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. 2000;9(3-4):155–160. doi: 10.1080/09629350020002895

Differential regulation of metalloproteinase production, proliferation and chemotaxis of human lung fibroblasts by PDGF, interleukin-1beta and TNF-alpha.

M Sasaki 1, M Kashima 1, T Ito 1, A Watanabe 1, N Izumiyama 1, M Sano 1, M Kagaya 1, T Shioya 1, M Miura 1
PMCID: PMC1781765  PMID: 11132772

Abstract

Fibroblast migration, proliferation, extracellular matrix protein synthesis and degradation, all of which play important roles in inflammation, are themselves induced by various growth factors and cytokines. Less is known about the interaction of these substances on lung fibroblast function in pulmonary fibrosis. The goal of this study was to investigate the effects of PDGF alone and in combination with IL-1beta and TNF-alpha on the production of human lung fibroblast matrix metalloproteinases, proliferation, and the chemotactic response. The assay for MMPs activity against FITC labeled type I and IV collagen was based on the specificity of the enzyme cleavage of collagen. Caseinolytis and gelatinolytic activities of secreted proteinases were analyzed by zymography. Fibronectin in conditioned media was measured using human lung fibronectin enzyme immunoassay. Cell proliferation was measured by 3H-Thymidine incorporation assay. Cell culture supernatants were tested for PGE2 content by ELISA. Chemotactic activity was measured using the modified Boyden chamber. Matrix metalloproteinase assay indicated that IL-1beta, TNF-alpha and PDGF induced intestitial collagenase (MMP-1) production. MMP assay also indicated that IL-1beta and TNF-alpha had inhibitory effects on MMP-2,9(gelatinaseA,B) production. Casein zymography confirmed that IL-1beta stimulated stromlysin (matrix metalloproteinase 3; MMP-3) and gelatin zymography demonstrated that TNF-alpha induced MMP-9 production in human lung fibroblast, whereas PDGF alone did not. PDGF in combination with IL-1beta and TNF-alpha induced MMP-3 and MMP-9 activity, as demonstrated by zymography. PDGF stimulated lung fibroblast proliferation in a concentration-dependent manner, whereas IL-1beta and TNF-alpha alone had no effect. In contrast, the proliferation of human lung fibroblasts by PDGF was inhibited in the presence of IL-1beta and TNF-alpha, and this inhibition was not a consequence of any elevation of PGE2. PDGF stimulated fibroblast chemotaxis in a concentration-dependent manner, and this stimulation was augmented by combining PDGF with IL-1beta and TNF-alpha. These findings suggested that PDGF differentially regulated MMPs production in combination with cytokines, and further that MMP assay and zymography had differential sensitivity for detecting MMPs. The presence of cytokines with PDGF appears to modulate the proliferation and chemotaxis of human lung fibroblasts.

