Skip to main content
SpringerLink
Log in

New multiple labelling method for improved satellite cell identification in human muscle: application to a cohort of power-lifters and sedentary men

  • Original Paper
  • Published:
Histochemistry and Cell Biology Aims and scope Submit manuscript

Abstract

Presently applied methods to identify and quantify human satellite cells (SCs) give discrepant results. We introduce a new immunofluorescence method that simultaneously monitors two SC markers (NCAM and Pax7), the basal lamina and nuclei. Biopsies from power-lifters, power-lifters using anabolic substances and untrained subjects were re-examined. Significantly different results from those with staining for NCAM and nuclei were observed. There were three subtypes of SCs; NCAM+/Pax7+ (94%), NCAM+/Pax7− (4%) and NCAM−/Pax7+ (1%) but large individual variability existed. The proportion of SCs per nuclei within the basal lamina of myofibres (SC/N) was similar for all groups reflecting a balance between the number of SCs and myonuclei to maintain homeostasis. We emphasise that it is important to quantify both SC/N and the number of SCs per fibre. Our multiple marker method is more reliable for SC identification and quantification and can be used to evaluate other markers of muscle progenitor cells.

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

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8
Fig. 9

Similar content being viewed by others

References

  • Allen DL, Roy RR, Edgerton VR (1999) Myonuclear domains in muscle adaptation and disease. Muscle Nerve 22:1350–1360

    Article  PubMed  CAS  Google Scholar 

  • Bischoff R (1975) Regeneration of single skeletal muscle fibers in vitro. Anat Rec 182:215–235

    Article  PubMed  CAS  Google Scholar 

  • Buckingham M (2006) Myogenic progenitor cells and skeletal myogenesis in vertebrates. Curr Opin Genet Dev 16:525–532

    Article  PubMed  CAS  Google Scholar 

  • Buckingham M, Montarras D (2008) Skeletal muscle stem cells. Curr Opin Genet Dev 18:330–336

    Article  PubMed  CAS  Google Scholar 

  • Cashman NR, Covault J, Wollman RL, Sanes JR (1987) Neural cell adhesion molecule in normal, denervated, and myopathic human muscle. Ann Neurol 21:481–489

    Article  PubMed  CAS  Google Scholar 

  • Charifi N, Kadi F, Feasson L, Denis C (2003) Effects of endurance training on satellite cell frequency in skeletal muscle of old men. Muscle Nerve 28:87–92

    Article  PubMed  Google Scholar 

  • Cooper RN, Butler-Browne GS, Mouly V (2006) Human muscle stem cells. Curr Opin Pharmacol 6:295–300

    Article  PubMed  CAS  Google Scholar 

  • Crameri RM, Langberg H, Magnusson P, Jensen CH, Schroder HD, Olesen JL, Suetta C, Teisner B, Kjaer M (2004) Changes in satellite cells in human skeletal muscle after a single bout of high intensity exercise. J Physiol 558:333–340

    Article  PubMed  CAS  Google Scholar 

  • Crameri RM, Aagaard P, Qvortrup K, Langberg H, Olesen J, Kjaer M (2007) Myofibre damage in human skeletal muscle: effects of electrical stimulation vs voluntary contraction. J Physiol 583:365–380

    Article  PubMed  CAS  Google Scholar 

  • Day K, Shefer G, Richardson JB, Enikolopov G, Yablonka-Reuveni Z (2006) Nestin-GFP reporter expression defines the quiescent state of skeletal muscle satellite cells. Dev Biol 304:246–259

    Article  PubMed  CAS  Google Scholar 

  • Dreyer HC, Blanco CE, Sattler FR, Schroeder ET, Wiswell RA (2006) Satellite cell numbers in young and older men 24 hours after eccentric exercise. Muscle Nerve 33:242–253

    Article  PubMed  Google Scholar 

  • Eriksson A (2006) Strength training and anabolic steroids. The Department of Integrative Medical Biology, Section for Anatomy, Umeå University, Sweden and The Department of Health Science, Section for Medical Science, Luleå University of Technology, Sweden

    Google Scholar 

  • Eriksson A, Kadi F, Malm C, Thornell LE (2005) Skeletal muscle morphology in power-lifters with and without anabolic steroids. Histochem Cell Biol 124:167–175

