scholarly journals Collagen synthesis by human fibroblasts. Regulation by transforming growth factor-β in the presence of other inflammatory mediators

1989 ◽  
Vol 260 (2) ◽  
pp. 463-469 ◽  
Author(s):  
A S Narayanan ◽  
R C Page ◽  
J Swanson
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor Beta ◽  
Collagen Synthesis ◽  
Transforming Growth Factor ◽  
Human Fibroblasts ◽  
Transforming Growth Factor Β ◽  
Tgf Beta ◽  
Collagen Production ◽  
Collagen Mrna ◽  
In Cells

We have examined the combined effects of transforming growth factor-beta (TGF-beta), serum and gamma-interferon (gamma-IFN) on collagen synthesis by fibroblasts and compared the response of fibroblast subpopulations to TGF-beta. Human diploid fibroblasts were treated with TGF-beta alone and with serum of gamma-IFN. Cells were labelled with radioactive amino acids, and collagen production was measured as collagenase-digestible radioactivity. Collagen mRNA was determined by a solution-hybridization assay using procollagen-alpha 1[I] cDNA clone HF 677. The results showed that either serum or TGF-beta increased incorporation, collagen production and mRNA by fibroblasts approx. 2-fold; however, collagen synthesis relative to total protein synthesis and collagen mRNA relative to total polyadenylated [poly(A)+] RNA were not affected. Only serum activated cell growth. Collagen production increased approx. 4-fold in cells exposed to both TGF-beta and serum, and this increase was equal to that expected for an additive effect by both components. Treatment with gamma-IFN decreased collagen production and collagen mRNA to 44 and 40% respectively, whereas total incorporation and poly(A)+ RNA were affected only marginally. Cells exposed simultaneously to both gamma-IFN and TGF-beta produced less collagen and contained less mRNA than did those treated with TGF-beta alone. The gamma-IFN decreased collagen synthesis in control and TGF-beta-treated cultures to a similar extent, and TGF-beta increased collagen synthesis 2-fold in cells pre-treated with gamma-IFN. Fibroblast strains obtained in medium containing plasma-derived serum synthesized approximately half as much collagen as did cells derived from the same explant in the presence of fresh human serum, and TGF-beta stimulated collagen production and mRNA in both cell strains. We conclude that TGF-beta, serum and gamma-IFN regulate collagen synthesis by independent mechanisms, and that the combined action of these components plays a significant role in regulating collagen synthesis during wound healing and tissue repair.

scholarly journals Stimulation of the chemotactic migration of human fibroblasts by transforming growth factor beta.

1987 ◽  
Vol 165 (1) ◽  
pp. 251-256 ◽  
Author(s):  
A E Postlethwaite ◽  
J Keski-Oja ◽  
H L Moses ◽  
A H Kang
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Human Fibroblasts ◽  
Critical Role ◽  
Dermal Fibroblasts ◽  
Type I ◽  
Tgf Beta ◽  
Chemotactic Migration ◽  
Growth Factor Beta

Transforming growth factor beta (TGF-beta) is a potent chemoattractant in vitro for human dermal fibroblasts. Intact disulfide and perhaps the dimeric structure of TGF-beta is essential for its ability to stimulate chemotactic migration of fibroblasts, since reduction with 2-ME results in a marked loss of its potency as a chemoattractant. Although epidermal growth factor (EGF) appears to be capable of modulating some effects of TGF-beta, it does not alter the chemotactic response of fibroblasts to TGF-beta. Specific polyvalent rabbit antibodies to homogeneously pure TGF-beta block its chemotactic activity but has no effect on the other chemoattractants tested (platelet-derived growth factor, fibronectin, and denatured type I collagen). Since TGF-beta is secreted by a variety of neoplastic and normal cells including platelets, monocytes/macrophages, and lymphocytes, it may play a critical role in vivo in embryogenesis, host response to tumors, and the repair response that follows damage to tissues by immune and nonimmune reactions.

