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Trans-arachidonic acids generated during nitrative stress induce a thrombospondin-1–dependent microvascular degeneration

Abstract

Nitrative stress has an important role in microvascular degeneration leading to ischemia in conditions such as diabetic retinopathy and retinopathy of prematurity. Thus far, mediators of nitrative stress have been poorly characterized. We recently described that trans-arachidonic acids are major products of NO2-mediated isomerization of arachidonic acid within the cell membrane, but their biological relevance is unknown. Here we show that trans-arachidonic acids are generated in a model of retinal microangiopathy in vivo in a NO-dependent manner. They induce a selective time- and concentration-dependent apoptosis of microvascular endothelial cells in vitro, and result in retinal microvascular degeneration ex vivo and in vivo. These effects are mediated by an upregulation of the antiangiogenic factor thrombospondin-1, independently of classical arachidonic acid metabolism. Our findings provide new insight into the molecular mechanisms of nitrative stress in microvascular injury and suggest new therapeutic avenues in the management of disorders involving nitrative stress, such as ischemic retinopathies and encephalopathies.

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Figure 1: TAA levels rise secondary to nitrative stress and induce retinal microvascular degeneration.
Figure 2: TAAs induce selective neuromicrovascular endothelial cell death by apoptosis.
Figure 3: TAAs induce endothelial cell death by an ERK 1/2-dependent upregulation of TSP-1.
Figure 4: TAAs induce a TSP-1–dependent microvascular degeneration and inhibit of angiogenesis in tissue explants.
Figure 5: TAA-induced upregulation of TSP-1 in vivo and importance in hyperoxia-induced retinal microvascular degeneration.

References

  1. Lee, P., Wang, C.C. & Adamis, A.P. Ocular neovascularization: an epidemiologic review. Surv. Ophthalmol. 43, 245–269 (1998).

    Article  CAS  Google Scholar 

  2. Hardy, P. et al. Oxidants, nitric oxide and prostanoids in the developing ocular vasculature: a basis for ischemic retinopathy. Cardiovasc. Res. 47, 489–509 (2000).

    Article  CAS  Google Scholar 

  3. Campochiaro, P.A. Retinal and choroidal neovascularization. J. Cell. Physiol. 184, 301–310 (2000).

    Article  CAS  Google Scholar 

  4. Alon, T. et al. Vascular endothelial growth factor acts as a survival factor for newly formed retinal vessels and has implications for retinopathy of prematurity. Nat. Med. 1, 1024–1028 (1995).

    Article  CAS  Google Scholar 

  5. Pierce, E.A., Foley, E.D. & Smith, L.E. Regulation of vascular endothelial growth factor by oxygen in a model of retinopathy of prematurity. Arch. Ophthalmol. 114, 1219–1228 (1996).

    Article  CAS  Google Scholar 

  6. Smith, L.E. et al. Regulation of vascular endothelial growth factor-dependent retinal neovascularization by insulin-like growth factor-1 receptor. Nat. Med. 5, 1390–1395 (1999).

    Article  CAS  Google Scholar 

  7. Shih, S.C., Ju, M., Liu, N. & Smith, L.E. Selective stimulation of VEGFR-1 prevents oxygen-induced retinal vascular degeneration in retinopathy of prematurity. J. Clin. Invest. 112, 50–57 (2003).

    Article  CAS  Google Scholar 

  8. Wang, S., Wu, Z., Sorenson, C.M., Lawler, J. & Sheibani, N. Thrombospondin-1-deficient mice exhibit increased vascular density during retinal vascular development and are less sensitive to hyperoxia-mediated vessel obliteration. Dev. Dyn. 228, 630–642 (2003).

    Article  CAS  Google Scholar 

  9. Spierer, A., Rabinowitz, R., Pri-Chen, S. & Rosner, M. An increase in superoxide dismutase ameliorates oxygen-induced retinopathy in transgenic mice. Eye 19, 86–91 (2005).

