Journal of Neurology

, Volume 258, Issue 5, pp 762–769

Monitoring systemic oxidative stress in an animal model of amyotrophic lateral sclerosis

  • Francisco Javier Miana-Mena
  • Cristina González-Mingot
  • Pilar Larrodé
  • María Jesús Muñoz
  • Sara Oliván
  • Lorena Fuentes-Broto
  • Enrique Martínez-Ballarín
  • Russel J. Reiter
  • Rosario Osta
  • Joaquín José García
Original Communication


A mutant form of the ubiquitous copper/zinc superoxide dismutase (SOD1) protein has been found in some patients with amyotrophic lateral sclerosis (ALS). We monitored oxidative stress in an animal model of ALS, the SODG93A mouse, which develops a disease similar to ALS with an accelerated course. The aim of this work was to show that ALS damages several organs and tissues, from an oxidative stress point of view. We measured lipid and protein oxidative damage in different tissue homogenates of SODG93A mice. The biomarkers that we analyzed were malondialdehyde + 4-hydroxyalkenal (MDA + 4-HDA) and carbonyls, respectively. The spinal cord and brain of SODG93A mice showed increased lipid peroxidation after 100 or 130 days compared to age-matched littermate controls. The CNS was most affected, but lipid peroxidation was also detected in the skeletal muscle and liver on day 130. No changes were observed in protein carbonylation in the homogenates. Our results are consistent with a multisystem etiology of ALS and suggest that oxidative stress may play a primary role in ALS pathogenesis. Thus, oxidative stress represents a potential biomarker that might be useful in developing new therapeutic strategies for ALS.


Amyotrophic lateral sclerosis Oxidative stress G93A Lipid peroxidation Protein oxidation 


