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Oxidative Damage in Muscular Dystrophy Correlates with the Severity of the Pathology: Role of Glutathione Metabolism

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Abstract

Muscular dystrophies (MDs) such as Duchenne muscular dystrophy (DMD), sarcoglycanopathy (Sgpy) and dysferlinopathy (Dysfy) are recessive genetic neuromuscular diseases that display muscle degeneration. Although these MDs have comparable endpoints of muscle pathology, the onset, severity and the course of these diseases are diverse. Different mechanisms downstream of genetic mutations might underlie the disparity in these pathologies. We surmised that oxidative damage and altered antioxidant function might contribute to these differences. The oxidant and antioxidant markers in the muscle biopsies from patients with DMD (n = 15), Sgpy (n = 15) and Dysfy (n = 15) were compared to controls (n = 10). Protein oxidation and lipid peroxidation was evident in all MDs and correlated with the severity of pathology, with DMD, the most severe dystrophic condition showing maximum damage, followed by Sgpy and Dysfy. Oxidative damage in DMD and Sgpy was attributed to the depletion of glutathione (GSH) and lowered antioxidant activities while loss of GSH peroxidase and GSH-S-transferase activities was observed in Dysfy. Lower GSH level in DMD was due to lowered activity of gamma-glutamyl cysteine ligase, the rate limiting enzyme in GSH synthesis. Similar analysis in cardiotoxin (CTX) mouse model of MD showed that the dystrophic muscle pathology correlated with GSH depletion and lipid peroxidation. Depletion of GSH prior to CTX exposure in C2C12 myoblasts exacerbated oxidative damage and myotoxicity. We deduce that the pro and anti-oxidant mechanisms could be correlated to the severity of MD and might influence the dystrophic pathology to a different extent in various MDs. On a therapeutic note, this could help in evolving novel therapies that offer myoprotection in MD.

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Abbreviations

NMD:

Neuromuscular disease

MD:

Muscular dystrophy

DMD:

Duchenne muscular dystrophy

Sgpy:

Sarcoglycanopathy

Dysfy:

Dysferlinopathy

GSH:

Glutathione

SOD:

Superoxide dismutase

MDA:

Malondialdehyde

3-NT:

3-Nitrotyrosine

GCL:

Gamma glutamyl cysteine ligase

GPx:

Glutathione peroxidase

GR:

Glutathione reductase

GST:

Glutathione-s-transferase

CTX:

Cardiotoxin

ROS:

Reactive oxygen species

MTT:

3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide

DNPH:

Dinitrophenyl hydrazine

PN:

Peroxynitrite

References

  1. Dubowitz V, Sewry CA (2007) Muscle biopsy—a practical approach, 3rd edn. Saunders Elsevier, London

  2. Munoz P, Rosemblatt M, Testar X, Palacin M, Zorzano A (1995) Isolation and characterization of distinct domains of sarcolemma and T-tubules from rat skeletal muscle. Biochem J 307(Pt 1):273–280

    PubMed  CAS  Google Scholar 

  3. Rando TA (2001) The dystrophin-glycoprotein complex, cellular signaling, and the regulation of cell survival in the muscular dystrophies. Muscle Nerve 24(12):1575–1594

    Article  PubMed  CAS  Google Scholar 

  4. Emery AEH (1999) Neuromuscular disorders: clinical and molecular genetics. Wiley, England

    Google Scholar 

  5. Burr IM, Asayama K, Fenichel GM (1987) Superoxide dismutases, glutathione peroxidase, and catalase in neuromuscular disease. Muscle Nerve 10(2):150–154. doi:10.1002/mus.880100208

    Article  PubMed  CAS  Google Scholar 

  6. Kar NC, Pearson CM (1979) Catalase, superoxide dismutase, glutathione reductase and thiobarbituric acid-reactive products in normal and dystrophic human muscle. Clin Chim Acta 94(3):277–280

    Article  PubMed  CAS  Google Scholar 

  7. Jongpiputvanich S, Sueblinvong T, Norapucsunton T (2005) Mitochondrial respiratory chain dysfunction in various neuromuscular diseases. J Clin Neurosci 12(4):426–428. doi:10.1016/j.jocn.2004.06.014

    Article  PubMed  CAS  Google Scholar 

  8. Haslbeck KM, Friess U, Schleicher ED, Bierhaus A, Nawroth PP, Kirchner A, Pauli E, Neundorfer B, Heuss D (2005) The RAGE pathway in inflammatory myopathies and limb girdle muscular dystrophy. Acta Neuropathol 110(3):247–254. doi:10.1007/s00401-005-1043-3

    Article  PubMed  CAS  Google Scholar 

  9. Burri BJ, Chan SG, Berry AJ, Yarnell SK (1980) Blood levels of superoxide dismutase and glutathione peroxidase in Duchenne muscular dystrophy. Clin Chim Acta 105(2):249–255

