Role of cell polarity dynamics and motility in pattern formation due to contact-dependent signalling

A key challenge in biology is to understand how spatio-temporal patterns and structures arise during the development of an organism. An initial aggregate of spatially uniform cells develops and forms the differentiated structures of a fully developed organism. On the one hand, contact-dependent cell–cell signalling is responsible for generating a large number of complex, self-organized, spatial patterns in the distribution of the signalling molecules. On the other hand, the motility of cells coupled with their polarity can independently lead to collective motion patterns that depend on mechanical parameters influencing tissue deformation, such as cellular elasticity, cell–cell adhesion and active forces generated by actin and myosin dynamics. Although modelling efforts have, thus far, treated cell motility and cell–cell signalling separately, experiments in recent years suggest that these processes could be tightly coupled. Hence, in this paper, we study how the dynamics of cell polarity and migration influence the spatiotemporal patterning of signalling molecules. Such signalling interactions can occur only between cells that are in physical contact, either directly at the junctions of adjacent cells or through cellular protrusional contacts. We present a vertex model which accounts for contact-dependent signalling between adjacent cells and between non-adjacent neighbours through long protrusional contacts that occur along the orientation of cell polarization. We observe a rich variety of spatiotemporal patterns of signalling molecules that is influenced by polarity dynamics of the cells, relative strengths of adjacent and non-adjacent signalling interactions, range of polarized interaction, signalling activation threshold, relative time scales of signalling and polarity orientation, and cell motility. Though our results are developed in the context of Delta–Notch signalling, they are sufficiently general and can be extended to other contact dependent morpho-mechanical dynamics.     

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.     

Orthogonal (transverse) arrangements of actin in endothelia and fibroblasts.

Though actin filaments running across the cell (transverse actin) have been occasionally reported for epithelial cells in groups and for cells growing on fibres, there has been no report heretofore of transverse actin in cells grown on planar substrata. This paper describes evidence in support of this possibility derived from actin staining, polarization microscopy and force measurements. The paper introduces two new methods for detecting the orientation and activity of contractile elements in cells. The orthogonal actin is most obvious in cells grown on groove ridge structures, but can be detected in cells grown on flat surfaces.     

The effect of remodelling and contractility of the actin cytoskeleton on the shear resistance of single cells: a computational and experimental investigation

The biomechanisms that govern the response of chondrocytes to mechanical stimuli are poorly understood. In this study, a series of in vitro tests are performed, in which single chondrocytes are subjected to shear deformation by a horizontally moving probe. Dramatically different probe force–indentation curves are obtained for untreated cells and for cells in which the actin cytoskeleton has been disrupted. Untreated cells exhibit a rapid increase in force upon probe contact followed by yielding behaviour. Cells in which the contractile actin cytoskeleton was removed exhibit a linear force–indentation response. In order to investigate the mechanisms underlying this behaviour, a three-dimensional active modelling framework incorporating stress fibre (SF) remodelling and contractility is used to simulate the in vitro tests. Simulations reveal that the characteristic force–indentation curve observed for untreated chondrocytes occurs as a result of two factors: (i) yielding of SFs due to stretching of the cytoplasm near the probe and (ii) dissociation of SFs due to reduced cytoplasm tension at the front of the cell. In contrast, a passive hyperelastic model predicts a linear force–indentation curve similar to that observed for cells in which the actin cytoskeleton has been disrupted. This combined modelling–experimental study offers a novel insight into the role of the active contractility and remodelling of the actin cytoskeleton in the response of chondrocytes to mechanical loading.     

