scholarly journals The extracellular matrix ligands fibronectin and tenascin collaborate in regulating collagenase gene expression in fibroblasts.

1994 ◽  
Vol 5 (4) ◽  
pp. 439-453 ◽  
Author(s):  
P Tremble ◽  
R Chiquet-Ehrismann ◽  
Z Werb
Keyword(s):  
Gene Expression ◽  
Extracellular Matrix ◽  
Type I Collagen ◽  
Serum Proteins ◽  
Synovial Fibroblasts ◽  
Type I ◽  
Mixed Substrate ◽  
Amino Terminal ◽  
Collagenase Gene ◽  
In Cells

Tenascin (TN) is a large oligomeric glycoprotein that is present transiently in the extracellular matrix (ECM) of cells and is involved in morphogenetic movements, tissue patterning, and tissue repair. It has multiple domains, both adhesive and anti-adhesive, that interact with cells and with fibronectin (FN) and other ECM macromolecules. We have studied the consequences of the interaction of TN with a FN matrix on gene expression in rabbit synovial fibroblasts. Fibroblasts plated on a mixed substrate of FN and TN, but not on FN alone, upregulated synthesis of four genes: collagenase, stromelysin, the 92-kDa gelatinase, and c-fos. Although the fibroblasts spread well on both FN and FN/TN substrates, nuclear c-Fos increased within 1 h only in cells that were plated on FN/TN. TN did not induce the expression of collagenase in cells plated on substrates of type I collagen or vitronectin (VN). Moreover, soluble TN added to cells adhering to a FN substrate or to serum proteins had no effect, suggesting that TN has an effect only in the context of mixed substrates of FN and TN. Collagenase increased within 4 h of plating on a FN/TN substrate and exhibited kinetics similar to those for induction of collagenase gene expression by signaling through the integrin FN receptor. Arg-Gly-Asp peptide ligands that recognize either the FN receptor or the VN receptor and function-perturbing anti-integrin monoclonal antibodies diminished the interaction of fibroblasts with a mixed substrate of FN, TN, and VN, but had no effect on the adhesion of fibroblasts to a substrate of FN and VN, suggesting that both receptors recognize the complex. Anti-TN68, an antibody that recognizes an epitope in the carboxyl-terminal type III repeats involved in the interaction of TN with both FN and cells, blocked the inductive effect of the FN/TN substrate, whereas anti-TNM1, an antibody that recognizes an epitope in the amino-terminal anti-adhesive region of epidermal growth factor-like repeats, had no effect. These data suggest that transient alteration of the composition of ECM by addition of proteins like TN may regulate the expression of genes involved in cell migration, tissue remodeling, and tissue invasion, in regions of tissue undergoing phenotypic changes.

Induction of vascular smooth muscle cell tenascin-C gene expression by denatured type I collagen is dependent upon a beta3 integrin-mediated mitogen-activated protein kinase pathway and a 122-base pair promoter element

1999 ◽  
Vol 112 (4) ◽  
pp. 435-445 ◽  
Author(s):  
P.L. Jones ◽  
F.S. Jones ◽  
B. Zhou ◽  
M. Rabinovitch
Keyword(s):  
Gene Expression ◽  
Extracellular Matrix ◽  
Smooth Muscle ◽  
Smooth Muscle Cell ◽  
Muscle Cell ◽  
Type I Collagen ◽  
Type I ◽  
C Protein ◽  
Tenascin C ◽  
Beta3 Integrin

