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Results for “Extracellular Matrix”

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PubMed1986

Effects of thrombin on the fibronectin-containing extracellular matrix.

The effects of human alpha-thrombin on the incorporation of exogenous human fibronectin into the extracellular matrix of cultured human skin fibroblasts were studied. Thrombin had no effect on the saturable cell-surface receptor for soluble fibronectin. Thrombin, however, did digest fibronectin bound reversibly to the cell surface and fibronectin incorporated irreversibly into the extracellular matrix. The products of thrombin digestion of cell surface and matrix-bound fibronectin were similar to the products of thrombin digestion of soluble fibronectin. These results indicate that fibronectin can potentially modulate some of the effects of thrombin on cells.

Cells, Cultured↗

PubMed2001

Differential regulation of liver-specific and ubiquitously-expressed genes in primary rat hepatocytes by the extracellular matrix.

Primary rat hepatocytes were cultured with an extracellular matrix (ECM) overlay, in order to investigate the effect of an ECM on gene expression in hepatocytes. When hepatocytes, isolated by the collagenase-perfusion method, were cultured on type I collagen-coated dishes, the mRNA levels of liver-specific genes (aldolase B, tyrosine aminotransferase and albumin) decreased continuously, while those of ubiquitously-expressed genes (glyceraldehyde 3-phosphate dehydrogenase and beta-actin) increased. When a dilute ECM derived from the Engelbreth-Holm-Swarm mouse sarcoma (an EHS gel) was added to the above hepatocytes 3 days after plating, the mRNA levels of liver-specific genes increased, while those of ubiquitously-expressed genes decreased. The effects of a rat liver biomatrix (a physiological ECM for rat hepatocytes) on gene expression in primary hepatocytes were similar to those of the EHS gel. A nuclear run-on assay, and 5,6-dichloro-1-beta-d-ribofuranosylbenzimidazole or actinomycin D treatments revealed that the transcriptional rates of liver-specific genes were enhanced by the EHS gel overlay, while the apparent stability of the corresponding mRNAs were unchanged. In contrast, the transcriptional rates of ubiquitously-expressed genes were not greatly affected by an EHS gel overlay, while the apparent stability of their mRNAs were decreased. These data suggest that the ECM plays an important role in the maintenance of the differentiated characteristics of primary hepatocytes by inducing the transcription of liver-specific genes and, also, by destabilizing the mRNAs of ubiquitously-expressed genes.

Actins↗

PubMed1989

The E8 subfragment of laminin promotes locomotion of myoblasts over extracellular matrix.

The locomotion of murine myoblasts over the extracellular matrix components laminin and fibronectin was analyzed using quantitative videomicroscopy, and the organization of the cytoskeleton was observed in parallel immunofluorescence studies. Cells plated on the laminin-nidogen complex locomoted twice as fast as on laminin alone. The main form of translocation on laminin was a jerky cycle of prolonged lamellipod extension followed by rapid (approximately 200- less than 500 microh h-1) movement of the cell body into the extended lamellipod. The locomotion-stimulating activity of laminin resides in the elastase digestion fragment E8, part of the laminin long arm, while the E1-4 fragment containing the three short arms is inactive. Myoblasts moved poorly over fibronectin irrespective of whether high, intermediate, or low coating concentrations were used (approximately 5,000- approximately 10 fmol cm-2). In contrast, the locomotory responses both to laminin and to E8 peaked sharply at coating concentrations approximately 20-50 fmol cm-2 and decreased at higher concentrations. This response corresponds to that expected for a haptotactic stimulant. When cells locomoted over a mixed substrate of laminin and fibronectin, the fibronectin effects appeared to predominate. The cytoskeleton has been implicated in many cellular motile processes. Within 6 h on fibronectin many cells expressed vinculin-containing focal contacts, elaborated stress fibers and had periodically organized alpha actinin, whereas on laminin, most cells showed diffuse vinculin and alpha actinin and a fine meshlike actin cytoskeleton. We conclude that the poor locomotion of cells over fibronectin is because of the cytoskeletal stabilization it induces.