Transforming growth factor beta increases cell surface binding and assembly of exogenous (plasma) fibronectin by normal human fibroblasts

1988 ◽  
Vol 8 (10) ◽  
pp. 4234-4242
Author(s):  
B L Allen-Hoffmann ◽  
C L Crankshaw ◽  
D F Mosher
Keyword(s):  
Extracellular Matrix ◽  
Growth Factor ◽  
Cell Surface ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Human Fibroblasts ◽  
Tgf Beta ◽  
Surface Binding ◽  
Amino Terminal ◽  
Cell Surface Binding

Transforming growth factor beta (TGF-beta) enhances the cell surface binding of 125I-fibronectin by cultured human fibroblasts. The effect of TGF-beta on cell surface binding was maximal after 2 h of exposure to TFG-beta and did not require epidermal growth factor or protein synthesis. The enhancement was dose dependent and was found with the 125I-labeled 70-kilodalton amino-terminal fragment of fibronectin as well as with 125I-fibronectin. Treatment of cultures with TGF-beta for 6 h resulted in a threefold increase in the estimated number of fibronectin binding sites. The increase in number of binding sites was accompanied by an increased accumulation of labeled fibronectin in detergent-insoluble extracellular matrix. The effect of TGF-beta was biphasic; after 6 h of exposure, less labeled fibronectin bound to treated cultures than to control cultures. Exposure of cells to TGF-beta for greater than 6 h caused a two- to threefold increase in the accumulation of cellular fibronectin in culture medium as detected by a quantitative enzyme-linked immunosorbent assay. The second phase of the biphasic effect and the increase in soluble cellular fibronectin were blocked by cycloheximide. Immunofluorescence staining of fibroblast cultures with antifibronectin revealed that TGF-beta caused a striking increase in fibronectin fibrils. The 70-kilodalton amino-terminal fragment of fibronectin, which blocks incorporation of fibronectin into extracellular matrix, blocked anchorage-independent growth of NRK-49F cells in the presence of epidermal growth factor. Our results show that an increase in the binding and rate of assembly of exogenous fibronectin is an early event preceding the increase in expression of extracellular matrix proteins. Such an early increase in cell surface binding of exogenous fibronectin may be a mechanism whereby TGF-beta can modify extracellular matrix characteristics rapidly after tissue injury or during embryonic morphogenesis.

scholarly journals The types II and III transforming growth factor-beta receptors form homo-oligomers.

1994 ◽  
Vol 126 (1) ◽  
pp. 139-154 ◽  
Author(s):  
Y I Henis ◽  
A Moustakas ◽  
H Y Lin ◽  
H F Lodish
Keyword(s):  
Growth Factor ◽  
Cell Surface ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Type Ii ◽  
Tgf Beta ◽  
Double Labeling ◽  
Beta Receptors ◽  
Growth Factor Beta ◽  
In Cells

Affinity-labeling experiments have detected hetero-oligomers of the types I, II, and III transforming growth factor beta (TGF-beta) receptors which mediate intracellular signaling by TGF-beta, but the oligomeric state of the individual receptor types remains unknown. Here we use two types of experiments to show that a major portion of the receptor types II and III forms homo-oligomers both in the absence and presence of TGF-beta. Both experiments used COS-7 cells co-transfected with combinations of these receptors carrying different epitope tags at their extracellular termini. In immunoprecipitation experiments, radiolabeled TGF-beta was bound and cross-linked to cells co-expressing two differently tagged type II receptors. Sequential immunoprecipitations using anti-epitope monoclonal antibodies showed that type II TGF-beta receptors form homo-oligomers. In cells co-expressing epitope-tagged types II and III receptors, a low level of co-precipitation of the ligand-labeled receptors was observed, indicating that some hetero-oligomers of the types II and III receptors exist in the presence of ligand. Antibody-mediated cross-linking studies based on double-labeling immunofluorescence explored co-patching of the receptors at the cell surface on live cells. In cells co-expressing two differently tagged type II receptors or two differently tagged type III receptors, forcing one receptor into micropatches by IgG induced co-patching of the receptor carrying the other tag, labeled by noncross-linking monovalent Fab'. These studies showed that homo-oligomers of the types II and III receptors exist on the cell surface in the absence or presence of TGF-beta 1 or -beta 2. In cells co-expressing types II and III receptors, the amount of heterocomplexes at the cell surface was too low to be detected in the immunofluorescence co-patching experiments, confirming that hetero-oligomers of the types II and III receptors are minor and probably transient species.