Contact inhibition of locomotion probabilities drive solitary versus collective cell migration

Contact inhibition of locomotion (CIL) is the process whereby cells collide, cease migrating in the direction of the collision, and repolarize their migration machinery away from the collision. Quantitative analysis of CIL has remained elusive because cell-to-cell collisions are infrequent in traditional cell culture. Moreover, whereas CIL predicts mutual cell repulsion and ‘scattering’ of cells, the same cells in vivo are observed to undergo CIL at some developmental times and collective cell migration at others. It remains unclear whether CIL is simply absent during collective cell migration, or if the two processes coexist and are perhaps even related. Here, we used micropatterned stripes of extracellular matrix to restrict cell migration to linear paths such that cells polarized in one of two directions and collisions between cells occurred frequently and consistently, permitting quantitative and unbiased analysis of CIL. Observing repolarization events in different contexts, including head-to-head collision, head-to-tail collision, collision with an inert barrier, or no collision, and describing polarization as a two-state transition indicated that CIL occurs probabilistically, and most strongly upon head-to-head collisions. In addition to strong CIL, we also observed ‘trains’ of cells moving collectively with high persistence that appeared to emerge from single cells. To reconcile these seemingly conflicting observations of CIL and collective cell migration, we constructed an agent-based model to simulate our experiments. Our model quantitatively predicted the emergence of collective migration, and demonstrated the sensitivity of such emergence to the probability of CIL. Thus CIL and collective migration can coexist, and in fact a shift in CIL probabilities may underlie transitions between solitary cell migration and collective cell migration. Taken together, our data demonstrate the emergence of persistently polarized, collective cell movement arising from CIL between colliding cells.     

Magnetic manipulation of actin orientation, polymerization, and gliding on myosin using superparamagnetic iron oxide particles

The actin cytoskeleton controls cell shape, motility, as well as intracellular molecular trafficking. The ability to remotely manipulate actin is therefore highly desirable as a tool to probe and manipulate biological processes at the molecular level. We demonstrate actin manipulation by labeling actin filaments with superparamagnetic iron oxide particles (IOPs) and applying a uniform magnetic field to affect actin orientation, polymerization and gliding on myosin. We show for the first time magnetic manipulation of magnetizable actin filaments at the molecular level while gliding on a bed of myosin molecules and during polymerization. A model for the magnetic alignment and guiding mechanism is proposed based on the torque from the induced molecular anisotropy due to interactions between neighboring IOPs distributed along magnetically labeled actin molecules.     

Cell Behavior within Nanogrooved Sandwich Culture Systems

Grooved topography and inherent cell contact guidance has shown promising results regarding cell proliferation, morphology, and lineage‐specific differentiation. Yet these approaches are limited to 2D applications. Sandwich‐culture conditions are developed to bridge the gap between 2D and 3D culture, enabling both ventral and dorsal cell surface stimulation. The effect of grooved surface topography is accessed on cell orientation and elongation in a highly controlled manner, with simultaneous and independent stimuli on two cell sides. Nanogrooved and non‐nanogrooved substrates are assembled into quasi‐3D systems with variable relative orientations. A plethora of sandwich‐culture conditions are created by seeding cells on lower, upper, or both substrates. Software image analysis demonstrates that F‐actin of cells acquires the orientation of the substrate on which cells are initially seeded, independently from the orientation of the second top substrate. Contrasting cell morphologies are observed, with a higher elongation for nanogrooved 2D substrates than nanogrooved sandwich‐culture conditions. Correlated with an increased pFAK activity and vinculin staining for sandwich‐culture conditions, these results point to an enhanced cell surface stimulation versus control conditions. The pivotal role of initial cell‐biomaterial contact on cellular alignment is highlighted, providing important insights for tissue engineering strategies aiming to guide cellular response through mechanotransduction approaches.     

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.     

Theory of domain wall motion induced by microwave magnetic fields

A general theory is developed that applies to arbitrary polarization and takes account of damping and of the dipolar interaction between domains. The effect of the microwave field on the domain structure can be characterized by a pressure on the domain walls and by an alignment energy, both of which are proportional to the square of the rf magnetic field and become large in the vicinity of a resonance. For circular polarization the pressure tends to decrease the Larmor-domains (domains in which the imposed sense of polarization coincides with the sense of the natural spin precession) for frequencies outside the resonance region. Inside the resonance region, however, the pressure tends to increase the Larmor-domains. A linearly polarized field also exerts a pressure on the domain walls, with the polarity dependent upon the orientation of the field to the wall normal. In a linearly polarized magnetic field the domain walls tend to become aligned parallel to the rf field at frequencies ω near the low-frequency resonance (ω =γHa, γ = gyromagnetic ratio, Ha= anisotropy field) and perpendicular to the rf field at frequencies near the high-frequency resonance (ω = γ[Ha(Ha+ 4πM0)]^1/2, M0= saturation magnetization).     