    Article  PubMed  CAS  Google Scholar 

  10. Grosso S, Perrone S, Longini M, Bruno C, Minetti C, Gazzolo D, Balestri P, Buonocore G (2008) Isoprostanes in dystrophinopathy: evidence of increased oxidative stress. Brain Dev 30(6):391–395. doi:10.1016/j.braindev.2007.11.005

    Article  PubMed  Google Scholar 

  11. Haycock JW, Mac Neil S, Mantle D (1998) Differential protein oxidation in Duchenne and Becker muscular dystrophy. Neuro Report 9(10):2201–2207

    CAS  Google Scholar 

  12. Nakae Y, Stoward PJ, Kashiyama T, Shono M, Akagi A, Matsuzaki T, Nonaka I (2004) Early onset of lipofuscin accumulation in dystrophin-deficient skeletal muscles of DMD patients and mdx mice. J Mol Histol 35(5):489–499

    Article  PubMed  CAS  Google Scholar 

  13. Dudley RW, Khairallah M, Mohammed S, Lands L, Des Rosiers C, Petrof BJ (2006) Dynamic responses of the glutathione system to acute oxidative stress in dystrophic mouse (mdx) muscles. Am J Physiol Regul Integr Comp Physiol 291(3):704–710. doi:10.1152/ajpregu.00031.2006

    Article  Google Scholar 

  14. Ragusa RJ, Chow CK, Porter JD (1997) Oxidative stress as a potential pathogenic mechanism in an animal model of Duchenne muscular dystrophy. Neuromuscul Disord 7(6–7):379–386

    Article  PubMed  CAS  Google Scholar 

  15. Dudley RW, Danialou G, Govindaraju K, Lands L, Eidelman DE, Petrof BJ (2006) Sarcolemmal damage in dystrophin deficiency is modulated by synergistic interactions between mechanical and oxidative/nitrosative stresses. Am J Pathol 168(4): 1276–1287; quiz 1404-1275. doi:10.2353/ajpath.2006.050683

    Google Scholar 

  16. Murphy ME, Kehrer JP (1986) Activities of antioxidant enzymes in muscle, liver and lung of chickens with inherited muscular dystrophy. Biochem Biophys Res Commun 134(2):550–556

    Article  PubMed  CAS  Google Scholar 

  17. Salminen A, Kihlstrom M (1989) Increased susceptibility to lipid peroxidation in skeletal muscles of dystrophic hamsters. Experientia 45(8):747–749

    Article  PubMed  CAS  Google Scholar 

  18. 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 72:248–254

    Article  PubMed  CAS  Google Scholar 

  19. Renjini R, Gayathri N, Nalini A, Srinivas Bharath MM (2011) Analysis of Calpain 3 protein in muscle biopsies of different muscular dystrophies from India. Indian J Med Res (in press)

  20. Hsu DK, Guo Y, Alberts GF, Copeland NG, Gilbert DJ, Jenkins NA, Peifley KA, Winkles JA (1996) Identification of a murine TEF-1-related gene expressed after mitogenic stimulation of quiescent fibroblasts and during myogenic differentiation. J Biol Chem 271(23):13786–13795

    Article  PubMed  CAS  Google Scholar 

  21. Ardite E, Barbera JA, Roca J, Fernandez-Checa JC (2004) Glutathione depletion impairs myogenic differentiation of murine skeletal muscle C2C12 cells through sustained NF-kappaB activation. Am J Pathol 165(3):719–728

    Article  PubMed  CAS  Google Scholar 

  22. Vali S, Mythri RB, Jagatha B, Padiadpu J, Ramanujan KS, Andersen JK, Gorin F, Bharath MM (2007) Integrating glutathione metabolism and mitochondrial dysfunction with implications for Parkinson’s disease: a dynamic model. Neuroscience 149(4):917–930. doi:10.1016/j.neuroscience.2007.08.028

    Article  PubMed  CAS  Google Scholar 

  23. Ohashi T, Kakimoto K, Sokawa Y, Taketani S (2002) Semi-quantitative estimation of heme/hemoprotein with dichlorodihydrofluorescin diacetate. Anal Biochem 308(2):392–395

    Article  PubMed  CAS  Google Scholar 

  24. Harish G, Venkateshappa C, Mythri RB, Dubey SK, Mishra K, Singh N, Vali S, Bharath MM (2010) Bioconjugates of curcumin display improved protection against glutathione depletion mediated oxidative stress in a dopaminergic neuronal cell line: implications for Parkinson’s disease. Bioorg Med Chem 18(7):2631–2638. doi:10.1016/j.bmc.2010.02.029