Tenascin-C is an extracellular matrix glycoprotein, the expression of which is upregulated in remodeling arteries. In previous studies we showed that the presence of tenascin-C alters vascular smooth muscle cell shape and amplifies their proliferative response by promoting growth factor receptor clustering and phosphorylation. Moreover, we demonstrated that denatured type I collagen induces smooth muscle cell tenascin-C protein production via beta3 integrins. In the present study, we examine the pathway by which beta3 integrins stimulate expression of tenascin-C, and define a promoter sequence that is critical for its induction. On native collagen, A10 smooth muscle cells adopt a stellate morphology and produce low levels of tenascin-C mRNA and protein, whereas on denatured collagen they spread extensively and produce high levels of tenascin-C mRNA and protein, which is incorporated into an elaborate extracellular matrix. Increased tenascin-C synthesis on denatured collagen is associated with elevated protein tyrosine phosphorylation, including activation of extracellular signal-regulated kinases 1 and 2 (ERK1 and ERK2). beta3 integrin function-blocking antibodies attenuate ERK1/2 activation and tenascin-C protein synthesis. Consistent with these findings, treatment with the specific MEK inhibitor, PD 98059, results in suppression of tenascin-C protein synthesis. To investigate whether beta3 integrin-dependent activation of ERK1/2 regulates the tenascin-C promoter, we transfected A10 cells with a full-length (approx. 4 kb) mouse tenascin-C gene promoter-chloramphenicol acetyltransferse reporter construct and showed that, relative to native collagen, its activity is increased on denatured collagen. Next, to identify regions of the promoter involved, we examined a series of tenascin-C promoter constructs with 5′ deletions and showed that denatured collagen-dependent promoter activity was retained by a 122-base pair element, located -43 to -165 bp upstream of the RNA start site. Activation of this element was suppressed either by blocking beta3 integrins, or by preventing ERK1/2 activation. These observations demonstrate that smooth muscle cell binding to beta3 integrins activates the mitogen activated protein kinase pathway, which is required for the induction of tenascin-C gene expression via a potential extracellular matrix response element in the tenascin-C gene promoter. Our data suggest a mechanism by which remodeling of type I collagen modulates tenascin-C gene expression via a beta3 integrin-mediated signaling pathway, and as such represents a paradigm for vascular development and disease whereby smooth muscle cells respond to perturbations in extracellular matrix composition by altering their phenotype and patterns of gene expression.

scholarly journals Components of the nuclear signaling cascade that regulate collagenase gene expression in response to integrin-derived signals.

1995 ◽  
Vol 129 (6) ◽  
pp. 1707-1720 ◽  
Author(s):  
P Tremble ◽  
C H Damsky ◽  
Z Werb
Keyword(s):  
Gene Expression ◽  
Synovial Fibroblasts ◽  
Plasma Fibronectin ◽  
Reporter Construct ◽  
Fibronectin Receptor ◽  
Receptor Antibody ◽  
Fibronectin Fragment ◽  
Integrin Fibronectin Receptor ◽  
Collagenase Gene ◽  
In Cells

We have shown previously that the expression of collagenase is upregulated in rabbit synovial fibroblasts cultured on a substrate of antibody to the alpha 5 chain of the alpha 5 beta 1 integrin fibronectin receptor or on the 120-kD cell-binding chymotryptic fragment of plasma fibronectin, but remains at basal levels in cells plated on intact plasma fibronectin. We now have identified some of the components of a signaling pathway that couples the fibronectin receptor to the induction of collagenase transcription. We studied the control of collagenase gene expression in cells adhering to the 120-kD fragment of fibronectin, to antifibronectin receptor antibody, or to plasma fibronectin by transiently introducing promoter-reporter constructs into rabbit synovial fibroblasts before plating cells on these matrices. The constructs contained segments of the human collagenase promoter regulating transcription of chloramphenicol acyl transferase. Expression of constructs containing the -1200/-42-bp segment or the -139/-42-bp segment of the collagenase promoter inserted upstream from the reporter gene was induced to similar extents in cells plated on the 120-kD fragment of fibronectin or on anti-fibronectin receptor antibody, relative to that in fibroblasts plated on fibronectin. The expression of the construct containing the -66/-42-bp segment of the promoter was not regulated and was similar to that of the parent pBLCAT2 plasmid, suggesting that the -139/-67 region of the collagenase promoter, which contains PEA3- and AP1-binding sites, regulates the transcription of collagenase caused by integrin-derived signals. Expression of a reporter construct containing only the PEA3 and AP1 sites in the collagenase promoter (-90/-67) also increased in cells plated on the 120-kD fragment of fibronectin or on anti-fibronectin receptor antibody, relative to that in cells plated on fibronectin. Mutations in either the AP1 or PEA3 site of this minimal promoter abrogated its activity in cells plated on these inductive ligands. Expression of c-fos mRNA increased within 1 h of plating cells on the 120-kD fibronectin fragment or on anti-fibronectin receptor antibody, relative to that in cells plated on fibronectin. c-Fos protein accumulated in the nuclei of fibroblasts within 10 min of plating on the 120-kD fibronectin fragment. The increase in c-Fos was required for the increase in collagenase in cells plated on the 120-kD fibronectin fragment: incubation of cells with antisense, but not sense, c-fos oligonucleotides diminished both basal and induced expression of the -139/-42 collagenase promoter-reporter construct and decreased expression of the endogenous collagenase gene.(ABSTRACT TRUNCATED AT 400 WORDS)

