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. 1990 Mar 1;110(3):767–775. doi: 10.1083/jcb.110.3.767

Release of basic fibroblast growth factor-heparan sulfate complexes from endothelial cells by plasminogen activator-mediated proteolytic activity

PMCID: PMC2116040  PMID: 2137829

Abstract

Cultured bovine capillary endothelial (BCE) cells synthesize heparan sulfate proteoglycans (HSPG), which are both secreted into the culture medium and deposited in the cell layer. The nonsoluble HSPGs can be isolated as two predominant species: a larger 800-kD HSPG, which is recovered from preparations of extracellular matrix, and a 250-kD HSPG, which is solubilized by nonionic detergent extraction of the cells. Both HSPG species bind bFGF. 125I-bFGF bound to BCE cell cultures is readily released by either heparinase or plasmin. When released by plasmin, the growth factor is recovered from the incubation medium as a complex with the partly degraded high molecular mass HSPG. Endogenous bFGF activity is released by a proteolytic treatment of cultured BCE cells. The bFGF-binding HSPGs are also released when cultures are incubated with the inactive proenzyme plasminogen. Under such experimental conditions, the release of the extracellular proteoglycans can be enhanced by treating the cells either with bFGF, which increases the plasminogen activating activity expressed by the cells, or decreased by treating the cells with transforming growth factor beta, which decreases the plasminogen activating activity of the cells. Specific immune antibodies raised against bovine urokinase also block the release of HSPG from BCE cell cultures. We propose that this plasminogen activator-mediated proteolysis provides a mechanism for the release of biologically active bFGF-HSPG complexes from the extracellular matrix and that bFGF release can be regulated by the balance between factors affecting the pericellular proteolytic activity.