  1. 1.
    Agar J, Durham H (2003) Relevance of oxidative injury in the pathogenesis of motor neuron diseases. Amyotroph Lateral Scler Other Motor Neuron Disord 4:232–242PubMedCrossRefGoogle Scholar
  2. 2.
    Aguirre N, Beal MF, Matson WR, Bogdanov MB (2005) Increased oxidative damage to DNA in an animal model of amyotrophic lateral sclerosis. Free Radic Res 39:383–388PubMedCrossRefGoogle Scholar
  3. 3.
    Andrus PK, Fleck TJ, Gurney ME, Hall ED (1998) Protein oxidative damage in a transgenic mouse model of familial amyotrophic lateral sclerosis. J Neurochem 71:2041–2048PubMedCrossRefGoogle Scholar
  4. 4.
    Asai H, Hirano M, Udaka F, Shimada K, Oda M, Kubori T, Nishinaka K, Tsujimura T, Izumi Y, Konishi N, Matsumoto S, Kameyama M, Ueno S (2007) Sympathetic disturbances increase risk of sudden cardiac arrest in sporadic ALS. J Neurol Sci 254:78–83PubMedCrossRefGoogle Scholar
  5. 5.
    Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254PubMedCrossRefGoogle Scholar
  6. 6.
    Brooks KJ, Hill MD, Hockings PD, Reid DG (2004) MRI detects early hindlimb muscle atrophy in Gly93Ala superoxide dismutase-1 (G93A SOD1) transgenic mice, an animal model of familial amyotrophic lateral sclerosis. NMR Biomed 17:28–32PubMedCrossRefGoogle Scholar
  7. 7.
    Bruijn LI, Becher MW, Lee MK, Anderson KL, Jenkins NA, Copeland NG, Sisodia SS, Rothstein JD, Borchelt DR, Price DL, Cleveland DW (1997) ALS-linked SOD1 mutant G85R mediates damage to astrocytes and promotes rapidly progressive disease with SOD1-containing inclusions. Neuron 18:327–338PubMedCrossRefGoogle Scholar
  8. 8.
    Cheeseman KH, Slater TF (1993) An introduction to free radical biochemistry. Br Med Bull 49:481–493PubMedGoogle Scholar
  9. 9.
    Chiu AY, Zhai P, Dal Canto MC, Peters TM, Kwon YW, Prattis SM, Gurney ME (1995) Age-dependent penetrance of disease in a transgenic mouse model of familial amyotrophic lateral sclerosis. Mol Cell Neurosci 6:349–362PubMedCrossRefGoogle Scholar
  10. 10.
    Ciriza J, Moreno-Igoa M, Calvo AC, Yague G, Palacio J, Miana-Mena FJ, Munoz MJ, Zaragoza P, Brulet P, Osta R (2008) A genetic fusion GDNF-C fragment of tetanus toxin prolongs survival in a symptomatic mouse ALS model. Restor Neurol Neurosci 26:459–465PubMedGoogle Scholar
  11. 11.
    Clement AM, Nguyen MD, Roberts EA, Garcia ML, Boillee S, Rule M, McMahon AP, Doucette W, Siwek D, Ferrante RJ, Brown RH Jr, Julien JP, Goldstein LS, Cleveland DW (2003) Wild-type nonneuronal cells extend survival of SOD1 mutant motor neurons in ALS mice. Science 302:113–117PubMedCrossRefGoogle Scholar
  12. 12.
    Cleveland DW (1999) From Charcot to SOD1: mechanisms of selective motor neuron death in ALS. Neuron 24:515–520PubMedCrossRefGoogle Scholar
  13. 13.
    Corcia P, Pradat PF, Salachas F, Bruneteau G, Forestier N, Seilhean D, Hauw JJ, Meininger V (2008) Causes of death in a post-mortem series of ALS patients. Amyotroph Lateral Scler 9:59–62PubMedCrossRefGoogle Scholar
  14. 14.
    Dal Canto MC, Gurney ME (1995) Neuropathological changes in two lines of mice carrying a transgene for mutant human Cu, Zn SOD, and in mice overexpressing wild type human SOD: a model of familial amyotrophic lateral sclerosis (FALS). Brain Res 676:25–40PubMedCrossRefGoogle Scholar
  15. 15.
    Davies KJ, Delsignore ME, Lin SW (1987) Protein damage and degradation by oxygen radicals. II. Modification of amino acids. J Biol Chem 262:9902–9907PubMedGoogle Scholar
  16. 16.
    Deng HX, Hentati A, Tainer JA, Iqbal Z, Cayabyab A, Hung WY, Getzoff ED, Hu P, Herzfeldt B, Roos RP et al (1993) Amyotrophic lateral sclerosis and structural defects in Cu, Zn superoxide dismutase. Science 261:1047–1051PubMedCrossRefGoogle Scholar
  17. 17.
    Dobrowolny G, Aucello M, Rizzuto E, Beccafico S, Mammucari C, Boncompagni S, Belia S, Wannenes F, Nicoletti C, Del Prete Z, Rosenthal N, Molinaro M, Protasi F, Fano G, Sandri M, Musaro A (2008) Skeletal muscle is a primary target of SOD1G93A-mediated toxicity. Cell Metab 8:425–436PubMedCrossRefGoogle Scholar
  18. 18.