scholarly journals Platelet-derived growth factors-AA and -BB regulate collagen and collagenase gene expression differentially in human fibroblasts

1995 ◽  
Vol 310 (2) ◽  
pp. 585-588 ◽  
Author(s):  
E M L Tan ◽  
H Qin ◽  
S H Kennedy ◽  
S Rouda ◽  
J W Fox ◽  
...  
Keyword(s):  
Gene Expression ◽  
Time Course ◽  
Type I Collagen ◽  
Biological Activities ◽  
Human Fibroblasts ◽  
Dermal Fibroblasts ◽  
Type I ◽  
Mrna Levels ◽  
Interstitial Collagenase ◽  
Collagenase Gene

Platelet-derived growth factor (PDGF) is a mitogen associated with tissue repair, a process involving collagen synthesis and remodelling by interstitial collagenase. This study examines and compares the regulation of interstitial collagenase and collagen gene expression by PDGF-AA and -BB in human fibroblasts. Time-course analysis showed that neither PDGF-AA or -BB had a consistent effect on the expression of pro-alpha 1(I) or pro-alpha 2(I) type-I collagen genes. In contrast, interstitial collagenase gene expression was found to be consistently up-regulated severalfold by PDGF-BB. Enhanced expression of the collagenase gene was not apparently due to up-regulation of its promoter activity in human dermal fibroblasts, as indicated by transient and stable transfection experiments. Unlike PDGF-BB, PDGF-AA did not alter collagenase mRNA levels under low-serum culture conditions. Thus, the biological activities of the PDGF homodimers are different, with PDGF-BB being clearly more potent than PDGF-AA in its regulation of collagenase gene expression.

scholarly journals Dermal fibroblasts have different extracellular matrix profiles induced by TGF-β, PDGF and IL-6 in a model for skin fibrosis

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Pernille Juhl ◽  
Sandie Bondesen ◽  
Clare Louise Hawkins ◽  
Morten Asser Karsdal ◽  
Anne-Christine Bay-Jensen ◽  
...  
Keyword(s):  
Gene Expression ◽  
Extracellular Matrix ◽  
Type I Collagen ◽  
Skin Diseases ◽  
Dermal Fibroblast ◽  
Dermal Fibroblasts ◽  
Type I ◽  
Healthy Human ◽  
Clinical Model ◽  
Collagen Formation

Abstract Different stimulants might induce different extracellular matrix profiles. It is essential to gain an understanding and quantification of these changes to allow for focused anti-fibrotic drug development. This study investigated the expression of extracellular matrix by dermal fibroblast mimicking fibrotic skin diseases as SSc using clinically validated biomarkers. Primary healthy human dermal fibroblasts were grown in media containing FICOLL. The cells were stimulated with PDGF-AB, TGF-β1, or IL-6. Anti-fibrotic compounds (iALK-5, Nintedanib) were added together with growth factors. Biomarkers of collagen formation and degradation together with fibronectin were evaluated by ELISAs in the collected supernatant. Immunohistochemical staining was performed to visualize fibroblasts and proteins, while selected gene expression levels were examined through qPCR. TGF-β and PDGF, and to a lesser extent IL-6, increased the metabolic activity of the fibroblasts. TGF-β primarily increased type I collagen and fibronectin protein and gene expression together with αSMA. PDGF stimulation resulted in increased type III and VI collagen formation and gene expression. IL-6 decreased fibronectin levels. iALK5 could inhibit TGF-β induced fibrosis while nintedanib could halt fibrosis induced by TGF-β or PDGF. Tocilizumab could not inhibit fibrosis induced in this model. The extent and nature of fibrosis are dependent on the stimulant. The model has potential as a pre-clinical model as the fibroblasts fibrotic phenotype could be reversed by an ALK5 inhibitor and Nintedanib.

scholarly journals Extracellular matrix controls tubulin monomer levels in hepatocytes by regulating protein turnover.

