Role of the extracellular matrix during neural crest cell migration

Once specified to become neural crest (NC), cells occupying the dorsal portion of the neural tube disrupt their cadherin-mediated cell-cell contacts, acquire motile properties, and embark upon an extensive migration through the embryo to reach their ultimate phenotype-specific sites. The understanding of how this movement is regulated is still rather fragmentary due to the complexity of the cellular and molecular interactions involved. An additional intricate aspect of the regulation of NC cell movement is that the timings, modes and patterns of NC cell migration are intimately associated with the concomitant phenotypic diversification that cells undergo during their migratory phase and the fact that these changes modulate the way that moving cells interact with their microenvironment. To date, two interplaying mechanisms appear central for the guidance of the migrating NC cells through the embryo: one involves secreted signalling molecules acting through their cognate protein kinase/phosphatase-type receptors and the other is contributed by the multivalent interactions of the cells with their surrounding extracellular matrix (ECM). The latter ones seem fundamental in light of the central morphogenetic role played by the intracellular signals transduced through the cytoskeleton upon integrin ligation, and the convergence of these signalling cascades with those triggered by cadherins, survival/growth factor receptors, gap junctional communications, and stretch-activated calcium channels. The elucidation of the importance of the ECM during NC cell movement is presently favoured by the augmenting knowledge about the macromolecular structure of the specific ECM assembled during NC development and the functional assaying of its individual constituents via molecular and genetic manipulations. Collectively, these data propose that NC cell migration may be governed by time- and space-dependent alterations in the expression of inhibitory ECM components; the relative ratio of permissive versus non-permissive ECM components; and the supramolecular assembly of permissive ECM components. Six multidomain ECM constituents encoded by a corresponding number of genes appear to date the master ECM molecules in the control of NC cell movement. These are fibronectin, laminin isoforms 1 and 8, aggrecan, and PG-M/version isoforms V0 and V1. This review revisits a number of original observations in amphibian and avian embryos and discusses them in light of more recent experimental data to explain how the interaction of moving NC cells with these ECM components may be coordinated to guide cells toward their final sites during the process of organogenesis.     

Fibronectin-rich fibrillar extracellular matrix controls cell migration during amphibian gastrulation

We have reviewed the evidence supporting the notion that the fibrillar extracellular matrix on the basal surface of the blastocoel roof in amphibian embryos directs and guides mesodermal cell migration during gastrulation. Based on extensive experimental evidence in several different systems, we conclude the following: (i) the fibrillar extracellular matrix contains fibronectin (FN) and laminin. (ii) The fibrils are oriented in such a way as to promote directional migration of mesodermal cells during migration. (iii) We have used several different probes to disrupt the interaction between migrating mesodermal cells and the fibrillar extracellular matrix. These probes include: (a) nucleocytoplasmic and interspecific hybridization. Such embryos have defects in FN synthesis and gastrulation. (b) Fab' fragments of anti-FN and anti-integrin VLA-5 IgGs prohibit mesodermal cell adhesion both in vitro and in vivo and gastrulation is arrested. (c) Peptides containing the RGDS sequence specifically inhibit interactions between migrating mesodermal cells and the FN-fibrillar matrix. (d) Tenascin blocks cell adhesion to FN in vitro and gastrulation in vivo. (e) Antibodies against the cytoplasmic domain of beta 1 integrin, when injected into blastomeres, prevent FN-fibrillogenesis in progeny of injected blastomeres and delay mesodermal cell migration selectively in the progeny of injected blastomeres but not in the uninjected blastomere progeny.     

