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Home / Equine Research Bank / J Muscle Res Cell Motil / Analysis of myosin heavy chains at the protein level in hors...
Journal of muscle research and cell motility1999; 20(2); 211-221; doi: 10.1023/a:1005461214800

Analysis of myosin heavy chains at the protein level in horse skeletal muscle.

Abstract: Combined methodologies of enzyme-linked immunosorbent assay (ELISA), sodium dodecyl sulphate polyacrilamide gel electrophoresis (SDS-PAGE), immunoblotting, traditional myofibrillar ATPase (mATPase) histochemistry and immunocytochemistry of whole biopsied samples were used to study myosin heavy chain (MHC) isoforms in the equine gluteus medius muscle. The ELISA technique allowed the quantification of the three MHC isoforms known to be present in different horse muscles: slow (MHC-I) and two fast (termed MHC-IIA and MCH-IIX). The SDS-PAGE method resolved MHCs in three bands: MHC-I, MHC-IIX and MHC-IIA from the fastest to the slowest migrating band and a quantification by densitometry for each MHC isoform was also possible. The identity of these three MHCs was confirmed by immunoblots with specific monoclonal antibodies. Five fibre types were defined immunohistochemically according to their MHC content: I, I + IIA, IIA, the hybrid IIAX and IIX. When quantitative data obtained with the four different methodologies were combined and compared, they were consistent and, when considered together, showed significant correlation. Nevertheless, the percentage of MHC-IIA histochemically derived was underestimated, while that of MHC-IIX was overestimated in comparison with the immunocytochemical determination of these MHC isoforms. The percentage of MHC-I obtained by ELISA technique was underestimated. In short, these integrated methods for the analysis of MHCs at the protein level demonstrate that equine skeletal muscle does not express the MHC-IIB, so type II fibres have been misclassified in numerous previous studies based upon the vary traditional mATPase histochemistry. They also offer new prospects for muscle fibre typing in equine experimental studies and veterinary medicine.
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Publication Date: 1999-07-21 PubMed ID: 10412092DOI: 10.1023/a:1005461214800Google Scholar: Lookup
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  • Journal Article

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

This study investigates the different forms of myosin heavy chain (MHC) proteins in horse muscles using various techniques to better inform muscle fibre typing.

Methods Used in the Study

  • The researchers utilized different techniques including enzyme-linked immunosorbent assay (ELISA), sodium dodecyl sulphate polyacrilamide gel electrophoresis (SDS-PAGE), immunoblotting, traditional myofibrillar ATPase (mATPase) histochemistry and immunocytochemistry. These techniques were used on biopsied samples from the gluteus medius muscle of a horse.
  • ELISA, a quantitative biochemical technique, was used to quantify three known MHC isoforms in horse muscles. These include a slow variant (MHC-I) and two fast variants (MHC-IIA and MCH-IIX).
  • SDS-PAGE, a method for separating proteins based on their electrophoretic mobility, was used to isolate the three MHC isoforms. The identities of these isoforms were then confirmed through immunoblots using specific monoclonal antibodies, which react against specific antigens.

Results of the Analysis

  • Using immunohistochemistry, five fibre types categorised by their MHC content were identified: I, I + IIA, IIA, the hybrid IIAX and IIX.
  • When combining data obtained from the various methodologies, results were consistent and showed significant correlation.
  • However, there were discrepancies in the quantification of certain isoforms. The amount of MHC-IIA was underestimated using histochemistry, and the MHC-IIX was overestimated in comparison to the immunocytochemical determination of these MHC isoforms. In addition, the ELISA technique underestimated MHC-I values.
  • The study also found that MHC-IIB does not occur in equine skeletal muscle, thereby correcting previous misclassifications in muscle fibre typing that relied solely on traditional mATPase histochemistry.

Significance of the Study

  • This study provides a more comprehensive approach to understanding muscle fibre types in horses and correcting previous misconceptions.
  • These integrated methods of analyzing MHCs at protein level offer new opportunities for experimental studies in horses and provide valuable insights for veterinary medicine.

