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Results for “Extracellular Matrix”

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At least 325 records · Page 18Linked to original sources

PubMed2007

Conjugation of extracellular matrix proteins to basal lamina analogs enhances keratinocyte attachment.

The dermal-epidermal junction of skin contains extracellular matrix proteins that are involved in initiating and controlling keratinocyte signaling events such as attachment, proliferation, and terminal differentiation. To characterize the relationship between extracellular matrix proteins and keratinocyte attachment, a biomimetic design approach was used to precisely tailor the surface of basal lamina analogs with biochemistries that emulate the native biochemical composition found at the dermal-epidermal junction. A high-throughput screening device was developed by our laboratory that allows for the simultaneous investigation of the conjugation of individual extracellular matrix proteins (e.g. collagen type I, collagen type IV, laminin, or fibronectin) as well as their effect on keratinocyte attachment, on the surface of an implantable collagen membrane. Fluorescence microscopy coupled with quantitative digital image analyses indicated that the extracellular matrix proteins adsorbed to the collagen-GAG membranes in a dose-dependent manner. To determine the relationship between extracellular matrix protein signaling cues and keratinocyte attachment, cells were seeded on protein-conjugated collagen-GAG membranes and a tetrazolium-based colorimetric assay was used to quantify viable keratinocyte attachment. Our results indicate that keratinocyte attachment was significantly enhanced on the surfaces of collagen membranes that were conjugated with fibronectin and type IV collagen. These findings define a set of design parameters that will enhance keratinocyte binding efficiency on the surface of collagen membranes and ultimately improve the rate of epithelialization for dermal equivalents.

Basement Membrane↗

PubMedDate not supplied

Three-dimensional reconstructions of extracellular matrix polymers using automated electron tomography.

The extracellular matrix is an intricate network of macromolecules which provides support for cells and a framework for tissues. The detailed structure and organisation of most matrix polymers is poorly understood. These polymers have a complex ultrastructure, and it has proved a major challenge both to define their structural organisation and to relate this to their biological function. However, new approaches using automated electron tomography are beginning to reveal important insights into the molecular assembly and structural organisation of two of the most abundant polymer systems in the extracellular matrix. We have generated three-dimensional reconstructions of collagen fibrils from bovine cornea and fibrillin microfibrils from ciliary zonules. Analysis of these data has provided new insights into the organisation and function of these large macromolecular assemblies.

Animals↗

PubMed1994

Inhibition of T cell activation by the extracellular matrix protein tenascin.

Tenascin (TN) is an extracellular matrix protein that is expressed widely in the fetus and sparingly in the adult, but reappears at high levels in certain areas of tissue insult such as tumor matrices and sites of wound healing. We show here that soluble TN inhibits proliferation of human T cells in response to alpha CD3 Ab co-immobilized with the extracellular matrix protein fibronectin (FN). TN also inhibits proliferation driven by alpha CD3/IL-2 or by phorbol ester/IL-2, and it prevents high level induction of IL-2R. The presence of TN in culture medium does not detectably alter the pattern of tyrosine phosphorylation resulting from T cell triggering with alpha CD3, but at later time points prevents the appearance of functional NF-AT1 transcription factor complexes in T cell nuclear extracts. These findings are consistent with the postulated role for TN as a natural antagonist to FN action, and suggest that T cell responses occurring at tissue sites in which TN is expressed could be influenced by its presence.

Base Sequence↗

PubMed1994

A line of rat ovarian surface epithelium provides a continuous source of complex extracellular matrix.

