Cell adhesion and proteoglycans. I. The effect of exogenous proteoglycans on the attachment of chick embryo fibroblasts to tissue culture plastic and collagen

1979 ◽  
Vol 40 (1) ◽  
pp. 77-88
Author(s):  
P. Knox ◽  
P. Wells
Keyword(s):  
Tissue Culture ◽  
Hyaluronic Acid ◽  
Cell Adhesion ◽  
Chick Embryo ◽  
Structural Integrity ◽  
Chondroitin Sulphate ◽  
Cell Types ◽  
Tissue Culture Plastic ◽  
Keratan Sulphate ◽  
Covalently Linked

Proteoglycan was isolated from cartilage and freed from contaminating glycoproteins and hyaluronic acid. The macromolecule consists of a protein core covalently linked to a number of glycosaminoglycan side chains, namely chondroitin sulphate and keratan sulphate. This proteoglycan retards the attachment of a variety of cell types to tissue culture plastic and to collagen. Glycosaminoglycans alone, have no significant effect on rates of attachment. Similarly, trypsinized proteoglycan is without effect. It is concluded that the structural integrity of the proteoglycan macromolecule is essential for its effect on cell adhesion.

Healing of Circular Defects in the Rabbit Medial Meniscus Can Occur Spontaneously and Is not Improved by Intra-Articular Hyaluronic Acid

1996 ◽  
Vol 09 (02) ◽  
pp. 60-5 ◽  
Author(s):  
N. Hope ◽  
P. Ghosh ◽  
S. Collier
Keyword(s):  
Hyaluronic Acid ◽  
Medial Meniscus ◽  
Chondroitin Sulphate ◽  
Rabbit Model ◽  
Type Ii Collagen ◽  
Type Ii ◽  
Keratan Sulphate ◽  
Meniscal Healing ◽  
Meniscus Healing ◽  
Made In

SummaryThe aim of this study was to determine the effects of intra-articular hyaluronic acid on meniscal healing. Circular defects, 1.0 mm in diameter, were made in the anterior third of the medial meniscus in rabbits. In one joint, 0.4 ml hyaluronic acid (Healon®) was instilled, and in the contralateral (control) joint, 0.4 ml Ringer’s saline. Four rabbits were killed after four, eight and 12 weeks and the menisci examined histologically. By eight weeks most of the lesions had healed by filling with hyaline-like cartilage. Healing was not improved by hyaluronic acid treatment. The repair tissue stained strongly with alcian blue, and the presence of type II collagen, keratan sulphate, and chondroitin sulphate was demonstrated by immunohistochemical localisation. In contrast to the circular defects, longitudinal incisions made in the medial menisci of a further six rabbits did not show any healing after 12 weeks, indicating that the shape of the lesion largely determined the potential for healing.The effect of hyaluronic acid on meniscal healing was tested in a rabbit model. With one millimeter circular lesions in the medial meniscus, healing by filling with hyalinelike cartilage was not significantly affected by the application of hyaluronic acid intra-articularly at the time of surgery, compared to saline controls, as assessed histologically four, eight and 12 weeks after the operation.

The Uptake and Localization of Catecholamines in Chick Embryo Sympathetic Neurons in Tissue Culture

1969 ◽  
Vol 4 (3) ◽  
pp. 677-691
Author(s):  
J. M. ENGLAND ◽  
M. N. GOLDSTEIN
Keyword(s):  
Tissue Culture ◽  
Chick Embryo ◽  
Ganglion Cells ◽  
Sympathetic Neurons ◽  
Neuronal Cell ◽  
Cell Types ◽  
Axon Terminals ◽  
Dense Core Granules ◽  
Insoluble Compounds ◽  
Dorsal Root Ganglion Cells