Cite This Article

APA
Rivero JL, Serrano AL, Barrey E, Valette JP, Jouglin M. (1999). Analysis of myosin heavy chains at the protein level in horse skeletal muscle. J Muscle Res Cell Motil, 20(2), 211-221. https://doi.org/10.1023/a:1005461214800

Publication

Journal of muscle research and cell motility
ISSN: 0142-4319
NlmUniqueID: 8006298
Country: Netherlands
Language: English
Volume: 20
Issue: 2
Pages: 211-221

Researcher Affiliations

Rivero, J L
  • Department of Anatomy, Faculty of Veterinary Science, University of Cordoba, Spain. anllorij@lucano.uco.es
Serrano, A L
    Barrey, E
      Valette, J P
        Jouglin, M

          MeSH Terms

          • Animals
          • Electrophoresis, Polyacrylamide Gel
          • Enzyme-Linked Immunosorbent Assay
          • Horses
          • Immunohistochemistry
          • Male
          • Muscle, Skeletal / chemistry
          • Myosin Heavy Chains / analysis

          References

          This article includes 28 references
          1. López-Rivero JL, Serrano AL, Diz AM, Galisteo AM. Variability of muscle fibre composition and fibre size in the horse gluteus medius: an enzyme-histochemical and morphometric study.. J Anat 1992 Aug;181 ( Pt 1)(Pt 1):1-10.
            pubmed: 1284127
          2. Snow DH, Billeter R, Jenny E. Myosin types in equine skeletal muscle fibres.. Res Vet Sci 1981 May;30(3):381-2.
            pubmed: 6454947
          3. Bottinelli R, Betto R, Schiaffino S, Reggiani C. Maximum shortening velocity and coexistence of myosin heavy chain isoforms in single skinned fast fibres of rat skeletal muscle.. J Muscle Res Cell Motil 1994 Aug;15(4):413-9.
            pubmed: 7806635doi: 10.1007/BF00122115google scholar: lookup
          4. Rivero JL, Talmadge RJ, Edgerton VR. Myosin heavy chain isoforms in adult equine skeletal muscle: an immunohistochemical and electrophoretic study.. Anat Rec 1996 Oct;246(2):185-94.
            pubmed: 8888960doi: 10.1002/(SICI)1097-0185(199610)246:2<185::AID-AR5>3.0.CO;2-0google scholar: lookup
          5. Pette D, Staron RS. Mammalian skeletal muscle fiber type transitions.. Int Rev Cytol 1997;170:143-223.
            pubmed: 9002237doi: 10.1016/s0074-7696(08)61622-8google scholar: lookup
          6. Staron RS. Correlation between myofibrillar ATPase activity and myosin heavy chain composition in single human muscle fibers.. Histochemistry 1991;96(1):21-4.
            pubmed: 1834618doi: 10.1007/BF00266756google scholar: lookup
          7. Biral D, Betto R, Danieli-Betto D, Salviati G. Myosin heavy chain composition of single fibres from normal human muscle.. Biochem J 1988 Feb 15;250(1):307-8.
            pubmed: 3355518doi: 10.1042/bj2500307google scholar: lookup
          8. Rossini K, Rizzi C, Sandri M, Bruson A, Carraro U. High-resolution sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunochemical identification of the 2X and embryonic myosin heavy chains in complex mixtures of isomyosins.. Electrophoresis 1995 Jan;16(1):101-4.
            pubmed: 7737081doi: 10.1002/elps.1150160118google scholar: lookup
          9. Talmadge RJ, Roy RR. Electrophoretic separation of rat skeletal muscle myosin heavy-chain isoforms.. J Appl Physiol (1985) 1993 Nov;75(5):2337-40.
            pubmed: 8307894doi: 10.1152/jappl.1993.75.5.2337google scholar: lookup
          10. Galler S, Schmitt TL, Pette D. Stretch activation, unloaded shortening velocity, and myosin heavy chain isoforms of rat skeletal muscle fibres.. J Physiol 1994 Aug 1;478 Pt 3(Pt 3):513-21.
            pubmed: 7965861doi: 10.1113/jphysiol.1994.sp020270google scholar: lookup
          11. Sant'ana Pereira JA, Wessels A, Nijtmans L, Moorman AF, Sargeant AJ. New method for the accurate characterization of single human skeletal muscle fibres demonstrates a relation between mATPase and MyHC expression in pure and hybrid fibre types.. J Muscle Res Cell Motil 1995 Feb;16(1):21-34.
            pubmed: 7751402doi: 10.1007/BF00125307google scholar: lookup
          12. Hu P, Zhang KM, Feher JJ, Wang SW, Wright LD, Wechsler AS, Spratt JA, Briggs FN. Salbutamol and chronic low-frequency stimulation of canine skeletal muscle.. J Physiol 1996 Oct 1;496 ( Pt 1)(Pt 1):221-7.
            pubmed: 8910210doi: 10.1113/jphysiol.1996.sp021679google scholar: lookup
          13. Brooke MH, Kaiser KK. Muscle fiber types: how many and what kind?. Arch Neurol 1970 Oct;23(4):369-79.
