Skip to main content
SpringerLink
Log in

Disruption of Thiol Homeostasis and Nitrosative Stress in the Cerebrospinal Fluid of Patients with Active Multiple Sclerosis: Evidence for a Protective Role of Acetylcarnitine

  • Published:
Neurochemical Research Aims and scope Submit manuscript

Abstract

Recent studies suggest that NO and its reactive derivative peroxynitrite are implicated in the pathogenesis of multiple sclerosis (MS). Patients dying with MS demonstrate increased astrocytic inducible nitric oxide synthase activity, as well as increased levels of iNOS mRNA. Peroxynitrite is a strong oxidant capable of damaging target tissues, particularly the brain, which is known to be endowed with poor antioxidant buffering capacity. Inducible nitric oxide synthase is upregulated in the central nervous system (CNS) of animals with experimental allergic encephalomyelitis (EAE) and in patients with MS. We have recently demonstrated in patients with active MS a significant increase of NOS activity associated with increased nitration of proteins in the cerebrospinal fluid (CSF). Acetylcarnitine is proposed as a therapeutic agent for several neurodegenerative disorders. Accordingly, in the present study, MS patients were treated for 6 months with acetylcarnitine and compared with untreated MS subjects or with patients noninflammatory neurological conditions, taken as controls. Western blot analysis showed in MS patients increased nitrosative stress associated with a significant decrease of reduced glutathione (GSH). Increased levels of oxidized glutathione (GSSG) and nitrosothiols were also observed. Interestingly, treatment of MS patients with acetylcarnitine resulted in decreased CSF levels of NO reactive metabolites and protein nitration, as well as increased content of GSH and GSH/GSSG ratio. Our data sustain the hypothesis that nitrosative stress is a major consequence of NO produced in MS-affected CNS and implicate a possible important role for acetylcarnitine in protecting brain against nitrosative stress, which may underlie the pathogenesis of MS.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

REFERENCES

  1. Hellings, N., Raus, J., and Stinissen, P. 2002. Insights into the immunopathogenesis of multiple sclerosis. Immunol. Res. 25: 27-51.

    Google Scholar 

  2. Smith, K. J. and McDonald, W. I. 1999. The pathophysiology of multiple sclerosis: The mechanisms underlying the production of symptoms and the natural history of the disease. Philos. Trans. R. Soc. Lond. B. Biol. Sci. 354:1649-1673.

    Google Scholar 

  3. Bagasra, O., Michaels, F. H., Zheng, Y. M., Bobroski, L. E., Spitsin, S. V., Fu, Z. F., Tawadros, R., and Koprowski, H. 1995 Activation of the inducible form of nitric oxide synthase in the brains of patients with multiple sclerosis. Proc. Natl. Acad. Sci. USA 92:12041-12045.

    Google Scholar 

  4. Cross, A. H., Manning, P. T., Keeling, R. M., Schmidt, R. E., and Misko, T. P. 1998. Peroxynitrite formation within the central nervous system in active multiple sclerosis. J. Neuroimmunol. 88:45-56.

    Google Scholar 

  5. Cross, A. H., Manning, P. T., Stern, M. K., and Misko, T. P. 1997. Evidence for the production of peroxynitrite in inflammatory CNS demyelination. J. Neuroimmunol. 80:121-130.

    Google Scholar 

  6. Tran, E. H., Hardin, P. H., Verge, G., and Owens, T. 1997. Astrocytes and microglia express inducible nitric oxide synthase in mice with experimental allergic encephalomyelitis. J. Neuroimmunol. 74:121-129.

    Google Scholar 

  7. Heales, S. J., Davies, S. E., Bates, T. E., and Clark, J. B. 1995. Depletion of brain glutathione is accompanied by impaired mitochondrial function and decreased N-acetyl aspartate concentration. Neurochem. Res. 20:31-38.

    Google Scholar 

  8. Bates, T. E., Heales, S. J., Davies, S. E., Boakye, P., and Clark, J. B. 1994. Effects of 1-Methyl-4-phenylpyridinium on isolated rat brain mitochondria: Evidence for a primary involvement of energy depletion. J. Neurochem. 63:640-648.

    Google Scholar 

  9. Martin, R. and McFarland, H. F. 1995. Immunological aspects of experimental allergic encephalomyelitis and multiple sclerosis. Crit. Rev. Clin. Lab. Sci. 32:121-182.

    Google Scholar 

  10. Lindsey, J. W., Kerman, R. H., and Wolinsky, J. S. 1997. T cell-T cell activation in multiple sclerosis. Mult. Scler. 3:238-242.