    Article  CAS  Google Scholar 

  10. Kowluru, R.A., Tang, J. & Kern, T.S. Abnormalities of retinal metabolism in diabetes and experimental galactosemia. VII. Effect of long-term administration of antioxidants on the development of retinopathy. Diabetes 50, 1938–1942 (2001).

    Article  CAS  Google Scholar 

  11. Penn, J.S., Tolman, B.L. & Bullard, L.E. Effect of a water-soluble vitamin E analog, trolox C, on retinal vascular development in an animal model of retinopathy of prematurity. Free Radic. Biol. Med. 22, 977–984 (1997).

    Article  CAS  Google Scholar 

  12. Raju, T.N., Langenberg, P., Bhutani, V. & Quinn, G.E. Vitamin E prophylaxis to reduce retinopathy of prematurity: a reappraisal of published trials. J. Pediatr. 131, 844–850 (1997).

    Article  CAS  Google Scholar 

  13. Squadrito, G.L. & Pryor, W.A. Oxidative chemistry of nitric oxide: the roles of superoxide, peroxynitrite, and carbon dioxide. Free Radic. Biol. Med. 25, 392–403 (1998).

    Article  CAS  Google Scholar 

  14. Kroncke, K.D. Mechanisms and biological consequences of nitrosative stress. Biol. Chem. 384, 1341 (2003).

    Article  Google Scholar 

  15. Gu, X. et al. Hyperoxia induces retinal vascular endothelial cell apoptosis through formation of peroxynitrite. Am. J. Physiol. Cell Physiol. 285, C546–C554 (2003).

    Article  CAS  Google Scholar 

  16. Beauchamp, M.H. et al. Redox-dependent effects of nitric oxide on microvascular integrity in oxygen-induced retinopathy. Free Radic. Biol. Med. 37, 1885–1894 (2004).

    Article  CAS  Google Scholar 

  17. El-Remessy, A.B., Abou-Mohamed, G., Caldwell, R.W. & Caldwell, R.B. High glucose-induced tyrosine nitration in endothelial cells: role of eNOS uncoupling and aldose reductase activation. Invest. Ophthalmol. Vis. Sci. 44, 3135–3143 (2003).

    Article  Google Scholar 

  18. Brooks, S.E. et al. Reduced severity of oxygen-induced retinopathy in eNOS-deficient mice. Invest. Ophthalmol. Vis. Sci. 42, 222–228 (2001).

    CAS  PubMed  Google Scholar 

  19. El-Remessy, A.B. et al. Experimental diabetes causes breakdown of the blood-retina barrier by a mechanism involving tyrosine nitration and increases in expression of vascular endothelial growth factor and urokinase plasminogen activator receptor. Am. J. Pathol. 162, 1995–2004 (2003).

    Article  CAS  Google Scholar 

  20. Jiang, H. et al. Nitrogen dioxide induces cis-trans-isomerization of arachidonic acid within cellular phospholipids. Detection of trans-arachidonic acids in vivo. J. Biol. Chem. 274, 16235–16241 (1999).

    Article  CAS  Google Scholar 

  21. Balazy, M. & Poff, C.D. Biological nitration of arachidonic acid. Curr. Vasc. Pharmacol. 2, 81–93 (2004).

    Article  CAS  Google Scholar 

  22. Balazy, M. Trans-arachidonic acids: new mediators of inflammation. J. Physiol. Pharmacol. 51, 597–607 (2000).

    CAS  PubMed  Google Scholar 

  23. Kirsch, M., Korth, H.G., Sustmann, R. & de Groot, H. The pathobiochemistry of nitrogen dioxide. Biol. Chem. 383, 389–399 (2002).

    Article  CAS  Google Scholar 

  24. Prutz, W.A., Monig, H., Butler, J. & Land, E.J. Reactions of nitrogen dioxide in aqueous model systems: oxidation of tyrosine units in peptides and proteins. Arch. Biochem. Biophys. 243, 125–134 (1985).

    Article  CAS  Google Scholar 

  25. Chan-Ling, T., Gock, B. & Stone, J. The effect of oxygen on vasoformative cell division. Evidence that 'physiological hypoxia' is the stimulus for normal retinal vasculogenesis. Invest. Ophthalmol. Vis. Sci. 36, 1201–1214 (1995).