A spontaneously immortalized, yet non-tumorigenic rat ovarian surface epithelial (ROSE 199) cell line, deposits large amounts of extracellular matrix (ECM) in response to crowding. The characteristics and components of ROSE 199-derived cell-free ECM were compared after three different preparative techniques: treatment with 20 mM ammonium hydroxide, with 1% sodium deoxycholate, or by repeated freeze-thaws. The ECMs were analyzed by histochemistry, immunofluorescence, electron microscopy, and Western immunoblotting. Components of ROSE 199 ECM included laminin, fibronectin, and collagen types I and III. Even though ROSE 199 is an epithelial cell line, striated collagen fibers formed a major part of its matrix. Thus, ROSE 199 matrix consists of both basement membrane and stromal matrix components. This matrix supported the adhesion, spreading, and growth of several cell types without altering their morphology or growth pattern, and enhanced the attachment of some cell types that spread on plastic only with difficulty. Immunofluorescence, electron microscopy, and dry weight determinations indicated that a greater proportion of matrix was retained in preparations obtained by ammonium hydroxide or freeze thaw techniques than after sodium deoxycholate treatment. Ammonium hydroxide and freeze-thaw treated matrices were also superior to sodium deoxycholate preparations as evidenced by enhanced initial cellular adhesion and spreading compared to cells plated on plastic. Residual nuclear material did not seem to affect the biological activity of this matrix. ROSE 199 extracellular matrix provides a novel, complex substratum for cell culture and for studies of matrix functions and synthesis.

Actins↗

PubMed2006

Extracellular matrix metalloproteinase inducer in interstitial pneumonias.

Extracellular matrix metalloproteinase inducer (EMMPRIN), a glycosylated transmembrane protein that induces matrix metalloproteinases (MMPs), is minimally expressed in the normal adult lung. We previously reported that it is up-regulated in murine bleomycin-induced lung injury. In this study, we determined the expression of EMMPRIN and its association with MMP-2, MMP-7, and MMP-9 in interstitial pneumonias (IPs). We performed immunohistochemistry for EMMPRIN and MMPs on lung tissue from 22 subjects with various IPs. We did bronchoalveolar lavage (BAL) on 9 of these subjects and 13 others with IPs to measure the soluble EMMPRIN in BAL fluid. For comparison, immunohistochemistry or BAL was done on 14 subjects without IPs. The staining intensity for each protein was scored from 0 to 3 in various epithelial cell types. Soluble EMMPRIN in BAL fluid was measured by an enzyme-linked immunosorbent assay. Extracellular matrix metalloproteinase inducer was prominent in abnormal epithelial cells. It was more prominent in hyperplastic type II cells, compared with epithelium in alveolar bronchiolization. It was also elevated in BAL fluid from the subjects with IPs. Matrix metalloproteinases were expressed in cells expressing EMMPRIN, although the profile of MMPs varied among the different abnormal epithelial cell types with MMP-2 and MMP-7 in hyperplastic type II cells and MMP-7 and MMP-9 in cells showing squamous metaplasia and cells comprising bronchiolization. These results suggest a role of EMMPRIN in reepithelialization in IPs.

Basigin↗

PubMed2003

Melanin in the extracellular matrix of germlings of Botrytis cinerea.

Previous work on the composition of the extracellular matrix of germlings of the plant pathogenic fungus Botrytis cinerea demonstrated the presence of carbohydrate, protein, and simple lipids; which, together, comprised 50-60% of the dry weight. Here we show that most of the remaining mass of the extracellular matrix consists of a chemically inert dark pigment with the electron paramagnetic resonance characteristics of a melanin. Scanning electron micrographs of the purified pigment, and transmission electron micrographs of thin sections made using the pigment indicate that it has a filamentous structure. We conclude that melanin is an important component of the extracellular matrix of germlings of B. cinerea. This is the first report of a melanin present in the extracellular matrix of a plant pathogenic fungus.

Botrytis↗

PubMed1990

[Immunomorphological research on the formation of the extracellular matrix in epithelial cell cultures].