Actins↗

PubMed2003

Glucocorticoid enhances transforming growth factor-beta effects on extracellular matrix protein expression in human placental mesenchymal cells.

Extracellular matrix (ECM) proteins synthesized by human placental mesenchymal cells (PMCs) provide structural support for the villus. Aberrant expression of ECM proteins by PMCs has been associated with intrauterine growth restriction (IUGR). To provide insight into the mechanisms of ECM protein regulation in the stroma of the placental villus, in the current study, we examined the interaction of glucocorticoid (GC) and transforming growth factor-beta (TGFbeta) in the modulation of ECM proteins in cultures of PMCs isolated from human term placentas. Initial results obtained by ELISA showed that combined treatment with dexamethasone (DEX) and TGFbeta enhanced oncofetal fibronectin (FFN) protein levels in serum-free culture medium severalfold in a dose-dependent manner. Northern blotting and real-time polymerase chain reaction (PCR) analyses revealed a similar enhancement in levels of FN mRNA in cells treated with TGFbeta and DEX. Real-time PCR results also revealed that DEX and TGFbeta enhanced collagen (Col) I and Col IV expression, but did not affect levels of Col III or laminin, indicative of selective stimulation of ECM proteins. Hypoxic treatment moderately enhanced FFN levels in control cells but not in those treated with DEX and TGFbeta. In contrast with the results obtained with PMCs, we noted that DEX treatment suppressed FFN levels in untreated and TGFbeta-treated cytotrophoblasts, suggesting that GC and TGFbeta modulate FFN expression in placenta in a cell-type-specific manner. We conclude that GC and TGFbeta are key regulators of ECM protein synthesis in PMCs, suggesting a role in modulating placental architecture in uncomplicated pregnancies and those associated with aberrant ECM protein expression.

Blotting, Northern↗

PubMed1999

Regulatory role of extracellular matrix components in expression of matrix metalloproteinases in cultured hepatic stellate cells.

Hepatic stellate cells (HSCs) were changed in their morphology, proliferative activity, and functions by culturing on type I collagen gel, as compared to the culture on polystyrene surface. HSCs have been found to produce extracellular matrix components and matrix metalloproteinases (MMPs). In this study, we have assessed the effects of several types of substrata on the expression of MMPs in HSC culture. MMP-1 expression was detectable in HSC culture on polystyrene surface and on type I collagen gel by immunofluorescence staining and reverse transcriptase-polymerase chain reaction (RT-PCR). The results from in situ zymography revealed the presence of interstitial collagenase activity around HSCs and along their cellular processes. Although proMMP-2 and proMMP-9 were detectable by gelatin zymography in the conditioned medium from both cultures using type I collagen gel and Matrigel as substratum, an active form of MMP-2 but not of MMP-9 was detected only in the culture using type I collagen as a substratum. Tissue inhibitor of metalloproteinase-2 expression was observed by RT-PCR in HSCs cultured on or in type I collagen gel, suggesting the suppression of MMP-2 activity detected in HSC culture using type I collagen. These results indicate a differential expression of MMP activity, hence the remodeling of extracellular matrix components is dependent on the substratum used for HSC culture. The HSC culture using several types of substrata appears to be a useful in vitro model to study the mechanism of extracellular matrix remodeling.

Actins↗

PubMed1997

Extracellular matrix composition and gene expression in collagenous colitis.

BACKGROUND & AIMS: Collagenous colitis is a rare diarrheal disease of unknown pathophysiology that is histologically defined by subepithelial bandlike structures. The objective of this study was to elucidate the biochemical composition and the origin of the bandlike structures in collagenous colitis. METHODS: Immunohistochemical and in situ hybridization analyses were performed on endoscopic specimens using specific antibodies and riboprobes for collagen types I, III, IV, and VI and for the glycoprotein tenascin. RESULTS: In collagenous colitis, the mucosal matrix with the exception of the bands retained a normal architecture and extracellular matrix composition. The bands stained most prominently for type VI collagen and tenascin. Less abundant staining for both proteins was also found in the subepithelial matrix of the normal mucosa. In situ hybridization showed no significant increase in collagen type VI messenger RNA expression in cells around and entrapped in the bands in collagenous colitis compared with normal specimens. CONCLUSIONS: The results support the suggestion that collagenous colitis is a localized alteration of the extracellular matrix, which involves the pericryptal-subepithelial myofibroblast sheath. The data suggest that reduced matrix degradation and not overactivation of matrix synthesis may be the reason for the subepithelial accumulation of matrix proteins.

