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The anti-apoptotic activity of albumin for endothelium is inhibited by advanced glycation end products restricting intramolecular movement

Abstract

Human serum albumin (HSA) inhibits endothelial apoptosis in a highly specific manner. CNBr fragmentation greatly increases the effectiveness of this activity, suggesting that this type of protection is mediated by a partially cryptic albumin domain which is transiently exposed by intramolecular movement. Advanced glycation end-product (AGE) formation in HSA greatly reduces its intra-molecular movement. This study aimed to determine if this inhibits the anti-apoptotic activity of HSA, and if such inactivation could be reversed by CNBr fragmentation. HSA-AGE was prepared by incubating HSA with glucose, and assessed using the fructosamine assay, mass spectrometry, SDS-PAGE and fluorometry. Low levels of AGE in the HSA had little effect upon its anti-apoptotic activity, but when the levels of AGE were high and the intra-molecular movement was reduced, endothelial cell survival was also found to be reduced to levels equivalent to those in cultures without HSA or serum (p > 0.001). Survival was restored by the inclusion of native HSA, despite the presence of HSA with high levels of AGE. Also, CNBr fragmentation of otherwise inactive HSA-AGE restored the anti-apoptotic activity for endothelium. Apoptosis was confirmed by DNA gel electrophoresis, transmission electron microscopy and fluorescence-activated cell sorting analysis, and there was no evidence for direct toxicity in the HSA-AGE preparations. The results are consistent with the proposed role of intra-molecular movement in exposing the anti-apoptotic domain in HSA for endothelium. The levels of AGE formation required to inhibit the anti-apoptotic activity of HSA exceeded those reported for diabetes. Nonetheless, the data from this study seems to be the first example of reduced protein function due to AGE-restricted intra-molecular movement.

Abbreviations

AGE:

advanced glycation end product

HSA:

human serum albumin

HUVEC:

human umbilical vein endothelial cells

PBS:

phosphate-buffered saline

SCS:

supplemented calf serum

References

  1. Araki, S., Shimada, Y., Kaji, K. and Hayashi, H. Role of protein kinase C in the inhibition by fibroblast growth factor of apoptosis in serum-deprived endothelial cells. Biochem. Biophys. Res. Commun. 172 (1990) 1081–1085.

    PubMed  Article  CAS  Google Scholar 

  2. Zoellner, H., Hou, J.Y., Lovery, M., Kingham, J., Srivastava, M., Bielek, E., Vanyek, E. and Binder, B.R. Inhibition of microvascular endothelial apoptosis in tissue explants by serum albumin. Microvasc. Res. 57 (1999) 162–173.

    PubMed  Article  CAS  Google Scholar 

  3. Zoellner, H., Hofler, M., Beckmann, R., Hufnagl, P., Vanyek, E., Bielek, E., Wojta, J., Fabry, A., Lockie, S. and Binder, B.R. Serum albumin is a specific inhibitor of apoptosis in human endothelial cells. J. Cell Sci. 109 (1996) 2571–2580.

    PubMed  CAS  Google Scholar 

  4. Bolitho, C., Bayl, P., Hou, J.Y., Lynch, G., Hassel, A.J., Wall, A.J. and Zoellner, H. The anti-apoptotic activity of albumin for endothelium is mediated by a partially cryptic protein domain and reduced by inhibitors of G-coupled protein and PI-3 kinase, but is independent of radical scavenging or bound lipid. J. Vasc. Res. 44 (2007) 313–324.

    PubMed  Article  CAS  Google Scholar 

  5. Djousse, L., Rothman, K.J., Cupples, L.A., Levy, D. and Ellison, R.C. Serum albumin and risk of myocardial infarction and all-cause mortality in the Framingham Offspring Study. Circulation 106 (2002) 2919–2924.

    PubMed  Article  CAS  Google Scholar 

  6. Schillinger, M., Exner, M., Mlekusch, W., Amighi, J., Sabeti, S., Schlager, O., Wagner, O. and Minar, E. Serum albumin predicts cardiac adverse events in patients with advanced atherosclerosis — interrelation with traditional cardiovascular risk factors. Thromb. Haemost. 91 (2004) 610–618.