            pubmed: 4248905doi: 10.1001/archneur.1970.00480280083010google scholar: lookup
          14. Serrano AL, Petrie JL, Rivero JL, Hermanson JW. Myosin isoforms and muscle fiber characteristics in equine gluteus medius muscle.. Anat Rec 1996 Apr;244(4):444-51.
            pubmed: 8694280doi: 10.1002/(SICI)1097-0185(199604)244:4<444::AID-AR3>3.0.CO;2-Vgoogle scholar: lookup
          15. Sinha AK, Rose RJ, Pozgaj I, Hoh JF. Indirect myosin immunocytochemistry for the identification of fibre types in equine skeletal muscle.. Res Vet Sci 1992 Jul;53(1):25-31.
            pubmed: 1410814doi: 10.1016/0034-5288(92)90079-hgoogle scholar: lookup
          16. Barrey E, Valette JP, Jouglin M, Picard B, Geay Y, Robelin J. Enzyme-linked immunosorbent assay for myosin heavy chains in the horse.. Reprod Nutr Dev 1995;35(6):619-28.
            pubmed: 8534356doi: 10.1051/rnd:19950602google scholar: lookup
          17. Rome LC, Sosnicki AA, Goble DO. Maximum velocity of shortening of three fibre types from horse soleus muscle: implications for scaling with body size.. J Physiol 1990 Dec;431:173-85.
            pubmed: 2100306doi: 10.1113/jphysiol.1990.sp018325google scholar: lookup
          18. DeNardi C, Ausoni S, Moretti P, Gorza L, Velleca M, Buckingham M, Schiaffino S. Type 2X-myosin heavy chain is coded by a muscle fiber type-specific and developmentally regulated gene.. J Cell Biol 1993 Nov;123(4):823-35.
            pubmed: 8227143doi: 10.1083/jcb.123.4.823google scholar: lookup
          19. Talmadge RJ, Grossman EJ, Roy RR. Myosin heavy chain composition of adult feline (Felis catus) limb and diaphragm muscles.. J Exp Zool 1996 Aug 15;275(6):413-20.
            pubmed: 8795286doi: 10.1002/(SICI)1097-010X(19960815)275:6<413::AID-JEZ3>3.0.CO;2-Rgoogle scholar: lookup
          20. Pereira Sant'Ana JA, Ennion S, Sargeant AJ, Moorman AF, Goldspink G. Comparison of the molecular, antigenic and ATPase determinants of fast myosin heavy chains in rat and human: a single-fibre study.. Pflugers Arch 1997 Dec;435(1):151-63.
            pubmed: 9359915doi: 10.1007/s004240050495google scholar: lookup
          21. Smerdu V, Karsch-Mizrachi I, Campione M, Leinwand L, Schiaffino S. Type IIx myosin heavy chain transcripts are expressed in type IIb fibers of human skeletal muscle.. Am J Physiol 1994 Dec;267(6 Pt 1):C1723-8.
            pubmed: 7545970doi: 10.1152/ajpcell.1994.267.6.C1723google scholar: lookup
          22. Schiaffino S, Gorza L, Sartore S, Saggin L, Ausoni S, Vianello M, Gundersen K, Lømo T. Three myosin heavy chain isoforms in type 2 skeletal muscle fibres.. J Muscle Res Cell Motil 1989 Jun;10(3):197-205.
            pubmed: 2547831doi: 10.1007/BF01739810google scholar: lookup
          23. Blanco CE, Sieck GC, Edgerton VR. Quantitative histochemical determination of succinic dehydrogenase activity in skeletal muscle fibres.. Histochem J 1988 Apr;20(4):230-43.
            pubmed: 3209423doi: 10.1007/BF01747468google scholar: lookup
          24. Nwoye L, Mommaerts WF, Simpson DR, Seraydarian K, Marusich M. Evidence for a direct action of thyroid hormone in specifying muscle properties.. Am J Physiol 1982 Mar;242(3):R401-8.
            pubmed: 6461259doi: 10.1152/ajpregu.1982.242.3.R401google scholar: lookup
          25. Bär A, Pette D. Three fast myosin heavy chains in adult rat skeletal muscle.. FEBS Lett 1988 Aug 1;235(1-2):153-5.
            pubmed: 3402594doi: 10.1016/0014-5793(88)81253-5google scholar: lookup
          26. Winkelmann DA, Lowey S, Press JL. Monoclonal antibodies localize changes on myosin heavy chain isozymes during avian myogenesis.. Cell 1983 Aug;34(1):295-306.
            pubmed: 6192935doi: 10.1016/0092-8674(83)90160-5google scholar: lookup
          27. Ennion S, Sant'ana Pereira J, Sargeant AJ, Young A, Goldspink G. Characterization of human skeletal muscle fibres according to the myosin heavy chains they express.. J Muscle Res Cell Motil 1995 Feb;16(1):35-43.
            pubmed: 7751403doi: 10.1007/BF00125308google scholar: lookup
          28. Andersen JL, Klitgaard H, Saltin B. Myosin heavy chain isoforms in single fibres from m. vastus lateralis of sprinters: influence of training.. Acta Physiol Scand 1994 Jun;151(2):135-42.
            pubmed: 7942047doi: 10.1111/j.1748-1716.1994.tb09730.xgoogle scholar: lookup