scholarly journals Phosphorylation of the human-transforming-growth-factor-β-binding protein endoglin

1994 ◽  
Vol 301 (3) ◽  
pp. 765-768 ◽  
Author(s):  
P Lastres ◽  
J Martín-Perez ◽  
C Langa ◽  
C Bernabéu
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β ◽  
Mouse Fibroblast ◽  
Tgf Beta ◽  
Kinase C ◽  
Protein Kinase C Inhibitor ◽  
A Minor ◽  
Minor Extent

Endoglin is an homodimeric membrane antigen with capacity to bind transforming growth factor-beta (TGF-beta). Phosphorylation of human endoglin was demonstrated in endothelial cells as well as in mouse fibroblast transfectants expressing two isoforms, L-endoglin or S-endoglin, with distinct cytoplasmic domains. The extent of L-endoglin phosphorylation was found to be 8-fold higher than that of S-endoglin, and phosphopeptide analyses revealed at least three different phosphorylation sites for L-endoglin, whereas S-endoglin produces only one phosphopeptide. The immunoprecipitated L-endoglin was found to be phosphorylated mainly on serine, and, to a minor extent, on threonine, residues. Treatment of the cells with TGF-beta 1 or the protein kinase C inhibitor H-7 resulted in a reduction of the levels of endoglin phosphorylation.

scholarly journals Transforming growth factor beta increases cell surface binding and assembly of exogenous (plasma) fibronectin by normal human fibroblasts.

1988 ◽  
Vol 8 (10) ◽  
pp. 4234-4242 ◽  
Author(s):  
B L Allen-Hoffmann ◽  
C L Crankshaw ◽  
D F Mosher
Keyword(s):  
Extracellular Matrix ◽  
Growth Factor ◽  
Cell Surface ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Human Fibroblasts ◽  
Tgf Beta ◽  
Surface Binding ◽  
Amino Terminal ◽  
Cell Surface Binding

Transforming growth factor beta (TGF-beta) enhances the cell surface binding of 125I-fibronectin by cultured human fibroblasts. The effect of TGF-beta on cell surface binding was maximal after 2 h of exposure to TFG-beta and did not require epidermal growth factor or protein synthesis. The enhancement was dose dependent and was found with the 125I-labeled 70-kilodalton amino-terminal fragment of fibronectin as well as with 125I-fibronectin. Treatment of cultures with TGF-beta for 6 h resulted in a threefold increase in the estimated number of fibronectin binding sites. The increase in number of binding sites was accompanied by an increased accumulation of labeled fibronectin in detergent-insoluble extracellular matrix. The effect of TGF-beta was biphasic; after 6 h of exposure, less labeled fibronectin bound to treated cultures than to control cultures. Exposure of cells to TGF-beta for greater than 6 h caused a two- to threefold increase in the accumulation of cellular fibronectin in culture medium as detected by a quantitative enzyme-linked immunosorbent assay. The second phase of the biphasic effect and the increase in soluble cellular fibronectin were blocked by cycloheximide. Immunofluorescence staining of fibroblast cultures with antifibronectin revealed that TGF-beta caused a striking increase in fibronectin fibrils. The 70-kilodalton amino-terminal fragment of fibronectin, which blocks incorporation of fibronectin into extracellular matrix, blocked anchorage-independent growth of NRK-49F cells in the presence of epidermal growth factor. Our results show that an increase in the binding and rate of assembly of exogenous fibronectin is an early event preceding the increase in expression of extracellular matrix proteins. Such an early increase in cell surface binding of exogenous fibronectin may be a mechanism whereby TGF-beta can modify extracellular matrix characteristics rapidly after tissue injury or during embryonic morphogenesis.

scholarly journals Macrophage production of transforming growth factor beta and fibroblast collagen synthesis in chronic pulmonary inflammation.