1994 ◽  
Vol 5 (12) ◽  
pp. 1281-1288 ◽  
Author(s):  
D J Mooney ◽  
L K Hansen ◽  
R Langer ◽  
J P Vacanti ◽  
D E Ingber
Keyword(s):  
Extracellular Matrix ◽  
Cell Shape ◽  
Type I Collagen ◽  
Cell Attachment ◽  
Rat Hepatocytes ◽  
Defined Medium ◽  
Type I ◽  
Mrna Levels ◽  
Mechanical Loads ◽  
In Cells

Cells have evolved an autoregulatory mechanism to dampen variations in the concentration of tubulin monomer that is available to polymerize into microtubules (MTs), a process that is known as tubulin autoregulation. However, thermodynamic analysis of MT polymerization predicts that the concentration of free tubulin monomer must vary if MTs are to remain stable under different mechanical loads that result from changes in cell adhesion to the extracellular matrix (ECM). To determine how these seemingly contradictory regulatory mechanisms coexist in cells, we measured changes in the masses of tubulin monomer and polymer that resulted from altering cell-ECM contacts. Primary rat hepatocytes were cultured in chemically defined medium on bacteriological petri dishes that were precoated with different densities of laminin (LM). Increasing the LM density from low to high (1-1000 ng/cm2), promoted cell spreading (average projected cell area increased from 1200 to 6000 microns2) and resulted in formation of a greatly extended MT network. Nevertheless, the steady-state mass of tubulin polymer was similar at 48 h, regardless of cell shape or ECM density. In contrast, round hepatocytes on low LM contained a threefold higher mass of tubulin monomer when compared with spread cells on high LM. Furthermore, similar results were obtained whether LM, fibronectin, or type I collagen were used for cell attachment. Tubulin autoregulation appeared to function normally in these cells because tubulin mRNA levels and protein synthetic rates were greatly depressed in round cells that contained the highest level of free tubulin monomer. However, the rate of tubulin protein degradation slowed, causing the tubulin half-life to increase from approximately 24 to 55 h as the LM density was lowered from high to low and cell rounding was promoted. These results indicate that the set-point for the tubulin monomer mass in hepatocytes can be regulated by altering the density of ECM contacts and changing cell shape. This finding is consistent with a mechanism of MT regulation in which the ECM stabilizes MTs by both accepting transfer of mechanical loads and altering tubulin degradation in cells that continue to autoregulate tubulin synthesis.

scholarly journals Interactions of thrombospondin with extracellular matrix proteins: selective binding to type V collagen.

1984 ◽  
Vol 98 (2) ◽  
pp. 646-652 ◽  
Author(s):  
S M Mumby ◽  
G J Raugi ◽  
P Bornstein
Keyword(s):  
Extracellular Matrix ◽  
Endothelial Cells ◽  
Immunosorbent Assay ◽  
Type I Collagen ◽  
Cultured Cells ◽  
Serum Proteins ◽  
Enzyme Linked Immunosorbent Assay ◽  
Type I ◽  
Type V Collagen ◽  
Type V

Thrombospondin (TS), a protein first described in platelets, was recently shown to be synthesized and secreted by endothelial cells, fibroblasts, and smooth muscle cells. The presence of TS in the extracellular matrix of cultured cells has prompted us to examine the associations of this protein with matrix macromolecules. Interactions of TS with both matrix and serum proteins were tested using an enzyme-linked immunosorbent assay. With this assay we assessed the binding of TS in solution to proteins adsorbed to polystyrene microtiter plates. Among collagens, platelet TS bound to type V but not to types I, III, or IV. This selective interaction was confirmed in experiments using proteins linked to cyanogen bromide-activated Sepharose. TS released from platelets in response to thrombin activation, as well as that secreted by endothelial cells in culture, bound to type V but not to type I collagen-Sepharose. No binding was observed to denatured type V collagen-Sepharose. The binding region for type V collagen was located in a chymotrypsin-produced fragment of TS with chains of Mr = 70,000, after reduction. Interactions of TS with a number of other proteins, including fibronectin, fibrinogen, and laminin, could be demonstrated using the enzyme-linked immunosorbent assay technique but the interpretation of these findings is difficult since comparable binding to protein-Sepharose was not always observed. Our findings suggest that both the extravascular distribution and function of TS in vivo may involve an interaction with type V collagen.