Adult↗

PubMed2006

Repopulation of osteosarcoma cells after treatment with doxorubicin in the presence of extracellular matrix biopolymers.

PURPOSE: To elucidate the role of extracellular matrix (ECM) in repopulation capacity of osteosarcoma cells after doxorubicin treatment. METHODS: OSCORT cells established in our laboratory from a human osteosarcoma, were treated with doxorubicin in monolayer for 4 h, then cells were further incubated either in monolayer or in ECM-containing three-dimensional cell-culture (3-DCC), apoptosis induction and changes in cell number were measured. Alkaline comet assay was applied to estimate DNA damage, immunoblot technique and immunocytochemistry were used to investigate p53 protein synthesis, and the repopulating capacity in monolayer culture and in ECM-based 3-DCC, after doxorubicin treatment was measured. In addition to OSCORT culture five other human cell lines (HT-1080, PC-3, MDA-MB231, A-431 and ZR-75-1) were used to compare the antimigratory and antiproliferative effects of doxorubicin. RESULTS: The apoptotic index, the extent of DNA damage and the representation of p53 were much lower in the OSCORT cell cultures if the cells were exposed to ECM after treatment with doxorubicin. The doxorubicin-treated OSCORT cells transferred from the monolayer culture were not able to proliferate at all, at the same time, the cytoprotection provided by ECM prevailed upon transferring the cells into plastic dish, and resulted in potent repopulation capacity of the cells. CONCLUSIONS: Present data indicate that ECM contributes to failure in therapy of human osteosarcoma in clinical situation. Overall, the application of ECM-based 3-DCC could be suggested as an appropriate model system for the better understanding of antitumor drug action and hereby to set the stage for promising novel pharmacological approaches in cancer therapy.

Antibiotics, Antineoplastic↗

PubMed1996

Human trophoblast cell adhesion to extracellular matrix protein, entactin.

PROBLEM: Trophoblast interaction with endometrial extracellular matrix (ECM) is crucial during human embryo implantation and placentation. Entactin, a ubiquitous basement membrane glycoprotein, plays a central role in ECM assembly, cell attachment, and chemotaxis. The present study was conducted to examine the possible role of entactin in promoting human trophoblast adhesion. METHODS: Using an extended life span first trimester trophoblast cell line HTR-8/SVneo (HTR) and a cell adhesion assay, we measured the adherence of human first trimester trophoblasts to recombinant entactin and its domains. Also, we used flow cytometry and indirect immunofluorescence to detect the presence of integrins that may be involved in human trophoblast-entactin interaction; these methods were used to analyze HTR cells, as well as tissue sections and freshly isolated human trophoblasts from first trimester and term placenta. RESULTS: We found that first trimester trophoblast cells were highly adherent to entactin and its E and G2 domains but not to G1 or G3 domains. Using indirect immunofluorescence and flow cytometry, we found that both beta 1 and beta 3 integrin subunits were expressed on the surface of HTR trophoblast cells adhering to entactin; in contrast, beta 2 and beta 4 integrin subunits were not detected. In addition, we found that alpha v beta 3 was expressed on freshly isolated villous cytotrophoblasts and cytotrophoblast and syncytiotrophoblasts in tissue sections from term placenta. The beta 3 integrin subunit was expressed in cytotrophoblasts and syncytiotrophoblasts in villi of first trimester placental tissue sections. CONCLUSION: Recombinant entactin promotes human trophoblast cell adhesion through both its E and G2 domains and these specific adhesive interactions may be mediated by beta 1 and/or beta 3 class integrins.

Antigens, CD↗

PubMed2005

The effect of extracellular matrix on embryonic stem cell-derived cardiomyocytes.