1989 ◽  
Vol 170 (3) ◽  
pp. 727-737 ◽  
Author(s):  
N Khalil ◽  
O Bereznay ◽  
M Sporn ◽  
A H Greenberg
Keyword(s):  
Extracellular Matrix ◽  
Growth Factor ◽  
Transforming Growth Factor Beta ◽  
Collagen Synthesis ◽  
Transforming Growth Factor ◽  
Pulmonary Inflammation ◽  
Late Phase ◽  
Primary Role ◽  
Tgf Beta ◽  
Growth Factor Beta

A rat model of bleomycin-induced pulmonary inflammation and fibrosis was used to examine the relationship between collagen synthesis and transforming growth factor beta (TGF-beta) production, and cellular distribution. Total lung TGF-beta was elevated within 2 h of intratracheal bleomycin administration and peaked 7 d later at levels 30-fold higher than controls. This was followed by a gradual decline with lower but persistent levels of production in the late phase of the response between 21 and 28 d later. The peak TGF-beta levels preceded the maximum collagen and noncollagen protein synthesis measured by [3H]proline incorporation into lung fibroblast explants of bleomycin-treated rats. The pattern of immunohistochemical staining localized TGF-beta initially in the cytoplasm of bronchiolar epithelium cells and subepithelial extracellular matrix. The peak of lung TGF-beta levels at 7 d coincided with intense TGF-beta staining of macrophages dispersed in the alveolar interstitium and in organized clusters. Later in the course of the response. TGF-beta was primarily associated with extracellular matrix in regions of increased cellularity and tissue repair, and coincided with the maximum fibroblast collagen synthesis. This temporal and spatial relationship between collagen production and TGF-beta production by macrophages suggests an important if not primary role for TGF-beta in the pathogenesis of the pulmonary fibrosis.

scholarly journals Inhibition of Nb2 T-lymphoma cell growth by transforming growth factor-β

1988 ◽  
Vol 253 (1) ◽  
pp. 295-298 ◽  
Author(s):  
E J Rayhel ◽  
D A Prentice ◽  
P S Tabor ◽  
W H Flurkey ◽  
R W Geib ◽  
...  
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Interleukin 2 ◽  
Transforming Growth Factor Β ◽  
Tgf Beta ◽  
Inhibitory Effects ◽  
Nb2 Cells ◽  
Growth Factor Beta ◽  
T Lymphoma

Transforming growth factor-beta (TGF-beta) inhibits proliferation of Nb2 cells, a rat T lymphoma, in response to lactogens and interleukin-2. Prostaglandins may play an important role in the pathway through which TGF-beta exerts its inhibitory actions, because prostaglandin E2 also inhibits proliferation of Nb2 cells, and indomethacin, an inhibitor of prostaglandin synthesis, reverses the inhibitory effects of TGF-beta on Nb2 cell proliferation.

scholarly journals Non-uniform influence of transforming growth factor-β on the biosynthesis of different forms of small chondroitin sulphate/dermatan sulphate proteoglycan

1990 ◽  
Vol 269 (2) ◽  
pp. 551-554 ◽  
Author(s):  
B Breuer ◽  
G Schmidt ◽  
H Kresse
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Core Protein ◽  
Chondroitin Sulphate ◽  
Transforming Growth Factor Β ◽  
Cell Types ◽  
Osteosarcoma Cell ◽  
Tgf Beta ◽  
Small Proteoglycan

The influence of transforming growth factor-beta (TGF-beta) on the expression of different forms of small proteoglycans was investigated in human skin fibroblasts and in a human osteosarcoma cell line. TGF-beta was not found to act as a general stimulator of small proteoglycan biosynthesis. In both cell types, an increased expression of the core protein of proteoglycan I was found. However, there was a profound decrease in the expression of a 106 kDa core protein, and either no alteration or a small decrease in the biosynthesis of the collagen-binding small proteoglycan II core protein. These results show that the production of individual members of the small proteoglycan family is differentially regulated.

scholarly journals Latent transforming growth factor-beta 1 and its binding protein are components of extracellular matrix microfibrils.