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Selected References

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  1. Antoniades H. N., Bravo M. A., Avila R. E., Galanopoulos T., Neville-Golden J., Maxwell M., Selman M. Platelet-derived growth factor in idiopathic pulmonary fibrosis. J Clin Invest. 1990 Oct;86(4):1055–1064. doi: 10.1172/JCI114808. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Battegay E. J., Raines E. W., Colbert T., Ross R. TNF-alpha stimulation of fibroblast proliferation. Dependence on platelet-derived growth factor (PDGF) secretion and alteration of PDGF receptor expression. J Immunol. 1995 Jun 1;154(11):6040–6047. [PubMed] [Google Scholar]
  3. Battegay E. J., Raines E. W., Colbert T., Ross R. TNF-alpha stimulation of fibroblast proliferation. Dependence on platelet-derived growth factor (PDGF) secretion and alteration of PDGF receptor expression. J Immunol. 1995 Jun 1;154(11):6040–6047. [PubMed] [Google Scholar]
  4. Bienkowski R. S., Gotkin M. G. Control of collagen deposition in mammalian lung. Proc Soc Exp Biol Med. 1995 Jun;209(2):118–140. doi: 10.3181/00379727-209-43886a. [DOI] [PubMed] [Google Scholar]
  5. Butler D. M., Leizer T., Hamilton J. A. Stimulation of human synovial fibroblast DNA synthesis by platelet-derived growth factor and fibroblast growth factor. Differences to the activation by IL-1. J Immunol. 1989 May 1;142(9):3098–3103. [PubMed] [Google Scholar]
  6. Chin J. R., Murphy G., Werb Z. Stromelysin, a connective tissue-degrading metalloendopeptidase secreted by stimulated rabbit synovial fibroblasts in parallel with collagenase. Biosynthesis, isolation, characterization, and substrates. J Biol Chem. 1985 Oct 5;260(22):12367–12376. [PubMed] [Google Scholar]
  7. Circolo A., Welgus H. G., Pierce G. F., Kramer J., Strunk R. C. Differential regulation of the expression of proteinases/antiproteinases in fibroblasts. Effects of interleukin-1 and platelet-derived growth factor. J Biol Chem. 1991 Jul 5;266(19):12283–12288. [PubMed] [Google Scholar]
  8. Duncan M. R., Berman B. Differential regulation of collagen, glycosaminoglycan, fibronectin, and collagenase activity production in cultured human adult dermal fibroblasts by interleukin 1-alpha and beta and tumor necrosis factor-alpha and beta. J Invest Dermatol. 1989 May;92(5):699–706. doi: 10.1111/1523-1747.ep12696891. [DOI] [PubMed] [Google Scholar]
  9. Elias J. A., Freundlich B., Adams S., Rosenbloom J. Regulation of human lung fibroblast collagen production by recombinant interleukin-1, tumor necrosis factor, and interferon-gamma. Ann N Y Acad Sci. 1990;580:233–244. doi: 10.1111/j.1749-6632.1990.tb17932.x. [DOI] [PubMed] [Google Scholar]
  10. Elias J. A., Gustilo K., Baeder W., Freundlich B. Synergistic stimulation of fibroblast prostaglandin production by recombinant interleukin 1 and tumor necrosis factor. J Immunol. 1987 Jun 1;138(11):3812–3816. [PubMed] [Google Scholar]
  11. Erkell L. J., Schirrmacher V. Quantitative in vitro assay for tumor cell invasion through extracellular matrix or into protein gels. Cancer Res. 1988 Dec 1;48(23):6933–6937. [PubMed] [Google Scholar]
  12. Fukuda Y., Ishizaki M., Masuda Y., Kimura G., Kawanami O., Masugi Y. The role of intraalveolar fibrosis in the process of pulmonary structural remodeling in patients with diffuse alveolar damage. Am J Pathol. 1987 Jan;126(1):171–182. [PMC free article] [PubMed] [Google Scholar]
  13. Gadek J. E., Kelman J. A., Fells G., Weinberger S. E., Horwitz A. L., Reynolds H. Y., Fulmer J. D., Crystal R. G. Collagenase in the lower respiratory tract of patients with idiopathic pulmonary fibrosis. N Engl J Med. 1979 Oct 4;301(14):737–742. doi: 10.1056/NEJM197910043011401. [DOI] [PubMed] [Google Scholar]
  14. Grotendorst G. R., Seppä H. E., Kleinman H. K., Martin G. R. Attachment of smooth muscle cells to collagen and their migration toward platelet-derived growth factor. Proc Natl Acad Sci U S A. 1981 Jun;78(6):3669–3672. doi: 10.1073/pnas.78.6.3669. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Harris E. D., Jr, Vater C. A. Vertebrate collagenases. Methods Enzymol. 1982;82(Pt A):423–452. doi: 10.1016/0076-6879(82)82077-6. [DOI] [PubMed] [Google Scholar]
  16. Landgren E., Eriksson A., Wennström S., Kanda S., Claesson-Welsh L. Induction of fibroblast growth factor receptor-1 mRNA and protein by platelet-derived growth factor BB. Exp Cell Res. 1996 Mar 15;223(2):405–411. doi: 10.1006/excr.1996.0095. [DOI] [PubMed] [Google Scholar]
  17. Lin L. L., Lin A. Y., DeWitt D. L. Interleukin-1 alpha induces the accumulation of cytosolic phospholipase A2 and the release of prostaglandin E2 in human fibroblasts. J Biol Chem. 1992 Nov 25;267(33):23451–23454. [PubMed] [Google Scholar]
  18. Martinet Y., Rom W. N., Grotendorst G. R., Martin G. R., Crystal R. G. Exaggerated spontaneous release of platelet-derived growth factor by alveolar macrophages from patients with idiopathic pulmonary fibrosis. N Engl J Med. 1987 Jul 23;317(4):202–209. doi: 10.1056/NEJM198707233170404. [DOI] [PubMed] [Google Scholar]