Role of the extracellular matrix in lymphocyte migration

The extracellular matrix (ECM) exists in various biochemical and structural forms that can act either as a barrier to migrating leukocytes, in the case of basement membranes, or provide a physical scaffold supporting or guiding migration (interstitial matrix). This review focuses on basement membranes and our current knowledge of the way that leukocytes transmigrate this protein barrier, with emphasis on T lymphocytes. Recent data suggest that the classical concept of cell-matrix adhesion requires revision with respect to leukocyte-ECM interactions. Whereas specific receptors may be required for leukocyte recognition of ECM molecules or three-dimensional structural domains, the role of adhesion in migration as perceived from the traditional studies of adherent cell-ECM interactions is less clear. Further, the indirect effects of ECM such as the binding and presentation of cytokines or chemotactic factors may more profoundly influence the directed migration of normally non-adherent leukocytes than the migration of adherent cells such as epithelial cells or fibroblasts. Proteases (in particular matrix metalloproteinases) released at sites of inflammation can selectively process ECM, cell surface molecules or soluble factors, which may result in the release of bioactive fragments that can function as chemoattractants for different leukocyte subsets or may modulate the activity/function of resident mesenchymal and immune cells. Current findings suggest that different leukocyte types employ different mechanisms to migrate across or through the ECM; this might be determined by the composition and organization of the ECM itself.     

Neural crest cell migration in relation to extracellular matrix organization in the embryonic axolotl trunk

Temporal and regional aspects of early neural crest cell migration in relation to extracellular matrix (ECM) organization and distribution in the embryonic axolotl trunk were studied by light microscopy, TEM, and SEM. The dominating structure of the interstitial ECM is a complex network of fibrils, which are indicated by ruthenium red staining to consist of collagen in association with ruthenium red-positive components, probably including glycosaminoglycans. The ECM fibrils, which are largely used as substratum for locomotion by the crest cells, have a temporally and regionally specific organization and distribution. Increase in ECM fibrils on the neural tube, ahead of the crest cell front, is correlated with initiation of crest cell emigration, and it is suggested that the fibrils may stimulate this process by providing a suitable substratum for cell locomotion. An increase in ECM fibrils in extracellular spaces surrounding the crest cell population is correlated with an expansion of these spaces and with progressing crest cell migration into them. It is proposed that the spatial organization of the ECM fibrils influences crest cell shape and orientation during early migration.     

Mouse mutants provide new insights into the role of extracellular matrix in cell migration and differentiation

Migratory cell populations in the developing embryo disperse, localize and eventually differentiate in environments rich in extracellular matrix material. The extracellular matrix provides both a substratum for migration and a source of differentiative cues for the developing cells. Mutations in mice and other animals that alter embryonic interstitial environments are now providing information about the role of the extracellular matrix in these early developmental processes.     

Bves and NDRG4 regulate directional epicardial cell migration through autocrine extracellular matrix deposition

Directional cell movement is universally required for tissue morphogenesis. Although it is known that cell/matrix interactions are essential for directional movement in heart development, the mechanisms governing these interactions require elucidation. Here we demonstrate that a novel protein/protein interaction between blood vessel epicardial substance (Bves) and N-myc downstream regulated gene 4 (NDRG4) is critical for regulation of epicardial cell directional movement, as disruption of this interaction randomizes migratory patterns. Our studies show that Bves/NDRG4 interaction is required for trafficking of internalized fibronectin through the “autocrine extracellular matrix (ECM) deposition” fibronectin recycling pathway. Of importance, we demonstrate that Bves/NDRG4-mediated fibronectin recycling is indeed essential for epicardial cell directional movement, thus linking these two cell processes. Finally, total internal reflectance fluorescence microscopy shows that Bves/NDRG4 interaction is required for fusion of recycling endosomes with the basal cell surface, providing a molecular mechanism of motility substrate delivery that regulates cell directional movement. This is the first evidence of a molecular function for Bves and NDRG4 proteins within broader subcellular trafficking paradigms. These data identify novel regulators of a critical vesicle-docking step required for autocrine ECM deposition and explain how Bves facilitates cell-microenvironment interactions in the regulation of epicardial cell–directed movement.     

Glycosaminoglycans modulate C6 glioma cell adhesion to extracellular matrix components and alter cell proliferation and cell migration

Background  

Adhesion to extracellular matrix (ECM) components has been implicated in the proliferative and invasive properties of tumor cells. We investigated the ability of C6 glioma cells to attach to ECM components in vitro and described the regulatory role of glycosaminoglycans (GAGs) on their adhesion to the substrate, proliferation and migration.     