          Citations

          This article has been cited 18 times.
          1. Busse NI, Gonzalez ML, Wagner AL, Johnson SE. Short Communication: Supplementation with calcium butyrate causes an increase in the percentage of oxidative fibers in equine gluteus medius muscle.. J Anim Sci 2022 Aug 1;100(8).
            doi: 10.1093/jas/skac108pubmed: 35908781google scholar: lookup
          2. Latham CM, Owen RN, Dickson EC, Guy CP, White-Springer SH. Skeletal Muscle Adaptations to Exercise Training in Young and Aged Horses.. Front Aging 2021;2:708918.
            doi: 10.3389/fragi.2021.708918pubmed: 35822026google scholar: lookup
          3. Cappelli K, Amadori M, Mecocci S, Miglio A, Antognoni MT, Razzuoli E. Immune Response in Young Thoroughbred Racehorses under Training.. Animals (Basel) 2020 Oct 5;10(10).
            doi: 10.3390/ani10101809pubmed: 33027949google scholar: lookup
          4. Camerino C. The New Frontier in Oxytocin Physiology: The Oxytonic Contraction.. Int J Mol Sci 2020 Jul 21;21(14).
            doi: 10.3390/ijms21145144pubmed: 32708109google scholar: lookup
          5. Camerino C, Conte E, Carratù MR, Fonzino A, Lograno MD, Tricarico D. Oxytocin/Osteocalcin/IL-6 and NGF/BDNF mRNA Levels in Response to Cold Stress Challenge in Mice: Possible Oxytonic Brain-Bone-Muscle-Interaction.. Front Physiol 2019;10:1437.
            doi: 10.3389/fphys.2019.01437pubmed: 31827442google scholar: lookup
          6. Rooney MF, Porter RK, Katz LM, Hill EW. Skeletal muscle mitochondrial bioenergetics and associations with myostatin genotypes in the Thoroughbred horse.. PLoS One 2017;12(11):e0186247.
            doi: 10.1371/journal.pone.0186247pubmed: 29190290google scholar: lookup
          7. Ravara B, Gobbo V, Carraro U, Gelbmann L, Pribyl J, Schils S. Functional Electrical Stimulation as a Safe and Effective Treatment for Equine Epaxial Muscle Spasms: Clinical Evaluations and Histochemical Morphometry of Mitochondria in Muscle Biopsies.. Eur J Transl Myol 2015 Mar 11;25(2):4910.
            doi: 10.4081/ejtm.2015.4910pubmed: 26913151google scholar: lookup
          8. Brown DM, Brameld JM, Parr T. Expression of the myosin heavy chain IIB gene in porcine skeletal muscle: the role of the CArG-Box promoter response element.. PLoS One 2014;9(12):e114365.
            doi: 10.1371/journal.pone.0114365pubmed: 25469802google scholar: lookup
          9. Kohn TA. Insights into the skeletal muscle characteristics of three southern African antelope species.. Biol Open 2014 Oct 17;3(11):1037-44.
            doi: 10.1242/bio.20149241pubmed: 25326514google scholar: lookup
          10. Hyytiäinen HK, Mykkänen AK, Hielm-Björkman AK, Stubbs NC, McGowan CM. Muscle fibre type distribution of the thoracolumbar and hindlimb regions of horses: relating fibre type and functional role.. Acta Vet Scand 2014 Jan 27;56(1):8.