The uptake of exogenous [3H]dopamine, [3H]norepinephrine,[3H]epinephrine by dissociated chick embryo sympathetic neurons growing in tissue culture was studied by autoradiography. The neurons, growing in a medium containing nerve growth factor, rapidly and specifically took up all three catecholamines for at least 60 days, while no uptake was observed in several other cell types, including satellite cells and chick dorsal-root ganglion cells. The uptake was dependent on the concentration of the catecholamine and the duration of the pulse and was inhibited by cocaine and several sympathomimetic amines. Labelling was visualized only with fixatives which react with catecholamines to form water-insoluble compounds. Autoradiographs showed that the label was much denser over the axons than the cell bodies. The label was distributed uniformly along the axons and did not seem to be preferentially localized at the axon terminals or varicosities which contain aggregates of dense core granules. These observations indicate that a large portion of the exogenous 3[H]catecholamine is localized in an extragranular compartment and suggest that the differentiated function of the sympathetic neuronal cell membrane, which plays an important role in uptake, is retained after prolonged tissue culture.

scholarly journals Studies on protein–polysaccharides from pig laryngeal cartilage. Extraction and purification

1969 ◽  
Vol 113 (5) ◽  
pp. 879-884 ◽  
Author(s):  
C. P. Tsiganos ◽  
Helen Muir
Keyword(s):  
Hyaluronic Acid ◽  
Chloride Solution ◽  
Sodium Acetate ◽  
Serum Proteins ◽  
Chondroitin Sulphate ◽  
Calcium Chloride Solution ◽  
Keratan Sulphate ◽  
Laryngeal Cartilage ◽  
Extraction And Purification ◽  
Degradative Enzymes

1. Protein–polysaccharides of chondroitin sulphate were extracted from fresh laryngeal cartilage at pH6·8 by two procedures. Procedure I consisted of brief low-speed homogenization in 0·15m (iso-osmotic) sodium acetate and procedure II consisted of longer homogenization followed by prolonged extraction in 10% calcium chloride solution. 2. The protein–polysaccharides in both extracts were isolated and purified by precipitation with 9-aminoacridine hydrochloride. They were free from serum proteins, collagen and nucleic acids and also of degradative enzymes. The absence of such enzymes was shown by viscosity measurements on solutions of protein–polysaccharides incubated for up to 24hr. at pH4 and 6·8. 3. Mannose, glucose or fucose were not detected by paper chromatography and only traces of sialic acid were present. 4. The yield with procedure II was twice that with procedure I and the products differed in their protein and glucosamine contents. 5. Hyaluronic acid was unlikely to have been precipitated at an acid pH, so the glucosamine was attributed to keratan sulphate, as serum proteins were absent. There was no free keratan sulphate in the preparation. 6. Both preparations were heterogeneous in the ultracentrifuge, showing at least three components.

scholarly journals A chondroitin sulphate proteoglycan from human cultured glial and glioma cells. Structural and functional properties

1984 ◽  
Vol 221 (3) ◽  
pp. 845-853 ◽  
Author(s):  
B Norling ◽  
B Glimelius ◽  
A Wasteson
Keyword(s):  
Hyaluronic Acid ◽  
Chondroitin Sulphate ◽  
Buoyant Density ◽  
Glioma Cells ◽  
Keratan Sulphate ◽  
Link Protein ◽  
Sulphate Proteoglycan ◽  
Chondroitin Sulphate Proteoglycan ◽  
Chondroitin Sulphate Proteoglycans ◽  
Structural And Functional Properties

A chondroitin sulphate proteoglycan capable of forming large aggregates with hyaluronic acid was identified in cultures of human glial and glioma cells. The glial- cell- and glioma-cell-derived products were mutually indistinguishable and had some basic properties in common with the analogous chondroitin sulphate proteoglycan of cartilage: hydrodynamic size, dependence on a minimal size of hyaluronic acid for recognition, stabilization of aggregates by link protein, and precipitability with antibodies raised against bovine cartilage chondroitin sulphate proteoglycan. However, they differed in some aspects: lower buoyant density, larger, but fewer, chondroitin sulphate side chains, presence of iduronic acid-containing repeating units, and absence (less than 1%) of keratan sulphate. Apparently the major difference between glial/glioma and cartilage chondroitin sulphate proteoglycans relates to the glycan rather than to the protein moiety of the molecule.