scholarly journals THE FEATURES OF THE FACE SKIN CONSTRUCTION THAT INFLUENCE ON THE FORMATION OF CICATRICAL TISSUES DURING SUGICAL INTERVENTIONS

2019 ◽  
pp. 19-23
Author(s):  
K.P. Lokes ◽  
D.S. Avetikov ◽  
S.O. Stavitsky ◽  
O.O. Rozkolupa ◽  
N.S. Lutsenko
Keyword(s):  
Extracellular Matrix ◽  
Head And Neck ◽  
Type I Collagen ◽  
Cell Attachment ◽  
Rapid Development ◽  
Maxillofacial Surgery ◽  
Type Iii Collagen ◽  
Type I ◽  
Amino Terminal ◽  
Functional Features

Formation of pathological scars of maxillofacial localization after surgery is a significant and widespread problem of modern surgical stomatology and maxillofacial surgery. A significant percentage of patients who needs planned and urgent surgical interventions cause rapid development of reconstructive-restorative surgery of the maxillofacial region. The analysis of domestic and foreign literary sources was devoted to the peculiarities of the structure of the skin of the head and neck and the optimization of the skin incisions of this localization. Functional features of human skin depend on the mechanical properties of the dermis, which provides elasticity and resistance to stretching. Changes in the biomechanics of the dermis occur during aging, excessive insolation, scarring, and fibrosis. In addition, mechanical changes in the extracellular matrix of the skin affect the activity and phenotype of the fibroblasts, which adapt the stiffness of the cytoskeleton. Extracellular matrix stiffness defines and maintains cell identity and influences the proliferation, differentiation, migration and expression of skin cells. The extracellular matrix has been regarded for a long time as a structure with simple architectonics. But, due to modern research, it is known that this complex formation is highly specialized. The different classes of macromolecules that make up the extracellular matrix determine its biological functions. For example, collagen proteins are responsible for the tensile strength of tissues, proteoglycans and glycosaminoglycan are important for hydration and compression resistance, and glycoproteins such as laminas facilitate cell attachment. The largest structures of the extracellular matrix are elastin fibers, which are mainly localized in tissues subject to high mechanical stress, such as skin, lungs, or arteries. These structures represent a very complex organization whose core consists of elastin surrounded by a mantle of microfibrils. Collagen proteins in the dermis contain mainly type I collagen (85% - 90%) with smaller amounts of type III collagen (10% - 15%). Skin fibroblasts synthesize individual collagen type I and III polypeptide chains as precursor molecules, called procollagens. During the formation of insoluble collagen fibrils, specific proteases break down the carboxy- and amino-terminal domains, forming pN-collagen (procollagen from which the carboxy-terminal domain propeptide is cleaved) and pC-collagen (procollagen, from which the amino-terminal propeptide is cleaved). Because type I and III procollagen, pN-collagen and pC-collagen are precursors of mature collagen molecules, their level usually reflects the level of collagen biosynthesis. Collagens and elastin contain highly abundant fibrils, each of which is repeated in a sequence enriched in the conformation of polyproline II, cross-linked, insoluble in assembly and resistant to the most photolytic enzymes. The monomeric block of type I collagen consists of two extended chains α1 and one chain α2, twisted together into a triple helix. The direction of collagen and elastin fibers, according to biomechanical studies, has a significant effect on the enlargement of the wound on the head skin and the tension when closing its edges. The overwhelming reduction of tension and accordingly the improvement of reparative processes in the skin occur when the incision lines correspond to the so-called "golden spiral". Conclusion. Thus, the analysis of domestic and foreign literature sources indicates the relevance of the selected topics, the need for further studies on the biomechanical and histological substantiation of incisions, which are due to the peculiarities of the structure of the skin in the head and neck to create optimal conditions for reparative regeneration.