Formation of extracellular matrix structures in cultures of rat liver epithelial nontransformed cell line IAR2 was studied with antisera to fibronectin, laminin and type IV collagen by immunofluorescence and immunoelectron microscopy of platinum replicas. Fibronectin formed peripheral spots of variable size some of which outlined free cell edges, as well as fibrils located towards the center of single cells or of cellular islands. Similarly distributed structures were seen in isolated matrices. Codistribution of fibronectin and actin was observed only for the peripheral line of fibronectin spots and marginal circular actin bundle. Basement membrane components. laminin and type IV collagen, formed mainly spots of variable size predominantly beneath the cell or each cell in an island. Occasional fibrils were seen also. Essentially the same results were obtained by immunofluorescence and immunogold electron microscopy. Cytochalasin D treated cells displayed spots of both fibronectin and laminin. The relevance of previously postulated receptor-mediated assembly of extracellular matrix structures to the epithelial cells is discussed.

Animals↗

PubMed1994

Assembly of amino-terminal fibronectin dimers into the extracellular matrix.

Fibronectin is a dimeric adhesion molecule that consists of three types of repeating modules. Adherent cells bind soluble fibronectin and incorporate it into insoluble fibrils in the extracellular matrix. The amino-terminal 70-kDa portion of fibronectin mediates binding to the cell surface, but amino-terminal fragments do not accumulate in the extracellular matrix. The ninth type I and first type III modules, the cell adhesion region, and the cysteines that form the interchain disulfide bonds have also been implicated in matrix assembly. To further define which regions of fibronectin are essential for matrix assembly, we generated a dimeric protein (d70 kDa) in which the 70-kDa amino terminus is directly linked to the last 51 amino acids of fibronectin, which contain the cysteines involved in interchain disulfide bonding. d70 kDa bound to cells and accumulated in the extracellular matrix. Incorporation of d70 kDa into the extracellular matrix was dependent upon protein synthesis; in cycloheximide-treated cultures that lacked a pre-existing matrix, d70 kDa accumulated in the extracellular matrix only in the presence of intact fibronectin. Monomeric 70-kDa protein was not incorporated into the matrix in the presence or absence of cycloheximide. These data indicate that fibronectin molecules containing only the amino-terminal 70-kDa region and the carboxyl-terminal 51 amino acids can become assembled into the extracellular matrix.

Animals↗

PubMed2002

The effect of external fractionated irradiation on the distribution pattern of extracellular matrix proteins in submandibular salivary glands of the rat.

INTRODUCTION: The aim of this study was to analyse the distribution pattern of extracellular matrix proteins in the irradiated and non-irradiated rat submandibular salivary gland in order to provide a more detailed profile of the radiation injury following radiotherapy of the head and neck. MATERIAL AND METHODS: External X-ray exposure, restricted to the left skull base and neck region, was performed in 60 female Wistar rats, fractionated to daily applications of 2 Gy, up to total dosages of 20, 40 or 60 Gy. Both submandibular glands were excised after supravital anaesthesia 6 months or 1 year after completion of the irradiation. Spatial and temporal patterns of extracellular matrix proteins were investigated histologically and immunohistochemically. RESULTS: The polyclonal anti-human antisera used, identified the same antigens in rat tissue as in human tissues. The alterations in staining patterns and staining intensities between irradiated and non-irradiated salivary glands showed statistically significant differences. Different structures in irradiated glands reacted with different intensities, e.g. nerve tissue and the basement membranes of excretory ducts were intensely laminin-positive, fibronectin was predominantly found around the excretory ducts with transition to the interstitial tissues. CONCLUSION: Irradiation leads to statistically significant differences in the amount and composition of the extracellular matrix in salivary glands. The amount of extracellular matrix proteins in irradiated glands is dose-dependent. The higher the dosage the more extracellular matrix can be expected. Consecutively, total dosage is associated with greater loss of acini. Scatter effects of irradiation have also to be recognized. Immunohistochemical studies on salivary glands have to consider the pretreatment status, in particular those studies that investigate degenerative changes.

Animals↗

PubMed1996

Articular chondrocyte tenascin-C production and assembly into de novo extracellular matrix.