    Google Scholar 

  11. Smith, K. J., Kapoor, R., and Felts, P. A. 1999. Demyelination: The role of reactive oxygen and nitrogen species. Brain Pathol. 9:69-92.

    Google Scholar 

  12. Moss, D. W. and Bates, T. E. 2001. Activation of murine microglial cell lines by lipopolysaccharide and interferon-χ causes NO-mediated decreases in mitochondrial and cellular function. Eur. J. Neurosci. 13:529-538.

    Google Scholar 

  13. Aquino, D. A., Capello, E., Weisstein, J., Sanders, V., Lopez, C., Tourtellotte, W. W., Brosnan, C. F., Raine, C. S., and Norton, W. T. 1997. Multiple sclerosis: Altered expression of 70-and 27-kDa heat shock proteins in lesions and myelin. J. Neuropathol. Exp. Neurol. 56:664-672.

    Google Scholar 

  14. Calabrese, V., Scapagnini, G., Giuffrida Stella, A. M., Bates, T. E., and Clark, J. B. 2001. Mitochondrial involvement in brain function and dysfunction: Relevance to aging, neurodegenerative disorders and longevity. Neurochem. Res. 26:739-764.

    Google Scholar 

  15. Calabrese, V., Scapagnini, G., Catalano, D., Dinotta, F., Bates, T. E., Calvani, M., and Giuffrida Stella, A. M. 2001. Effects of acetyl-L-carnitine on the formation of fatty acid ethyl esters in brain and peripheral organs after short-term ethanol administration in rat. Neurochem. Res. 67:1315-1341.

    Google Scholar 

  16. Calabrese, V., Raffaele, R., Cosentino, E., and Rizza, V. 1994. Changes in cerebrospinal fluid levels of malonaldehyde and glutathione reductase activity in multiple sclerosis. Int. J. Clin. Pharmacol. Res. 4:119-123.

    Google Scholar 

  17. Thompson, E. J. 1988. The CSF proteins: A biochemical approach (Pager 16-24). London, Elsevier.

    Google Scholar 

  18. Calabrese, V., Scapagnini, G., Ravagna, A., Bella, R., Foresti, R., Bates, T., Giuffrida Stella, A. M., and Pennisi, G. 2002. Nitric oxide synthase is present in the cerebrospinal fluid of patients with active multiple sclerosis and is associated with increases in CSF protein nitrotyrosine, S-nitrosothiols and with changes in glutathione levels. J. Neurosci. Res. 70:580-587.

    Google Scholar 

  19. Poser, C. M., Paty, D. W., Scheinberg, L., McDonald, W. I., Davis, F. A., Ebers, G. C., Johnson, K. P., Sibley, W. A., Silberberg, D. H., and Tourtellotte, W. W. 1983. New diagnostic criteria for multiple sclerosis: Guidelines for research protocols. Ann. Neurol. 13:227-231.

    Google Scholar 

  20. Lublin, F. D. and Reingold, S. C. 1996. Defining the clinical course of multiple sclerosis: Results of an international survey—National Multiple Sclerosis Society (USA) Advisory Committee on Clinical Trials of New Agents in Multiple Sclerosis. Neurology 46:907-911.

    Google Scholar 

  21. Kurtzke, J. F. 1983. Rating neurologic impairment in multiple sclerosis: An expanded disability status scale (EDSS). Neurology 33:1444-1452.

    Google Scholar 

  22. Laemmli, U. K. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680-685.

    Google Scholar 

  23. Adams, J. D., Lauterburg, B. H., and Mitchell, J. R. 1983. Plasma glutathione and glutathione disulfide in the rat: Regulation and response to oxidative stress. J. Pharmacol. Exp. Ther. 227: 749-754.

    Google Scholar 

  24. Radi, R., Cosgrove, T. P., Beckman, J. S., and Freeman, B. A. 1993. Peroxynitrite-induced luminol chemiluminescence. Biochem. J. 290:51-57.

    Google Scholar 

  25. Kojima, H., Kikuchi, K., Hirobe, M., and Nagano, T. 1997. Real-time measurement of nitric oxide production in rat brain by the combination of luminol-H2O2 chemiluminescence and microdialysis. Neurosci. Lett. 233:157-159.

    Google Scholar 

  26. Flecha, B., Llesuy, S., and Boveris, A. 1991. Hydroperoxide-initiated chemiluminescence: An assay for oxidative stress in biopsies of heart, liver, and muscle. Free Rad. Biol. Med. 10:93-96.

