Interactions between mesoderm cells and the extracellular matrix following gastrulation in the chick embryo

1991 ◽  
Vol 99 (2) ◽  
pp. 431-441
Author(s):  
A.J. Brown ◽  
E.J. Sanders
Keyword(s):  
Extracellular Matrix ◽  
Chick Embryo ◽  
Basement Membrane ◽  
Primitive Streak ◽  
Cell Binding ◽  
Fab Fragment ◽  
Fibronectin Receptor ◽  
In Vitro Methods

In the gastrulating chick embryo, the mesoderm cells arise from the epiblast layer by ingression through the linear accumulation of cells called the primitive streak. The mesoderm cells emerge from the streak with a fibroblastic morphology and proceed to move away from the mid-line of the embryo using, as a substratum, the basement membrane of the overlying epiblast and the extracellular matrix. We have investigated the roles of fibronectin and laminin as putative substrata for mesoderm cells using complementary in vivo and in vitro methods. We have microinjected agents into the tissue space adjacent to the primitive streak of living embryos and, after further incubation, we have examined the embryos for perturbation of the mesoderm tissue. These agents were: cell-binding regions from fibronectin (RGDS) and laminin (YIGSR), antibodies to these glycoproteins, and a Fab' fragment of the antibody to fibronectin. We find that RGDS, antibody to fibronectin, and the Fab' fragment cause a decrease in the number of mesoderm cells spread on the basement membrane, and a perturbation of cell shape suggesting locomotory impairment. No such influence was seen with YIGSR or antibodies to laminin. These results were extended using in vitro methods in which mesoderm cells were cultured in fibronectin-free medium on fibronectin or laminin in the presence of various agents. These agents were: RGDS; YIGSR; antibodies to fibronectin, fibronectin receptor, laminin and vitronectin; and a Fab' fragment of the fibronectin antiserum. We find that cell attachment and spreading on fibronectin is impaired by RGDS, antiserum to fibronectin, the Fab' fragment of fibronectin antiserum, and antiserum to fibronectin receptor. The results suggest that although the RGDS site in fibronectin is important, it is probably not the only fibronectin cell-binding site involved in mediating the behaviour of the mesoderm cells. Cells growing on laminin were perturbed by YIGSR, RGDS and antibodies to laminin, suggesting that mesoderm cells are able to recognise at least two sites in the laminin molecule. We conclude that the in vivo dependence of mesoderm cells on fibronectin is confirmed, but that although these cells have the ability to recognise sites in laminin as mediators of attachment and spreading, the in vivo role of this molecule in mesoderm morphogenesis is not yet certain.

scholarly journals Sitagliptin and the Blood-Retina Barrier: Effects on Retinal Endothelial Cells Manifested Only after Prolonged Exposure

2020 ◽  
Vol 2020 ◽  
pp. 1-16
Author(s):  
Anja Jäckle ◽  
Focke Ziemssen ◽  
Eva-Maria Kuhn ◽  
Jürgen Kampmeier ◽  
Gerhard K. Lang ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Endothelial Cells ◽  
Prolonged Exposure ◽  
Diabetic Patients ◽  
Barrier Dysfunction ◽  
Fibronectin Receptor ◽  
Retinal Endothelial Cells ◽  
Junction Protein

Inhibitors of dipeptidyl peptidase-4 (DPP-4) are widely used to treat diabetes mellitus, but data concerning their effects on the barrier stability of retinal endothelial cells (REC) in vivo and in vitro are inconsistent. Therefore, we studied whether the barrier properties of immortalized endothelial cells of the bovine retina (iBREC) were affected by the inhibitors of DPP-4 sitagliptin (10-1000 nM) and diprotin A (1-25 μM). Their effects were also investigated in the presence of VEGF-A165 because diabetic patients often develop macular edema caused by VEGF-A-induced permeability of REC. To detect even transient or subtle changes of paracellular and transcellular flow as well as adhesion of the cells to the extracellular matrix, we continuously monitored the cell index (CI) of confluent iBREC grown on gold electrodes. Initially, the CI remained stable but started to decline significantly and persistently at 40 h or 55 h after addition of sitagliptin or diprotin A, respectively. Both inhibitors did not modulate, prevent, or revert the persistent VEGF-A165-induced reduction of the CI. Interestingly, sitagliptin and diprotin A increased the expression of the tight-junction protein claudin-1 which is an important component of a functional barrier formed by iBREC. In contrast, expressions of CD29—a subunit of the fibronectin receptor—or of the tetraspanin CD9 were lower after extended treatment with the DPP-4 inhibitors; less of the CD9 was seen at the plasma membrane after prolonged exposure to sitagliptin. Because both associated proteins are important for adhesion of iBREC to the extracellular matrix, the observed low CI might be caused by weakened attachment of the cells. From our results, we conclude that extended inhibition of DPP-4 destabilizes the barrier formed by microvascular REC and that DPP-4 inhibitors like sitagliptin do not counteract or enhance a VEGF-A165-induced barrier dysfunction as frequently observed in DME.

