that move about laterally within an expanding curved sheet, subject to continuously evolving spatial and temporal gradients in strain. Discrete-cell simulations of this process compute the changing strain environment of all cells and predict cell trajectories by invoking simple rules for the motion of an individual cell in response to its strain environment. The rules balance a tendency for cells to enhance relative sliding motion against a tendency to maintain uniform cell–cell separation. The simulations account for observed waviness in the enamel microstructure, the speed and shape of the ‘commencement front’ that separates domains of migrating secretory-stage ameloblasts from those that are not yet migrating, the initiation and sustainment of layered, fracture-resistant decussation patterns (cross-plied microstructure) and the transition from decussating inner enamel to non-decussating outer enamel. All these characteristics can be correctly predicted with the use of a single scalar adjustable parameter.     

Computational modelling of epidermal stratification highlights the importance of asymmetric cell division for predictable and robust layer formation

Skin is a complex organ tasked with, among other functions, protecting the body from the outside world. Its outermost protective layer, the epidermis, is comprised of multiple cell layers that are derived from a single-layered ectoderm during development. Using a new stochastic, multi-scale computational modelling framework, the anisotropic subcellular element method, we investigate the role of cell morphology and biophysical cell–cell interactions in the formation of this layered structure. This three-dimensional framework describes interactions between collections of hundreds to thousands of cells and (i) accounts for intracellular structure and morphology, (ii) easily incorporates complex cell–cell interactions and (iii) can be efficiently implemented on parallel architectures. We use this approach to construct a model of the developing epidermis that accounts for the internal polarity of ectodermal cells and their columnar morphology. Using this model, we show that cell detachment, which has been previously suggested to have a role in this process, leads to unpredictable, randomized stratification and that this cannot be abrogated by adjustment of cell–cell adhesion interaction strength. Polarized distribution of cell adhesion proteins, motivated by epithelial polarization, can however eliminate this detachment, and in conjunction with asymmetric cell division lead to robust and predictable development.     

Peeling dynamics of fluid membranes bridged by molecular bonds: moving or breaking

Biological adhesion is a critical mechanical function of complex organisms. At the scale of cell–cell contacts, adhesion is remarkably tunable to enable both cohesion and malleability during development, homeostasis and disease. It is physically supported by transient and laterally mobile molecular bonds embedded in fluid membranes. Thus, unlike specific adhesion at solid–solid or solid–fluid interfaces, peeling at fluid–fluid interfaces can proceed by breaking bonds, by moving bonds or by a combination of both. How the additional degree of freedom provided by bond mobility changes the mechanics of peeling is not understood. To address this, we develop a theoretical model coupling diffusion, reactions and mechanics. Mobility and reaction rates determine distinct peeling regimes. In a diffusion-dominated Stefan-like regime, bond motion establishes self-stabilizing dynamics that increase the effective fracture energy. In a reaction-dominated regime, peeling proceeds by travelling fronts where marginal diffusion and unbinding control peeling speed. In a mixed reaction–diffusion regime, strengthening by bond motion competes with weakening by bond breaking in a force-dependent manner, defining the strength of the adhesion patch. In turn, patch strength depends on molecular properties such as bond stiffness, force sensitivity or crowding. We thus establish the physical rules enabling tunable cohesion in cellular tissues and in engineered biomimetic systems.     

Shear-induced orientational dynamics and spatial heterogeneity in suspensions of motile phytoplankton

Fluid flow, ubiquitous in natural and man-made environments, has the potential to profoundly impact the transport of microorganisms, including phytoplankton in aquatic habitats and bioreactors. Yet, the effect of ambient flow on the swimming behaviour of phytoplankton has remained poorly understood, largely owing to the difficulty of observing cell–flow interactions at the microscale. Here, we present microfluidic experiments where we tracked individual cells for four species of motile phytoplankton exposed to a spatially non-uniform fluid shear rate, characteristic of many flows in natural and artificial environments. We observed that medium-to-high mean shear rates (1–25 s⁻¹) produce heterogeneous cell concentrations in the form of regions of accumulation and regions of depletion. The location of these regions relative to the flow depends on the cells'' propulsion mechanism, body shape and flagellar arrangement, as captured by an effective aspect ratio. Species having a large effective aspect ratio accumulated in the high-shear regions, owing to shear-induced alignment of the swimming orientation with the fluid streamlines. Species having an effective aspect ratio close to unity exhibited little preferential accumulation at low-to-moderate flow rates, but strongly accumulated in the low-shear regions under high flow conditions, potentially owing to an active, behavioural response of cells to shear. These observations demonstrate that ambient fluid flow can strongly affect the motility and spatial distribution of phytoplankton and highlight the rich dynamics emerging from the interaction between motility, morphology and flow.     