Rear actomyosin contractility-driven directional cell migration in three-dimensional matrices: a mechano-chemical coupling mechanism

Cell migration is of vital importance in many biological processes, including organismal development, immune response and development of vascular diseases. For instance, migration of vascular smooth muscle cells from the media to intima is an essential part of the development of atherosclerosis and restenosis after stent deployment. While it is well characterized that cells use actin polymerization at the leading edge to propel themselves to move on two-dimensional substrates, the migration modes of cells in three-dimensional matrices relevant to in vivo environments remain unclear. Intracellular tension, which is created by myosin II activity, fulfils a vital role in regulating cell migration. We note that there is compelling evidence from theoretical and experimental work that myosin II accumulates at the cell rear, either isoform-dependent or -independent, leading to three-dimensional migration modes driven by posterior myosin II tension. The scenario is not limited to amoeboid migration, and it is also seen in mesenchymal migration in which a two-dimensional-like migration mode based on front protrusions is often expected, suggesting that there may exist universal underlying mechanisms. In this review, we aim to shed some light on how anisotropic myosin II localization induces cell motility in three-dimensional environments from a biomechanical view. We demonstrate an interesting mechanism where an interplay between mechanical myosin II recruitment and biochemical myosin II activation triggers directional migration in three-dimensional matrices. In the case of amoeboid three-dimensional migration, myosin II first accumulates at the cell rear to induce a slight polarization displayed as a uropod-like structure under the action of a tension-dependent mechanism. Subsequent biochemical signalling pathways initiate actomyosin contractility, producing traction forces on the adhesion system or creating prominent motile forces through blebbing activity, to drive cells to move. In mesenchymal three-dimensional migration, cells can also take advantage of the elastic properties of three-dimensional matrices to move. A minor myosin isoform, myosin IIB, is retained by relatively stiff three-dimensional matrices at the posterior side, then activated by signalling cascades, facilitating prominent cell polarization by establishing front–back polarity and creating cell rear. Myosin IIB initiates cell polarization and coordinates with the major isoform myosin IIA-assembled stress fibres, to power the directional migration of cells in the three-dimensional matrix.     

Effect of pore size of self-organized honeycomb-patterned polymer films on spreading, focal adhesion, proliferation, and function of endothelial cells

The design of nano- and microstructures based on self-organization is a key area of research in the search for new materials, and it has a variety of potential applications in tissue engineering scaffolds. We have reported a honeycomb-patterned polymer film (honeycomb film) with highly regular pores that is formed by self-organization. This study describes the behavior of vascular endothelial cells (ECs) on honeycomb films with four different pore sizes (5, 9, 12, and 16 microm) as well as on a flat film. We examined the influence of the honeycomb pattern and pore size on cell behavior. The changes in cell morphologies, actin filaments, vinculin clusters, cell proliferation, and secreted extracellular matrix (ECM) (fibronectin, laminin, type IV collagen, and elastin) production profiles were observed by using optical, fluorescence, and scanning electron microscopy. The ECs that adhered to the flat film showed an elongated morphology with random orientation; the actin filaments and focal adhesions were not conspicuous. On the other hand, the ECs on the honeycomb films exhibited greater spreading and flattening; the degree of spreading of the ECs increased with an increase in the pore size. The actin filaments and focal adhesions appeared conspicuous, and the focal adhesions localized along the edge of the honeycomb pores were distributed over the entire projected cell area. The honeycomb film with a pore size of 5 microm showed the highest cell proliferation and ECM production profiles. These results suggest that the honeycomb film is a suitable material for designing a new vascular device.     