scholarly journals The Dynamic Interaction between Extracellular Matrix Remodeling and Breast Tumor Progression

Cells ◽  
2021 ◽  
Vol 10 (5) ◽  
pp. 1046
Author(s):  
Jorge Martinez ◽  
Patricio C. Smith
Keyword(s):  
Extracellular Matrix ◽  
Type I Collagen ◽  
Cell Metabolism ◽  
Breast Tumors ◽  
Extracellular Matrix Remodeling ◽  
Type I ◽  
Matrix Remodeling ◽  
Desmoplastic Reaction ◽  
The Impact

Desmoplastic tumors correspond to a unique tissue structure characterized by the abnormal deposition of extracellular matrix. Breast tumors are a typical example of this type of lesion, a property that allows its palpation and early detection. Fibrillar type I collagen is a major component of tumor desmoplasia and its accumulation is causally linked to tumor cell survival and metastasis. For many years, the desmoplastic phenomenon was considered to be a reaction and response of the host tissue against tumor cells and, accordingly, designated as “desmoplastic reaction”. This notion has been challenged in the last decades when desmoplastic tissue was detected in breast tissue in the absence of tumor. This finding suggests that desmoplasia is a preexisting condition that stimulates the development of a malignant phenotype. With this perspective, in the present review, we analyze the role of extracellular matrix remodeling in the development of the desmoplastic response. Importantly, during the discussion, we also analyze the impact of obesity and cell metabolism as critical drivers of tissue remodeling during the development of desmoplasia. New knowledge derived from the dynamic remodeling of the extracellular matrix may lead to novel targets of interest for early diagnosis or therapy in the context of breast tumors.

scholarly journals Cruciate Ligament Cell Sheets Can Be Rapidly Produced on Thermoresponsive poly(glycidyl ether) Coating and Successfully Used for Colonization of Embroidered Scaffolds

Cells ◽  
2021 ◽  
Vol 10 (4) ◽  
pp. 877
Author(s):  
Ingrid Zahn ◽  
Daniel David Stöbener ◽  
Marie Weinhart ◽  
Clemens Gögele ◽  
Annette Breier ◽  
...  
Keyword(s):  
Gene Expression ◽  
Type I Collagen ◽  
Cruciate Ligament ◽  
Glycidyl Ether ◽  
Stable Gene ◽  
Type I ◽  
Cell Sheets ◽  
Thermoresponsive Polymers ◽  
Related Phenotype ◽  
Anterior Cruciate

Anterior cruciate ligament (ACL) cell sheets combined with biomechanically competent scaffolds might facilitate ACL tissue engineering. Since thermoresponsive polymers allow a rapid enzyme-free detachment of cell sheets, we evaluated the applicability of a thermoresponsive poly(glycidyl ether) (PGE) coating for cruciate ligamentocyte sheet formation and its influence on ligamentocyte phenotype during sheet-mediated colonization of embroidered scaffolds. Ligamentocytes were seeded on surfaces either coated with PGE or without coating. Detached ligamentocyte sheets were cultured separately or wrapped around an embroidered scaffold made of polylactide acid (PLA) and poly(lactic-co-ε-caprolactone) (P(LA-CL)) threads functionalized by gas-phase fluorination and with collagen foam. Ligamentocyte viability, protein and gene expression were determined in sheets detached from surfaces with or without PGE coating, scaffolds seeded with sheets from PGE-coated plates and the respective monolayers. Stable and vital ligamentocyte sheets could be produced within 24 h with both surfaces, but more rapidly with PGE coating. PGE did not affect ligamentocyte phenotype. Scaffolds could be colonized with sheets associated with high cell survival, stable gene expression of ligament-related type I collagen, decorin, tenascin C and Mohawk after 14 d and extracellular matrix (ECM) deposition. PGE coating facilitates ligamentocyte sheet formation, and sheets colonizing the scaffolds displayed a ligament-related phenotype.

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