Extracellular matrix (ECM) components significantly influence the growth characteristics of cardiomyocytes, development of spontaneous contractile activity and morphologic differentiation. The present study involves characterization of ES cell-derived cardiomyocytes cultured on different ECMs. We hypothesized that cardiogel, which is a naturally occurring ECM containing a complex mixture of laminin, fibronectin may better support ES cell-derived cardiomyocytes maturation in terms of chronotropic characteristics and subcellular structure development. Spontaneously differentiated cardiomyocytes from 7-day embryoid bodies (EBs) derived from mouse ES cells line Royan B1 (C57BL/6) were cultured on commercial ECM (matrigel), a fibroblast-derived ECM (cardiogel), and control (without ECM) for 7 + 21 days. The growth characteristics of cardiomyocytes were assessed by immunocytochemistry, reverse transcription polymerase chain reaction (RT-PCR), positive and negative chronotropic drugs and transmission electron microscopy (TEM). The spontaneously beating cells expressed markers characteristic of cardiomyocytes including alpha-actinin, desmin, troponin-I, sarcomeric myosin heavy chain (MHC), pan-cadherin, connexin-43, cardiac alpha-MHC, cardiac beta-MHC, atrial natriuretic factor (ANF) and myosin light chain isoform-2V (MLC-2V). In addition, spontaneous beating of cardiomyocytes on both ECMs was enhanced by treatment of cells with isopernaline, while reduced more on matrigel, by carbacol when compared with control and cardiogel. However, the change in beating was similar in all groups upon treatment with phenylephrine and/or Bay-K. Ultrastructure analysis showed myofibrillar bundle organization and exhibited intercalated discs, T-tubules, Z-, A-, I-, H- and M-bands and the showed earlier maturation of the cardiomyocytes on cardiogel. We conclude that substructurally matured cardiomyocytes can be produced from ES cells and the cardiac fibroblast-derived ECM (cardiogel) supported earlier maturation of ES-derived cardiomyocytes, in terms of chronotropic characteristics and subcellular structure development.

Animals↗

PubMed1994

Developmental regulation of glucocorticoid-mediated effects on extracellular matrix protein expression in the human placenta.

The extracellular matrix (ECM) protein fibronectin (FN) is a critical regulator of uterine-placental adherence. In the present report we compared the effects of glucocorticoids on FN expression in cytotrophoblast cultures isolated from human first trimester and term placentas to elucidate potential steroid-dependent cellular mechanisms associated with human parturition. Based on immunoassays, treatment of first trimester cytotrophoblasts with 10(-7) M dexamethasone (DEX) for 2 cr 4 days reduced medium levels of oncofetal FN (onfFN; i.e. FNs bearing an oncofetal epitope) to approximately 80% of control levels. Conversely, treatment of cytotrophoblasts isolated from term placentas with DEX dramatically reduced medium levels of onfFN to approximately 12% of control values. Treatment of both first trimester and term cells with 10(-6) M progestin, mineralocorticoid, or estrogen had no significant effect on onfFN expression in either cell type. Glucocorticoids specifically down-regulated medium levels of onfFN in term cells, but not in first trimester cells. In contrast, DEX treatment promoted an approximately 3- to 7-fold increase in levels of hCG in both first trimester and term cytotrophoblasts, suggesting that the effects of glucocorticoid on FN and hCG expression are elicited through independent cell-signaling pathways. In first trimester cells, DEX promoted a reduction in rates of FN and laminin synthesis to 60-70% of control levels. In term cells, DEX treatment reduced levels of FN and laminin synthesis to approximately 10% of control levels. Similarly, DEX treatment down-regulated levels of FN mRNA to approximately 60% and 10% of control values in first trimester and term cells, respectively. The first trimester of human pregnancy is associated with low levels of glucocorticoids and reduced glucocorticoid responsiveness. These conditions would favor high levels of placental ECM protein synthesis, thus stabilizing uterine-placental adherence. Conversely, elevated levels of glucocorticoids near parturition and increased glucocorticoid responsiveness would inhibit placental ECM protein synthesis, reducing uterine-placental adherence and promoting the separation of placenta from uterus.