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Selected References

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  1. Abraham J. A., Whang J. L., Tumolo A., Mergia A., Friedman J., Gospodarowicz D., Fiddes J. C. Human basic fibroblast growth factor: nucleotide sequence and genomic organization. EMBO J. 1986 Oct;5(10):2523–2528. doi: 10.1002/j.1460-2075.1986.tb04530.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Baird A., Ling N. Fibroblast growth factors are present in the extracellular matrix produced by endothelial cells in vitro: implications for a role of heparinase-like enzymes in the neovascular response. Biochem Biophys Res Commun. 1987 Jan 30;142(2):428–435. doi: 10.1016/0006-291x(87)90292-0. [DOI] [PubMed] [Google Scholar]
  3. Bashkin P., Doctrow S., Klagsbrun M., Svahn C. M., Folkman J., Vlodavsky I. Basic fibroblast growth factor binds to subendothelial extracellular matrix and is released by heparitinase and heparin-like molecules. Biochemistry. 1989 Feb 21;28(4):1737–1743. doi: 10.1021/bi00430a047. [DOI] [PubMed] [Google Scholar]
  4. Bergman B. L., Scott R. W., Bajpai A., Watts S., Baker J. B. Inhibition of tumor-cell-mediated extracellular matrix destruction by a fibroblast proteinase inhibitor, protease nexin I. Proc Natl Acad Sci U S A. 1986 Feb;83(4):996–1000. doi: 10.1073/pnas.83.4.996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Castellot J. J., Jr, Wright T. C., Karnovsky M. J. Regulation of vascular smooth muscle cell growth by heparin and heparan sulfates. Semin Thromb Hemost. 1987 Oct;13(4):489–503. doi: 10.1055/s-2007-1003525. [DOI] [PubMed] [Google Scholar]
  6. Danø K., Andreasen P. A., Grøndahl-Hansen J., Kristensen P., Nielsen L. S., Skriver L. Plasminogen activators, tissue degradation, and cancer. Adv Cancer Res. 1985;44:139–266. doi: 10.1016/s0065-230x(08)60028-7. [DOI] [PubMed] [Google Scholar]
  7. Deutsch D. G., Mertz E. T. Plasminogen: purification from human plasma by affinity chromatography. Science. 1970 Dec 4;170(3962):1095–1096. doi: 10.1126/science.170.3962.1095. [DOI] [PubMed] [Google Scholar]
  8. Dexter T. M. Stromal cell associated haemopoiesis. J Cell Physiol Suppl. 1982;1:87–94. doi: 10.1002/jcp.1041130414. [DOI] [PubMed] [Google Scholar]
  9. Edwards D. R., Murphy G., Reynolds J. J., Whitham S. E., Docherty A. J., Angel P., Heath J. K. Transforming growth factor beta modulates the expression of collagenase and metalloproteinase inhibitor. EMBO J. 1987 Jul;6(7):1899–1904. doi: 10.1002/j.1460-2075.1987.tb02449.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Folkman J., Haudenschild C. C., Zetter B. R. Long-term culture of capillary endothelial cells. Proc Natl Acad Sci U S A. 1979 Oct;76(10):5217–5221. doi: 10.1073/pnas.76.10.5217. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Folkman J., Klagsbrun M., Sasse J., Wadzinski M., Ingber D., Vlodavsky I. A heparin-binding angiogenic protein--basic fibroblast growth factor--is stored within basement membrane. Am J Pathol. 1988 Feb;130(2):393–400. [PMC free article] [PubMed] [Google Scholar]
  12. Gordon M. Y., Hibbin J. A., Kearney L. U., Gordon-Smith E. C., Goldman J. M. Colony formation by primitive haemopoietic progenitors in cocultures of bone marrow cells and stromal cells. Br J Haematol. 1985 May;60(1):129–136. doi: 10.1111/j.1365-2141.1985.tb07393.x. [DOI] [PubMed] [Google Scholar]
  13. Gordon M. Y., Riley G. P., Watt S. M., Greaves M. F. Compartmentalization of a haematopoietic growth factor (GM-CSF) by glycosaminoglycans in the bone marrow microenvironment. 1987 Mar 26-Apr 1Nature. 326(6111):403–405. doi: 10.1038/326403a0. [DOI] [PubMed] [Google Scholar]
  14. Gospodarowicz D., Ferrara N., Schweigerer L., Neufeld G. Structural characterization and biological functions of fibroblast growth factor. Endocr Rev. 1987 May;8(2):95–114. doi: 10.1210/edrv-8-2-95. [DOI] [PubMed] [Google Scholar]
  15. Gross J. L., Moscatelli D., Jaffe E. A., Rifkin D. B. Plasminogen activator and collagenase production by cultured capillary endothelial cells. J Cell Biol. 1982 Dec;95(3):974–981. doi: 10.1083/jcb.95.3.974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hascall V. C., Kimura J. H. Proteoglycans: isolation and characterization. Methods Enzymol. 1982;82(Pt A):769–800. doi: 10.1016/0076-6879(82)82102-2. [DOI] [PubMed] [Google Scholar]