    PubMed  CAS  Google Scholar 

  7. Peters, T. All about albumin–biochemistry, genetics and medical applications. San Diego, Academic Press, 1996.

    Google Scholar 

  8. Zoellner, H., Hou, J.Y., Hochgrebe, T., Poljak, A., Duncan, M.W., Golding, J., Henderson, T. and Lynch, G. Fluorometric and mass spectrometric analysis of nonenzymatic glycosylated albumin. Biochem. Biophys. Res. Commun. 284 (2001) 83–89.

    PubMed  Article  CAS  Google Scholar 

  9. Brownlee, M., Vlassara, H. and Cerami, A. Nonenzymatic glycosylation and the pathogenesis of diabetic complications. Ann. Intern. Med. 101 (1984) 527–537.

    PubMed  CAS  Google Scholar 

  10. John, W.G. and Lamb, E.J. The Maillard or browning reaction in diabetes. Eye 7 (Pt 2) (1993) 230–237.

    PubMed  Google Scholar 

  11. Drickamer, K. Diabetes: Breaking the curse of the AGEs. Nature 382 (1996) 211–212.

    PubMed  Article  CAS  Google Scholar 

  12. Lee, A.T. and Cerami, A. Role of glycation in aging. Ann. N. Y. Acad. Sci. 663 (1992) 63–70.

    PubMed  Article  CAS  Google Scholar 

  13. Ahmed, M.U., Thorpe, S.R. and Baynes, J.W. Identification of N epsilon-carboxymethyllysine as a degradation product of fructoselysine in glycated protein. J. Biol. Chem. 261 (1986) 4889–4894.

    PubMed  CAS  Google Scholar 

  14. Wolff, S.P., Jiang, Z.Y. and Hunt, J.V. Protein glycation and oxidative stress in diabetes mellitus and ageing. Free Radic. Biol. Med. 10 (1991) 339–352.

    PubMed  Article  CAS  Google Scholar 

  15. Johnson, R.N., Metcalf, P.A. and Baker, J.R. Fructosamine: a new approach to the estimation of serum glycosylprotein. An index of diabetic control. Clin. Chim. Acta 127 (1983) 87–95.

    PubMed  Article  CAS  Google Scholar 

  16. Emmanuel, C., Foo, E., Medbury, H., Matthews, J., Comis, A. and Zoellner, H. Synergistic induction of apoptosis in human endothelial cells by tumor necrosis factor-α and transforming growth factor-β. Cytokine 18 (2002) 237–241.

    PubMed  Article  CAS  Google Scholar 

  17. Smith, C.A., Williams, G.T., Kingston, R., Jenkinson, E.J. and Owen, J.J. Antibodies to CD3/T-cell receptor complex induce death by apoptosis in immature T-cells in thymic cultures. Nature 337 (1989) 181–184.

    PubMed  Article  CAS  Google Scholar 

  18. Darzynkiewicz, Z., Bruno, S., Del Bino, G., Gorczyca, W., Hotz, M.A., Lassota, P. and Traganos, F. Features of apoptotic cells measured by flow cytometry. Cytometry 13 (1992) 795–808.

    PubMed  Article  CAS  Google Scholar 

  19. Gerschenson, L.E. and Rotello, R.J. Apoptosis: a different type of cell death. FASEB J. 6 (1992) 2450–2455.

    PubMed  CAS  Google Scholar 

  20. Raff, M.C., Barres, B.A., Burne, J.F., Coles, H.S., Ishizaki, Y. and Jacobson, M.D. Programmed cell death and the control of cell survival: lessons from the nervous system. Science 262 (1993) 695–700.

    PubMed  Article  CAS  Google Scholar 

  21. Kumar, V., Abbas, A.K., Fausto, N. Robbins and Cotran Pathologic Basis of Disease. Philadelphia, W.B. Saunders Co., 2004.

    Google Scholar 

  22. Zoellner, H., Bielek, E., Vanyek, E., Fabry, A., Wojta, J., Hofler, M. and Binder, B.R. Canalicular fragmentation of apoptotic human endothelial cells. Endothelium 4 (1996) 177–188.

    Article  Google Scholar 

  23. Xu, W., Boadle, R., Dear, L., Cvejic, M., Emmanuel, C. and Zoellner, H. Ultrastructural changes in endothelium during apoptosis indicate low microembolic potential. J. Vasc. Res. 42 (2005) 377–387.

    PubMed  Article  Google Scholar 

  24. Chibber, R., Molinatti, P.A., Rosatto, N., Lambourne, B. and Kohner, E.M. Toxic action of advanced glycation end products on cultured retinal capillary pericytes and endothelial cells: relevance to diabetic retinopathy. Diabetologia 40 (1997) 156–164.