Cells, Cultured↗

PubMed1999

Cytokines modulate MIA PaCa 2 and CAPAN-1 adhesion to extracellular matrix proteins.

Variations in cancer cell adhesion to extracellular matrix (ECM) proteins might underlie an enhanced metastatic potential. ECM binding is mediated by cell-adhesion molecules, the membrane expression of which might be influenced by soluble mediators, such as cytokines. The aims of our study were to ascertain whether epidermal growth factor (EGF), transforming growth factor beta1 (TGF-beta1), interleukin 1alpha (IL-1alpha), or interleukin 1beta (IL-1beta) can modify MIA PaCa 2 (pancreatic cancer cell line) and CAPAN-1 (metastatic pancreatic cancer cell line) adhesion to fibronectin, laminin, or type I collagen, and whether these cytokines can shift the membrane expression of the hyaluronic acid receptor (CD44). EGF significantly enhanced MIA PaCa 2, but not CAPAN-1, adhesion to fibronectin, laminin, and type I collagen. TGF-beta1 reduced MIA PaCa 2 adhesion to type I collagen, but enhanced CAPAN-1 adhesion to fibronectin and laminin. IL-1alpha was found to enhance MIA PaCa 2 adhesion to fibronectin, while reducing adhesion to type I collagen, whereas IL-1beta reduced the adhesion to laminin. IL-1alpha enhanced CAPAN-1 adhesion to laminin in a dose-dependent manner; IL-1beta slightly increased the adhesion of these cells to laminin at low dosage, and to type I collagen at high dosage. Both IL-1alpha and IL-1beta reduced CD44 membrane expression of MIA PaCa 2, while TGF-beta1 increased the percentage of CD44-positive CAPAN-1 cells. We suggest that the effects on cell adhesion induced by different cytokines depend on the status of the target pancreatic cancer cell. EGF and, in part, IL-1alpha can favor nonmetastatic pancreatic cancer cell adhesion to ECM, possibly favoring tumor spread. Metastatic cells seem to lose the responsiveness to EGF, while becoming hyperresponsive to IL-1alpha. TGF-beta1 might exert an antidiffusive effect on primary, and a prodiffusive effect on metastatic pancreatic cancer cells. Only IL-1alpha, IL-1beta, and TGF-beta1 seem to influence CD44 membrane expression. All the results presented in this study were obtained in vitro, and in vivo studies are needed to verify whether the studied cytokines can favor or counteract pancreatic cancer spread.

Cell Adhesion↗

PubMed2005

Comparative effects of TGF-beta 1 and TGF-beta 2 on extracellular matrix production, proliferation, migration, and collagen contraction of human Tenon's capsule fibroblasts in pseudoexfoliation and primary open-angle glaucoma.

PURPOSE: To comparatively investigate the effects of TGF-beta(1) and TGF-beta(2) on extracellular matrix production, proliferation, migration, and collagen contraction of cultured human Tenon's capsule fibroblasts derived from patients with pseudoexfoliation (PEX) syndrome, PEX glaucoma, primary open-angle glaucoma (POAG), and cataract. METHODS: Tenon's capsule fibroblasts obtained from four groups of patients were cultured and stimulated with different concentrations (0.1-10 ng ml(-1)) of TGF-beta(1) or TGF-beta(2) for up to 14 days. Cell proliferation was determined with the WST-1 colorimetric assay, cell migration by using the Transwell assay system, and collagen contraction by computerised analysis of three-dimensional collagen lattices and immunohistochemistry for alpha-smooth muscle actin expression. Expression and synthesis of extracellular matrix components (fibronectin, collagen types I and III) was assessed by enzyme-linked immunosorbent assays, by real-time RT-PCR, and by transmission electron microscopy. RESULTS: Both TGF-beta(1) and TGF-beta(2) in pathophysiological concentrations of 0.1-5 ng ml(-1) stimulated cell proliferation, migration, collagen contraction, alpha-smooth muscle actin expression as well as mRNA expression and secretion of fibronectin, collagen type I, and collagen type III by Tenon's fibroblasts derived from all groups of patients. TGF-beta stimulation occurred in a concentration-dependent manner with different peak activities associated with different fibroblast functions. There was some variability among the different groups of patients with an increased response of cells derived from PEX and POAG patients as compared to cataract patients. Although no statistically significant differences were found between both TGF-beta isoforms, TGF-beta(1) had a more pronounced stimulatory effect on expression and synthesis of extracellular matrix components including the production of elastic microfibrils, particularly in cells derived from patients with PEX syndrome/glaucoma. CONCLUSIONS: These findings suggest a significant contribution of TGF-beta(1) in addition to TGF-beta(2) to the conjunctival scarring process following glaucoma filtration surgery. Due to its pronounced fibrogenic potential, TGF-beta(1) may become another focus for targeting drug therapy, particularly in patients with PEX glaucoma.