  17. Hedman K., Kurkinen M., Alitalo K., Vaheri A., Johansson S., Hök M. Isolation of the pericellular matrix of human fibroblast cultures. J Cell Biol. 1979 Apr;81(1):83–91. doi: 10.1083/jcb.81.1.83. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Iozzo R. V. Cell surface heparan sulfate proteoglycan and the neoplastic phenotype. J Cell Biochem. 1988 May;37(1):61–78. doi: 10.1002/jcb.240370107. [DOI] [PubMed] [Google Scholar]
  19. Jalkanen M., Rapraeger A., Bernfield M. Mouse mammary epithelial cells produce basement membrane and cell surface heparan sulfate proteoglycans containing distinct core proteins. J Cell Biol. 1988 Mar;106(3):953–962. doi: 10.1083/jcb.106.3.953. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Joseph-Silverstein J., Moscatelli D., Rifkin D. B. The development of a quantitative RIA for basic fibroblast growth factor using polyclonal antibodies against the 157 amino acid form of human bFGF. The identification of bFGF in adherent elicited murine peritoneal macrophages. J Immunol Methods. 1988 Jun 13;110(2):183–192. doi: 10.1016/0022-1759(88)90102-0. [DOI] [PubMed] [Google Scholar]
  21. Joseph-Silverstein J., Rifkin D. B. Endothelial cell growth factors and the vessel wall. Semin Thromb Hemost. 1987 Oct;13(4):504–513. doi: 10.1055/s-2007-1003526. [DOI] [PubMed] [Google Scholar]
  22. Keller R., Silbert J. E., Furthmayr H., Madri J. A. Aortic endothelial cell proteoheparan sulfate. I. Isolation and characterization of plasmamembrane-associated and extracellular species. Am J Pathol. 1987 Aug;128(2):286–298. [PMC free article] [PubMed] [Google Scholar]
  23. Kinsella M. G., Wight T. N. Structural characterization of heparan sulfate proteoglycan subclasses isolated from bovine aortic endothelial cell cultures. Biochemistry. 1988 Mar 22;27(6):2136–2144. doi: 10.1021/bi00406a048. [DOI] [PubMed] [Google Scholar]
  24. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  25. Laiho M., Saksela O., Keski-Oja J. Transforming growth factor-beta induction of type-1 plasminogen activator inhibitor. Pericellular deposition and sensitivity to exogenous urokinase. J Biol Chem. 1987 Dec 25;262(36):17467–17474. [PubMed] [Google Scholar]
  26. Levin E. G., Santell L. Association of a plasminogen activator inhibitor (PAI-1) with the growth substratum and membrane of human endothelial cells. J Cell Biol. 1987 Dec;105(6 Pt 1):2543–2549. doi: 10.1083/jcb.105.6.2543. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Mimuro J., Schleef R. R., Loskutoff D. J. Extracellular matrix of cultured bovine aortic endothelial cells contains functionally active type 1 plasminogen activator inhibitor. Blood. 1987 Sep;70(3):721–728. [PubMed] [Google Scholar]
  28. Moscatelli D. High and low affinity binding sites for basic fibroblast growth factor on cultured cells: absence of a role for low affinity binding in the stimulation of plasminogen activator production by bovine capillary endothelial cells. J Cell Physiol. 1987 Apr;131(1):123–130. doi: 10.1002/jcp.1041310118. [DOI] [PubMed] [Google Scholar]
  29. Moscatelli D. Metabolism of receptor-bound and matrix-bound basic fibroblast growth factor by bovine capillary endothelial cells. J Cell Biol. 1988 Aug;107(2):753–759. doi: 10.1083/jcb.107.2.753. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Moscatelli D., Presta M., Joseph-Silverstein J., Rifkin D. B. Both normal and tumor cells produce basic fibroblast growth factor. J Cell Physiol. 1986 Nov;129(2):273–276. doi: 10.1002/jcp.1041290220. [DOI] [PubMed] [Google Scholar]
  31. Moscatelli D., Presta M., Rifkin D. B. Purification of a factor from human placenta that stimulates capillary endothelial cell protease production, DNA synthesis, and migration. Proc Natl Acad Sci U S A. 1986 Apr;83(7):2091–2095. doi: 10.1073/pnas.83.7.2091. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Moscatelli D., Rifkin D. B. Membrane and matrix localization of proteinases: a common theme in tumor cell invasion and angiogenesis. Biochim Biophys Acta. 1988 Aug 3;948(1):67–85. doi: 10.1016/0304-419x(88)90005-4. [DOI] [PubMed] [Google Scholar]
  33. Moses H. L., Coffey R. J., Jr, Leof E. B., Lyons R. M., Keski-Oja J. Transforming growth factor beta regulation of cell proliferation. J Cell Physiol Suppl. 1987;Suppl 5:1–7. doi: 10.1002/jcp.1041330403. [DOI] [PubMed] [Google Scholar]
  34. Neufeld G., Gospodarowicz D. Basic and acidic fibroblast growth factors interact with the same cell surface receptors. J Biol Chem. 1986 Apr 25;261(12):5631–5637. [PubMed] [Google Scholar]
  35. Neufeld G., Gospodarowicz D. The identification and partial characterization of the fibroblast growth factor receptor of baby hamster kidney cells. J Biol Chem. 1985 Nov 5;260(25):13860–13868. [PubMed] [Google Scholar]
