Angiogenesis

, Volume 4, Issue 4, pp 277–288 | Cite as

Effect of glycation on basic fibroblast growth factor induced angiogenesis and activation of associated signal transduction pathways in vascular endothelial cells: Possible relevance to wound healing in diabetes

  • Y. Duraisamy
  • M. Slevin
  • N. Smith
  • J. Bailey
  • J. Zweit
  • C. Smith
  • N. Ahmed
  • J. Gaffney
Article

Abstract

Ineffectual wound healing in hyperglycaemic patients suffering from diabetes mellitus is characterised by a reduction in capillary reformation (angiogenesis). Basic fibroblast growth factor (FGF-2) is secreted by fibroblasts, macrophages and in particular endothelial cells (EC) in response to tissue injury and is important in promotion of neovascularisation. Recently, glycation of FGF-2 has been shown to significantly reduce its activity in vitro. We have examined the kinetics of FGF-2 glycation and compared its ability with that of native FGF-2 to activate mitogenesis, capillary formation and associated signal transduction in bovine aortic EC (BAEC). FGF-2 was exposed to 0.25 M glucose-6-phosphate (G-6-P) for 24–72 h and the degree of glycation determined by matrix assisted laser desorption ionisation mass spectrometry. Native FGF-2 was heterogeneous with Mw in the range 15,153.6–17,903 Da. After 24 h incubation with G-6-P there was evidence of glycation, and the mass increase corresponded to addition of 2.7 mol of G-6-P residues; after 48 h, 4 mol sugar was added and this increased to 8.7 after 72 h. Dimerisation of FGF-2 was observed after 72 h of treatment. Induction of mitogenesis in BAEC was significantly reduced by 25%–40% after treatment for 48–96 h with glycated (24 h) FGF-2 (gFGF-2;100 pg/ml–5 ng/ml; P < 0.05), whilst capillary tubule formation was significantly reduced by between 60% and 90% (100 pg/ml–1 ng/ml; P < 0.05) after 5 days compared to native FGF-2. Subsequent investigation of the signal transduction molecules associated with mitogenesis showed a reduction in FGF-2 induced tyrosine phosphorylated proteins of approximate Mw 20–150 kDa between 10 min and 24 h, in particular, mitogen activated protein kinase (MAPK)/early response kinase (ERK-1, ERK-2), after glycation. To determine the reason for reduced angiogenic activity of gFGF-2, we compared its binding characteristics to that of native FGF-2. Total binding of gFGF-2 to the cell surface was significantly reduced in BAEC analysed by FACS compared to native FGF-2 (P < 0.05). Further investigation using 125I-labelled differentially washed samples, demonstrated a significant reduction in gFGF-2 binding to the high affinity tyrosine kinase receptor (46%) compared to native FGF-2. In summary, glycation of FGF-2 in vitro occurs rapidly within 24 h in the presence of elevated levels of G-6-P. Glycation caused a significant reduction in the ability of FGF-2 to bind to the tyrosine kinase receptor and activate signal transduction pathways responsible for both mitogenesis and capillary formation in BAEC. These results could help to explain the mechanism behind impaired wound healing in patients with diabetes mellitus.

