Molecular and Cellular Biochemistry

, Volume 232, Issue 1–2, pp 113–119 | Cite as

Regulation of ecto-5′-nucleotidase by TNF-α in human endothelial cells

  • Kameljit Kalsi
  • Charlotte Lawson
  • Martin Dominguez
  • Patricia Taylor
  • Magdi H. Yacoub
  • Ryszard T. Smolenski
Article

Abstract

Ecto-5′-nucleotidase (E5′N, CD73) is key enzyme responsible for formation of anti-inflammatory and immunosuppressive adenosine from extracellular nucleotides as well as an important surface molecule involved in cellular signalling. In this study we provide evidence that the pro-inflammatory cytokine, tumour necrosis factor-α (TNF-α) may reduce the capacity of human endothelial cells to produce adenosine by a decrease in surface expression and in the activity of E5′N. Human umbilical vein endothelial cells incubated for 24 h with TNF-α lost 54% of the activity of E5′N while activities of the other enzymes involved in adenosine metabolism remained unaffected. Immunofluorescence staining with anti-E5′N (1E9) following exposure to TNF-α, showed reduced numbers of positive cells. TNF-α induced down-regulation of E5′N was prevented by addition of the PLC inhibitor neomycin, but not by inhibitors of MAPK-like pathways (MEK and p38). Therefore, we conclude that TNF-α through activation of endogenous PLC leads to cleavage of the GPI-linkage of E5′N resulting in loss of E5′N from the extracellular surface. This change may lead to decrease in formation of adenosine and could be an important mechanism of endothelial activation during inflammation.