Model for self-polarization and motility of keratocyte fragments

Computational modelling of cell motility on substrates is a formidable challenge; regulatory pathways are intertwined and forces that influence cell motion are not fully quantified. Additional challenges arise from the need to describe a moving deformable cell boundary. Here, we present a simple mathematical model coupling cell shape dynamics, treated by the phase-field approach, to a vector field describing the mean orientation (polarization) of the actin filament network. The model successfully reproduces the primary phenomenology of cell motility: discontinuous onset of motion, diversity of cell shapes and shape oscillations. The results are in qualitative agreement with recent experiments on motility of keratocyte cells and cell fragments. The asymmetry of the shapes is captured to a large extent in this simple model, which may prove useful for the interpretation of experiments.     

Loss of Vimentin Enhances Cell Motility through Small Confining Spaces

The migration of cells through constricting spaces or along fibrous tracks in tissues is important for many biological processes and depends on the mechanical properties of a cytoskeleton made up of three different filaments: F‐actin, microtubules, and intermediate filaments. The signaling pathways and cytoskeletal structures that control cell motility on 2D are often very different from those that control motility in 3D. Previous studies have shown that intermediate filaments can promote actin‐driven protrusions at the cell edge, but have little effect on overall motility of cells on flat surfaces. They are however important for cells to maintain resistance to repeated compressive stresses that are expected to occur in vivo. Using mouse embryonic fibroblasts derived from wild‐type and vimentin‐null mice, it is found that loss of vimentin increases motility in 3D microchannels even though on flat surfaces it has the opposite effect. Atomic force microscopy and traction force microscopy experiments reveal that vimentin enhances perinuclear cell stiffness while maintaining the same level of acto‐myosin contractility in cells. A minimal model in which a perinuclear vimentin cage constricts along with the nucleus during motility through confining spaces, providing mechanical resistance against large strains that could damage the structural integrity of cells, is proposed.     

Photo-induced molecular alignment of azo dye derivative

Photo-induced molecular alignment behavior of azo dye derivative (sodium 4,4′-bis (4-hydroxy-3-carboxy-phenylazo) benzidine-2,2′-disulphonate: SD1) was investigated using SD1 film prepared by spin-coating method. As-prepared SD1 film was composed of an amorphous layer with smooth surface. Upon linearly polarized UV light irradiation, the film surface was roughened slightly and X-ray diffraction measurement and transmittance electron microscope observation indicated that SD1 molecules crystallized with orientation. Molecular plane of SD1 aligned parallel to the substrate surface along the normal to the polarization direction of irradiated UV light through the trans-cis and cis-trans isomerizations of the azobenzene chromophore.     

Solitary Waves of Maxwell's Equations in Nonlinear Anisotropic Media

Abstract

The (1 + 1)-D solitary wave solutions of Maxwell's equations in nonlinearity induced anisotropic media (in liquids such as carbon disulphide, and in crystals, etc.) are investigated. We find that there is no arbitrarily linearly polarized (in the x-y plane perpendicular to the propagation direction z) soliton solution from Maxwell's equations except that with linear polarization either in alignment with or orthogonal to the geometric axis of the light induced refractive index change. This contradicts the prediction of the vector nonlinear Schroedinger equation (an approximation of Maxwell's equations) which yields soliton solutions with an arbitrary linear polarization. However, Maxwell's equations are found to admit stable elliptically polarized solitary wave solutions which reduce to the stable circularly polarized solitary wave solutions of the vector nonlinear Schroedinger equation when the induced refractive index change approaches zero.     

Label-free magnetic resonance imaging to locate live cells in three-dimensional porous scaffolds

Porous scaffolds are widely tested materials used for various purposes in tissue engineering. A critical feature of a porous scaffold is its ability to allow cell migration and growth on its inner surface. Up to now, there has not been a method to locate live cells deep inside a material, or in an entire structure, using real-time imaging and a non-destructive technique. Herein, we seek to demonstrate the feasibility of the magnetic resonance imaging (MRI) technique as a method to detect and locate in vitro non-labelled live cells in an entire porous material. Our results show that the use of optimized MRI parameters (4.7 T; repetition time = 3000 ms; echo time = 20 ms; resolution 39 × 39 µm) makes it possible to obtain images of the scaffold structure and to locate live non-labelled cells in the entire material, with a signal intensity higher than that obtained in the culture medium. In the current study, cells are visualized and located in different kinds of porous scaffolds. Moreover, further development of this MRI method might be useful in several three-dimensional biomaterial tests such as cell distribution studies, routine qualitative testing methods and in situ monitoring of cells inside scaffolds.     