            doi: 10.1186/1751-0147-56-8pubmed: 24468115google scholar: lookup
          11. Zhang M, Liu YL, Fu CY, Wang J, Chen SY, Yao J, Lai SJ. Expression of MyHC genes, composition of muscle fiber type and their association with intramuscular fat, tenderness in skeletal muscle of Simmental hybrids.. Mol Biol Rep 2014 Feb;41(2):833-40.
            doi: 10.1007/s11033-013-2923-6pubmed: 24374854google scholar: lookup
          12. Graziotti GH, Chamizo VE, Ríos C, Acevedo LM, Rodríguez-Menéndez JM, Victorica C, Rivero JL. Adaptive functional specialisation of architectural design and fibre type characteristics in agonist shoulder flexor muscles of the llama, Lama glama.. J Anat 2012 Aug;221(2):151-63.
            doi: 10.1111/j.1469-7580.2012.01520.xpubmed: 22625659google scholar: lookup
          13. Butcher MT, Chase PB, Hermanson JW, Clark AN, Brunet NM, Bertram JE. Contractile properties of muscle fibers from the deep and superficial digital flexors of horses.. Am J Physiol Regul Integr Comp Physiol 2010 Oct;299(4):R996-R1005.
            doi: 10.1152/ajpregu.00510.2009pubmed: 20702801google scholar: lookup
          14. Rhee HS, Steel CM, Derksen FJ, Robinson NE, Hoh JF. Immunohistochemical analysis of laryngeal muscles in normal horses and horses with subclinical recurrent laryngeal neuropathy.. J Histochem Cytochem 2009 Aug;57(8):787-800.
            doi: 10.1369/jhc.2009.953844pubmed: 19398607google scholar: lookup
          15. Moreno-Sánchez N, Díaz C, Carabaño MJ, Rueda J, Rivero JL. A comprehensive characterisation of the fibre composition and properties of a limb (flexor digitorum superficialis, membri thoraci) and a trunk (psoas major) muscle in cattle.. BMC Cell Biol 2008 Dec 15;9:67.
            doi: 10.1186/1471-2121-9-67pubmed: 19077313google scholar: lookup
          16. Rhee HS, Hoh JF. Immunohistochemical analysis of myosin heavy chain expression in laryngeal muscles of the rabbit, cat, and baboon.. J Histochem Cytochem 2008 Oct;56(10):929-50.
            doi: 10.1369/jhc.2008.951756pubmed: 18606609google scholar: lookup
          17. Zhong WW, Lucas CA, Hoh JF. Myosin isoforms and fibre types in limb muscles of Australian marsupials: adaptations to hopping and non-hopping locomotion.. J Comp Physiol B 2008 Jan;178(1):47-55.
            doi: 10.1007/s00360-007-0198-8pubmed: 17703312google scholar: lookup
          18. Serrano AL, Rivero JL. Myosin heavy chain profile of equine gluteus medius muscle following prolonged draught-exercise training and detraining.. J Muscle Res Cell Motil 2000 Apr;21(3):235-45.
            doi: 10.1023/a:1005642632711pubmed: 10952171google scholar: lookup
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