Pigment cell pattern formation in Taricha torosa: The role of the extracellular matrix in controlling pigment cell migration and differentiation

The neural crest is a population of highly migratory mesenchymal cells that ultimately localize in specific sites and differentiate into a variety of cell types. This report describes studies on the factors governing the migratory pathways, differentiation, and ultimate localization of the neural crest-derived pigment cells (black melanophores and yellow xanthophores) in the California newt, Taricha torosa. Melanophores first appear scattered in the dorsal portion of the lateral neural crest migratory pathway (between the somites and the ectoderm). These cells are eventually found in two stripes: a dorsal stripe that runs along the apex of the somites, and a midbody stripe near the somite-lateral plate mesoderm border. Melanophores are not seen in the dorsal fin of prehatching embryos. Xanthophores can be identified with the light microscope using NH₄OH-induced autofluorescence of pteridines and in the transmission electron microscope (TEM) by the presence of pterinosomes. Xanthophores first appear scattered among the melanophores over the surface of the somites; these cells eventually are found between the two melanophore stripes and in the dorsal fin. We were interested in determining the roles of the extracellular matrix (ECM) in controlling the formation of pigment cell patterns in T. torosa. Immunocytochemistry, Alcian blue staining of paraffin sections and ruthenium red staining of thin sections (accompanied by Streptomyces hyaluronidase and chondroitinase ABC digestion) were used to identify the composition and distribution of the ECM surrounding the pigment cells at various stages during development. The adhesive glycoprotein fibronectin is found in the dorsal portion of the lateral neural crest migratory pathway as well as in the dorsal fin matrix. Glycosaminoglycans (GAG) are found primarily in the dorsal fin and in the ECM surrounding the notochord. The dorsal fin ECM contains hyaluronate (HA), which was identified in the TEM as Streptomyces hyaluronidase-sensitive 3–5 nm microfibrils, as well as sulfated proteoglycan aggregates. We then confronted T. torosa neural crest cells in vitro with known ECM molecules. When neural folds are explanted onto tissue culture plastic in half-strength L-15 medium containing 10% fetal calf serum (FCS), cells migrate from the explant and differentiate into melanophores after 6 to 9 days. Xanthophores appear in the cultures 2 to 4 days after the appearance of melanophores. When cultured on three-dimensional collagen gels, xanthophores migrate significantly farther (P < 0.01) onto and into the collagen than melanophores (336 ± 183 vs 196 ± 160 μm from the edge of the explant). When 2.5 mg/ml chondroitin sulfate (CS) is present in the collagen gel, the distance that both pigment cell types migrate from the explant is reduced, with the result being that only xanthophores invade the GAG-rich matrix. When 1 mg/ml HA is present in the collagen gel, the differentiation of pigment cells is inhibited. Melanophores appear 48 hr later than in control gels without HA, and the number of melanophores in the explant after 10 days is significantly reduced (P < 0.01; 26.6 vs 1.1 melanophores/explant). When 1 mg/ml of HA is added to the FCS-enriched medium over neural crest cells spreading on tissue culture plastic, there is a similar delay and inhibition of pigment cell differentiation. With 2 mg/ml of CS there is no effect on pigment cell differentiation in vitro. Melanophores eventually appear in the dorsal fin of T. torosa several weeks after hatching. When fragments of dorsal fin that contain no apparent melanophores are transferred onto tissue culture plastic, melanophores appear in the explants after a few days in culture. These results suggest the following model of ECM-cell interactions during pigment cell pattern formation in T. torosa: Pigment cells differentiate in regions of the embryo that contain relatively little GAG. Xanthophores are able to invade the GAG-rich dorsal fin, but melanophores can not. The melanophores that eventually appear in the dorsal fin are derived from the neural crest cells that invaded the fin during early development, and were delayed in differentiating by the presence of HA.     