Aged↗

PubMed2000

Gene expression of TGF-beta, TGF-beta receptor, and extracellular matrix proteins during membranous bone healing in rats.

Poorly healing mandibular fractures and osteotomies can be troublesome complications of craniomaxillofacial trauma and reconstructive surgery. Gene therapy may offer ways of enhancing bone formation by altering the expression of desired growth factors and extracellular matrix molecules. The elucidation of suitable candidate genes for therapeutic intervention necessitates investigation of the endogenously expressed patterns of growth factors during normal (i.e., successful) fracture repair. Transforming growth factor beta1 (TGF-beta1), its receptor (Tbeta-RII), and the extracellular matrix proteins osteocalcin and type I collagen are thought to be important in long-bone (endochondral) formation, fracture healing, and osteoblast proliferation. However, the spatial and temporal expression patterns of these molecules during membranous bone repair remain unknown. In this study, 24 adult rats underwent mandibular osteotomy with rigid external fixation. In addition, four identically treated rats that underwent sham operation (i.e., no osteotomy) were used as controls. Four experimental animals were then killed at each time point (3, 5, 7, 9, 23, and 37 days after the procedure) to examine gene expression of TGF-beta1 and Tbeta-RII, osteocalcin, and type I collagen. Northern blot analysis was used to compare gene expression of these molecules in experimental animals with that in control animals (i.e., nonosteotomized; n = 4). In addition, TGF-beta1 and T-RII proteins were immunolocalized in an additional group of nine animals killed on postoperative days 3, 7, and 37. The results of Northern blot analysis demonstrated a moderate increase (1.7 times) in TGF-beta1 expression 7 days postoperatively; TGF-beta1 expression returned thereafter to near baseline levels. Tbeta-RII mRNA expression was downregulated shortly after osteotomy but then increased, reaching a peak of 1.8 times the baseline level on postoperative day 9. Osteocalcin mRNA expression was dramatically downregulated shortly after osteotomy and remained low during the early phases of fracture repair. Osteocalcin expression trended slowly upward as healing continued, reaching peak expression by day 37 (1.7 times the control level). In contrast, collagen type IalphaI mRNA expression was acutely downregulated shortly after osteotomy, peaked on postoperative days 5, and then decreased at later time points. Histologic samples from animals killed 3 days after osteotomy demonstrated TGF-beta1 protein localized to inflammatory cells and extracellular matrix within the fracture gap, periosteum, and peripheral soft tissues. On postoperative day 7, TGF-beta1 staining was predominantly localized to the osteotomized bone edges, periosteum, surrounding soft tissues, and residual inflammatory cells. By postoperative day 37, complete bony healing was observed, and TGF-beta1 staining was localized to the newly formed bone matrix and areas of remodeling. On postoperative day 3, Tbeta-RII immunostaining localized to inflammatory cells within the fracture gap, periosteal cells, and surrounding soft tissues. By day 7, Tbeta-RII staining localized to osteoblasts of the fracture gap but was most intense within osteoblasts and mesenchymal cells of the osteotomized bone edges. On postoperative day 37, Tbeta-RII protein was seen in osteocytes, osteoblasts, and the newly formed periosteum in the remodeling bone. These observations agree with those of previous in vivo studies of endochondral bone formation, growth, and healing. In addition, these results implicate TGF-beta1 biological activity in the regulation of osteoblast migration, differentiation, and proliferation during mandibular fracture repair. Furthermore, comparison of these data with gene expression during mandibular distraction osteogenesis may provide useful insights into the treatment of poorly healing fractures because distraction osteogenesis has been shown to be effective in the management of these difficult clinical cases.