    Google Scholar 

  27. Hevel, J. and Marletta, M. 1994. Oxygen radicals in biological systems. Methods Enzymol. 233:250-258.

    Google Scholar 

  28. Southan, G. J., Szabo, C., and Thiemermann, C. 1995. Isothioureas: Potent inhibitors of nitric oxide synthases with variable isoform selectivity. British J. Pharmacol. 114:510-516.

    Google Scholar 

  29. Griffith, O. W. and Stuehr, D. J. 1995. Nitric oxide synthases: Properties and catalytic mechanism. Annu. Rev. Physiol. 57: 707-736.

    Google Scholar 

  30. Granger, D. L., Taintor, R. R., Boockvar, K. S., and Hibbs, J. B. 1996 Measurement of nitrate and nitrite in biological samples using nitrate reductase and Griess reaction. Methods Enzymol. 268:142-151.

    Google Scholar 

  31. Smith, P. K., Krohn, R. I., Hermanson, G. T., Mallia, A. K., Gartner, F. H., Provenzano, M. D., Fujimoto, E. K., Goeke, N. M., Olson, B. J., and Klenk, D. C. 1985. Measurement of protein using bicinchoninic acid. Anal. Biochem. 150:76-85.

    Google Scholar 

  32. Butterfield, D. A. and Lauderback, C. M. 2002. Lipid peroxidation and protein oxidation in Alzheimer's disease brain: Potential causes and consequences involving amyloid β-peptide-associated free Radical oxidative stress. Free Rad. Biol. Med. 32:1050-1060

    Google Scholar 

  33. Drake, J., Kanski, J., Varadarajan, S., Tsoras, M., and Butterfield, D. A. 2002. Elevation of brain glutathione by γ-glutamylcysteine ethyl ester protects against peroxynitrite-induced oxidative stress. J. Neurosci. Res. 68:776-784.

    Google Scholar 

  34. Castegna, A., Aksenov, M., Aksenova, M., Thongboonkerd, V., Klein, J. B., Pierce, W. M., Booze, R., Markesbery, W. R., and Butterfield, D. A. 2002. Proteomic identification of oxidatively modified proteins in Alzheimer's disease brain: I. Creatine kinase BB, glutamine synthase, and ubiquitin carboxy-terminal hydrolase L-1. Free Rad. Biol. Med. 33:562-571.

    Google Scholar 

  35. Giovannoni, G. 1998. Cerebrospinal fluid and serum nitric oxide metabolites in patients with multiple sclerosis. Mult. Scler. 4: 27-30.

    Google Scholar 

  36. Meli, R., Nauser, T., Latal, P., and Koppenol, W. H. 2002. Reaction of peroxynitrite with carbon dioxide: Intermediates and determination of the yield of CO3*- and NO2*. J. Biol. Inorg. Chem. 7:31-36.

    Google Scholar 

  37. Calabrese, V., Copani, A., Testa, D., Ravagna, A., Spadaro, F., Tendi, E., Nicoletti, V. G., and Giuffrida Stella, A. M. 2000. Nitric oxide synthase induction in astroglial cell cultures: Effect on heat shock protein 70 synthesis and oxidant/antioxidant balance. J. Neurosci. Res. 60:613-622.

    Google Scholar 

  38. Calabrese, V., Scapagnini, G., Ravagna, A., Fariello, R. G., Giuffrida Stella, A. M., and Abraham, N. 2002. Regional distribution of heme oxygenase, hsp 70, and glutathione in brain: Relevance for endogenous oxidant/antioxidant balance and stress tolerance. J. Neurosci. Res. 68:65-75.

    Google Scholar 

  39. Calabrese, V., Bates, T. E., and Giuffrida Stella, A. M. 2000. NO synthase and NO-dependent signal pathways in brain aging and Neurodegenerative disorders: The role of oxidant/antioxidant balance. Neurochemical Research 25:1315-1341.

    Google Scholar 

  40. Butterfield, D. A., Castegna, A., Drake, J., Scapagnini, G., and Calabrese, V. 2002. Vitamin E and neurodegenerative disorders associated with oxidative stress. Nutr. Neurosci. 5:229-239

    Google Scholar 

  41. Calabrese, V., Scapagnini, G., Ravagna, A., Giuffrida Stella, A. M., and Butterfield, A. 2002. Molecular chaperones and their roles in neural cell ifferentiation. Dev. Neurosci. 24:40-56.

    Google Scholar 

  42. Calabrese, V., Renis, M., Calderone, A., Russo, A., Reale, S., Barcellona, M. L., and Rizza V. 1998. Stress proteins and SH-groups in oxidant-induced cell injury after chronic ethanol administration in rat. Free Rad. Biol. Med. 24:1159-1167

    Google Scholar 

  43. Butterfield, D., Castegna, A., Pocernich, C., Drake, J., Scapagnini, G., and Calabrese, V. 2002. Nutritional approaches to combat oxidative stress in Alzheimer's disease. J. Nutr. Biochem. 13: 444-461.