ecto-5′-nucleotidase TNF-α adenosine endothelium phospholipase C 

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References

  1. 1.
    Baron MD, Pope B, Luzio JP: The membrane topography of ecto-5′-nucleotidase in rat hepatocytes. Biochem J 236: 495–502, 1986Google Scholar
  2. 2.
    Naito Y, Lowenstein JM: 5′-Nucleotidase from rat heart. Biochemistry 20: 5188–5194, 1981Google Scholar
  3. 3.
    Berne RM: Cardiac nucleotides in hypoxia: Possible role in regulation of coronary blood flow. Am J Physiol 204: 317–322, 1963Google Scholar
  4. 4.
    Angielski S, Redlak M, Szczepanska-Konkel M: Intrarenal adenosine prevents hyperfiltration induced by atrial natriuretic factor. Miner Electrolyte Metab 16: 57–60, 1990Google Scholar
  5. 5.
    Daval JL, Nehlig A, Nicolas F: Physiological and pharmacological properties of adenosine: Therapeutic implications. Life Sci 49: 1435–1453, 1991Google Scholar
  6. 6.
    Edwards NL, Gelfand EW, Burk L, Dosch HM, Fox IH: Distribution of 5′-nucleotidase in human lymphoid tissues. Proc Natl Acad Sci USA 76: 3474–3476, 1979Google Scholar
  7. 7.
    Deguchi H, Takeya H, Urano H, Gabazza EC, Zhou H, Suzuki K: Adenosine regulates tissue factor expression on endothelial cells. Thromb Res 91: 57–64, 1998Google Scholar
  8. 8.
    Meldrum DR: Tumor necrosis factor in the heart. Am J Physiol 274: R577–R595, 1998Google Scholar
  9. 9.
    Muller G, Storz P, Bourteele S, Doppler H, Pfizenmaier K, Mischak H, Philipp A, Kaiser C, Kolch W: Regulation of Raf-1 kinase by TNF via its second messenger ceramide and cross-talk with mitogenic signalling. EMBO J 17: 732–742, 1998Google Scholar
  10. 10.
    Ghosh S, Strum JC, Bell RM: Lipid biochemistry: Functions of glycerolipids and sphingolipids in cellular signaling. FASEB J 11: 45–50, 1997Google Scholar
  11. 11.
    Rapuano BE, Bockman RS: Tumor necrosis factor-alpha stimulates phosphatidylinositol breakdown by phospholipase C to coordinately increase the levels of diacylglycerol, free arachidonic acid and prostaglandins in an osteoblast (MC3T3-E1) cell line. Biochim Biophys Acta 1091: 374–384, 1991Google Scholar
  12. 12.
    Jaffe EA, Nachman RL, Becker CG, Minick CR: Culture of human endothelial cells derived from umbilical veins. Identification by morphologic and immunologic criteria. J Clin Invest 52: 2745–2756, 1973Google Scholar
  13. 13.
    Hengstenberg C, Rose ML, Page C, Taylor PM, Yacoub MH: Immunocytochemical changes suggestive of damage to endothelial cells during rejection of human cardiac allografts. Transplantation 49: 895–899, 1990Google Scholar
  14. 14.
    Kochan Z, Smolenski RT, Yacoub MH, Seymour A-ML: Nucleotide and adenosine metabolism in different cell types in human and rat heart. J Mol Cell Cardiol 26: 1497–1503, 1994Google Scholar
  15. 15.
    Skladanowski AC, Smolenski RT, Tavenier M, de Jong JW, Yacoub MH, Seymour A M: Soluble forms of 5′-nucleotidase in rat and human heart. Am J Physiol 270: H1493–H1500, 1996Google Scholar
  16. 16.
    Smolenski RT, Lachno DR, Ledingham SJM, Yacoub MH: Determination of sixteen nucleotides, nucleosides and bases using high-performance liquid chromatography and its application to the study of purine metabolism in hearts for transplantation. J Chromatogr 527: 414–420, 1990Google Scholar
  17. 17.
    Bradford MM: A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72: 248–254, 1976Google Scholar
  18. 18.
    Wilson AC, Cahn RD, Kaplan NO: Functions of the two forms of lactic dehydrogenase in the breast muscle of birds. Nature 197: 331–334, 1963Google Scholar
  19. 19.
    Chomczynski P: One-hour downward alkaline capillary transfer for blotting of DNA and RNA. Anal Biochem 201: 134–139, 1992Google Scholar
  20. 20.
    Misumi Y, Ogata S, Hirose S, Ikehara Y: Primary structure of rat liver 5′-nucleotidase deduced from the cDNA. Presence of the COOH-terminal hydrophobic domain for possible post-translational modification by glycophospholipid. J Biol Chem 265: 2178–2183, 1990Google Scholar
  21. 21.
    Franco R, Casado V, Ciruela F, Saura C, Mallol J, Canela EI, Lluis C: Cell surface adenosine deaminase: Much more than an ectoenzyme. Prog Neurobiol 52: 283–294, 1997Google Scholar
  22. 22.
    Dudley DT, Pang L, Decker SJ, Bridges AJ, Saltiel AR: A synthetic inhibitor of the mitogen-activated protein kinase cascade. Proc Natl Acad Sci USA 92: 7686–7689, 1995Google Scholar
  23. 23.
    Cuenda A, Rouse J, Doza YN, Meier R, Cohen P, Gallagher TF, Young PR, Lee JC: SB 203580 is a specific inhibitor of a MAP kinase homologue which is stimulated by cellular stresses and interleukin-1. FEBS Lett 364: 229–233, 1995Google Scholar
  24. 24.
    Savic V, Stefanovic V, Ardaillou N, Ardaillou R: Induction of ecto-5′-nucleotidase of rat cultured mesangial cells by interleukin-1 beta and tumour necrosis factor-alpha. Immunology 70: 321–326, 1990Google Scholar
  25. 25.
    Klip A, Ramlal T, Douen AG, Burdett E, Young D, Cartee GD, Holloszy JO: Insulin-induced decrease in 5′-nucleotidase activity in skeletal muscle membranes. FEBS Lett 238: 419–423, 1988Google Scholar
  26. 26.
    Incerpi S, Baldini P, Lo BM, Luly P: Insulin-dependent release of 5′-nucleotidase and alkaline phosphatase from liver plasma membranes. Biosci Rep 12: 101–108, 1992Google Scholar
  27. 27.
    Sharom FJ, McNeil GL, Glover JR, Seier S: Modulation of the cleavage of glycosylphosphatidylinositol-anchored proteins by specific bacterial phospholipases. Biochem Cell Biol 74: 701–713, 1996Google Scholar
  28. 28.
    Lehto MT, Sharom FJ: Release of the glycosylphosphatidylinositol-anchored enzyme ecto-5′-nucleotidase by phospholipase C: Catalytic activation and modulation by the lipid bilayer. Biochem J 332: 101–109, 1998Google Scholar
  29. 29.
    Siegfried G, Amiel C, Friedlander G: Inhibition of ecto-5′-nucleotidase by nitric oxide donors. Implications in renal epithelial cells. J Biol Chem 271: 4659–4664, 1996Google Scholar
  30. 30.
    Bhagyalakshmi A, Berthiaume F, Reich KM, Frangos JA: Fluid shear stress stimulates membrane phospholipid metabolism in cultured human endothelial cells. J Vasc Res 29: 443–449, 1992Google Scholar
  31. 31.
    Yegutkin G, Bodin P, Burnstock G: Effect of shear stress on the release of soluble ecto-enzymes ATPase and 5′-nucleotidase along with endogenous ATP from vascular endothelial cells. Br J Pharmacol 129: 921–926, 2000Google Scholar
  32. 32.
    Cronstein BN, Levin RI, Philips M, Hirschhorn R, Abramson SB, Weissmann G: Neutrophil adherence to endothelium is enhanced via adenosine A1 receptors and inhibited via adenosine A2 receptors. J Immunol 148: 2201–2206, 1992Google Scholar
  33. 33.
    Kitakaze M, Hori M, Kamada T: Role of adenosine and its interaction with alpha adrenoceptor activity in ischaemic and reperfusion injury of the myocardium. Cardiovasc Res 27: 18–27, 1993Google Scholar
  34. 34.
    Lieberman GE, Lewis GP, Peters TJ: A membrane-bound enzyme in rabbit aorta capable of inhibiting adenosine-diphosphate-induced platelet aggregation. Lancet 2: 330–332, 1977Google Scholar
  35. 35.
    Imai M, Goepfert C, Kaczmarek E, Robson SC: CD39 modulates IL-1 release from activated endothelial cells. Biochem Biophys Res Commun 270: 272–278, 2000Google Scholar
  36. 36.
    Siwik DA, Chang DL, Colucci WS: Interleukin-1beta and tumor necrosis factor-alpha decrease collagen synthesis and increase matrix metalloproteinase activity in cardiac fibroblasts in vitro. Circ Res 86: 1259–1265, 2000Google Scholar
  37. 37.
    Mayne M, Shepel PN, Jiang Y, Geiger JD, Power C: Dysregulation of adenosine A1 receptor-mediated cytokine expression in peripheral blood mononuclear cells from multiple sclerosis patients. Ann Neurol 45: 633–639, 1999Google Scholar

Copyright information

© Kluwer Academic Publishers 2002

Authors and Affiliations

  • Kameljit Kalsi
  • Charlotte Lawson
  • Martin Dominguez
  • Patricia Taylor
  • Magdi H. Yacoub
  • Ryszard T. Smolenski

There are no affiliations available

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