1996 ◽  
Vol 44 (8) ◽  
pp. 875-889 ◽  
Author(s):  
J Taipale ◽  
J Saharinen ◽  
K Hedman ◽  
J Keski-Oja
Keyword(s):  
Extracellular Matrix ◽  
Growth Factor ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Binding Protein ◽  
Human Fibroblasts ◽  
Tgf Beta ◽  
Growth Factor Beta ◽  
Cellular Fibronectin

We studied the localization of latent transforming growth factor-beta 1 (TGF-beta 1) and its binding protein (LTBP-1) in the extracellular matrix of cultured human fibroblasts by immunofluorescence and immunoelectron microscopy. Immunofluorescence of confluent fibroblast cultures indicated that LTBP-1 localizes to extracellular fibrillar structures resembling fibronectin-collagen matrix. Similar fibrillar structures were detected in cells stained with antibodies specific for TGF-beta 1 propeptide (beta 1-LAP). Both LTBP-1 and beta 1-LAP colocalized with fibronectin in double immunofluorescence analysis. These fibrillar structures were resistant to extraction with sodium deoxycholate, which is further evidence that LTBP-1 and large latent TGF-beta 1 complexes are integral components of the extracellular matrix. SV-40-transformed human fibroblasts lacked extracellular LTBP-1 fibers. EM analysis revealed approximately 10-nm-thick microfibrils that were labeled by anti-LTBP at 90-140-nm intervals. In addition, LTBP-1 was found in structures that were heavily labeled for fibronectin. The accumulation of LTBP-1 in the fibronectin matrix could be reconstituted in vitro. When isolated matrix components were immobilized on nitrocellulose and incubated with fibroblast conditioned medium, LTBP-1 from the medium associated with cellular fibronectin but not with heparan or chondroitin sulfate, vitronectin, tenascin, laminin, or collagen I or IV. The association of LTBP-1 with cellular fibronectin was abolished by treatment of the medium with plasmin, which cleaves LTBP-1 and inhibits its assembly to matrix. The present results indicate that latent TGF-beta 1 complexes are components of the extracellular matrix and suggest that alterations of the pericellular matrix could result in aberrant TGF-beta signaling.

scholarly journals Transforming-growth-factor-β activation elements in the distal promoter regions of the rat α1 type I collagen gene

1991 ◽  
Vol 280 (1) ◽  
pp. 157-162 ◽  
Author(s):  
J D Ritzenthaler ◽  
R H Goldstein ◽  
A Fine ◽  
A Lichtler ◽  
D W Rowe ◽  
...  
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor Beta ◽  
Transcriptional Activation ◽  
Type I Collagen ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β ◽  
Collagen Type I ◽  
Type I ◽  
Tgf Beta ◽  
Mobility Shift

We have located a cis-acting element (alpha 1-TAE) within the promoter sequences of the rat collagen alpha 1(I) gene (COL1A1) 1600 bases upstream of the transcription start site which mediates transcriptional activation by transforming growth factor beta (TGF-beta). The functional significance of this region was established by (1) deletion analysis of the alpha 1(I) promoter cloned upstream of the bacterial chloramphenicol acetyltransferase (CAT) gene and (2) by co-transfection of promoter constructs with double-stranded oligonucleotides. DNA-mobility-shift assays with radiolabelled alpha 1-TAE demonstrated increased nuclear binding activity after TGF-beta stimulation. Oligonucleotides encoding the alpha 1-TAE, additional upstream regions within the alpha 1(I) promoter, as well as consensus nuclear-factor-1 (NF-1) sequences, competed with the alpha 1-TAE sequence. The two collagen type I genes are stimulated by TGF-beta through different regions of their promoters.

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