Myogel, a Novel, Basement Membrane-Rich, Extracellular Matrix Derived from Skeletal Muscle, Is Highly Adipogenic in vivo and in vitro

2008 ◽  
Vol 188 (4) ◽  
pp. 347-358 ◽  
Author(s):  
K.M. Abberton ◽  
S.K. Bortolotto ◽  
A.A. Woods ◽  
M. Findlay ◽  
W.A. Morrison ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Extracellular Matrix ◽  
Basement Membrane

scholarly journals Invasion of a basement membrane matrix by chick embryo primitive streak cells in vitro

1989 ◽  
Vol 92 (3) ◽  
pp. 497-504 ◽  
Author(s):  
E.J. Sanders ◽  
S. Prasad
Keyword(s):  
Chick Embryo ◽  
Basement Membrane ◽  
Primitive Streak ◽  
Lamina Densa ◽  
Invasive Potential ◽  
Tissue Space ◽  
Epithelial Sheet ◽  
Mesenchymal Transformation

At the time of gastrulation in the chick embryo the upper epiblast layer penetrates its own basement membrane at the primitive streak so that its cells may invade the underlying tissue space. In so forming the primary mesoderm, the cells undergo a concomitant epithelial-to-mesenchymal transformation. In this study, epiblast tissue has been explanted onto a basement membrane gel in order to examine its invasive potential. Fully ingressed primary mesoderm cells were able to penetrate the gel as individual cells, during the course of which the texture of the gel was disrupted. By contrast, epiblast tissue taken from the immediate vicinity of the primitive streak penetrated the gel, but only as a coherent tongue of cells and without gel disruption. These tongues of cells did not undergo the epithelial-to-mesenchymal transformation, and consequently spread as a epithelial sheet when replated on glass. Thus, the absence of gel disruption correlated with the failure of transformation, suggesting that these two events may be linked and that they may require in situ cell interactions for their manifestation. Tissue from the lateral epiblast failed to penetrate the gel. Instead, this tissue either spread on the gel surface or rounded up into a hollow sphere with the basal surface of the cells innermost. In the former case, despite the cell spreading, no lamina densa was organized beneath the sheet, but in the latter case polarity reversal occurred with the formation of a new lamina densa at the cell-gel interface.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Sertoli cell binding to isolated testicular basement membrane.

1986 ◽  
Vol 103 (3) ◽  
pp. 1109-1119 ◽  
Author(s):  
G C Enders ◽  
J H Henson ◽  
C F Millette
Keyword(s):  
Basement Membrane ◽  
Sertoli Cell ◽  
Basal Lamina ◽  
Sertoli Cells ◽  
Matrix Material ◽  
Three Dimensional ◽  
Myoid Cells ◽  
Cell Binding

We have examined the adhesion of primary Sertoli cells to a seminiferous tubule basement membrane (STBM) preparation in vitro. The STBM isolation procedure (Watanabe, T.K., L.J. Hansen, N.K. Reddy, Y.S. Kanwar, and J.K. Reddy, 1984, Cancer Res., 44:5361-5368) yields segments of STBM that retain their histotypic form in both three-dimensional tubular geometry and ultrastructural appearance. The STBM sleeves contain two laminae: a thick, inner basal lamina that was formed in vivo between Sertoli cells and peritubular myoid cells; and a thinner, outer basal lamina that was formed between myoid cells and sinusoidal endothelial cells. Characterization by immunofluorescence and SDS PAGE revealed that the isolated STBM retained fibronectin, laminin, and putative type IV collagen among its many components. When the STBM sleeves were gently shaken with an enriched fraction of primary Sertoli cells, the Sertoli cells bound preferentially to the lumenal basal lamina at the ends of the STBM sleeves. Few Sertoli cells bound to either the outer basal lamina of the STBM sleeves or to vascular extracellular matrix material which contaminated the STBM preparation. 3T3 cells, in contrast, bound to all surfaces of the STBM sleeves. Pretreatment of the STBM sleeves with proteases, 0.1 M Na metaperiodate, 4 M guanidine HCl, or heating to 80 degrees-90 degrees C inhibited lumenal Sertoli cell binding, but binding was not inhibited by chondroitinase ABC, heparinase, hyaluronidase, or 4 M NaCl. The lumenal Sertoli cell binding occurred in the presence or absence of added soluble laminin, but not fibronectin. The addition of soluble laminin, but not fibronectin, restored random binding of Sertoli cells to trypsinized STBM sleeves. Our in vitro model system indicates that Sertoli cells recognize differences in two basal laminae produced in vivo on either side of myoid cells.