Conventional Cells—The Last Step Toward General Acceptance of Standard Conventional Cells for the Reporting of Crystallographic Data

In 1969, a seminal section on reduced forms and conventional cells was published in the International Tables for X-Ray Crystallography. The section contains a table that gives a metric classification of the 44 reduced forms. In 2001, this table with appropriate revisions was republished in the Journal of Research of the National Institute of Standards and Technology. An especially valuable feature of the table is that it defines and allows the user to determine a standard conventional cell. Since 1969, there has been an evolution toward acceptance and widespread use of such conventional cells. An inspection of the articles in key crystallographic journals reveals that most cells follow the conventions. However, one major exception remains—the centered monoclinic lattices. In approximately one-third of these cases, non-conventional C-centered cells are used, apparently to avoid the use of I-centered cells. It is recommended that the crystallographic community routinely use the I-centered conventional cell in such cases.     

Graphene Oxide Nanosheets Retard Cellular Migration via Disruption of Actin Cytoskeleton

Graphene and graphene‐based nanomaterials are broadly used for various biomedical applications due to their unique physiochemical properties. However, how graphene‐based nanomaterials interact with biological systems has not been thoroughly studied. This study shows that graphene oxide (GO) nanosheets retard A549 lung carcinoma cell migration through nanosheet‐mediated disruption of intracellular actin filaments. After GO nanosheets treatment, A549 cells display slower migration and the structure of the intracellular actin filaments is dramatically changed. It is found that GO nanosheets are capable of absorbing large amount of actin and changing the secondary structures of actin monomers. Large‐scale all‐atom molecular dynamics simulations further reveal the interactions between GO nanosheets and actin filaments at molecular details. GO nanosheets can insert into the interstrand gap of actin tetramer (helical repeating unit of actin filament) and cause the separation of the tetramer which eventually leads to the disruption of actin filaments. These findings offer a novel mechanism of GO nanosheet induced biophysical responses and provide more insights into their potential for biomedical applications.     

Emergence of polar order and cooperativity in hydrodynamically coupled model cilia

As a model of ciliary beat, we use two-state oscillators that have a defined direction of oscillation and have strong synchronization properties. By allowing the direction of oscillation to vary according to the interaction with the fluid, with a timescale longer than the timescale of synchronization, we show in simulations that several oscillators can align in a direction set by the geometrical configuration of the system. In this system, the alignment depends on the state of synchronization of the system, and is therefore linked to the beat pattern of the model cilia. By testing various configurations from two to 64 oscillators, we deduce empirically that, when the synchronization state of neighbouring oscillators is in phase, the angles of the oscillators align in a configuration of high hydrodynamic coupling. In arrays of oscillators that break the planar symmetry, a global direction of alignment emerges reflecting this polarity. In symmetric configurations, where several directions are geometrically equivalent, the array still displays strong internal cooperative behaviour. It also appears that the shape of the array is more important than the lattice type and orientation in determining the preferred direction.     

Fabrication of a thermoresponsive cell culture dish: a key technology for cell sheet tissue engineering

This article reviews the properties and characterization of an intelligent thermoresponsive surface, which is a key technology for cell sheet-based tissue engineering. Intelligent thermoresponsive surfaces grafted with poly(N-isopropylacrylamide) exhibit hydrophilic/hydrophobic alteration in response to temperature change. Cultured cells are harvested on thermoresponsive cell culture dishes by decreasing the temperature without the use of digestive enzymes or chelating agents. Our group has developed cell sheet-based tissue engineering for therapeutic uses with single layer or multilayered cell sheets, which were recovered from the thermoresponsive cell culture dish. Using surface derivation techniques, we developed a new generation of thermoresponsive cell culture dishes to improve culture conditions. We also designed a new methodology for constructing well-defined organs using microfabrication techniques.