Timing in the regulation of neural crest cell migration: retarded "maturation" of regional extracellular matrix inhibits pigment cell migration in embryos of the white axolotl mutant

In larvae of the white axolotl mutant (Ambystoma mexicanum), contrary to normal dark ones, trunk pigmentation is restricted because the epidermis is unable to support subepidermal migration of pigment cells from the neural crest (NC). This study examines whether the subepidermal extracellular matrix (ECM) is the defective component which prevents pigment cell migration in the white embryo. We transplanted subepidermal ECM, adsorbed in vivo on membrane microcarriers, from and to white and dark embryos in various combinations. White embryos have demonstrated normal NC cell migration along the medioventral pathway, and in order to test the effects of medial ECM on subepidermal migration, this ECM was similarly transplanted. Carriers with ECM attached were inserted subepidermally in host embryos at a premigratory NC stage. Control carriers without ECM and carriers with subepidermal ECM from white donors did not affect NC cell migration in white or dark embryos. In contrast, subepidermal ECM from dark donors triggered NC cell migration in the subepidermal space of both white and dark hosts. Remarkably, subepidermal ECM from white donors which were older than those normally used also stimulated migration in embryos of both strains. Likewise, medial ECM from white donors elicited migration in white as well as dark hosts. Pigment cells occurred among those NC cells that were stimulated to migrate in response to contact with ECM on carriers. These results indicate that the subepidermal ECM of the white embryo is transiently defective as a substrate for pigment cell migration, implying that "maturation" of the ECM is retarded beyond the times during which pigment cells are able to respond. In contrast, the medial ECM of the white embryo appears to mature normally. These findings suggest that the effect of the d gene is expressed regionally through the subepidermal ECM during a limited period of development. Hence, the action of the d gene seems to retard ECM maturation, bringing it out of phase with the migratory capability of the pigment cells. We propose that such a shift in relative timing of the developmental phenomena involved inhibits pigment cell migration in embryos of the white axolotl mutant and, accordingly, that the restricted pigmentation of the mutant larva is generated through heterochrony.     

Repetitive proline-rich proteins in the extracellular matrix of the plant cell

Several classes of hydroxyproline-rich proteins have been found in the cell walls of plants. Monomeric forms of the proteins can be solubilized from the walls, but the majority of the proteins are insolubilized by as yet unidentified crosslinks. The proteins have repeated sequences and are often rich in basic amino acids. The repeat proline-rich region may be serving to generate an elongated rod-like structure while the regularly spaced lysines probably interact with the acidic pectins. Several of the genes coding for these proteins have been isolated, and their expression has been found in some cases to be tissue specific and in others inducible by hormones and various types of stress.     

The extracellular matrix: a new turn-of-the-screw for anti-angiogenic strategies

Anti-angiogenic therapy is currently one of most active fields in cancer research. The initial strategies, which were aimed at inhibiting tumor vascularization, included upregulation of endogenous inhibitors and blocking of the signals delivered by angiogenic factors. However, interactions between endothelial cells and their surrounding extracellular matrix also play a crucial role in modulation of the angiogenic process. Compounds that target either the integrins implicated in these interactions or the proteases responsible for matrix remodeling have been shown to halt tumor growth in murine models and are now in clinical trials. However, little attention has been paid to integrin ligands, the extracellular matrix components that support endothelial cell survival, movement and reorganization. Here, we summarize the current knowledge about these angiogenesis inhibitors and propose a novel therapeutic approach based on the blocking of crucial binding sites present in the extracellular matrix.     

Diffusion and reaction in the cell glycocalyx and the extracellular matrix

Many biologically important macromolecular reactions are assembled and catalyzed at the cell lipid-surface and thus, the extracellular matrix and the glycocalyx layer mediate transfer and exchange of reactants and products between the flowing blood and the catalytic lipid-surface. This paper presents a mathematical model of reaction–diffusion equations that simply describes the transfer process and explores its influence on surface reactivity for a prototypical pathway, the tissue factor (Tf) pathway of blood coagulation. The progressively increasing friction offered by the matrix and glycocalyx to reactants and to the product (coagulation factors X, VIIa and Xa) approaching the reactive surface is simulated and tested by solving the equations numerically with both, monotonically decreasing and constant diffusion profiles. Numerical results show that compared to isotropic transfer media, the anisotropic structure of the matrix and glycocalyx sharply decreases overall reaction rates and significantly increases the mean transit time of reactants; this implies that the anisotropy modifies the distribution of reactants. Results also show that the diffusional transfer, whether isotropic or anisotropic, influences reaction rates according to the order at which the reactants arrive at the boundary. Faster rates are observed when at least one of the reactants is homogeneously distributed before the other arrives at the boundary than when both reactants transfer simultaneously from the boundary.     