Cost–benefit analysis of the mechanisms that enable migrating cells to sustain motility upon changes in matrix environments

Cells can move through extracellular environments with varying geometries and adhesive properties. Adaptation to these differences is achieved by switching between different modes of motility, including lamellipod-driven and blebbing motility. Further, cells can modulate their level of adhesion to the extracellular matrix (ECM) depending on both the level of force applied to the adhesions and cell intrinsic biochemical properties. We have constructed a computational model of cell motility to investigate how motile cells transition between extracellular environments with varying surface continuity, confinement and adhesion. Changes in migration strategy are an emergent property of cells as the ECM geometry and adhesion changes. The transition into confined environments with discontinuous ECM fibres is sufficient to induce shifts from lamellipod-based to blebbing motility, while changes in confinement alone within a continuous geometry are not. The geometry of the ECM facilitates plasticity, by inducing shifts where the cell has high marginal gain from a mode change, and conserving persistency where the cell can continue movement regardless of the motility mode. This regulation of cell motility is independent of global changes in cytoskeletal properties, but requires locally higher linkage between the actin network and the plasma membrane at the cell rear, and changes in internal cell pressure. In addition to matrix geometry, we consider how cells might transition between ECM of different adhesiveness. We find that this requires positive feedback between the forces cells apply on the adhesion points, and the strength of the cell–ECM adhesions on those sites. This positive feedback leads to the emergence of a small number of highly adhesive cores, similar to focal adhesions. While the range of ECM adhesion levels the cell can invade is expanded with this feedback mechanism; the velocities are lowered for conditions where the positive feedback is not vital. Thus, plasticity of cell motility sacrifices the benefits of specialization, for robustness.     

Collective cell migration: leadership,invasion and segregation

A number of biological processes, such as embryo development, cancer metastasis or wound healing, rely on cells moving in concert. The mechanisms leading to the emergence of coordinated motion remain however largely unexplored. Although biomolecular signalling is known to be involved in most occurrences of collective migration, the role of physical and mechanical interactions has only been recently investigated. In this study, a versatile framework for cell motility is implemented in silico in order to study the minimal requirements for the coordination of a group of epithelial cells. We find that cell motility and cell–cell mechanical interactions are sufficient to generate a broad array of behaviours commonly observed in vitro and in vivo. Cell streaming, sheet migration and susceptibility to leader cells are examples of behaviours spontaneously emerging from these simple assumptions, which might explain why collective effects are so ubiquitous in nature. The size of the population and its confinement appear, in particular, to play an important role in the coordination process. In all cases, the complex response of the population can be predicted from the knowledge of the correlation length of the velocity field measured in the bulk of the epithelial layer. This analysis provides also new insights into cancer metastasis and cell sorting, suggesting, in particular, that collective invasion might result from an emerging coordination in a system where single cells are mechanically unable to invade.     

Modelling epigenetic regulation of gene expression in 12 human cell types reveals combinatorial patterns of cell‐type‐specific genes