    PubMed  Article  CAS  Google Scholar 

  25. Min, C., Kang, E., Yu, S., Shinn, S. and Kim, Y. Advanced glycation end products induce apoptosis and procoagulant activity in cultured human umbilical vein endothelial cells. Diabetes Res. Clin. Pract. 46 (1999) 197–202.

    PubMed  Article  CAS  Google Scholar 

  26. Kowluru, R.A. Effect of advanced glycation end products on accelerated apoptosis of retinal capillary cells under in vitro conditions. Life Sci. 76 (2005) 1051–1060.

    PubMed  Article  CAS  Google Scholar 

  27. Xiang, M., Yang, M., Zhou, C., Liu, J., Li, W. and Qian, Z. Crocetin prevents AGEs-induced vascular endothelial cell apoptosis. Pharmacol. Res. 54 (2006) 268–274.

    PubMed  Article  CAS  Google Scholar 

  28. Stefani, M. Protein misfolding and aggregation: new examples in medicine and biology of the dark side of the protein world. Biochim. Biophys. Acta 1739 (2004) 5–25.

    PubMed  CAS  Google Scholar 

  29. Cantara, S., Ziche, M. and Donnini, S. Opposite effects of beta amyloid on endothelial cell survival: role of fibroblast growth factor-2 (FGF-2). Pharmacol. Rep. 57 Suppl (2005) 138–143.

    PubMed  Google Scholar 

  30. Cecchi, C., Pensalfini, A., Baglioni, S., Fiorillo, C., Caporale, R., Formigli, L., Liguri, G. and Stefani, M. Differing molecular mechanisms appear to underlie early toxicity of prefibrillar HypF-N aggregates to different cell types. FEBS J. 273 (2006) 2206–2222.

    PubMed  Article  CAS  Google Scholar 

  31. Bouma, B., Kroon-Batenburg, L.M., Wu, Y.P., Brunjes, B., Posthuma, G., Kranenburg, O., de Groot, P.G., Voest, E.E. and Gebbink, M.F. Glycation induces formation of amyloid cross-beta structure in albumin. J. Biol. Chem. 278 (2003) 41810–41819.

    PubMed  Article  CAS  Google Scholar 

  32. Rondeau, P., Singh, N.R., Caillens, H., Tallet, F., and Bourdon, E. Oxidative stress induced by glycoxidized human or bovine serum albumin on human monocytes. Free Radic. Biol. Med. 45 (2008) 799–812.

    PubMed  Article  CAS  Google Scholar 

  33. Hunt, J.V., Bottoms, M.A., and Mitchinson, M.J. Oxidative alterations in experimental glycation model of diabetes mellitus are due to protein-glucose adduct oxidation. Some fundamental differences in proposed mechanisms of glucose oxidation and oxidant production. Biochem. J. 291 (1993) 529–535.

    PubMed  CAS  Google Scholar 

  34. Darby, I.A., Bisucci, T., Hewitson, T.D. and MacLellan, D.G. Apoptosis is increased in a model of diabetes-impaired wound healing in genetically diabetic mice. Int. J. Biochem. Cell Biol. 29 (1997) 191–200.

    PubMed  Article  CAS  Google Scholar 

  35. Mizutani, M., Kern, T.S. and Lorenzi, M. Accelerated death of retinal microvascular cells in human and experimental diabetic retinopathy. J. Clin. Invest. 97 (1996) 2883–2890.

    PubMed  Article  CAS  Google Scholar 

  36. Thornalley, P.J., Argirova, M., Ahmed, N., Mann, V.M., Argirov, O. and Dawnay, A. Mass spectrometric monitoring of albumin in uremia. Kidney Int. 58 (2000) 2228–2234.

    PubMed  Article  CAS  Google Scholar 

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Correspondence to Hans Zoellner.

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Zoellner, H., Siddiqui, S., Kelly, E. et al. The anti-apoptotic activity of albumin for endothelium is inhibited by advanced glycation end products restricting intramolecular movement. Cell Mol Biol Lett 14, 575–586 (2009). https://doi.org/10.2478/s11658-009-0021-5

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  • DOI: https://doi.org/10.2478/s11658-009-0021-5

Key words

  • Apoptosis
  • Cryptic domain
  • Endothelium
  • HSA
  • HSA-AGE