scholarly journals αvβ3Integrin Mediates the Cell-adhesive Capacity and Biological Activity of Basic Fibroblast Growth Factor (FGF-2) in Cultured Endothelial Cells

1997 ◽  
Vol 8 (12) ◽  
pp. 2449-2461 ◽  
Author(s):  
Marco Rusnati ◽  
Elena Tanghetti ◽  
Patrizia Dell’Era ◽  
Anna Gualandris ◽  
Marco Presta
Keyword(s):  
Tissue Culture ◽  
Endothelial Cells ◽  
Cell Adhesion ◽  
Growth Factor ◽  
Fibroblast Growth Factor ◽  
Amino Acid Sequences ◽  
Dependent Manner ◽  
Fibroblast Growth ◽  
Tissue Culture Plastic ◽  
Cell Adhesive

Fibroblast growth factor-2 (FGF-2) immobilized on non-tissue culture plastic promotes adhesion and spreading of bovine and human endothelial cells that are inhibited by anti-FGF-2 antibody. Heat-inactivated FGF-2 retains its cell-adhesive activity despite its incapacity to bind to tyrosine-kinase FGF receptors or to cell-surface heparan sulfate proteoglycans. Recombinant glutathione-S-transferase-FGF-2 chimeras and synthetic FGF-2 fragments identify two cell-adhesive domains in FGF-2 corresponding to amino acid sequences 38–61 and 82–101. Both regions are distinct from the FGF-receptor-binding domain of FGF-2 and contain a DGR sequence that is the inverse of the RGD cell-recognition sequence. Calcium deprivation, RGD-containing eptapeptides, soluble vitronectin (VN), but not fibronectin (FN), inhibit cell adhesion to FGF-2. Conversely, soluble FGF-2 prevents cell adhesion to VN but not FN, thus implicating VN receptor in the cell-adhesive activity of FGF-2. Accordingly, monoclonal and polyclonal anti-αvβ3antibodies prevent cell adhesion to FGF-2. Also, purified human αvβ3binds to immobilized FGF-2 in a cation-dependent manner, and this interaction is competed by soluble VN but not by soluble FN. Finally, anti-αvβ3monoclonal and polyclonal antibodies specifically inhibit mitogenesis and urokinase-type plasminogen activator (uPA) up-regulation induced by free FGF-2 in endothelial cells adherent to tissue culture plastic. These data demonstrate that FGF-2 interacts with αvβ3integrin and that this interaction mediates the capacity of the angiogenic growth factor to induce cell adhesion, mitogenesis, and uPA up-regulation in endothelial cells.

scholarly journals The presence of a cartilage-like proteoglycan in the adult human meniscus

1981 ◽  
Vol 197 (1) ◽  
pp. 77-83 ◽  
Author(s):  
P J Roughley ◽  
D McNicol ◽  
V Santer ◽  
J Buckwalter
Keyword(s):  
Hyaluronic Acid ◽  
Chondroitin Sulphate ◽  
Gradient Centrifugation ◽  
Large Subunit ◽  
Dermatan Sulphate ◽  
Keratan Sulphate ◽  
Adult Human ◽  
Cscl Density Gradient ◽  
Cartilage Proteoglycans ◽  
Aggregate Structures

Proteoglycans were extracted from the adult human meniscus under dissociative conditions and purified by CsCl-density-gradient centrifugation. The preparations of highest density contained proteoglycan that possessed the ability to interact with hyaluronic acid, was of large subunit size and was composed of chondroitin sulphate, keratan sulphate and sialic acid-containing oligosaccharides. This ‘cartilage-like’ proteoglycan also exhibited subunit and aggregate structures analogous to those of hyaline-cartilage proteoglycans when examined by electron microscopy. However, the composition of this proteoglycan was more comparable with proteoglycans from immature cartilage than from age-matched cartilage. The preparations from lower density, which were enriched in dermatan sulphate, contained smaller proteoglycan that was not able to interact with hyaluronic acid. This non-aggregating proteoglycan may be structurally distinct from the ‘cartilage-like’ proteoglycan, which does not contain dermatan sulphate.

