Collagen matrices attenuate the collagen-synthetic response of cultured fibroblasts to TGF-beta.

Transforming growth factor-beta, a potent modulator of cell function, induces fibroblasts cultured on plastic to increase collagen synthesis. In 5- and 7-day porcine skin wounds, which have minimal to moderate collagen matrix, respectively, transforming growth factor-beta and type I procollagen were coordinately expressed throughout the granulation tissue. However, in 10-day collagen-rich granulation tissue type I procollagen expression diminished despite persistence of transforming growth factor-beta. To investigate whether collagen matrix attenuates the collagen-synthetic response of fibroblasts to transforming growth factor-beta, we cultured human dermal fibroblasts in conditions that simulate collagen-rich granulation tissue. Therefore, human dermal fibroblasts were suspended in attached collagen gels and collagen and noncollagen production was assayed in the absence and presence of transforming growth factor-beta. Although transforming growth factor-beta stimulated collagen synthesis by fibroblasts cultured in the collagen gels, these fibroblasts consistently produced less collagen than similarly treated fibroblasts cultured on plastic. This phenomenon was not secondary to nonspecific binding of transforming growth factor-beta to the collagen matrix. Fibroblasts cultured in a fibrin gel responded to transforming growth factor-beta similarly to fibroblasts cultured on plastic. Using immunofluorescence probes to type I procollagen, we observed that transforming growth factor-beta increased type I procollagen expression in most fibroblasts cultured on plastic, but only in occasional fibroblasts cultured in collagen gels. From these data we conclude that collagen matrices attenuate the collagen synthetic response of fibroblast to transforming growth factor-beta in vitro and possibly in vivo.

[1]  T. Krieg,et al.  Collagenase gene expression in fibroblasts is regulated by a three‐dimensional contact with collagen , 1989, FEBS letters.

[2]  Z. Werb,et al.  Signal transduction by integrin receptors for extracellular matrix: cooperative processing of extracellular information. , 1992 .

[3]  R. Weinberg,et al.  Expression cloning and characterization of the TGF-beta type III receptor. , 1991, Cell.

[4]  C. Heldin,et al.  Role for carbohydrate structures inTGF-β1 latency , 1989, Nature.

[5]  R. Clark,et al.  Optimization of immunohistochemical techniques to detect extracellular matrix proteins in fixed skin specimens. , 1989, The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society.

[6]  R. Ross,et al.  Effects of growth factors in vivo. I. Cell ingrowth into porous subcutaneous chambers. , 1987, The American journal of pathology.

[7]  M. P. Welch,et al.  Temporal relationships of F-actin bundle formation, collagen and fibronectin matrix assembly, and fibronectin receptor expression to wound contraction , 1990, The Journal of cell biology.

[8]  R. Weinberg,et al.  Expression cloning and characterization of the TGF-β type III receptor , 1991, Cell.

[9]  F. Grinnell,et al.  Spatial organization of extracellular matrix and fibroblast activity: effects of serum, transforming growth factor beta, and fibronectin. , 1990, Experimental cell research.

[10]  E. Kohn,et al.  Extracellular matrix promotes mammary epithelial growth and differentiation in vitro. , 1982, Proceedings of the National Academy of Sciences of the United States of America.

[11]  H. Moses,et al.  Proteolytic activation of latent transforming growth factor-beta from fibroblast-conditioned medium , 1988, The Journal of cell biology.

[12]  T. Krieg,et al.  A defective cell surface collagen-binding protein in dermatosparactic sheep fibroblasts , 1988, The Journal of cell biology.

[13]  Frederick Grinnell,et al.  Fibroblasts, myofibroblasts, and wound contraction , 1994, The Journal of cell biology.

[14]  T. Broekelmann,et al.  A monoclonal antibody to the carboxyterminal domain of procollagen type I visualizes collagen-synthesizing fibroblasts. Detection of an altered fibroblast phenotype in lungs of patients with pulmonary fibrosis. , 1986, The Journal of clinical investigation.

[15]  M. Sporn,et al.  Transforming growth factor type beta: rapid induction of fibrosis and angiogenesis in vivo and stimulation of collagen formation in vitro. , 1986, Proceedings of the National Academy of Sciences of the United States of America.

[16]  William,et al.  Characterization of the class III collagen receptor, a phosphorylated, transmembrane glycoprotein expressed in nucleated human cells. , 1988, The Journal of biological chemistry.

[17]  M. Sporn,et al.  Polypeptide transforming growth factors isolated from bovine sources and used for wound healing in vivo. , 1983, Science.

[18]  T. Krieg,et al.  Regulation of collagen synthesis in fibroblasts within a three-dimensional collagen gel. , 1988, Experimental cell research.

[19]  R. Derynck,et al.  Inactivation of the type II receptor reveals two receptor pathways for the diverse TGF-beta activities. , 1993, Science.

[20]  E. Hay,et al.  Response of basal epithelial cell surface and Cytoskeleton to solubilized extracellular matrix molecules , 1981, The Journal of cell biology.

[21]  J. McPherson,et al.  Collagen fibrillogenesis in vitro: a characterization of fibril quality as a function of assembly conditions. , 1985, Collagen and related research.

[22]  S. Haskill,et al.  Signal transduction from the extracellular matrix , 1993, The Journal of cell biology.

