Differential morphofunctional characteristics and gene expression in fast and slow muscle of rats with monocrotaline-induced heart failure

  • Raquel Santilone Bertaglia
  • Joyce Reissler
  • Francis Silva Lopes
  • Walter Luiz Garrido Cavalcante
  • Fernanda Regina Carani
  • Carlos Roberto Padovani
  • Sergio Augusto Rodrigues
  • Antônio Carlos Cigogna
  • Robson Francisco Carvalho
  • Ana Angélica Henrique Fernandes
  • Marcia Gallacci
  • Maeli Dal Pai Silva
Original Paper


Heart failure (HF) is characterized by limited exercise tolerance, skeletal muscle atrophy, a shift toward fast muscle fiber, and myogenic regulatory factor (MRF) changes. Reactive oxygen species (ROS) also contribute to target organ damage in this syndrome. In this study, we investigated and compared morphofunctional characteristics and gene expression in Soleus (SOL—oxidative and slow twitching muscle) and in Extensor Digitorum Longus (EDL—glycolytic and fast twitching muscle) during HF. Two groups of rats were used: control (CT) and heart failure (HF), induced by a single injection of monocrotaline. MyoD and myogenin gene expression were determined by RT-qPCR, and MHC isoforms by SDS–PAGE; muscle fiber type frequency and cross sectional area (CSA) were analyzed by mATPase. A biochemical study was performed to determine lipid hydroperoxide (LH), glutathione peroxidase (GSH-Px), and superoxide dismutase (SOD); myography was used to determine amplitude, rise time, fall time, and fatigue resistance in both muscles. HF showed SOL and EDL muscle atrophy in all muscle fiber types; fiber frequency decreased in type IIC and muscle contraction fall time increased only in SOL muscle. Myogenin mRNA expression was lower in SOL and myoD decreased in HF EDL muscle. LH increased, and SOD and GSH-Px activity decreased only in HF SOL muscle. HF EDL muscle did not present changes in MHC distribution, contractile properties, HL concentration, and antioxidant enzyme activity. In conclusion, our results indicate that monocrotaline induced HF promoted more prominent biochemical, morphological and functional changes in SOL (oxidative and slow twitching muscle). Although further experiments are required to better determine the mechanisms involved in HF pathophysiology, our results contribute to understanding the muscle-specific changes that occur in this syndrome.


Skeletal muscle Oxidative stress Fiber types Myogenic regulatory factors Monocrotaline 



This study was supported by FAPESP (Fundação de Amparo to Pesquisa do Estado de São Paulo) Process no 2009/51060-8 and CAPES. This work is part of the M.Sc. Thesis presented by R.S.B. to São Paulo State University (UNESP) in 2011.


  1. Allen DL, Sartorius CA, Sycuro LK, Leinwands LA (2001) Different pathway regulate of the skeletal myosin heavy chain. J Biol Chem 274:43524–43533CrossRefGoogle Scholar
  2. Alway SE, Degens H, Lowe DA, Krishnamurthy G (2002) Increased myogenic repressor Id mRNA and protein levels in hind limb muscles of aged rats. Am J Physiol Regul Integr Comp Physiol 282:R411–R422PubMedGoogle Scholar
  3. Anker SD, Ponikowski PP, Clark AL, Leyva F, Rauchhaus M, Kemp M, Teixeira MM, Hellewell PG, Hooper J, Poole-Wilson PA, Coats AJ (1999) Cytokines and neurohormones relating to body composition alterations in the wasting syndrome of chronic heart failure. Eur Heart J 20:683–693PubMedCrossRefGoogle Scholar
  4. Barclay CJ (1992) Effects of fatigue on rate of isometric force development in mouse fast- and slow- twitch muscle. Am J Physiol 263:C1065–C1072 (Cell Physiol 32)PubMedGoogle Scholar
  5. Bärr A, Pette D (1988) Three fast myosin heavy chain in adult rat skeletal muscle. FEBS Lett 233:153–155CrossRefGoogle Scholar