    CAS  PubMed  Google Scholar 

  26. Smith, L.E. Pathogenesis of retinopathy of prematurity. Growth Horm. IGF Res. 14 Suppl A, 140–4 (2004).

    Article  Google Scholar 

  27. Caffe, A.R. et al. Mouse retina explants after long-term culture in serum free medium. J. Chem. Neuroanat. 22, 263–273 (2001).

    Article  CAS  Google Scholar 

  28. Beauchamp, M.H. et al. Role of thromboxane in retinal microvascular degeneration in oxygen-induced retinopathy. J. Appl. Physiol. 90, 2279–2288 (2001).

    Article  CAS  Google Scholar 

  29. Sennlaub, F. et al. Cyclooxygenase-2 in human and experimental ischemic proliferative retinopathy. Circulation 108, 198–204 (2003).

    Article  CAS  Google Scholar 

  30. Roy, U., Loreau, O. & Balazy, M. Cytochrome P450/NADPH-dependent formation of trans epoxides from trans-arachidonic acids. Bioorg. Med. Chem. Lett. 14, 1019–1022 (2004).

    Article  CAS  Google Scholar 

  31. Nor, J.E. et al. Thrombospondin-1 induces endothelial cell apoptosis and inhibits angiogenesis by activating the caspase death pathway. J. Vasc. Res. 37, 209–218 (2000).

    Article  CAS  Google Scholar 

  32. Armstrong, L.C. & Bornstein, P. Thrombospondins 1 and 2 function as inhibitors of angiogenesis. Matrix Biol. 22, 63–71 (2003).

    Article  CAS  Google Scholar 

  33. Guo, N., Krutzsch, H.C., Inman, J.K. & Roberts, D.D. Thrombospondin 1 and type I repeat peptides of thrombospondin 1 specifically induce apoptosis of endothelial cells. Cancer Res. 57, 1735–1742 (1997).

    CAS  PubMed  Google Scholar 

  34. Dawson, D.W. et al. CD36 mediates the in vitro inhibitory effects of thrombospondin-1 on endothelial cells. J. Cell Biol. 138, 707–717 (1997).

    Article  CAS  Google Scholar 

  35. Jimenez, B. et al. Signals leading to apoptosis-dependent inhibition of neovascularization by thrombospondin-1. Nat. Med. 6, 41–48 (2000).

    Article  CAS  Google Scholar 

  36. Swerlick, R.A., Lee, K.H., Wick, T.M. & Lawley, T.J. Human dermal microvascular endothelial but not human umbilical vein endothelial cells express CD36 in vivo and in vitro. J. Immunol. 148, 78–83 (1992).

    CAS  PubMed  Google Scholar 

  37. Wada, T. & Penninger, J.M. Mitogen-activated protein kinases in apoptosis regulation. Oncogene 23, 2838–2849 (2004).

    Article  CAS  Google Scholar 

  38. Ishikawa, Y. & Kitamura, M. Dual potential of extracellular signal-regulated kinase for the control of cell survival. Biochem. Biophys. Res. Commun. 264, 696–701 (1999).

    Article  CAS  Google Scholar 

  39. Gauld, S.B., Blair, D., Moss, C.A., Reid, S.D. & Harnett, M.M. Differential roles for extracellularly regulated kinase-mitogen-activated protein kinase in B cell antigen receptor-induced apoptosis and CD40-mediated rescue of WEHI-231 immature B cells. J. Immunol. 168, 3855–3864 (2002).

    Article  CAS  Google Scholar 

  40. Zghibeh, C.M., Raj Gopal, V., Poff, C.D., Falck, J.R. & Balazy, M. Determination of trans-arachidonic acid isomers in human blood plasma. Anal. Biochem. 332, 137–144 (2004).

    Article  CAS  Google Scholar 

  41. Llorens, S. & Nava, E. Cardiovascular diseases and the nitric oxide pathway. Curr. Vasc. Pharmacol. 1, 335–346 (2003).