Animals↗

PubMed1990

Enhanced expression of the extracellular matrix molecule J1/tenascin in the regenerating adult mouse sciatic nerve.

We have investigated the expression of J1/tenascin in the sciatic nerve of the adult mouse under normal and regenerating conditions by immunocytological and immunochemical methods. In the normal nerve, J1/tenascin expression was confined to the extracellular matrix at the node of Ranvier and in the perineurium. At 2 days after nerve transection, J1/tenascin was detectable in the fibroblast-containing caps of the distal and proximal nerve stumps, in the distal nerve stump along its entire length and in the distal end of the proximal nerve stump. In the nerve stumps immunoreactivity was predominantly associated with extracellular matrix consisting of collagen fibrils and Schwann cell basal laminae. Approximately 7 days after transection, the caps of the nerve stumps had usually grown together forming a bridge. This bridge consisted of a J1/tenascin-negative perineurium-like structure and an inner part of predominantly fibroblasts, endothelial cells and macrophages. All cell types in this inner part were embedded in a J1/tenascin-positive matrix of collagen fibrils indicating the prospective direction of growth of neural elements. A few days later, J1/tenascin in the bridge was confined to the extracellular matrix around small Schwann cell-containing nerve fascicles. In nerves chronically denervated for 19 days, J1/tenascin was poorly detectable in the cap of the distal stump, although Schwann cells had infiltrated this cap. Approximately 19 days after the lesion, J1/tenascin expression returned to control levels in the proximal nerve stump. In the distal nerve stump, J1/tenascin immunoreactivity reached a peak at approximately 14 days after nerve transection and vanished only at approximately 35 days, thus correlating with the time of active regrowth of axons into the distal nerve stump. This reduction was prevented by chronic denervation, suggesting that reinnervation of target structures may be related to the down-regulation of J1/tenascin. These combined observations suggest that J1/tenascin is differentially regulated in the individual parts of the regenerating nerve, possibly triggered by different cellular and molecular signals.

Animals↗

PubMed2006

[Experimental study on protective effects of curcumin on exaggerated extracellular matrix accumulation of pulmonary fibrosis rats].

OBJECTIVE: To study the protective effects of curcumin on exaggerated extracellular matrix accumulation of pulmonary fibrosis rats. METHOD: One hundred and forty-four male Sprague-Dawley rats were randomly divided into 6 groups (24 rats in each group). Rats in the model control group, positive medicine group, and high, moderate and low curcumin groups were injected with a single dose of bleomycin by trachea, and rats in sham-model control group with same volume normal saline. One day after the injection, curcumin solution of different dosages (200, 100, 50 mg x kg(-1) x d(-1)) was respectively given to rats in the high, moderate and low curcumin group daily by gastrogavage, while equal volume of normal saline was given to those in the sham-model control group and model control group, and an equal volume of prednisone (0.56 mg x kg(-1) x d(-1)) was given to those in positive medicine control group. On the 7, 14, 28 days, 8 rats per treatment group were randomly killed, the levels of III-collagen, IV-collagen, laminin and hyaluronic acid in the serum were determined, the determination of hydroxyproline in lung homogenates was analyzed, and the lung was incised to make pathological sections which were stained with HE and Mallory. RESULT: Curcumin could decreas the levels of III-collagen, IV-collagen, laminin and hyaluronic acid in the serum, and inhihit the proliferation of fibrous tissue. CONCLUSION: Curcumin may play its therapetuic role by leveling down the content of extracellular matrix in rats with pulmonary fibrosis induced by bleomycin.

Animals↗

PubMed1992

Role of the extracellular matrix in age-related modifications of the rat aorta. Ultrastructural, morphometric, and enzymatic evaluations.