Tenascin-C is an oligomeric glycoprotein of the extracellular matrix that is expressed in a variety of processes including development, tissue remodeling, wound healing, cell adhesion/antiadhesion, and cell/matrix interactions. Tenascin has recently been acknowledged as a component of the extracellular matrix of articular cartilage, but its function remains unclear. In this study, bovine articular chondrocytes were grown in alginate beads for 35 days to examine the kinetics of tenascin synthesis and incorporation into de novo extracellular matrix. During the culture period, 6 harvest days were established in which culture medium was recovered, alginate beads were dissociated with an EDTA solution, and chondrocytes were collected and lysed by sonication. Total DNA determination performed on the cell lysates demonstrated chondrocyte survival and proliferation. Western blotting performed on the medium, EDTA/alginate, and lysate samples demonstrated the production of both the 220 and 320 kDa tenascin size variants and their differential compartmentalization within the culture system. Tenascin was incorporated into the alginate bead matrix at a constant rate of 3.8 micrograms/day. The 320 kDa variant was produced in higher quantity, but the 220 kDa fragment was twice as likely to be incorporated into the de novo matrix. Methylene blue/acid fuchsin staining and tenascin immunohistochemistry demonstrated the incorporation of tenascin into a progressively expanding matrix surrounding the chondrocytes. The results suggest a role for tenascin in the assembly of the chondrocyte matrix and as a soluble mediator of chondrocytes with possible diverse functions for the tenascin size variants.

Animals↗

PubMed2000

Defective extracellular matrix reorganization by chronic wound fibroblasts is associated with alterations in TIMP-1, TIMP-2, and MMP-2 activity.

Chronic leg wounds are characterized by defective remodeling of the extracellular matrix, failure of reepithelialization, and prolonged inflammation. The hypothesis that this defective extracellular matrix remodeling is associated with phenotypic differences in the activity of the matrix metalloproteinases and tissue inhibitors of metalloproteinases was studied in chronic wound and patient-matched normal fibroblasts in three-dimensional collagen lattice systems. Chronic wound fibroblasts exhibited no differences in morphology or proliferation (p > 0.1) compared with patient-matched uninvolved dermal fibroblasts. The ability of chronic wound fibroblasts to reorganize extracellular matrix was significantly impaired, however, in comparison to the uninvolved dermal fibroblasts (p < 0.01). This difference in extracellular matrix reorganization was not related to differences in proliferation within the collagen lattices (p > 0.05) or attachment to type I collagen (p > 0.1). Marked differences were evident in matrix metalloproteinase-2 activity between chronic wound and patient-matched normal fibroblasts. Whereas levels of pro-matrix metalloproteinase-2 were similar between the two fibroblast populations (p > 0.1), the chronic wound fibroblasts exhibited significantly decreased levels of the 62 kDa active form of matrix metalloproteinase-2 (p < 0.01). Reverse zymography and enzyme-linked immunosorbent assay demonstrated that the decreased matrix metalloproteinase-2 activity was associated with increased production of tissue inhibitors of metalloproteinase-1 and -2 by the chronic wound fibroblasts (p < 0.05). Increased production of tissue inhibitors of metalloproteinases in chronic wound fibroblasts was also reflected in decreased levels of matrix metalloproteinase-1 (p < 0.005). These data suggest that the impaired ability of chronic wound fibroblasts to reorganize extracellular matrix in vitro is related to decreased levels of active matrix metalloproteinase-2 and matrix metalloproteinase-1 resulting from increased production of tissue inhibitors of metalloproteinase-1 and -2 by chronic wound fibroblasts. These findings provide a mechanism to explain the impaired cellular responses and extracellular matrix reorganization observed in chronic leg wounds in vivo.

Cell Adhesion↗

PubMed1997

[Research on development of hybrid artificial liver utilizing reconstituted extracellular matrix].