    Dupuis L, di Scala F, Rene F, de Tapia M, Oudart H, Pradat PF, Meininger V, Loeffler JP (2003) Up-regulation of mitochondrial uncoupling protein 3 reveals an early muscular metabolic defect in amyotrophic lateral sclerosis. FASEB J 17:2091–2093PubMedGoogle Scholar
  19. 19.
    Ferrante RJ, Shinobu LA, Schulz JB, Matthews RT, Thomas CE, Kowall NW, Gurney ME, Beal MF (1997) Increased 3-nitrotyrosine and oxidative damage in mice with a human copper/zinc superoxide dismutase mutation. Ann Neurol 42:326–334PubMedCrossRefGoogle Scholar
  20. 20.
    Frey D, Schneider C, Xu L, Borg J, Spooren W, Caroni P (2000) Early and selective loss of neuromuscular synapse subtypes with low sprouting competence in motoneuron diseases. J Neurosci 20:2534–2542PubMedGoogle Scholar
  21. 21.
    Fridovich I (1995) Superoxide radical and superoxide dismutases. Annu Rev Biochem 64:97–112PubMedCrossRefGoogle Scholar
  22. 22.
    Genova ML, Pich MM, Bernacchia A, Bianchi C, Biondi A, Bovina C, Falasca AI, Formiggini G, Castelli GP, Lenaz G (2004) The mitochondrial production of reactive oxygen species in relation to aging and pathology. Ann NY Acad Sci 1011:86–100PubMedCrossRefGoogle Scholar
  23. 23.
    Gil J, Funalot B, Verschueren A, Danel-Brunaud V, Camu W, Vandenberghe N, Desnuelle C, Guy N, Camdessanche JP, Cintas P, Carluer L, Pittion S, Nicolas G, Corcia P, Fleury MC, Maugras C, Besson G, Le Masson G, Couratier P (2008) Causes of death amongst French patients with amyotrophic lateral sclerosis: a prospective study. Eur J Neurol 15:1245–1251PubMedCrossRefGoogle Scholar
  24. 24.
    Gurney ME, Pu H, Chiu AY, Dal Canto MC, Polchow CY, Alexander DD, Caliendo J, Hentati A, Kwon YW, Deng HX et al (1994) Motor neuron degeneration in mice that express a human Cu, Zn superoxide dismutase mutation. Science 264:1772–1775PubMedCrossRefGoogle Scholar
  25. 25.
    Hall ED, Andrus PK, Oostveen JA, Fleck TJ, Gurney ME (1998) Relationship of oxygen radical-induced lipid peroxidative damage to disease onset and progression in a transgenic model of familial ALS. J Neurosci Res 53:66–77PubMedCrossRefGoogle Scholar
  26. 26.
    Halliwell B, Aruoma OI (1991) DNA damage by oxygen-derived species. Its mechanism and measurement in mammalian systems. FEBS Lett 281:9–19PubMedCrossRefGoogle Scholar
  27. 27.
    Halliwell B, Gutteridge JM (1999) Free radicals, ‘‘reactive species’’ and toxicology. In: Editor OU (ed) Free radicals in biology and medicine. Oxford University Press, New York, pp 544–616Google Scholar
  28. 28.
    Jokic N, Di Scala F, Dupuis L, Rene F, Muller A, Gonzalez De Aguilar JL, Loeffler JP (2003) Early activation of antioxidant mechanisms in muscle of mutant Cu/Zn-superoxide dismutase-linked amyotrophic lateral sclerosis mice. Ann NY Acad Sci 1010:552–556PubMedCrossRefGoogle Scholar
  29. 29.
    Jonsson PA, Bergemalm D, Andersen PM, Gredal O, Brannstrom T, Marklund SL (2008) Inclusions of amyotrophic lateral sclerosis-linked superoxide dismutase in ventral horns, liver, and kidney. Ann Neurol 63:671–675PubMedCrossRefGoogle Scholar
  30. 30.
    Kanner J, German JB, Kinsella JE (1987) Initiation of lipid peroxidation in biological systems. Crit Rev Food Sci Nutr 25:317–364PubMedCrossRefGoogle Scholar
  31. 31.
    Leclerc N, Ribera F, Zoll J, Warter JM, Poindron P, Lampert E, Borg J (2001) Selective changes in mitochondria respiratory properties in oxidative or glycolytic muscle fibers isolated from G93AhumanSOD1 transgenic mice. Neuromuscul Disord 11:722–727PubMedCrossRefGoogle Scholar
  32. 32.
    Levine RL, Williams JA, Stadtman ER, Shacter E (1994) Carbonyl assays for determination of oxidatively modified proteins. Methods Enzymol 233:346–357PubMedCrossRefGoogle Scholar
  33. 33.
    Lino MM, Schneider C, Caroni P (2002) Accumulation of SOD1 mutants in postnatal motoneurons does not cause motoneuron pathology or motoneuron disease. J Neurosci 22:4825–4832PubMedGoogle Scholar
  34. 34.
    Mahoney DJ, Kaczor JJ, Bourgeois J, Yasuda N, Tarnopolsky MA (2006) Oxidative stress and antioxidant enzyme upregulation in SOD1-G93A mouse skeletal muscle. Muscle Nerve 33:809–816PubMedCrossRefGoogle Scholar
  35. 35.
    Miana-Mena FJ, Munoz MJ, Yague G, Mendez M, Moreno M, Ciriza J, Zaragoza P, Osta R (2005) Optimal methods to characterize the G93A mouse model of ALS. Amyotroph Lateral Scler Other Motor Neuron Disord 6:55–62PubMedCrossRefGoogle Scholar