Connective tissues such as blood vessels are known to be greatly affected by age because of impaired functional properties and increased susceptibility to diseases. With the aim of providing further information on the role of the extracellular matrix in age-related modifications, we investigated the aorta in the rat model from birth to senescence by means of morphological and morphometric observations and by evaluation of lysyl oxidase activity. Results focused on the dramatic vascular rearrangements due to progressive fibrosis of the extracellular matrix and on prominent elastin modifications. The presence of lysyl oxidase activity, even in the oldest animals, might be at least partly responsible for the increased stiffness of the aging extracellular matrix. The striking age-related remodeling of the aortic architecture and the alterations of the interactions between cellular and extracellular compartments might greatly influence the functional properties of the arterial wall in senescence, at least contributing to the consequences of some apparently age-related vascular disorders.

Aging↗

PubMed1999

Three-dimensional environments preserve extracellular matrix compartments of ovarian follicles and increase FSH-dependent growth.

In this study we performed a systematic comparative analysis of two culture environments-flat/adhesive liquid and three-dimensional collagen gel-upon in vitro ovarian follicle development. We paid particular attention to the effects of in vitro environments upon the preservation of follicular structure and of peri- and intra-follicular extracellular matrix. We show that flat/adhesive environment leads to an obvious distortion of follicle morphology, marked extracellular matrix modifications and high rates of spontaneous, i.e., FSH-independent, follicle disruption. In contrast, three-dimensional collagen gel environments are able to maintain follicular structure with an in vivo-like basal lamina architecture, minimizing spontaneous disruption. Follicle distortions found in flat/adhesive culture systems include a pronounced flattening, causing the follicle horizontal diameters not to adequately reflect follicle volume. Our volume data, based on three-axis follicle diameter measurements, indicate that three-dimensional collagen gel environments increase follicle growth, particularly in response to FSH. This study demonstrates that preservation of both peri- and intra-follicular extracellular matrix compartments during the in vitro growth and differentiation of ovarian follicles is highly desirable, and is now possible through the use of appropriate three-dimensional collagen gel culture environments. This system allows a better understanding of the specific roles played by each of the follicle compartments during development.

Animals↗

PubMed1987

Immunoelectronmicroscopic localization of extracellular matrix components produced by bovine corneal endothelial cells in vitro.

Bovine corneal endothelial cells deposit an extracellular matrix in short-term cultures, which contains various morphologically distinct structures when analysed by electron microscopy after negative staining. Amongst these were long-spacing fibers with a 150 nm periodicity, which appeared also to be assembled into more complex hexagonal lattices. Another structure was fine filaments, 10-40 nm in diameter, which occasionally exhibited 67 nm periodic cross-striation. Non-striated 10-20 nm filaments sometimes formed radially oriented bundles arranged in networks and fuzzy granular material was associated with the filaments in the bundles. Often, these bundles extended into solitary filaments, 10-20 nm in diameter, with a smooth surface. In addition, amorphous patches were seen, which contained dense aggregates of fibrillar and granular material. In longer-term cultures, some of the structures coalesced to form large fibrillar bundles. By using specific antibodies to various extracellular matrix components and immunolabeling with gold some of these structures could be identified as to their protein composition. Whereas fibronectin antibodies labeled a variety of structures--fine filaments with granular materials, radially oriented bundles, patchy amorphous aggregates and small granular material scattered throughout the background--type III collagen antibody predominantly labeled filaments with periodic banding (10-40 nm in diameter). A small amount of type III specific labeling was also observed over the networks of radially oriented fibrils and fine filaments associated with granular material. Type IV collagen and laminin antibodies localized in areas of the patchy amorphous aggregates. Type VI collagen antibodies, on the other hand, labeled fine filaments and the gold particles showed a pattern of 100 nm periodicity. Many of the fine 10-20 nm filaments exhibited a tubular appearance on cross-section, but they were not reactive with any of the antibodies used. Also negative were the long-spacing fibers and assemblies--including hexagonal lattices--containing this structural element.

Animals↗