We investigated culture of adult rat hepatocytes on reconstituted extracellular matrix, glycosaminoglycans containing collagen gel and on a porous gelatin sponge (gelform) for the maintenance of the liver-specific functions of hepatocytes. In the culture on reconstituted extracellular matrix, only heparin containing collagen cultures could significantly sustain albumin synthesis for over 3 weeks. On the other hand, in the culture of rat hepatocytes on a porous gelatin sponge, the morphology of the hepatocytes immobilized on the support was close to that in vivo and the secretion of albumin and bile acids was stable for over 12 days. These results demonstrate that culture on two- and three-dimensional culture with reconstituted extracellular matrix is one of the promising hepatocytes cultures for development of artificial liver.

Albumins↗

PubMedDate not supplied

Matrix metalloproteinase expression in malignant melanomas: tumor-extracellular matrix interactions in invasion and metastasis.

Structural changes in the extracellular matrix (ECM) are necessary for cell migration during tissue remodeling and tumor invasion. The matrix metalloproteinases (MMPs) and their inhibitors have been shown to be critical modulators of ECM composition and are, thus, crucial in neoplastic cell invasion and metastasis. Expression of MMP-2, -3, -9, -10, and -13 was investigated in human malignant melanomas (MMs) employing an indirect alkaline phosphatase conjugated immunocytochemical technique. Evaluation of the results was based on (a) the percent of neoplastically transformed cells/surrounding stroma that reacted positively and (b) a measure of staining intensity [graded from A (highest) to D]. The two forms of stromelysin, MMP-3 and -10, share 82% sequence homology, but exhibit differences in cellular synthesis and inducibility by cytokines and growth factors in vitro. Strong overall expression of MMP-3 and -10 was found in MMs, especially in the ECM adjacent to blood vessels. Positive immunoreactivity could be seen for these two MMPs in the ECM surrounding over 90% of the neoplastically transformed cells (++++), and the staining intensity was also the strongest possible (A,B). Focal (+), high intensity (A,B) staining could be detected for MMP-2, -9, and -13. Thus, it seems that the stromelysins are involved in the generalized growth and expansion of the neoplastic cell mass, while MMP-2, -9 and -13 are involved in the neoangiogenic and focal clonal selection and expansion phenomena associated with in situ tumor progression, invasion of the microvasculature, and metastasis.

Antibodies, Monoclonal↗

PubMed2001

[The current situation and future of extracellular matrix materials for bone tissue engineering].

The requirements of ideal extracellular matrix materials for bone tissue engineering were stated, and the advantages and disadvantages of bioceramics, biodegradable synthetic polymers and natural polymers were analyzed. Based on these, we highlight a point that the ideal extracellular matrix materials for bone tissue engineering should be made up of bioceramics materials, synthetic polymers or natural polymers. These materials should possess a morphological structure of three-dimensional foam and have a self-mediated drug delivery system of bone growth factors. The design and manufacture of such compound extracellular matrix materials for bone tissue engineering is a very important and urgent challenge.

Biocompatible Materials↗

PubMed1991

Expression of antisense transcripts encoding an extracellular matrix protein by stably transfected vascular smooth muscle cells.

The ability to selectively inhibit the expression of various extracellular matrix proteins would provide an approach towards elucidating not only the function of these molecules but allow some insight to be gained into the mechanisms by which these molecules influence various aspects of vascular smooth muscle cell behavior such as growth, migration, morphology, and extracellular matrix assembly. The present study explores the feasibility of employing an antisense RNA approach to study vascular smooth muscle extracellular matrix proteoglycan function. Vascular smooth muscle cells were transfected with a test plasmid designed to express an antisense transcript encoding an extracellular matrix protein. The test plasmid was constructed by inserting the cDNA encoding the majority of the rat chondrosarcoma extracellular matrix link protein in an antisense orientation relative to the rat metallothionein-I promoter. Several stably transfected cell lines were isolated and characterized with respects to link antisense transcript expression, morphology, growth, and smooth muscle cytoskeletal protein expression. Results from these studies have shown that vascular smooth muscle cells could be stably transfected and link antisense transcript could be expressed at high levels by these cells. In addition, the rat metallothionein-I promoter was found to be a strong inducible promoter in vascular smooth muscle cells. Cell biology characterization of the various transfected vascular smooth muscle cell lines generated in this study has shown that one type of transfected vascular smooth muscle cell line expressed cellular characteristics indicative of early passaged vascular smooth muscle cells while a second type of transfected cell line was morphologically similar to vascular smooth muscle cells recovered from neonatal rat aortas and mechanically injured arteries obtained from adult rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1989

Comparison of the amino acid composition of the intracellular and extracellular matrix protein fractions isolated from avian skeletal muscles.