  36. Presta M., Moscatelli D., Joseph-Silverstein J., Rifkin D. B. Purification from a human hepatoma cell line of a basic fibroblast growth factor-like molecule that stimulates capillary endothelial cell plasminogen activator production, DNA synthesis, and migration. Mol Cell Biol. 1986 Nov;6(11):4060–4066. doi: 10.1128/mcb.6.11.4060. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Rapraeger A., Bernfield M. Cell surface proteoglycan of mammary epithelial cells. Protease releases a heparan sulfate-rich ectodomain from a putative membrane-anchored domain. J Biol Chem. 1985 Apr 10;260(7):4103–4109. [PubMed] [Google Scholar]
  38. Roberts R., Gallagher J., Spooncer E., Allen T. D., Bloomfield F., Dexter T. M. Heparan sulphate bound growth factors: a mechanism for stromal cell mediated haemopoiesis. Nature. 1988 Mar 24;332(6162):376–378. doi: 10.1038/332376a0. [DOI] [PubMed] [Google Scholar]
  39. Saksela O., Moscatelli D., Rifkin D. B. The opposing effects of basic fibroblast growth factor and transforming growth factor beta on the regulation of plasminogen activator activity in capillary endothelial cells. J Cell Biol. 1987 Aug;105(2):957–963. doi: 10.1083/jcb.105.2.957. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Saksela O., Moscatelli D., Sommer A., Rifkin D. B. Endothelial cell-derived heparan sulfate binds basic fibroblast growth factor and protects it from proteolytic degradation. J Cell Biol. 1988 Aug;107(2):743–751. doi: 10.1083/jcb.107.2.743. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Saksela O. Plasminogen activation and regulation of pericellular proteolysis. Biochim Biophys Acta. 1985 Nov 12;823(1):35–65. doi: 10.1016/0304-419x(85)90014-9. [DOI] [PubMed] [Google Scholar]
  42. Saksela O., Rifkin D. B. Cell-associated plasminogen activation: regulation and physiological functions. Annu Rev Cell Biol. 1988;4:93–126. doi: 10.1146/annurev.cb.04.110188.000521. [DOI] [PubMed] [Google Scholar]
  43. Salonen E. M., Tervo T., Törmä E., Tarkkanen A., Vaheri A. Plasmin in tear fluid of patients with corneal ulcers: basis for new therapy. Acta Ophthalmol (Copenh) 1987 Feb;65(1):3–12. doi: 10.1111/j.1755-3768.1987.tb08482.x. [DOI] [PubMed] [Google Scholar]
  44. San Antonio J. D., Winston B. M., Tuan R. S. Regulation of chondrogenesis by heparan sulfate and structurally related glycosaminoglycans. Dev Biol. 1987 Sep;123(1):17–24. doi: 10.1016/0012-1606(87)90422-2. [DOI] [PubMed] [Google Scholar]
  45. Sato Y., Rifkin D. B. Autocrine activities of basic fibroblast growth factor: regulation of endothelial cell movement, plasminogen activator synthesis, and DNA synthesis. J Cell Biol. 1988 Sep;107(3):1199–1205. doi: 10.1083/jcb.107.3.1199. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Schor A. M., Schor S. L. Inhibition of endothelial cell morphogenetic interactions in vitro by alpha- and beta-xylosides. In Vitro Cell Dev Biol. 1988 Jul;24(7):659–668. doi: 10.1007/BF02623603. [DOI] [PubMed] [Google Scholar]
  47. Schweigerer L., Neufeld G., Friedman J., Abraham J. A., Fiddes J. C., Gospodarowicz D. Capillary endothelial cells express basic fibroblast growth factor, a mitogen that promotes their own growth. Nature. 1987 Jan 15;325(6101):257–259. doi: 10.1038/325257a0. [DOI] [PubMed] [Google Scholar]
  48. Shing Y., Folkman J., Sullivan R., Butterfield C., Murray J., Klagsbrun M. Heparin affinity: purification of a tumor-derived capillary endothelial cell growth factor. Science. 1984 Mar 23;223(4642):1296–1299. doi: 10.1126/science.6199844. [DOI] [PubMed] [Google Scholar]
  49. Sommer A., Brewer M. T., Thompson R. C., Moscatelli D., Presta M., Rifkin D. B. A form of human basic fibroblast growth factor with an extended amino terminus. Biochem Biophys Res Commun. 1987 Apr 29;144(2):543–550. doi: 10.1016/s0006-291x(87)80001-3. [DOI] [PubMed] [Google Scholar]
  50. Thesleff I., Jalkanen M., Vainio S., Bernfield M. Cell surface proteoglycan expression correlates with epithelial-mesenchymal interaction during tooth morphogenesis. Dev Biol. 1988 Oct;129(2):565–572. doi: 10.1016/0012-1606(88)90401-0. [DOI] [PubMed] [Google Scholar]
  51. Vlodavsky I., Folkman J., Sullivan R., Fridman R., Ishai-Michaeli R., Sasse J., Klagsbrun M. Endothelial cell-derived basic fibroblast growth factor: synthesis and deposition into subendothelial extracellular matrix. Proc Natl Acad Sci U S A. 1987 Apr;84(8):2292–2296. doi: 10.1073/pnas.84.8.2292. [DOI] [PMC free article] [PubMed] [Google Scholar]

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