    Article  PubMed  CAS  Google Scholar 

  25. Kherif S, Lafuma C, Dehaupas M, Lachkar S, Fournier JG, Verdiere-Sahuque M, Fardeau M, Alameddine HS (1999) Expression of matrix metalloproteinases 2 and 9 in regenerating skeletal muscle: a study in experimentally injured and mdx muscles. Dev Biol 205(1):158–170. doi:10.1006/dbio.1998.9107

    Article  PubMed  CAS  Google Scholar 

  26. Ohkawa H, Ohishi N, Yagi K (1979) Assay for lipid peroxides in animal tissues by thiobarbituric acid reaction. Anal Biochem 95(2):351–358

    Article  PubMed  CAS  Google Scholar 

  27. Mythri RB, Venkateshappa C, Harish G, Mahadevan A, Muthane UB, Yasha TC, Srinivas Bharath MM, Shankar SK (2011) Evaluation of markers of oxidative stress, antioxidant function and astrocytic proliferation in the striatum and frontal cortex of Parkinson’s disease brains. Neurochem Res 36(8):1452–1463. doi:10.1007/s11064-011-0471-9

    Article  PubMed  CAS  Google Scholar 

  28. Levine RL, Garland D, Oliver CN, Amici A, Climent I, Lenz AG, Ahn BW, Shaltiel S, Stadtman ER (1990) Determination of carbonyl content in oxidatively modified proteins. Methods Enzymol 186:464–478

    Article  PubMed  CAS  Google Scholar 

  29. Mythri RB, Jagatha B, Pradhan N, Andersen J, Bharath MM (2007) Mitochondrial complex I inhibition in Parkinson’s disease: how can curcumin protect mitochondria? Antioxid Redox Signal 9(3):399–408. doi:10.1089/ars.2007.9.ft-25

    Article  PubMed  CAS  Google Scholar 

  30. Sambrook J, Russell DW (2001) Molecular cloning, a laboratory manual, 3rd ed edn. Cold spring Harbor Laboratory Press, New York

    Google Scholar 

  31. Bagnyukova TV, Storey KB, Lushchak VI (2003) Induction of oxidative stress in Rana ridibunda during recovery from winter hibernation. J Therm Biol 28(1):21–28. doi:10.1016/s0306-4565(02)00031-1

    Article  CAS  Google Scholar 

  32. Aebi H (1984) Catalase in vitro. Methods Enzymol 105:121–126

    Article  PubMed  CAS  Google Scholar 

  33. Flohe L, Gunzler WA (1984) Assays of glutathione peroxidase. Methods Enzymol 105:114–121

    Article  PubMed  CAS  Google Scholar 

  34. Carlberg I, Mannervik B (1985) Glutathione reductase. Methods Enzymol 113:484–490

    Article  PubMed  CAS  Google Scholar 

  35. Guthenberg C, Alin P, Mannervik B (1985) Glutathione transferase from rat testis. Methods Enzymol 113:507–510

    Article  PubMed  CAS  Google Scholar 

  36. Seelig GF, Meister A (1985) Glutathione biosynthesis; gamma-glutamylcysteine synthetase from rat kidney. Methods Enzymol 113:379–390

    Article  PubMed  CAS  Google Scholar 

  37. Hill KE, McCollum GW, Burk RF (1997) Determination of thioredoxin reductase activity in rat liver supernatant. Anal Biochem 253(1):123–125. doi:10.1006/abio.1997.2373

    Article  PubMed  CAS  Google Scholar 

  38. Brancaccio P, Lippi G, Maffulli N (2010) Biochemical markers of muscular damage. Clin Chem Lab Med 48(6):757–767. doi:10.1515/cclm.2010.179

    Article  PubMed  CAS  Google Scholar 

  39. Degl’Innocenti D, Rosati F, Iantomasi T, Vincenzini MT, Ramponi G (1999) GSH system in relation to redox state in dystrophic skin fibroblasts. Biochimie 81(11):1025–1029

    Article  PubMed  Google Scholar 

  40. Khouzami L, Bourin MC, Christov C, Damy T, Escoubet B, Caramelle P, Perier M, Wahbi K, Meune C, Pavoine C, Pecker F (2010) Delayed cardiomyopathy in dystrophin deficient mdx mice relies on intrinsic glutathione resource. Am J Pathol 177(3):1356–1364. doi:10.2353/ajpath.2010.090479

    Article  PubMed  CAS  Google Scholar 

  41. Spassov A, Gredes T, Gedrange T, Pavlovic D, Lupp A, Kunert-Keil C (2010) Increased oxidative stress in dystrophin deficient (mdx) mice masticatory muscles. Exp Toxicol Pathol. doi:10.1016/j.etp.2010.04.006