Intracellular and extracellular skeletal muscle protein fractions were isolated from the legs and breasts of young and adult White Leghorn chickens and quantified by detailed amino acid analysis. This involved repeated homogenization in the presence of 50 mM CaCl2, neutral phosphate-buffered saline (pH 7.4), solubilization by 2% sodium dodecyl sulfate (SDS), and centrifugation to separate all intracellular muscle proteins from the extracellular matrix. The total SDS-soluble intracellular muscle proteins in the adult and young birds ranged respectively from 93.2 to 94.5% in the leg and from 93.5 to 94.1% in the breast muscles. Collagen and collagen-like proteins in the extracellular matrix protein fractions were calculated from the amounts of 5-hydroxylysine found in their 96-h acid hydrolysates and elastin content from the amounts of desmosine present. Total collagen ranged from 3.42 to 5.18% in legs and from 2.91 to 3.89% in breasts. The elastin content of leg muscles represents only .061% of the total muscle protein. The calculated protein efficiency ratios for intracellular avian muscle proteins averaged 3.2 compared with a mean value of 1.4 for the extracellular matrix.

Amino Acids↗

PubMed2000

Ets factors and regulation of the extracellular matrix.

Ets factors are critical mediators of extracellular matrix (ECM) remodelling. As the spectrum of Ets-regulated target genes widens, so does their role in various pathological and physiological processes. Regulation of matrix degrading proteases by Ets factors in tumor invasion and metastasis is well established. Emerging evidence suggests that they may also play a role in the pathology of autoimmune diseases. Newly characterized Ets target genes such as tenascin-C and collagen type I suggest their role in diseases characterized by aberrant collagen deposition (fibrosis). Ets function is also critical in bone and cartilage development. There is increasing knowledge of the complex regulatory mechanisms involved in transcription of Ets target genes. Ets factors may function as activators or as repressors via association with specific cofactors depending on the promoter context. Signaling pathways can modulate the activation status of Ets factors and their transcriptional partners. Precise understanding of the role of Ets factors in the complex cellular network governing the expression of ECM proteins and the enzymes that degrade them will be a focus of future studies.

3T3 Cells↗

PubMed1998

[Relationship between extracellular matrix and progressive growth of malignant tumor].

OBJECTIVE: To explore the role of extracellular matrix proteins in tumor cell invasion and metastasis. METHODS: Mouse pulmonary adenocarcinoma cell line LA795 and human nasopharyngeal carcinoma cell line CNE-2Z were implanted subcutaneously and under the renal capsule in T739 and nude mice, respectively. At different time intervals, expression of fibronectin(FN), laminin(LN) and type IV collagen(IV C) in the tumor extracellular matrix and in tumor adjacent tissues was examined by indirect immunoperoxidase and immunofluorescence techniques. In addition, various in vitro methods (random migration of tumor cells in agarose droplet, growth of tumor cell colonies in soft agar, dot blotting, etc.) were used to analyse the effect of exogenous LN, FN or IV C. RESULTS: As tumor grew, the expression of FN, LN or IV C increased and presented different distribution patterns. Exogenous FN, LN and IV C enhanced the mobility of tumor cells and the growth of the tumor cell colonies. CONCLUSION: The distribution difference and the changes in synthesis and degradation of the extracellular matrix proteins are valuable in predicting the biologic behaviour. Tumor implanted under the renal capsule of mice offers a better experimental model to study the relationship between extracellular matrix and tumor cell invasion.

Adenocarcinoma↗