scholarly journals Immunochemical analysis of cartilage proteoglycans. Antigenic determinants of substructures

1979 ◽  
Vol 179 (1) ◽  
pp. 35-45 ◽  
Author(s):  
J Wieslander ◽  
D Heinegård
Keyword(s):  
Hyaluronic Acid ◽  
Chondroitin Sulphate ◽  
Antigenic Site ◽  
Trypsin Digestion ◽  
Antigenic Determinants ◽  
Binding Region ◽  
Keratan Sulphate ◽  
Link Protein ◽  
Before And After ◽  
Hyaluronic Acid Binding

Antibodies were raised in rabbits by injection of cartilage proteoglycan monomers, isolated hyaluronic acid-binding region, polysaccharide-peptides prepared by trypsin digestion of proteoglycans and link-protein. The rabbits injected with the proteoglycan monomers made antibodies reacting with the intact proteoglycan. The antiserum contained antibodies specific for, and also reacting with, the isolated hyaluronic acid-binding region and the keratan sulphate-rich region. In addition there were probably antibodies reacting with other structures of the proteoglycan monomer. When isolated hyaluronic acid-binding region was used for immunization the antibodies obtained reacted specifically with the hyaluronic acid-binding region. The antibodies obtained from rabbits immunized with the polysaccharide-peptides reacted with the proteoglycan monomers and showed a reaction identical with that of the chondroitin sulphate-peptides isolated after trypsin digestion of proteoglycans. The antibodies prepared with the link-protein as the antigen reacted only with the link-protein and not with any preparation from the proteoglycan monomer. Neither did any of the antisera raised against the proteoglycan monomer or its substructures react with the link-protein. Separately it was shown that the peptide ‘maps’ prepared from trypsin digests of the link-protein and the hyaluronic acid-binding region were different. Therefore it appears that the link-protein is not structurally related to the proteoglycan or the hyaluronic acid-binding region. Digestion of proteoglycan monomers or isolated hyaluronic acid-binding region with trypsin did not destroy the antigenic sites of the hyaluronic acid-binding region. In contrast trypsin digests of previously reduced and alkylated preparations did not react with the anti-(hyaluronic acid-binding region). The trypsin digests, however, reacted with both the antibodies directed against the chondroitin sulphate-peptides and those against the keratan sulphate-peptides. Trypsin digestion of the link-proteins destroyed the antigenic site and the reactivity with the antibodies. By combining immunoassay of proteoglycan preparations before and after trypsin digestion it is feasible to quantitatively determine its substructures by using the antisera described above.

Morphological Changes in Tissue-Cultured Cells Due to Variation in Growth Matrices

1988 ◽  
Vol 46 ◽  
pp. 252-253
Author(s):  
M. E. Hogan ◽  
D. H. DeGaetano ◽  
K. L. Klomparens
Keyword(s):  
Tissue Culture ◽  
Cultured Cells ◽  
Cell Attachment ◽  
Morphological Changes ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Cell Types ◽  
Growth Surface ◽  
Textured Surface ◽  
Tissue Culture Plastic

As the use of isolated cell cultures increases as an experimental model, there has been a proportional increase in the number of matrices used for cell growth support. These matrices vary in shape, texture and porosity, and as a result, affect the cells grown on them.The ultrastructural characteristics of several of these growth matrices were examined using two cell types chosen for their distinct growth habits. Chinese Hamster Ovary cells and Balb 3T3 Mouse Fibroblasts were grown on flat substrates (glass, tissue culture plastic, Millipore filters) as well as spherical (glass, tissue culture plastic, cross-linked dextran) matrices. Cells were plated maintaining egual densities and growth surface area. Once the majority of the cells reached confluency (approx. one week), the cell morphology on each matrix was examined using scanning electron microscopy and digital analysis of cell attachment area.Variation in cell shape was dramatic between matrices, being most noticeable between a textured surface (filter, dextran bead), and that of a smooth (glass) surface (Figs. 1 and 2). Even within smooth surfaces, some variation was observed.