The maintenance of the diverse cell types in a multicellular organism is one of the fundamental mysteries of biology. Modelling the dynamic regulatory relationships between the histone modifications and the gene expression across the diverse cell types is essential for the authors to understand the mechanisms of the epigenetic regulation. Here, the authors thoroughly assessed the histone modification enrichment profiles at the promoters and constructed quantitative models between the histone modification abundances and the gene expression in 12 human cell types. The author''s results showed that the histone modifications at the promoters exhibited remarkably cell‐type‐dependent variability in the cell‐type‐specific (CTS) genes. They demonstrated that the variable profiles of the modifications are highly predictive for the dynamic changes of the gene expression across all the cell types. Their findings revealed the close relationship between the combinatorial patterns of the histone modifications and the CTS gene expression. They anticipate that the findings and the methods they used in this study could provide useful information for the future studies of the regulatory roles of the histone modifications in the CTS genes.Inspec keywords: cellular biophysics, genetics, genomics, physiological models, proteinsOther keywords: CTS gene expression, variable profiles, cell‐type‐dependent variability, histone modification abundances, constructed quantitative models, promoters, histone modification enrichment profiles, dynamic regulatory relationship modelling, biology, multicellular organism, cell‐type‐specific genes, combinatorial patterns, human cell types, epigenetic regulation modelling     

Shear-induced force transmission in a multicomponent,multicell model of the endothelium

Haemodynamic forces applied at the apical surface of vascular endothelial cells (ECs) provide the mechanical signals at intracellular organelles and through the inter-connected cellular network. The objective of this study is to quantify the intracellular and intercellular stresses in a confluent vascular EC monolayer. A novel three-dimensional, multiscale and multicomponent model of focally adhered ECs is developed to account for the role of potential mechanosensors (glycocalyx layer, actin cortical layer, nucleus, cytoskeleton, focal adhesions (FAs) and adherens junctions (ADJs)) in mechanotransmission and EC deformation. The overriding issue addressed is the stress amplification in these regions, which may play a role in subcellular localization of mechanotransmission. The model predicts that the stresses are amplified 250–600-fold over apical values at ADJs and 175–200-fold at FAs for ECs exposed to a mean shear stress of 10 dyne cm⁻². Estimates of forces per molecule in the cell attachment points to the external cellular matrix and cell–cell adhesion points are of the order of 8 pN at FAs and as high as 3 pN at ADJs, suggesting that direct force-induced mechanotransmission by single molecules is possible in both. The maximum deformation of an EC in the monolayer is calculated as 400 nm in response to a mean shear stress of 1 Pa applied over the EC surface which is in accord with measurements. The model also predicts that the magnitude of the cell–cell junction inclination angle is independent of the cytoskeleton and glycocalyx. The inclination angle of the cell–cell junction is calculated to be 6.6° in an EC monolayer, which is somewhat below the measured value (9.9°) reported previously for ECs subjected to 1.6 Pa shear stress for 30 min. The present model is able, for the first time, to cross the boundaries between different length scales in order to provide a global view of potential locations of mechanotransmission.     

Synergy of cell–cell repulsion and vacuolation in a computational model of lumen formation

A key step in blood vessel development (angiogenesis) is lumen formation: the hollowing of vessels for blood perfusion. Two alternative lumen formation mechanisms are suggested to function in different types of blood vessels. The vacuolation mechanism is suggested for lumen formation in small vessels by coalescence of intracellular vacuoles, a view that was extended to extracellular lumen formation by exocytosis of vacuoles. The cell–cell repulsion mechanism is suggested to initiate extracellular lumen formation in large vessels by active repulsion of adjacent cells, and active cell shape changes extend the lumen. We used an agent-based computer model, based on the cellular Potts model, to compare and study both mechanisms separately and combined. An extensive sensitivity analysis shows that each of the mechanisms on its own can produce lumens in a narrow region of parameter space. However, combining both mechanisms makes lumen formation much more robust to the values of the parameters, suggesting that the mechanisms may work synergistically and operate in parallel, rather than in different vessel types.     