    Google Scholar 

  44. de Andres, C. and Lledo, A. 1997. Fatty diet and multiple sclerosis. Rev. Neurol. 25:2032-2035.

    Google Scholar 

  45. Szeinberg, A., Golan, R., Ben-Ezzer, J., Sarova-Pinhas, I., and Kindler, D. 1981. Glutathione peroxidase activity in various types of blood cells in multiple sclerosis. Acta Neurol. Scand. 63:67-75.

    Google Scholar 

  46. Calabrese, V., Bella, R., Testa, D., Spadaro, F., Scrofani, A., Rizza, V., and Pennisi, G. 1998. Increased cerebrospinal fluid and plasma levels of ultraweak chemiluminescence are associated with changes in the thiol pool and lipid-soluble fluorescence in multiple sclerosis: The pathogenic role of oxidative stress. Drugs Exp. Clin. Res. 24:125-131.

    Google Scholar 

  47. Scapagnini, G., Foresti, R., Calabrese, V., Giuffrida Stella, A. M., Green, C. J., and Motterlini, R. 2002. Caffeic acid phenethyl ester and curcumin: A novel class of heme oxygenase-l inducers. Mol. Pharmacol. 61:554-561.

    Google Scholar 

  48. Garthwaite, G., Goodwin, D. A., Batchelor, A. M., Leeming, K., and Garthwaite, J. 2002. Nitric oxide toxicity in CNS white matter: An in vitro study using rat optic nerve. Neuroscience. 109:145-155.

    Google Scholar 

  49. Boerrigter, M. E., Franceschi, C., Arrigoni-Martelli, E., Wei, J. Y., and Vijg, J. 1993. The effect of L-carnitine and acetyl-L-carnitine on the disappearance of DNA single-strand breaks in human peripheral blood lymphocytes. Carcinogenesis 14:2131-2136.

    Google Scholar 

  50. Calabrese, V. and Rizza, V. 1999. Formation of propionate after short-term ethanol treatment and its interaction with the carnitine pool in rat. Alcohol 19:169-176.

    Google Scholar 

  51. Calvani, M. and Arrigoni-Martelli, E. 1999. Attenuation by acetyl-L-carnitine of neurological damage and biochemical derangement following brain ischemia and reperfusion. Int. J. Tissue React. 21:1-6.

    Google Scholar 

  52. Hagen, T. M., Wehr, C. M., and Ames, B. N. 1998. Mitochondrial decay in aging: Reversal through supplementation of acetyl-L-carnitine and N-tert-butyl-alpha-phenyl-nitrone. Ann. N. Y. Acad. Sci. 854:214-223.

    Google Scholar 

  53. Fernandez, E., Pallini, R., Tamburrini, G., Lauretti, L., Tancredi, A., and La Marca, F. 1995. Effects of levo-acetylcarnitine on second motoneuron survival after axotomy. Neurol. Res. 5:373-376.

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Chemistry, Section of Biochemistry and Molecular Biology, Italy

    V. Calabrese, A. Ravagna & A. M. Giuffrida Stella

  2. Faculty of Medicine, University of Catania, Catania, Italy

    V. Calabrese, A. Ravagna & A. M. Giuffrida Stella

  3. Blanchette Rockefeller Neurosciences Institute, West Virginia University, Rockville, Maryland

    G. Scapagnini & R. Bella

  4. Department of Chemistry, Center of Membrane Sciences, USA

    D. A. Butterfield

  5. anders-Brown Center on Aging, University of Kentucky, Lexington, Kentucky

    D. A. Butterfield

  6. Department of Physiology, University of Tor Vergata, Rome, Italy

    M. Calvani

  7. Department of Neurology, Faculty of Medicine, University of Catania, Catania, Italy

    G. Pennisi

Authors
  1. V. Calabrese
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. G. Scapagnini
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. A. Ravagna
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. R. Bella
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. D. A. Butterfield
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. M. Calvani
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. G. Pennisi
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. A. M. Giuffrida Stella
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to V. Calabrese.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Calabrese, V., Scapagnini, G., Ravagna, A. et al. Disruption of Thiol Homeostasis and Nitrosative Stress in the Cerebrospinal Fluid of Patients with Active Multiple Sclerosis: Evidence for a Protective Role of Acetylcarnitine. Neurochem Res 28, 1321–1328 (2003). https://doi.org/10.1023/A:1024984013069

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1023/A:1024984013069

Search

Navigation