scholarly journals The core molecule from type H proteoglycan. Release of mannose-containing oligosaccharides by digestion with N-oligosaccharide glycopeptidase

1985 ◽  
Vol 229 (3) ◽  
pp. 561-571 ◽  
Author(s):  
N Takahashi ◽  
H Ishihara ◽  
S Tejima ◽  
Y Oike ◽  
K Kimata ◽  
...  
Keyword(s):  
Chick Embryo ◽  
Chondroitin Sulphate ◽  
Keratan Sulphate ◽  
Pepsin Digestion ◽  
The Core ◽  
Proteoglycan Release

Chick-embryo cartilage contains a unique set of proteoglycans. Type H proteoglycan (PG-H) is the most abundant, constituting over 90% of the total cartilage hexuronate. We previously showed that treatment of PG-H with chondroitinase ACII and keratanase yields a protein-enriched core molecule [PG(-CS, KS)] with enzymically modified linkage oligosaccharides of the chondroitin sulphate and keratan sulphate chains. We report here that further treatment of PG(-CS, KS) with pepsin and N-oligosaccharide glycopeptidase (almond glycopeptidase) released four distinct types of mannose-containing oligosaccharide. Two of them were shown to be: (Formula: see text). Of the mannose-containing glycopeptides formed by pepsin digestion, about 40% (as mannose) were resistant to N-oligosaccharide glycopeptidase. Since the resistant fraction was enriched in keratan sulphate remnants, it is suggest that the mannose-containing oligosaccharides in this fraction represent those located in a keratan sulphate-enriched region of PG-H.

scholarly journals Variations in the composition of bovine hip articular cartilage with distance from the articular surface

1981 ◽  
Vol 195 (3) ◽  
pp. 535-543 ◽  
Author(s):  
A Franzén ◽  
S Inerot ◽  
S O Hejderup ◽  
D Heinegård
Keyword(s):  
Hyaluronic Acid ◽  
Articular Cartilage ◽  
Chondroitin Sulphate ◽  
Articular Surface ◽  
Full Thickness ◽  
Rich Region ◽  
Keratan Sulphate ◽  
Average Size ◽  
Almost All ◽  
Cartilage Proteoglycans

Punch biopsies of bovine hip articular cartilage was sectioned according to depth and the proteoglycans were isolated. The mid-sections of the cartilage contained more proteoglycans than did either the superficial or the deepest portions of the cartilage proteoglycans than did either the superficial or the deepest portions of the cartilage. The most superficial 40 micrometer of the cartilage contained relatively more glucosaminoglycans compared with the remainder of the cartilage. The proteoglycans recovered from the surface 200 micrometer layer contained less chondroitin sulphate, were smaller and almost all of these molecules were able to interact with hyaluronic acid to form aggregates. From about 200 micrometer and down to 1040 micrometer from the surface, the proteoglycans became gradually somewhat smaller, probably owing to decreasing size of the chondroitin sulphate-rich region. The proportion of molecules that were able to interact with the hyaluronic acid was about 90% and remained constant with depth. The proteoglycans from the deepest layer near the cartilage-bone junction contained a large proportion of non-aggregating molecules, and the average size of the proteoglycans was somewhat larger. The alterations of proteoglycan structure observed with increasing depth of the articular cartilage beneath the surface layer (to 200 micrometer) are of the same nature as those observed with increasing age in full-thickness articular cartilage. The articular-cartilage proteoglycans were smaller and had much higher keratan sulphate and protein contents that did molecules isolated from bovine nasal or tracheal cartilage.

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