    Article  PubMed  CAS  Google Scholar 

  • Eriksson A, Lindstrom M, Carlsson L, Thornell LE (2006) Hypertrophic muscle fibers with fissures in power-lifters; fiber splitting or defect regeneration? Histochem Cell Biol 126:409–417

    Article  PubMed  CAS  Google Scholar 

  • Gnocchi VF, White RB, Ono Y, Ellis JA, Zammit PS (2009) Further characterisation of the molecular signature of quiescent and activated mouse muscle satellite cells. PLoS ONE 4:e5205

    Article  PubMed  CAS  Google Scholar 

  • Gundersen HJ (1986) Stereology of arbitrary particles. A review of unbiased number and size estimators and the presentation of some new ones, in memory of William R. Thompson. J Microsc 143:3–45

    PubMed  CAS  Google Scholar 

  • Illa I, Leon-Monzon M, Dalakas MC (1992) Regenerating and denervated human muscle fibers and satellite cells express neural cell adhesion molecule recognized by monoclonal antibodies to natural killer cells. Ann Neurol 31:46–52

    Article  PubMed  CAS  Google Scholar 

  • Irintchev A, Zeschnigk M, Starzinski-Powitz A, Wernig A (1994) Expression pattern of M-cadherin in normal, denervated, and regenerating mouse muscles. Dev Dyn 199:326–337

    PubMed  CAS  Google Scholar 

  • Ishido M, Uda M, Kasuga N, Masuhara M (2009) The expression patterns of Pax7 in satellite cells during overload-induced rat adult skeletal muscle hypertrophy. Acta Physiol (Oxf) 195:459–469

    Article  CAS  Google Scholar 

  • Kadi F (2000) Adaptation of human skeletal muscle to training and anabolic steroids. Acta Physiol Scand Suppl 646:1–52

    PubMed  CAS  Google Scholar 

  • Kadi F (2008) Cellular and molecular mechanisms responsible for the action of testosterone on human skeletal muscle. A basis for illegal performance enhancement. Br J Pharmacol 154:522–528

    Article  PubMed  CAS  Google Scholar 

  • Kadi F, Thornell LE (2000) Concomitant increases in myonuclear and satellite cell content in female trapezius muscle following strength training. Histochem Cell Biol 113:99–103

    Article  PubMed  CAS  Google Scholar 

  • Kadi F, Eriksson A, Holmner S, Butler-Browne GS, Thornell LE (1999a) Cellular adaptation of the trapezius muscle in strength-trained athletes. Histochem Cell Biol 111:189–195

    Article  PubMed  CAS  Google Scholar 

  • Kadi F, Eriksson A, Holmner S, Thornell LE (1999b) Effects of anabolic steroids on the muscle cells of strength-trained athletes. Med Sci Sports Exerc 31:1528–1534

    Article  PubMed  CAS  Google Scholar 

  • Kadi F, Ahlgren C, Waling K, Sundelin G, Thornell LE (2000) The effects of different training programs on the trapezius muscle of women with work-related neck and shoulder myalgia. Acta Neuropathol 100:253–258

    Article  PubMed  CAS  Google Scholar 

  • Kadi F, Charifi N, Denis C, Lexell J (2004a) Satellite cells and myonuclei in young and elderly women and men. Muscle Nerve 29:120–127

    Article  PubMed  Google Scholar 

  • Kadi F, Schjerling P, Andersen LL, Charifi N, Madsen JL, Christensen LR, Andersen JL (2004b) The effects of heavy resistance training and detraining on satellite cells in human skeletal muscles. J Physiol 558:1005–1012

    Article  PubMed  CAS  Google Scholar 

  • Kadi F, Charifi N, Henriksson J (2006) The number of satellite cells in slow and fast fibres from human vastus lateralis muscle. Histochem Cell Biol 126:83–87

    Article  PubMed  CAS  Google Scholar 

  • Kuang S, Rudnicki MA (2008) The emerging biology of satellite cells and their therapeutic potential. Trends Mol Med 14:82–91