Spatial gradients in kinase cascade regulation

The spatiotemporal kinetics of proteins and other substrates regulate cell fate and signaling. In this study, we consider a reaction-diffusion model of interaction of membrane receptors with a two-step kinase cascade. The receptors activate the 'up-stream' kinase, which may diffuse over cell volume and activate the 'down-stream' kinase, which is also diffusing. Both kinase species and receptors are inactivated by uniformly distributed phosphatases. The positive feedback, key to the considered dynamics, arises since the up-stream kinase activates the receptors. Such a mutual interaction is characteristic for immune cell receptors. Based on the proposed model, we demonstrated that cell sensitivity (measured as a critical value of phosphatase activity at which cell maybe activated) increases with decreasing motility of receptor-interacting kinases and with increasing polarity of receptors distribution. These two effects are cooperating, the effect of receptors localisation close to one pole of the cell grows with the decreasing kinase diffusion and vanishes in the infinite diffusion limit. As the cell sensitivity increases with decreasing diffusion of receptor-interacting kinase, the overall activity of the down-stream kinase increases with its diffusion. In conclusion, the analysis of the proposed model shows that, for the fixed substrate interaction rates, spatial distribution of the surface receptors together with the motility of intracellular kinases control cell signalling and sensitivity to extracellular signals. The increase of the cell sensitivity can be achieved by (i) localisation of receptors in a small subdomain of the cell membrane, (ii) lowering the motility of receptor-interacting kinase, (iii) increasing the motility of down-stream kinases which distribute the signal over the whole cell.     

Actin flow and talin dynamics govern rigidity sensing in actin–integrin linkage through talin extension

At cell–substrate adhesion sites, the linkage between actin filaments and integrin is regulated by mechanical stiffness of the substrate. Of potential molecular regulators, the linker proteins talin and vinculin are of particular interest because mechanical extension of talin induces vinculin binding with talin, which reinforces the actin–integrin linkage. For understanding the molecular and biophysical mechanism of rigidity sensing at cell–substrate adhesion sites, we constructed a simple physical model to examine a role of talin extension in the stiffness-dependent regulation of actin–integrin linkage. We show that talin molecules linking between retrograding actin filaments and substrate-bound integrin are extended in a manner dependent on substrate stiffness. The model predicts that, in adhesion complexes containing ≈30 talin links, talin is extended enough for vinculin binding when the substrate is stiffer than 1 kPa. The lifetime of talin links needs to be 2–5 s to achieve an appropriate response of talin extension against substrate stiffness. Furthermore, changes in actin velocity drastically shift the range of substrate stiffness that induces talin–vinculin binding. Our results suggest that talin extension is a key step in sensing and responding to substrate stiffness at cell adhesion sites.     

Cell type-specific aspects in biocompatibility testing: the intercellular contact in vitro as an indicator for endothelial cell compatibility

Endothelial cells cover the inner surface of blood vessels and form the interface between the blood and the tissues. Endothelial cells are involved in regulating barrier function, which is maintained by the interendothelial cell contacts. These interendothelial cell contacts are established by the interaction of different molecules. The maintenance of the barrier requires an appropriate signalling between these molecules. Thus, a number of different signalling pathways are integrated within interendothelial contacts. Since endothelial cells are important in tissue-implant interactions (especially for stent materials) this study examines the expression pattern of different interendothelial contact molecules to determine the usefulness in the analysis of biocompatibility in vitro. The effects of different pro-inflammatory and toxic stimuli and contact of human microvascular endothelial cells to metallic surfaces were examined for their impact on the pattern of interendothelial contact molecules. Striking modifications in the arrangement of these molecules were induced and the mode of modification was dependent on the tested compound. Thus, examining the pattern of expression of specific interendothelial contact molecules in vitro may be useful for testing the endothelial cell compatibility of biomaterials and their corrosion products.     

Mechanics regulates ATP-stimulated collective calcium response in fibroblast cells

Cells constantly sense their chemical and mechanical environments. We study the effect of mechanics on the ATP-induced collective calcium response of fibroblast cells in experiments that mimic various tissue environments. We find that closely packed two-dimensional cell cultures on a soft polyacrylamide gel (Young''s modulus E = 690 Pa) contain more cells exhibiting calcium oscillations than cultures on a rigid substrate (E = 36 000 Pa). Calcium responses of cells on soft substrates show a slower decay of calcium level relative to those on rigid substrates. Actin enhancement and disruption experiments for the cell cultures allow us to conclude that actin filaments determine the collective Ca²⁺ oscillatory behaviour in the culture. Inhibition of gap junctions results in a decrease of the oscillation period and reduced correlation of calcium responses, which suggests additional complexity of signalling upon cell–cell contact. Moreover, the frequency of calcium oscillations is independent of the rigidity of the substrate but depends on ATP concentration. We compare our results with those from similar experiments on individual cells. Overall, our observations show that collective chemical signalling in cell cultures via calcium depends critically on the mechanical environment.     