scholarly journals Cell based strain stiffening of a non-fibrous matrix as organizing principle for morphogenesis

2019 ◽  
Author(s):  
Daniel Rüdiger ◽  
Kerstin Kick ◽  
Andriy Goychuk ◽  
Angelika M. Vollmar ◽  
Erwin Frey ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Pattern Formation ◽  
Basement Membrane ◽  
Self Assembly ◽  
Matrix Material ◽  
Cell Contacts ◽  
Cell Contractility ◽  
Cell Cell

AbstractEndothelial tube formation on a reconstituted extracellular matrix (Matrigel) is a well-established in vitro model for studying the processes of angiogenesis and vasculogenesis. However, to date, the organizing principles that underlie the morphogenesis of this network, and that shape the initial process of cell-cell finding remain elusive. Furthermore, it is unclear how in vitro results extrapolate to in vivo morphogenesis. Here, we identify a mechanism that allows cells to form networks by mechanically reorganizing and stiffening their extracellular matrix, independent of chemical guidance cues. Interestingly, we find that this cellular self-organization strongly depends on the connectivity and topology of the surrounding matrix, as well as on cell contractility and cell density. Cells rearrange the matrix, and form bridges of matrix material that are stiffer than their surroundings, thus creating a durotactic track for the initiation of cell-cell contacts. This contractility-based communication via strain stiffening and matrix rearrangement might be a general organizing principle during tissue development or regeneration.Significance StatementIn addition to chemotactic gradients, biomechanical cues are important for guiding biological pattern formation. Self-assembly of cells has often been ascribed to reorganization of collagen fibres in the extracellular matrix. However, the basement membrane surrounding vascular cells, is per se non-fibrous. Here, we find that this difference in matrix topology can crucially influence cell behaviour and pattern formation. In a homogeneously elastic environment like the basement membrane, endothelial cells rearrange extracellular matrix proteins by contractile force, forming stiff intercellular bridges as tracks for cell-cell contacts. Our findings shine some light why there is a lot of merit in having multiple approaches to matrix elasticity (like continuum theories or dilated network approaches). Our observations might help to understand why vascular nets look different in different tissues and after rearrangement of the extracellular matrix during disease.

Extracellular matrix modulation of endothelial cell shape and motility following injury in vitro

1985 ◽  
Vol 73 (1) ◽  
pp. 19-32
Author(s):  
W.C. Young ◽  
I.M. Herman
Keyword(s):  
Extracellular Matrix ◽  
Basement Membrane ◽  
Fluorescence Microscopy ◽  
Type Iv Collagen ◽  
Type I ◽  
Post Injury ◽  
Type Iv ◽  
Interstitial Collagens

We utilized fluorescence microscopy and affinity-purified antibodies to probe the form and function of cytoplasmic actin in endothelial cells (EC) recovering from injury and grown on extracellular matrices in vitro. Bovine aortic EC were seeded onto glass microscope coverslips that had been coated with either BSA, fibronectin, type I and III (interstitial) collagens, type IV (basement membrane) collagen or gelatin. After EC that had been grown on glass, glass-BSA or extracellular matrix-coated coverslips reached confluence, a 300–400 micron zone of cells was mechanically removed to stimulate EC migration and proliferation. Post-injury EC movements were monitored with time-lapse, phase-contrast videomicrography before fixation for actin localization with fluorescence microscopy using affinity-purified antibodies. We found that the number of stress fibres within EC was inversely proportional to the rate of movement; and, the rates of movement for EC grown on glass or glass-BSA were approximately eight times faster than EC grown on gelatin or type IV collagen (X velocity = 0.5 micron/min versus 0.06 micron/min). EC movements on fibronectin and interstitial collagens were similar (X velocity = 0.2 micron/min). These results suggest that extracellular matrix molecules modulate EC stress fibre expression, thereby producing alterations in the cytoskeleton and the resultant EC movements that follow injury in vitro. Moreover, the induction of stress fibres in the presence of basement membrane (type IV) collagen may explain the failure of aortic EC to migrate and repopulate wounded regions of intima during atherogenesis in vivo.