  42. Jackson MJ (2008) Redox regulation of skeletal muscle. IUBMB Life 60(8):497–501. doi:10.1002/iub.72

    Article  PubMed  CAS  Google Scholar 

  43. Dorchies OM, Wagner S, Buetler TM, Ruegg UT (2009) Protection of dystrophic muscle cells with polyphenols from green tea correlates with improved glutathione balance and increased expression of 67LR, a receptor for (-)-epigallocatechin gallate. Biofactors 35(3):279–294. doi:10.1002/biof.34

    Article  PubMed  CAS  Google Scholar 

  44. Jackson MJ, Brooke MH, Kaiser K, Edwards RH (1991) Glutathione depletion during experimental damage to rat skeletal muscle and its relevance to Duchenne muscular dystrophy. Clin Sci (Lond) 80(6):559–564

    CAS  Google Scholar 

  45. Lesgards JF, Gauthier C, Iovanna J, Vidal N, Dolla A, Stocker P (2011) Effect of reactive oxygen and carbonyl species on crucial cellular antioxidant enzymes. Chem Biol Interact 190(1):28–34. doi:10.1016/j.cbi.2010.12.028

    Article  PubMed  CAS  Google Scholar 

  46. Kramerova I, Kudryashova E, Wu B, Germain S, Vandenborne K, Romain N, Haller RG, Verity MA, Spencer MJ (2009) Mitochondrial abnormalities, energy deficit and oxidative stress are features of calpain 3 deficiency in skeletal muscle. Hum Mol Genet 18(17):3194–3205. doi:10.1093/hmg/ddp257

    Article  PubMed  CAS  Google Scholar 

  47. Dioszeghy P, Imre S, Mechler F (1989) Lipid peroxidation and superoxide dismutase activity in muscle and erythrocytes in adult muscular dystrophies and neurogenic atrophies. Eur Arch Psychiatry Neurol Sci 238(3):175–177

    Article  PubMed  CAS  Google Scholar 

  48. Potgieter M, Pretorius E, Van der Merwe CF, Beukes M, Vieira WA, Auer RE, Auer M, Meyer S (2011) Histological assessment of SJL/J mice treated with the antioxidants coenzyme Q10 and resveratrol. Micron 42(3):275–282. doi:10.1016/j.micron.2010.10.001

    Article  PubMed  CAS  Google Scholar 

  49. Bharath S, Hsu M, Kaur D, Rajagopalan S, Andersen JK (2002) Glutathione, iron and Parkinson’s disease. Biochem Pharmacol 64(5–6):1037–1048

    Article  PubMed  CAS  Google Scholar 

  50. Tidball JG, Wehling-Henricks M (2007) The role of free radicals in the pathophysiology of muscular dystrophy. J Appl Physiol 102(4):1677–1686

    Article  PubMed  CAS  Google Scholar 

  51. Vidal B, Serrano AL, Tjwa M, Suelves M, Ardite E, De Mori R, Baeza-Raja B, Martinez de Lagran M, Lafuste P, Ruiz-Bonilla V, Jardi M, Gherardi R, Christov C, Dierssen M, Carmeliet P, Degen JL, Dewerchin M, Munoz-Canoves P (2008) Fibrinogen drives dystrophic muscle fibrosis via a TGFbeta/alternative macrophage activation pathway. Genes Dev 22(13):1747–1752. doi:10.1101/gad.465908

    Article  PubMed  CAS  Google Scholar 

  52. Yuasa K, Hagiwara Y, Ando M, Nakamura A, Takeda S, Hijikata T (2008) MicroRNA-206 is highly expressed in newly formed muscle fibers: implications regarding potential for muscle regeneration and maturation in muscular dystrophy. Cell Struct Funct 33(2):163–169

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

This work was financially supported by the Department of Science and Technology, India. RR is a senior research fellow of the Council for Scientific and Industrial Research, India. The authors thank all the patients and their families for the muscle biopsies. RR and MMSB conceived the experiments. RR and NG carried out the experiments. AN carried out the clinical analysis. MMSB, RR and NG analyzed the data. RR and MMSB wrote the paper.

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Authors and Affiliations

  1. Department of Neurochemistry, National Institute of Mental Health and Neurosciences (NIMHANS), P.B # 2900, Hosur Road, Bangalore, 560029, Karnataka, India

    R. Renjini & M. M. Srinivas Bharath

  2. Department Neuropathology, National Institute of Mental Health and Neurosciences (NIMHANS), Bangalore, 560029, Karnataka, India

    N. Gayathri

  3. Department Neurology, National Institute of Mental Health and Neurosciences (NIMHANS), Bangalore, 560029, Karnataka, India

    A. Nalini

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  1. R. Renjini
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Correspondence to M. M. Srinivas Bharath.

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Renjini, R., Gayathri, N., Nalini, A. et al. Oxidative Damage in Muscular Dystrophy Correlates with the Severity of the Pathology: Role of Glutathione Metabolism. Neurochem Res 37, 885–898 (2012). https://doi.org/10.1007/s11064-011-0683-z

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