  36. 36.
    Moges H, Vasconcelos OM, Campbell WW, Borke RC, McCoy JA, Kaczmarczyk L, Feng J, Anders JJ (2009) Light therapy and supplementary Riboflavin in the SOD1 transgenic mouse model of familial amyotrophic lateral sclerosis (FALS). Lasers Surg Med 41:52–59PubMedCrossRefGoogle Scholar
  37. 37.
    Orrell RW (2000) Amyotrophic lateral sclerosis: copper/zinc superoxide dismutase (SOD1) gene mutations. Neuromuscul Disord 10:63–68PubMedCrossRefGoogle Scholar
  38. 38.
    Pramatarova A, Laganiere J, Roussel J, Brisebois K, Rouleau GA (2001) Neuron-specific expression of mutant superoxide dismutase 1 in transgenic mice does not lead to motor impairment. J Neurosci 21:3369–3374PubMedGoogle Scholar
  39. 39.
    Puttaparthi K, Wojcik C, Rajendran B, DeMartino GN, Elliott JL (2003) Aggregate formation in the spinal cord of mutant SOD1 transgenic mice is reversible and mediated by proteasomes. J Neurochem 87:851–860PubMedCrossRefGoogle Scholar
  40. 40.
    Reaume AG, Elliott JL, Hoffman EK, Kowall NW, Ferrante RJ, Siwek DF, Wilcox HM, Flood DG, Beal MF, Brown RH Jr, Scott RW, Snider WD (1996) Motor neurons in Cu/Zn superoxide dismutase-deficient mice develop normally but exhibit enhanced cell death after axonal injury. Nat Genet 13:43–47PubMedCrossRefGoogle Scholar
  41. 41.
    Rosen DR, Siddique T, Patterson D, Figlewicz DA, Sapp P, Hentati A, Donaldson D, Goto J, O’Regan JP, Deng HX et al (1993) Mutations in Cu/Zn superoxide dismutase gene are associated with familial amyotrophic lateral sclerosis. Nature 362:59–62PubMedCrossRefGoogle Scholar
  42. 42.
    Sau D, De Biasi S, Vitellaro-Zuccarello L, Riso P, Guarnieri S, Porrini M, Simeoni S, Crippa V, Onesto E, Palazzolo I, Rusmini P, Bolzoni E, Bendotti C, Poletti A (2007) Mutation of SOD1 in ALS: a gain of a loss of function. Hum Mol Genet 16:1604–1618PubMedCrossRefGoogle Scholar
  43. 43.
    Shaw IC, Fitzmaurice PS, Mitchell JD, Lynch PG (1995) Studies on cellular free radical protection mechanisms in the anterior horn from patients with amyotrophic lateral sclerosis. Neurodegeneration 4:391–396PubMedCrossRefGoogle Scholar
  44. 44.
    Simpson EP, Henry YK, Henkel JS, Smith RG, Appel SH (2004) Increased lipid peroxidation in sera of ALS patients: a potential biomarker of disease burden. Neurology 62:1758–1765PubMedGoogle Scholar
  45. 45.
    Turner BJ, Lopes EC, Cheema SS (2003) Neuromuscular accumulation of mutant superoxide dismutase 1 aggregates in a transgenic mouse model of familial amyotrophic lateral sclerosis. Neurosci Lett 350:132–136PubMedCrossRefGoogle Scholar
  46. 46.
    Weydt P, Hong S, Witting A, Moller T, Stella N, Kliot M (2005) Cannabinol delays symptom onset in SOD1 (G93A) transgenic mice without affecting survival. Amyotroph Lateral Scler Other Motor Neuron Disord 6:182–184PubMedCrossRefGoogle Scholar
  47. 47.
    Wong PC, Pardo CA, Borchelt DR, Lee MK, Copeland NG, Jenkins NA, Sisodia SS, Cleveland DW, Price DL (1995) An adverse property of a familial ALS-linked SOD1 mutation causes motor neuron disease characterized by vacuolar degeneration of mitochondria. Neuron 14:1105–1116PubMedCrossRefGoogle Scholar
  48. 48.
    Zhou C, Zhao CP, Zhang C, Wu GY, Xiong F (2007) A method comparison in monitoring disease progression of G93A mouse model of ALS. Amyotroph Lateral Scler 8:366–372PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  • Francisco Javier Miana-Mena
    • 1
  • Cristina González-Mingot
    • 1
    • 2
  • Pilar Larrodé
    • 2
  • María Jesús Muñoz
    • 1
    • 3
  • Sara Oliván
    • 3
  • Lorena Fuentes-Broto
    • 1
  • Enrique Martínez-Ballarín
    • 1
  • Russel J. Reiter
    • 4
  • Rosario Osta
    • 3
  • Joaquín José García
    • 1
  1. 1.Departamento Farmacología y FisiologíaUniversidad de ZaragozaZaragozaSpain
  2. 2.Neurology ServiceUniversitary Hospital Lozano Blesa of ZaragozaZaragozaSpain
  3. 3.LAGENBIOUniversidad de ZaragozaZaragozaSpain
  4. 4.Department of Cellular and Structural BiologyUniversity of Texas Health Science CenterSan AntonioUSA

Personalised recommendations