Cytokeratin expression in rat mesothelial cells in vitro is controlled by the extracellular matrix

1990 ◽  
Vol 95 (1) ◽  
pp. 97-107
Author(s):  
A.M. Mackay ◽  
R.P. Tracy ◽  
J.E. Craighead
Keyword(s):  
Extracellular Matrix ◽  
Basement Membrane ◽  
Intermediate Filament ◽  
Culture Conditions ◽  
Mesothelial Cells ◽  
Intermediate Filament Protein ◽  
Cellular Contact ◽  
Filament Protein

Rat mesothelial cells co-express vimentin and the simple epithelial cytokeratins. While cytokeratins predominate in situ, under most culture conditions vimentin is the major intermediate filament protein of the cells. This loss of cytokeratin production upon culture can be partly prevented by growing mesothelial cells on a basement membrane matrix. However, the basement membrane-promoted persistence of cytokeratin synthesis is not accompanied by expression of cytokeratin G (no. 19), the major acidic cytokeratin of mesothelium in vivo. While cells grown on plastic establish a prominent juxtanuclear assemblage of tonofilaments, those cultured on basement membrane exhibit cytokeratin filaments which are distributed throughout the cytoplasm and attach to neighboring cells at the plasma membrane. This latter pattern resembles that seen in the intact mesothelium. Intermediate filaments are markers of cellular differentiation, but their roles are obscure. The response of cultured mesothelial cells to different growth substrata supports the hypothesis that intermediate filament synthesis is influenced by cellular contact with the extracellular matrix.

scholarly journals Extracellular matrix components of the mouse thymus microenvironment: ontogenetic studies and modulation by glucocorticoid hormones.

1991 ◽  
Vol 39 (11) ◽  
pp. 1539-1546 ◽  
Author(s):  
J Lannes-Vieira ◽  
M Dardenne ◽  
W Savino
Keyword(s):  
Extracellular Matrix ◽  
Basement Membrane ◽  
Type I Collagen ◽  
Basement Membranes ◽  
Thymic Epithelial Cell ◽  
Epithelial Cell Line ◽  
Type I ◽  
Extracellular Matrix Components

The present investigation was an ontogenetic study on the distribution of extracellular matrix (ECM) components in the thymic microenvironment of C57BL/6 mice (comprising young and old adults and developing embryos) and NZB mice. In addition, we evaluated the in vivo and in vitro influence of hydrocortisone treatment on basement membrane protein production by a thymic epithelial cell line. In young normal animals, Type I collagen was restricted to the interstitial spaces of the capsule and septa, where Type IV collagen, fibronectin, and laminin could be detected in the basement membranes. In addition, fibronectin-containing fibers were seen within the medulla of the thymic lobules. The ECM distribution pattern in the developing embryos was distinct from that observed in adults, since a fine meshwork of basement membrane-containing proteins was clearly seen throughout the parenchyma. Moreover, aging normal and NZB mice exhibited a denser ECM pattern than young adult normal animals. Treatment with hydrocortisone, both in vivo and in vitro, resulted in enhancement of ECM expression, detected in mice as early as 2 hr post injection and lasting for several days. Considering that the fluctuations of ECM expression parallel important events in thymocyte differentiation, we discuss the possibility that the two phenomena may be associated.

scholarly journals Traversing the basement membrane in vivo: A diversity of strategies

2014 ◽  
Vol 204 (3) ◽  
pp. 291-302 ◽  
Author(s):  
Laura C. Kelley ◽  
Lauren L. Lohmer ◽  
Elliott J. Hagedorn ◽  
David R. Sherwood
Keyword(s):  
Extracellular Matrix ◽  
Basement Membrane ◽  
Metastatic Disease ◽  
Tissue Level ◽  
In Vitro Studies ◽  
In Vivo Studies ◽  
Leukocyte Trafficking ◽  
Diverse Range

The basement membrane is a dense, highly cross-linked, sheet-like extracellular matrix that underlies all epithelia and endothelia in multicellular animals. During development, leukocyte trafficking, and metastatic disease, cells cross the basement membrane to disperse and enter new tissues. Based largely on in vitro studies, cells have been thought to use proteases to dissolve and traverse this formidable obstacle. Surprisingly, recent in vivo studies have uncovered a remarkably diverse range of cellular- and tissue-level strategies beyond proteolysis that cells use to navigate through the basement membrane. These fascinating and unexpected mechanisms have increased our understanding of how cells cross this matrix barrier in physiological and disease settings.

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