    Article  CAS  Google Scholar 

  42. Liu, L. et al. Essential roles of S-nitrosothiols in vascular homeostasis and endotoxic shock. Cell 116, 617–628 (2004).

    Article  CAS  Google Scholar 

  43. Radi, R., Rodriguez, M., Castro, L. & Telleri, R. Inhibition of mitochondrial electron transport by peroxynitrite. Arch. Biochem. Biophys. 308, 89–95 (1994).

    Article  CAS  Google Scholar 

  44. Tabuchi, A, Oh, E, Taoka, A, Sakurai, H, Tsuchiya, T & Tsuda . Rapid attenuation of AP-1 transcriptional factors associated with nitric oxide (NO)-mediated neuronal cell death. J Biol. Chem. 271, 31061–7 (1996).

    Article  CAS  Google Scholar 

  45. Dameron, K.M., Volpert, O.V., Tainsky, M.A. & Bouck, N. Control of angiogenesis in fibroblasts by p53 regulation of thrombospondin-1. Science 265, 1582–1584 (1994).

    Article  CAS  Google Scholar 

  46. Okuno, M., Arimoto, E., Nishizuka, M., Nishihara, T. & Imagawa, M. Isolation of up- or down-regulated genes in PPARgamma-expressing NIH-3T3 cells during differentiation into adipocytes. FEBS Lett. 519, 108–112 (2002).

    Article  CAS  Google Scholar 

  47. Nielsen, J.C., Naash, M.I. & Anderson, R.E. The regional distribution of vitamins E and C in mature and premature human retinas. Invest. Ophthalmol. Vis. Sci. 29, 22–26 (1988).

    CAS  PubMed  Google Scholar 

  48. Flynn, J.T. et al. A cohort study of transcutaneous oxygen tension and the incidence and severity of retinopathy of prematurity. N. Engl. J. Med. 326, 1050–1054 (1992).

    Article  CAS  Google Scholar 

  49. Mann, R.M., Riva, C.E., Stone, R.A., Barnes, G.E. & Cranstoun, S.D. Nitric oxide and choroidal blood flow regulation. Invest. Ophthalmol. Vis. Sci. 36, 925–930 (1995).

    CAS  PubMed  Google Scholar 

  50. Lahaie, I. et al. A novel mechanism for vasoconstrictor action of 8-isoprostaglandin F2 alpha on retinal vessels. Am. J. Physiol. 274, R1406–R1416 (1998).

    CAS  PubMed  Google Scholar 

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Acknowledgements

The authors wish to thank H. Fernandez for her technical skills and help. This work was supported by grants from the Canadian Institutes of Health Research, the March of Dimes Birth Defects Foundation, the Heart and Stroke Foundation of Québec, the Fonds de la Recherche en Santé du Québec, Le Réseau de Recherche en Santé de la Vision and La Fondation du NO. E.K.-D. is recipient of a fellowship award from the 'Association des Juniors en Pédiatrie/Gallia' (France). F.S. and S.C. are recipients of fellowship and scientist awards, respectively, from the Canadian Institutes of Health Research. S.B. and M.S. are recipients of studentships from the Canadian Institutes of Health Research and Heart and Stroke Foundation of Canada, respectively. P.H. is supported by grants from the Hospital For Sick Children Foundation and Fonds de la Recherche en Santé du Québec. M.B. is supported by grants from the US National Institutes of Health (R01 GM62453) and Philip Morris USA, Inc. S.C. also holds a Canada Research Chair (perinatology). The authors thank M. Febbraio and J. Lawler, who provided the CD36 and TSP-1 knockout animals, respectively.

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Correspondence to Michael Balazy or Sylvain Chemtob.

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Supplementary Fig. 1

TAA-induced endothelial cell death is independent of classic arachidonic acid pathways. (PDF 53 kb)

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Kermorvant-Duchemin, E., Sennlaub, F., Sirinyan, M. et al. Trans-arachidonic acids generated during nitrative stress induce a thrombospondin-1–dependent microvascular degeneration. Nat Med 11, 1339–1345 (2005). https://doi.org/10.1038/nm1336

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