  6. Barreiro E, de la Puente B, Minguella J, Corominas JM, Serrano S, Hussain SNA, Gea J (2005) Oxidative stress and respiratory muscle dysfunction in severe chronic obstructive pulmonary disease. Am J Respir Crit Care Med 171:1116–1124PubMedCrossRefGoogle Scholar
  7. Brooke MH, Kaiser KK (1970) Three “myosin adenosine triphosphatase” systems: the nature of their pH lability and sulfhydryl dependence. J Histochem Cytochem 18:670–672PubMedCrossRefGoogle Scholar
  8. Brunotte F, Thompson CH, Adamopoulos S, Coats A, Unitt J, Lindsay D, Kaklamanis L, Radda GK, Rajagopalan B (1995) Rat skeletal muscle metabolism in experimental heart failure: effects of physical training. Acta Physiol Scand 154:439–447PubMedCrossRefGoogle Scholar
  9. Campos GE, Luecke TJ, Wendeln HK, Toma K, Hagerman FC, Murray TF, Ragg KE, Ratamess NA, Kraemer WJ, Staron RS (2002) Muscular adaptations in response to three different resistance-training regimens: specificity of repetition maximum training zones. Eur J Appl Physiol 88:50–60PubMedCrossRefGoogle Scholar
  10. Carvalho RF, Cicogna AC, Campos GE, De Assis JM, Padovani CR, Okoshi MP, Dal Pai-Silva M (2003) Myosin heavy chain expression and atrophy in rat skeletal muscle during transition from cardiac hypertrophy to heart failure. Int J Exp Pathol 84:201–206PubMedCrossRefGoogle Scholar
  11. Carvalho RF, Cicogna AC, Campos GE, Lopes FS, Sugizaki MM, Nogueira CR, Dal Pai-Silva M (2006) Heart failure alters MyoD and MRF4 expression in rat skeletal muscle. Int J Exp Pathol 87:219–225PubMedCrossRefGoogle Scholar
  12. Carvalho RF, Castan EP, Coelho CA, Lopes FS, Almeida FL, Michelin A, de Souza RW, Araújo JP Jr, Cicogna AC, Dal Pai-Silva M (2010) Heart failure increases atrogin-1 and MuRF1 gene expression in skeletal muscle with fiber type-specific atrophy. J Mol Histol 41:81–87PubMedCrossRefGoogle Scholar
  13. Coats AJS, Clark AL, Piepoli M, Volterrani M, Poole-Wilson PA (1994) Symptoms and quality of life in heart failure. The muscle hypothesis. Br Heart J 72:36–39CrossRefGoogle Scholar
  14. Coirault C, Guellich A, Barbry T, Samuel JL, Riou B, Lecar-pentier Y (2007) Oxidative stress of myosin contributes to skeletal muscle dysfunction in rats with chronic heart failure. Am J Physiol Heart Circ Physiol 292:H1009–H1017PubMedCrossRefGoogle Scholar
  15. Dalla Libera L, Sabbadini R, Renken C, Ravara B, Sandri M, Betto R, Angelini A, Vescovo G (2001) Apoptosis in the skeletal muscle of rats with heart failure is associated with increased serum levels of TNF-alpha and sphingosine. J Mol Cell Cardiol 33:1871–1878PubMedCrossRefGoogle Scholar
  16. Dalla Libera L, Ravara B, Volterrani M, Gobbo V, Della Barbera M, Angelini A (2004) Beneficial effects of GH/IGF-1 on skeletal muscle atrophy and function in experimental heart failure. Am J Physiol Cell Physiol 286:C138–C144PubMedCrossRefGoogle Scholar
  17. Dalla Libera L, Ravara B, Gobbo V, Betto DD, Germinario E, Angelini A, Vescovo G (2005) Skeletal muscle myofibrillar protein oxidation in heart failure and the protective of Carvedilol. J Mol Cell Cardiol 38:803–807PubMedCrossRefGoogle Scholar
  18. Dalla Libera L, Ravara B, Gobbo V, Betto DD, Germinario E, Angelini A, Evangelista S, Vescovo G (2010) Skeletal muscle protein oxidation in chronic right heart failure in rat: can different beta-blockers prevent it to the same degree? Int J Cardiol 143:192–199PubMedCrossRefGoogle Scholar
  19. De Sousa E, Veksler V, Bigard X, Mateo P, Ventura-Clapier R (2000) Heart failure affects mitochondrial but not intrinsic properties of skeletal muscle. Circulation 102:1847–1853PubMedGoogle Scholar
  20. De Sousa E, Veksler V, Bigard X, Mateo P, Serrurier B, Ventura-Clapier R (2001) Dual influence of disease and increased load on diaphragm muscle in heart failure. J Mol Cell Cardiol 33:699–710PubMedCrossRefGoogle Scholar