Stimulation of initial neural crest cell migration in the axolotl embryo by tissue grafts and extracellular matrix transplanted on microcarriers

The present experiments were designed to test whether the onset of neural crest cell migration in the embryonic axolotl trunk is stimulated by surrounding tissues and their associated extracellular matrix (ECM). Tissue grafts, or embryonic ECM adsorbed in vivo onto inert "microcarriers" prepared from Nuclepore filters, were placed close to the premigratory neural crest cells, and the embryos were then incubated to a specific stage. The experiments were evaluated with light microscopy, SEM, and TEM. It was found that grafts from the dorsal epidermis were especially effective in locally stimulating initial neural crest cell migration in the region under the graft. The microcarrier experiments showed that the subepidermal ECM alone could initiate neural crest cell migration, implying that the ECM of the epidermal grafts was the stimulating factor. These results indicate that the premigratory neural crest cells along the trunk have migratory capability but that they need to be triggered from the environment, probably from the surrounding ECM, to start migration. It is proposed that ECM, as substrate for cell locomotion, initiates and regulates the onset of neural crest cell migration.     

LKB1 loss in melanoma disrupts directional migration toward extracellular matrix cues

Somatic inactivation of the serine/threonine kinase gene STK11/LKB1/PAR-4 occurs in a variety of cancers, including ∼10% of melanoma. However, how the loss of LKB1 activity facilitates melanoma invasion and metastasis remains poorly understood. In LKB1-null cells derived from an autochthonous murine model of melanoma with activated Kras and Lkb1 loss and matched reconstituted controls, we have investigated the mechanism by which LKB1 loss increases melanoma invasive motility. Using a microfluidic gradient chamber system and time-lapse microscopy, in this paper, we uncover a new function for LKB1 as a directional migration sensor of gradients of extracellular matrix (haptotaxis) but not soluble growth factor cues (chemotaxis). Systematic perturbation of known LKB1 effectors demonstrated that this response does not require canonical adenosine monophosphate–activated protein kinase (AMPK) activity but instead requires the activity of the AMPK-related microtubule affinity-regulating kinase (MARK)/PAR-1 family kinases. Inhibition of the LKB1–MARK pathway facilitated invasive motility, suggesting that loss of the ability to sense inhibitory matrix cues may promote melanoma invasion.     

Modelling actin polymerization: the effect on confined cell migration

The aim of this work is to model cell motility under conditions of mechanical confinement. This cell migration mode may occur in extravasation of tumour and neutrophil-like cells. Cell migration is the result of the complex action of different forces exerted by the interplay between myosin contractility forces and actin processes. Here, we propose and implement a finite element model of the confined migration of a single cell. In this model, we consider the effects of actin and myosin in cell motility. Both filament and globular actin are modelled. We model the cell considering cytoplasm and nucleus with different mechanical properties. The migration speed in the simulation is around 0.1 μm/min, which is in agreement with existing literature. From our simulation, we observe that the nucleus size has an important role in cell migration inside the channel. In the simulation the cell moves further when the nucleus is smaller. However, this speed is less sensitive to nucleus stiffness. The results show that the cell displacement is lower when the nucleus is stiffer. The degree of adhesion between the channel walls and the cell is also very important in confined migration. We observe an increment of cell velocity when the friction coefficient is higher.

     

3D mesenchymal cell migration is driven by anterior cellular contraction that generates an extracellular matrix prestrain

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Transchromosomic cell model of Down syndrome shows aberrant migration, adhesion and proteome response to extracellular matrix

Background  

Down syndrome (DS), caused by trisomy of human chromosome 21 (HSA21), is the most common genetic birth defect. Congenital heart defects (CHD) are seen in 40% of DS children, and >50% of all atrioventricular canal defects in infancy are caused by trisomy 21, but the causative genes remain unknown.     

Modelling of endothelial cell migration and angiogenesis in microfluidic cell culture systems

Biomechanics and Modeling in Mechanobiology - Tumour-induced angiogenesis is a complex biological process that involves growth of new blood vessels within the tumour microenvironment and is an...