basic fibroblast growth factor bovine aortic endothelial cells diabetes glycated basic fibroblast growth factor signal transduction wound healing 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Most RS, Sinnock P. The epidemiology of lower extremity amputations in diabetic individuals. Diabetes Care 1983; 6: 87–91.PubMedGoogle Scholar
  2. 2.
    Fiddes JC, Hebda PA, Hayward P et al. Preclinical wound-healing studies with recombinant basic fibroblast growth factor. Ann NY Acad Sci 1991; 638: 316–28.PubMedGoogle Scholar
  3. 3.
    Tanaka E, Ase K, Okuda T et al. Mechanism of acceleration of wound healing by basic fibroblast growth factor in genetically diabetic mice. Biol Pharm Bull 1996; 19: 1141–48.PubMedGoogle Scholar
  4. 4.
    Akimoto S, Ishikawa O, Iijima C et al. Expression of basic fibroblast growth factor and its receptor by fibroblasts, macrophages and mast cells in hypertrophic scar. Eur J Dermatol 1999; 9: 357–62.PubMedGoogle Scholar
  5. 5.
    Schott RJ, Morrow LA. Growth factors and angiogenesis. Cardiovasc Res 1993; 27: 1155–61.PubMedGoogle Scholar
  6. 6.
    Slevin M, Kumar S, Xiaotong H et al. Physiological concentrations of the gangliosides GM1, GM2 and GM3 differentially modify basic fibroblast growth factor induced mitogenesis and the associated signalling pathways in endothelial cells. Int J Cancer 1999; 82: 412–23.PubMedCrossRefGoogle Scholar
  7. 7.
    Okumura M, Okuda T, Nakamura T et al. Effect of basic fibroblast growth factor on wound healing in healing-impaired animal models. Drug Res 1996a; 46: 547–51.Google Scholar
  8. 8.
    Okumura M, Okuda T, Nakamura T et al. Acceleration of wound healing in diabetic mice by basic fibroblast growth factor. Biol Pharm Bull 1996b; 19: 530–35.PubMedGoogle Scholar
  9. 9.
    Pierce GF, Tarpley JE, Yanagihara D et al. Platelet derived growth factor (BB homodimer), transforming growth factor-b1, and basic fibroblast growth factor in dermal wound healing. Am J Pathol 1992; 140: 1375–88.PubMedGoogle Scholar
  10. 10.
    Teixeira AS, Andrade SP. Glucose-induced inhibition of angiogenesis in the rat sponge granuloma is prevented by aminoguanidine. Life Sci 1999; 64: 655–62.PubMedCrossRefGoogle Scholar
  11. 11.
    Furth AJ. Glycated proteins in diabetes. Br J Biomed Sci 1997; 54: 192–200.PubMedGoogle Scholar
  12. 12.
    Lyons TJ, Jenkins AJ. Glycation, oxidation and lipoxidation in the development of the complications of diabetes: A carbonyl stress hypothesis. Diabetes Rev 1997; 5: 365–91.Google Scholar
  13. 13.
    Cohen MP, Ziyadeh F. Role of Amadori-modified non-enzymically glycated serum proteins in pathogenesis of diabetic nephropathy. J Am Soc Nephrol 1996; 7: 183–90.PubMedGoogle Scholar
  14. 14.
    Brownlee M. The pathological implications of protein glycation. Clin Invest Med 1995; 18: 275–87.PubMedGoogle Scholar
  15. 15.
    Florkiewicz RZ, Majack RA, Buechler RD et al. Quantitative export of FGF-2 occurs through an alternative, energy-dependent, non-er/golgi pathway. J Cell Physiol 1995; 162: 388–99.PubMedCrossRefGoogle Scholar
  16. 16.
    Rifkin DB, Moscatelli D, Flaumenhaft R et al. Mechanisms controlling the extracellular activity of basic fibroblasat growth factor and transforming growth factorb. Ann NY Acad Sci 1997; 57: 250–8.Google Scholar
  17. 17.
    Nissen N, Shankar R, Gamelli R et al. Heparan and heparan sulphate protect basic fibroblast growth factor from non-enzymic glycation. Biochem J 1999; 338: 637–42.PubMedCrossRefGoogle Scholar
  18. 18.
    Vainnika S, Joukou V, Wennistrom S et al. Signal transduction by fibroblast growth factor receptor-4 (FGFR-4): Comparison with FGFR-1. J Biol Chem 1994; 269: 18,320–6.Google Scholar
  19. 19.
    Giardino I, Edelstein D, Brownlee M. Nonenzymatic glycation invitro and in bovine endothelial cells alters basic fibroblast growth factor activity. A model for intracellular glycation in diabetes. J Clin Invest 1994; 94: 110–7.PubMedCrossRefGoogle Scholar
  20. 20.
    Sattar A, Kumar S, West D. Does hyaluronan have a role in endothelial cell proliferation. Semin Arthritis Rheu 1992; 22: 37–43.CrossRefGoogle Scholar
  21. 21.
    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; 131: 123–30.PubMedCrossRefGoogle Scholar
  22. 22.
    Raj DSC, Choudhury D, Welbourne TC et al. Advanced glycation end products: A nephrologist's perspective. Am J Kidney Dis 2000; 35: 365–80.PubMedGoogle Scholar
  23. 23.
    Faham S, Hileman RE, Fromm JR et al. Heparan structure and interactions with basic fibroblast growth factor. Science 1996; 271: 1116–20.PubMedGoogle Scholar
  24. 24.
    Seno M, Sasada R, Kurokawa T et al. Carboxyl terminal structure of basic fibroblast growth factor significantly contributes to its affinity for heparan. Eur J Biochem 1990; 188: 239–45.PubMedCrossRefGoogle Scholar
  25. 25.
    Baird A, Schubert D, Ling M et al. Receptor and heparan binding domains for basic fibroblast growth factor. Proc Natl Acad Sci USA 1988; 85: 2324–8.PubMedCrossRefGoogle Scholar
  26. 26.
    Krishnamurti U, Rondeau E, Sraer JD et al. Alterations in human glomerular epithelial cells interacting with nonenzymatically glycosylated matrix. J Biol Chem 1997; 272: 27,966–70.Google Scholar
  27. 27.
    Pugnaloni A, Tesei M, Amati S et al. Tyrosine phosphorylation in type-1 diabetes by immunogold detection: An in vitro human aortic endothelial cell (HAEC) study in the presence of diabetic low density lipoproteins (LDL). Eur J Histochem 1999; 43: 199–204.PubMedGoogle Scholar
  28. 28.
    Li LY, Safran M, Aviezer D et al. Diminished heparin-binding of basic fibroblast growth factor mutant is associated with reduced receptor-binding, mitogenesis, plasminogen-activator induction, and in vitro angiogenesis. Biochemistry 1994; 33: 10,999–11,007.Google Scholar

Copyright information

© Kluwer Academic Publishers 2001

Authors and Affiliations

  • Y. Duraisamy
    • 1
  • M. Slevin
    • 1
  • N. Smith
    • 2
  • J. Bailey
    • 2
  • J. Zweit
    • 2
  • C. Smith
    • 1
  • N. Ahmed
    • 1
  • J. Gaffney
    • 3
  1. 1.Department of Biological SciencesManchester Metropolitan UniversityManchesterUK
  2. 2.Patterson InstituteChristie HospitalManchesterUK
  3. 3.Department of Biological SciencesManchester Metropolitan UniversityManchesterUK. Tel:

Personalised recommendations