  21. Delp M, Duan C, Mattson JP, Musch TI (1997) Changes in skeletal muscle biochemistry and histology relative to fiber type in rats with heart failure. J Appl Physiol 83:1291–1299PubMedGoogle Scholar
  22. Drexler H (1992) Skeletal muscle failure in heart failure. Circulation 85:1621–1623PubMedGoogle Scholar
  23. Ekmark M, Gronevik E, Schjerling P, Gundersen K (2003) Myogenin induces higher oxidative capacity in pre-existing mouse muscle fibres after somatic DNA transfer. J Physiol 548(Pt1):259–269PubMedCrossRefGoogle Scholar
  24. Ertunc M, Sara Y, Onur R (2009) Differential Contractile Impairment of fast-and slow-twitch skeletal muscle in a rat model of doxorubicin-induced congestive heart failure. Pharnacology 84:240–248Google Scholar
  25. Ewing JF, Janero DR (1995) Microplate superoxide dismutase assay employing a nonenzymatic superoxide generation. Annal Biochem 232:243–248CrossRefGoogle Scholar
  26. Feuers RJ (1998) The effects of dietary restriction on mitochondrial dysfunction in aging. Annal NY Acad Sci 125:192–201CrossRefGoogle Scholar
  27. Filippatos GS, Anker SD, Kremastinos DT (2005) Pathophysiology of peripheral muscle wasting in cardiac cachexia. Curr Opin Clin Nutr Metab Care 8(3):249–254PubMedCrossRefGoogle Scholar
  28. Gallacci M, Oliveira AC (1994) Pre- and postsynaptic mechanisms involved in tetanic fade induced by pancuronium in the isolated rat muscle. Pharmacology 49:265–270PubMedCrossRefGoogle Scholar
  29. Gundersen K, Merlie JP (1994) Id-1 as a possible transcriptional mediator of muscle disuse atrophy. Proc Natl Acad Sci 91:3647–3651PubMedCrossRefGoogle Scholar
  30. Guth L, Samaha FJ (1969) Qualitative differences between acto myosin ATPase of slow and fast mammalian muscle. Exp Neurol 25:138–152PubMedCrossRefGoogle Scholar
  31. Hopkins J, Tudhope GR (1973) Glutathione peroxidase in human red cells in health and disease. Br J Haematol 25:563–575PubMedCrossRefGoogle Scholar
  32. Hughes SM, Taylor JM, Tapscott SJ, Gurley CM, Carter WJ, Peterson CA (1993) Selective accumulation of MyoD and Miogenin mRNAs in fast and slow muscle is controlled by innervation and hormones. Development 118:1137–1147PubMedGoogle Scholar
  33. Hughes SM, Chi MM, Lowry OH, Gundersen K (1999) Myogenin induces a shift of enzyme activity from glycolytic to oxidative metabolism in muscles of transgenic mice. J Cell Biol 145:633–642PubMedCrossRefGoogle Scholar
  34. Jiang ZY, Woollard ACS, Wolf SP (1991) Lipid hydroperoxide measurement by oxidation of Fe2+ in the presence of xylenol orange. Comparison with the TBA assay and an iodometric method. Lipids 26:853–856PubMedCrossRefGoogle Scholar
  35. Kaasik A, Minajeva A, De Sousa E, Ventura-Clapier R, Veksler V (1999) Nitric oxide inhibits cardiac energy production via inhibition of mitochondrial creatine kinase. FEBS Lett 444:75–77PubMedCrossRefGoogle Scholar
  36. Kinugawa S, Tsutsui H, Hayashidani S, Ide T, Suematsu N, Satoh S, Utsumi H, Takeshita A (2000) Treatment with dimethylthiourea prevents left ventricular remodeling and failure after experimental myocardial infarction in mice: role of oxidative stress. Circ Res 87:392–398PubMedGoogle Scholar
  37. Kuno S, Katsuta S, Anno I, Matsumoto K, Akisada M (1988) Relationship between MR relaxation time and muscle fiber composition. Radiology 169:567–568PubMedGoogle Scholar
  38. Lapu-Bula OfiliE (2007) From hypertension to heart failure: role of nitric oxide-mediated endothelial dysfunction and emerging insights from myocardial contrast echocardiography. Am J Cardiol 26:7–14CrossRefGoogle Scholar
  39. Leineweber K, Brandt K, Wludyka B, Beilfuss A, Ponicke K, Heinroth-Hoffmann I, Brodde OE (2002) Ventricular hypertrophy plus neurohumoral activation is necessary to alter the cardiac beta-adrenoceptor system in experimental heart failure. Circ Res 91:1056–1062PubMedCrossRefGoogle Scholar