[23]  S. Leibovich,et al.  Electron microscope studies of the effects of endo- and exopeptidase digestion on tropocollagen. A novel concept of the role of terminal regions in fibrillogenesis. , 1970, Biochimica et biophysica acta.

[24]  F. Grinnell,et al.  Decreased level of PDGF-stimulated receptor autophosphorylation by fibroblasts in mechanically relaxed collagen matrices , 1993, The Journal of cell biology.

[25]  K. Paigen,et al.  A simple, rapid, and sensitive DNA assay procedure. , 1980, Analytical biochemistry.

[26]  W. T. Chen,et al.  Regulation of fibronectin receptor distribution by transformation, exogenous fibronectin, and synthetic peptides , 1986, The Journal of cell biology.

[27]  W. Carter,et al.  The membrane glycoprotein Ia-IIa (VLA-2) complex mediates the Mg++- dependent adhesion of platelets to collagen , 1989, The Journal of cell biology.

[28]  Expression cloning of the TGF-beta type II receptor, a functional transmembrane serine/threonine kinase. , 1992, Cell.

[29]  D. Gospodarowicz,et al.  Permissive effect of the extracellular matrix on cell proliferation in vitro. , 1980, Proceedings of the National Academy of Sciences of the United States of America.

[30]  F. Grinnell,et al.  Extracellular matrix organization modulates fibroblast growth and growth factor responsiveness. , 1989, Experimental cell research.

[31]  K. Miyazono,et al.  Role for carbohydrate structures in TGF-beta 1 latency. , 1989, Nature.

[32]  B. Nusgens,et al.  Collagen biosynthesis by cells in a tissue equivalent matrix in vitro. , 1984, Collagen and related research.

[33]  U. K. Laemmli,et al.  Cleavage of structural proteins during , 1970 .

[34]  T. Borg,et al.  Beta 1 integrin-mediated collagen gel contraction is stimulated by PDGF. , 1990, Experimental cell research.

[35]  J. Massagué,et al.  Transforming growth factor-beta stimulates the expression of fibronectin and collagen and their incorporation into the extracellular matrix. , 1986, The Journal of biological chemistry.

[36]  T. Krieg,et al.  Integrin alpha 2 beta 1 is upregulated in fibroblasts and highly aggressive melanoma cells in three-dimensional collagen lattices and mediates the reorganization of collagen I fibrils , 1991, The Journal of cell biology.

[37]  J. Massagué,et al.  Betaglycan presents ligand to the TGFβ signaling receptor , 1993, Cell.

[38]  M. Sporn,et al.  Accelerated healing of incisional wounds in rats induced by transforming growth factor-beta. , 1987, Science.

[39]  M. Sporn,et al.  Transforming growth factor-beta 1: histochemical localization with antibodies to different epitopes , 1989, The Journal of cell biology.

[40]  J. McPherson,et al.  Cartilage-inducing factor-A. Apparent identity to transforming growth factor-beta. , 1986, The Journal of biological chemistry.

[41]  R. Ross,et al.  The Platelet Derived Growth Factor and Cell Proliferation , 1979 .

[42]  P. Henson,et al.  The immunologic release of constituents from neutrophil leukocytes. II. Mechanisms of release during phagocytosis, and adherence to nonphagocytosable surfaces. , 1971, Journal of immunology.

[43]  D. Rosen,et al.  Antibodies to the N-terminal portion of cartilage-inducing factor A and transforming growth factor beta. Immunohistochemical localization and association with differentiating cells. , 1986, The Journal of biological chemistry.

[44]  B. Sykes,et al.  The estimation of two collagens from human dermis by interrupted gel electrophoresis. , 1976, Biochemical and biophysical research communications.

[45]  W. Carter,et al.  The fibronectin receptor is organized by extracellular matrix fibronectin: implications for oncogenic transformation and for cell recognition of fibronectin matrices , 1989, The Journal of cell biology.

[46]  M. O’Connor-McCourt,et al.  Latent transforming growth factor-beta in serum. A specific complex with alpha 2-macroglobulin. , 1987, The Journal of biological chemistry.

[47]  R. Hynes,et al.  Relationships between fibronectin (LETS protein) and actin , 1978, Cell.

[48]  J. McPherson,et al.  Influence of the extracellular matrix on the proliferative response of human skin fibroblasts to serum and purified platelet‐derived growth factor , 1988, Journal of cellular physiology.

[49]  D. Rifkin,et al.  Recent developments in the cell biology of basic fibroblast growth factor , 1989, The Journal of cell biology.

[50]  A. Fine,et al.  The effect of transforming growth factor-beta on cell proliferation and collagen formation by lung fibroblasts. , 1987, The Journal of biological chemistry.

[51]  M. Sporn,et al.  Transforming growth factor-beta: recent progress and new challenges , 1992, The Journal of cell biology.

[52]  A. Eisen,et al.  Integrin α2β1 (VLA-2) mediates reorganization and contraction of collagen matrices by human cells , 1991, Cell.

[53]  F. Grinnell,et al.  Long-term culture of fibroblasts in contracted collagen gels: effects on cell growth and biosynthetic activity. , 1989, The Journal of investigative dermatology.

[54]  J. McPherson,et al.  Platelet isoforms of platelet-derived growth factor stimulate fibroblasts to contract collagen matrices. , 1989, The Journal of clinical investigation.