Multi-scale modelling of the dynamics of cell colonies: insights into cell-adhesion forces and cancer invasion from in silico simulations

Studying the biophysical interactions between cells is crucial to understanding how normal tissue develops, how it is structured and also when malfunctions occur. Traditional experiments try to infer events at the tissue level after observing the behaviour of and interactions between individual cells. This approach assumes that cells behave in the same biophysical manner in isolated experiments as they do within colonies and tissues. In this paper, we develop a multi-scale multi-compartment mathematical model that accounts for the principal biophysical interactions and adhesion pathways not only at a cell–cell level but also at the level of cell colonies (in contrast to the traditional approach). Our results suggest that adhesion/separation forces between cells may be lower in cell colonies than traditional isolated single-cell experiments infer. As a consequence, isolated single-cell experiments may be insufficient to deduce important biological processes such as single-cell invasion after detachment from a solid tumour. The simulations further show that kinetic rates and cell biophysical characteristics such as pressure-related cell-cycle arrest have a major influence on cell colony patterns and can allow for the development of protrusive cellular structures as seen in invasive cancer cell lines independent of expression levels of pro-invasion molecules.     

The importance of structured noise in the generation of self-organizing tissue patterns through contact-mediated cell�Ccell signalling

Lateral inhibition provides the basis for a self-organizing patterning system in which distinct cell states emerge from an otherwise uniform field of cells. The development of the microchaete bristle pattern on the notum of the fruitfly, Drosophila melanogaster, has long served as a popular model of this process. We recently showed that this bristle pattern depends upon a population of dynamic, basal actin-based filopodia, which span multiple cell diameters. These protrusions establish transient signalling contacts between non-neighbouring cells, generating a type of structured noise that helps to yield a well-ordered and spaced pattern of bristles. Here, we develop a general model of protrusion-based patterning to analyse the role of noise in this process. Using a simple asynchronous cellular automata rule-based model we show that this type of structured noise drives the gradual refinement of lateral inhibition-mediated patterning, as the system moves towards a stable configuration in which cells expressing the inhibitory signal are near-optimally packed. By analysing the effects of introducing thresholds required for signal detection in this model of lateral inhibition, our study shows how filopodia-mediated cell–cell communication can generate complex patterns of spots and stripes, which, in the presence of signalling noise, align themselves across a patterning field. Thus, intermittent protrusion-based signalling has the potential to yield robust self-organizing tissue-wide patterns without the need to invoke diffusion-mediated signalling.     

From cytotoxicity to biocompatibility testing in vitro: cell adhesion molecule expression defines a new set of parameters

Determination of potential cytotoxicity is a central issue in current biocompatibility testing standards such as ISO and ASTM. Most of these tests do not assess biocompatibility of a biomaterial with regard to cell function. This study was aimed at screening a number of potential parameters that could be included in assessment of cell functional aspects of biocompatibility. Human umbilical vein endothelial cells (HUVEC) were seeded directly on titanium, NiCr alloy, CoCr alloy, PMMA, PE, PU, PVC, and silicone, or were exposed to the material extracts. Cytotoxicity was assessed for these materials through MTT conversion, crystal violet protein determination and Ki67 expression. In addition, expression of the cell adhesion molecules E-selectin, cadherin-5 and PECAM, as well as of the adhesion-associated proteins fibronectin and vinculin (focal adhesions), was determined by immunocytochemistry and western blotting. Cytotoxicity was not detected with the material extracts. Cells were able to adhere to bare metals, but not polymers. Fibronectin preadsorption resulted in adhesion and spreading also on the polymers. Cells were able to establish cell–cell contacts and focal adhesions. Western blotting, in combination with differential detergent extraction, indicated that linkage of cell–cell adhesion markers to the cytoskeleton may be used as an additional parameter relevant to cell function.