  40. Lipkin DP, Jones DA, Round JM, Poole-Wilson PA (1988) Abnormalities of skeletal muscle in patients with chronic heart failure. Int J Cardiol 18:187–195PubMedCrossRefGoogle Scholar
  41. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods 25:402–408Google Scholar
  42. Lopes FS, Carvalho RF, Campos GER, Sugizaki MM, Padovani CR, Nogueira CR, Cicogna AC, Dal Pai-Silva M (2008) Down-regulation of MyoD gene expression in rat diaphragm muscle with heart failure. Int J Exp Path 89:216–222CrossRefGoogle Scholar
  43. Mancini DM, Coyle E, Coggan A, Beltz J, Ferraro N, Montain S, Wilson JR (1989) Contribution of intrinsic skeletal muscle changes to 31P NMR skeletal muscle metabolic abnormalities in patients with chronic heart failure. Circulation 80:1338–1346PubMedGoogle Scholar
  44. Mancini DM, Walter G, Reichek N, Lenkinski R, McCully KK, Mullen JL, Wilson JR (1992) Contribution of skeletal muscle atrophy to exercise intolerance and altered muscle metabolism in heart failure. Circulation 85:1364–1373PubMedGoogle Scholar
  45. Martinez PF, Okoshi K, Zornoff LA, Carvalho RF, Oliveira Junior SA, Lima AR, Campos DH, Damatto RL, Padovani CR, Nogueira CR, Dal Pai-Silva M, Okoshi MP (2010) Chronic heart failure-induced skeletal muscle atrophy, necrosis, and changes in myogenic regulatory factors. Med Sci Monit 16:BR374–BR383PubMedGoogle Scholar
  46. Megeney LA, Rudnicki MA (1995) Determination versus differentiation and the MyoD family of transcription factors. Biochem Cell Biol 73:723–732PubMedCrossRefGoogle Scholar
  47. Murre C, Mccaw PS, Vaessin H, Caudy M, Jan LY, Yan JN, Cabrera CV, Buskin JN, Hauschka SD, Lassar AB, Weintraub H, Baltimore D (1989) Interactions between heterologous helix-loop-helix proteins generate complexes that bind specifically to a common DNA sequence. Cell 58:537–544PubMedCrossRefGoogle Scholar
  48. Nishiyama Y, Ikeda H, Haramaki N, Yoshida N, Imaizumi T (1998) Oxidative stress is related to exercise intolerance in patients with heart failure. Am Heart J 135:115–120PubMedCrossRefGoogle Scholar
  49. Parker MH, Seale P, Rudnicki MA (2003) Looking back to the embryo: defining transcriptional networks in adult myogenesis. Nat Rev Genet 4:497–507PubMedCrossRefGoogle Scholar
  50. Powers SK, Ji LL, Leeuwenburgh C (1999) Exercise training-induced alterations in skeletal muscle antioxidant capacity: a brief review. Med Sci Sports Exerc 31:987–997PubMedCrossRefGoogle Scholar
  51. Reindel JF, Ganey JG, Wagner Rf, Slocombe RF, Roth AR (1990) Development of morphologic, hemodynamic, and biochemical changes in lung of rats given monocrotaline pirrole. Toxicol Appl Pharmacol 106:179–200PubMedCrossRefGoogle Scholar
  52. Schulze PC, Gielen S, Adams V, Linke A, Möbius-Winkler S, Erbs S, Kratzsch J, Hambrecht R, Schuler G (2003) Muscular levels of proinflammatory cytokines correlate with a reduced expression of insulin-like growth factor-1 in chronic heart failure. Basic Res Cardiol 98:267–274PubMedGoogle Scholar
  53. Semba RD, Lauretani F, Ferrucci L (2007) Carotenoids as protection against sarcopenia in older adults. Arch Biochem Biophys 458:141–145PubMedCrossRefGoogle Scholar
  54. Siu PM, Donley DA, Bryner RW, Alway SE (2004) Myogenin and oxidative enzyme gene expression levels are elevated in rat soleus muscles after endurance training. J Appl Physiol 97:277–285PubMedGoogle Scholar
  55. Spangenburg EE, Talmadge RJ, Musch TI, Pfeifer PC, McAllister RM, Williams JH (2002) Changes in skeletal muscle myosin heavy chain isoform content during congestive heart failure. Eur J Appl Physiol 87:182–186PubMedCrossRefGoogle Scholar