  19. Matrisian L. M., Bowden G. T. Stromelysin/transin and tumor progression. Semin Cancer Biol. 1990 Apr;1(2):107–115. [PubMed] [Google Scholar]
  20. Miyazaki K., Hattori Y., Umenishi F., Yasumitsu H., Umeda M. Purification and characterization of extracellular matrix-degrading metalloproteinase, matrin (pump-1), secreted from human rectal carcinoma cell line. Cancer Res. 1990 Dec 15;50(24):7758–7764. [PubMed] [Google Scholar]
  21. Pardo A., Selman M., Ramírez R., Ramos C., Montaño M., Stricklin G., Raghu G. Production of collagenase and tissue inhibitor of metalloproteinases by fibroblasts derived from normal and fibrotic human lungs. Chest. 1992 Oct;102(4):1085–1089. doi: 10.1378/chest.102.4.1085. [DOI] [PubMed] [Google Scholar]
  22. Paulsson Y., Austgulen R., Hofsli E., Heldin C. H., Westermark B., Nissen-Meyer J. Tumor necrosis factor-induced expression of platelet-derived growth factor A-chain messenger RNA in fibroblasts. Exp Cell Res. 1989 Feb;180(2):490–496. doi: 10.1016/0014-4827(89)90075-x. [DOI] [PubMed] [Google Scholar]
  23. Rennard S. I., Hunninghake G. W., Bitterman P. B., Crystal R. G. Production of fibronectin by the human alveolar macrophage: mechanism for the recruitment of fibroblasts to sites of tissue injury in interstitial lung diseases. Proc Natl Acad Sci U S A. 1981 Nov;78(11):7147–7151. doi: 10.1073/pnas.78.11.7147. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Rojas-Valencia L., Montiel F., Montaño M., Selman M., Pardo A. Expression of a 2.8-kb PDGF-B/c-sis transcript and synthesis of PDGF-like protein by human lung fibroblasts. Chest. 1995 Jul;108(1):240–245. doi: 10.1378/chest.108.1.240. [DOI] [PubMed] [Google Scholar]
  25. Schmidt J. A., Mizel S. B., Cohen D., Green I. Interleukin 1, a potential regulator of fibroblast proliferation. J Immunol. 1982 May;128(5):2177–2182. [PubMed] [Google Scholar]
  26. Selman M., Montaño M., Ramos C., Chapela R. Concentration, biosynthesis and degradation of collagen in idiopathic pulmonary fibrosis. Thorax. 1986 May;41(5):355–359. doi: 10.1136/thx.41.5.355. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Seppä H., Grotendorst G., Seppä S., Schiffmann E., Martin G. R. Platelet-derived growth factor in chemotactic for fibroblasts. J Cell Biol. 1982 Feb;92(2):584–588. doi: 10.1083/jcb.92.2.584. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Soma Y., Takehara K., Ishibashi Y. Alteration of the chemotactic response of human skin fibroblasts to PDGF by growth factors. Exp Cell Res. 1994 Jun;212(2):274–277. doi: 10.1006/excr.1994.1143. [DOI] [PubMed] [Google Scholar]
  29. Tingström A., Reuterdahl C., Lindahl P., Heldin C. H., Rubin K. Expression of platelet-derived growth factor-beta receptors on human fibroblasts. Regulation by recombinant platelet-derived growth factor-BB, IL-1, and tumor necrosis factor-alpha. J Immunol. 1992 Jan 15;148(2):546–554. [PubMed] [Google Scholar]
  30. Tsukamoto T., Matsui T., Nakata H., Ito M., Natazuka T., Fukase M., Fujita T. Interleukin-1 enhances the response of osteoblasts to platelet-derived growth factor through the alpha receptor-specific up-regulation. J Biol Chem. 1991 Jun 5;266(16):10143–10147. [PubMed] [Google Scholar]
  31. Umenishi F., Umeda M., Miyazaki K. Efficient purification of TIMP-2 from culture medium conditioned by human hepatoma cell line, and its inhibitory effects on metalloproteinases and in vitro tumor invasion. J Biochem. 1991 Aug;110(2):189–195. doi: 10.1093/oxfordjournals.jbchem.a123555. [DOI] [PubMed] [Google Scholar]
  32. Unemori E. N., Ehsani N., Wang M., Lee S., McGuire J., Amento E. P. Interleukin-1 and transforming growth factor-alpha: synergistic stimulation of metalloproteinases, PGE2, and proliferation in human fibroblasts. Exp Cell Res. 1994 Feb;210(2):166–171. doi: 10.1006/excr.1994.1025. [DOI] [PubMed] [Google Scholar]
  33. Ura H., Bonfil R. D., Reich R., Reddel R., Pfeifer A., Harris C. C., Klein-Szanto A. J. Expression of type IV collagenase and procollagen genes and its correlation with the tumorigenic, invasive, and metastatic abilities of oncogene-transformed human bronchial epithelial cells. Cancer Res. 1989 Aug 15;49(16):4615–4621. [PubMed] [Google Scholar]
  34. Vignaud J. M., Allam M., Martinet N., Pech M., Plenat F., Martinet Y. Presence of platelet-derived growth factor in normal and fibrotic lung is specifically associated with interstitial macrophages, while both interstitial macrophages and alveolar epithelial cells express the c-sis proto-oncogene. Am J Respir Cell Mol Biol. 1991 Dec;5(6):531–538. doi: 10.1165/ajrcmb/5.6.531. [DOI] [PubMed] [Google Scholar]
  35. Zeng G., Millis A. J. Differential regulation of collagenase and stromelysin mRNA in late passage cultures of human fibroblasts. Exp Cell Res. 1996 Jan 10;222(1):150–156. doi: 10.1006/excr.1996.0019. [DOI] [PubMed] [Google Scholar]

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