  56. Staron RS, Kraemer WJ, Hikida RS, Fry AC, Murray JD, Campos GE (1999) Fiber type composition of four hind limb muscles of adult Fisher 344 rats. Histochem Cell Biol 111:117–123PubMedCrossRefGoogle Scholar
  57. Sullivan MJ, Green HJ, Cobb FR (1990) Skeletal muscle biochemistry and histology in ambulatory patients with long-term heart failure. Circulation 81:518–527PubMedGoogle Scholar
  58. Toth MJ, Palmer BM, Lewinter MM (2006) Effect of heart failure on skeletal muscle myofibrillar protein content, isoform expression and calcium sensitivity. Int J Cardiol 107:211–219PubMedCrossRefGoogle Scholar
  59. Tsutsui H, Ide T, Hayashidani S, Suematsu N, Shiomi T, Wen J, Nakamura K, Ichikawa K, Utsumi H, Takeshita A (2001) Enhanced generation of reactive oxygen species in the limb skeletal muscles from a murine infarct model of heart failure. Circulation 104:134–136PubMedCrossRefGoogle Scholar
  60. Tsutsui H, Kinugawa S, Matsushima S (2008) Oxidative stress and mitochondrial DNA damage in heart failure (Suppl A):A31–A37Google Scholar
  61. Van Albada ME, Bartelds B, Wijnberg H, Mohaupt S, Dickinson MG, Schoemaker RG, Kooi KA, Gerbens F, Berger RM (2010) Gene expression profile in flow-associated pulmonary arterial hypertension with neointimal lesions. Am J Physiol Lung Cell Mol Physiol 298:L483–L491PubMedCrossRefGoogle Scholar
  62. Vandesompele J, De Preter K, Pattyn F, Poppe B, Van Roy N, De Paepe A, Speleman F (2002) Accurate normalization of Real-Time quantitative RT–PCR by geometric averaging of multiple internal control genes. Genome Biol 3(7):34.1–34.11CrossRefGoogle Scholar
  63. Vescovo G, Harding SE, Jones M, Dalla Libera L, Pessina AC, Poole-Wilson PA (1989) Contractile abnormalities of single right ventricular myocytes isolated from rats with right ventricular hypertrophy. J Mol Cell Cardiol 21(Suppl 5):103–111PubMedCrossRefGoogle Scholar
  64. Vescovo G, Ceconi C, Bernocchi P, Ferrari R, Carraro U, Ambrosio GB, Libera LD (1998) Skeletal muscle myosin heavy chain expression in rats with monocrotaline-induced cardiac hypertrophy and failure. Relation to blood flow and degree of muscle atrophy. Cardiovasc Res 39:233–241PubMedCrossRefGoogle Scholar
  65. Vescovo G, Ravara B, Libera LD (2008) Skeletal muscle myofibrillar protein oxidation and exercise capacity in heart failure. Basic Res Cardiol 103:285–290PubMedCrossRefGoogle Scholar
  66. Voytik SL, Przyborski M, Badylak SF, Konieczny SF (1993) Differential expression of muscle regulatory factor genes in normal and denervated adult rat hind limb muscle. Dev Dynam 198:214–224CrossRefGoogle Scholar
  67. Zar JH (1999) Biostatistical analysis. Prentice-Hall, New JerseyGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2011

Authors and Affiliations

  • Raquel Santilone Bertaglia
    • 1
  • Joyce Reissler
    • 1
  • Francis Silva Lopes
    • 2
  • Walter Luiz Garrido Cavalcante
    • 3
  • Fernanda Regina Carani
    • 1
  • Carlos Roberto Padovani
    • 4
  • Sergio Augusto Rodrigues
    • 4
  • Antônio Carlos Cigogna
    • 5
  • Robson Francisco Carvalho
    • 1
  • Ana Angélica Henrique Fernandes
    • 6
  • Marcia Gallacci
    • 3
  • Maeli Dal Pai Silva
    • 1
  1. 1.Department of Morphology, Institute of Biosciences, UNESPSão Paulo State UniversityBotucatuBrazil
  2. 2.Department of PhysiotherapyUNOESTEPresidente PrudenteBrazil
  3. 3.Department of Phamacology, Institute of Biosciences, UNESPSão Paulo State UniversityBotucatuBrazil
  4. 4.Department of Bioestatistics, Institute of Biosciences, UNESPSão Paulo State UniversityBotucatuBrazil
  5. 5.Department of Internal Medicine, School of Medicine, UNESPSão Paulo State UniversityBotucatuBrazil
  6. 6.Department of Chemistry and Biochemistry, Institute of Biosciences, UNESPSão Paulo State UniversityBotucatuBrazil

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