    Article  PubMed  CAS  Google Scholar 

  • Kuang S, Charge SB, Seale P, Huh M, Rudnicki MA (2006) Distinct roles for Pax7 and Pax3 in adult regenerative myogenesis. J Cell Biol 172:103–113

    Article  PubMed  CAS  Google Scholar 

  • Kuang S, Kuroda K, Le Grand F, Rudnicki MA (2007) Asymmetric self-renewal and commitment of satellite stem cells in muscle. Cell 129:999–1010

    Article  PubMed  CAS  Google Scholar 

  • Lanier LL, Testi R, Bindl J, Phillips JH (1989) Identity of Leu-19 (CD56) leukocyte differentiation antigen and neural cell adhesion molecule. J Exp Med 169:2233–2238

    Article  PubMed  CAS  Google Scholar 

  • Lipton BH, Schultz E (1979) Developmental fate of skeletal muscle satellite cells. Science 205:1292–1294

    Article  PubMed  CAS  Google Scholar 

  • Mackey AL, Esmarck B, Kadi F, Koskinen SO, Kongsgaard M, Sylvestersen A, Hansen JJ, Larsen G, Kjaer M (2007a) Enhanced satellite cell proliferation with resistance training in elderly men and women. Scand J Med Sci Sports 17:34–42

    PubMed  CAS  Google Scholar 

  • Mackey AL, Kjaer M, Dandanell S, Mikkelsen KH, Holm L, Dossing S, Kadi F, Koskinen SO, Jensen CH, Schroder HD, Langberg H (2007b) The influence of anti-inflammatory medication on exercise-induced myogenic precursor cell responses in humans. J Appl Physiol 103:425–431

    Article  PubMed  CAS  Google Scholar 

  • Mauro A (1961) Satellite cell of skeletal muscle fibers. J Biophys Biochem Cytol 9:493–495

    Article  PubMed  CAS  Google Scholar 

  • Mechtersheimer G, Staudter M, Moller P (1992) Expression of the natural killer (NK) cell-associated antigen CD56 (Leu-19), which is identical to the 140-kDa isoform of N-CAM, in neural and skeletal muscle cells and tumors derived therefrom. Ann N Y Acad Sci 650:311–316

    Article  PubMed  CAS  Google Scholar 

  • Olguin HC, Olwin BB (2004) Pax-7 up-regulation inhibits myogenesis and cell cycle progression in satellite cells: a potential mechanism for self-renewal. Dev Biol 275:375–388

    Article  PubMed  CAS  Google Scholar 

  • Olsen S, Aagaard P, Kadi F, Tufekovic G, Verney J, Olesen JL, Suetta C, Kjaer M (2006) Creatine supplementation augments the increase in satellite cell and myonuclei number in human skeletal muscle induced by strength training. J Physiol 573:525–534

    Article  PubMed  CAS  Google Scholar 

  • Oustanina S, Hause G, Braun T (2004) Pax7 directs postnatal renewal and propagation of myogenic satellite cells but not their specification. Embo J 23:3430–3439

    Article  PubMed  CAS  Google Scholar 

  • Peault B, Rudnicki M, Torrente Y, Cossu G, Tremblay JP, Partridge T, Gussoni E, Kunkel LM, Huard J (2007) Stem and progenitor cells in skeletal muscle development, maintenance, and therapy. Mol Ther 15:867–877

    Article  PubMed  CAS  Google Scholar 

  • Petrella JK, Kim JS, Cross JM, Kosek DJ, Bamman MM (2006) Efficacy of myonuclear addition may explain differential myofiber growth among resistance-trained young and older men and women. Am J Physiol Endocrinol Metab 291:E937–E946

    Article  PubMed  CAS  Google Scholar 

  • Petrella JK, Kim JS, Mayhew DL, Cross JM, Bamman MM (2008) Potent myofiber hypertrophy during resistance training in humans is associated with satellite cell-mediated myonuclear addition: a cluster analysis. J Appl Physiol 104:1736–1742

    Article  PubMed  Google Scholar 

  • Reimann J, Brimah K, Schroder R, Wernig A, Beauchamp JR, Partridge TA (2004) Pax7 distribution in human skeletal muscle biopsies and myogenic tissue cultures. Cell Tissue Res 315:233–242

    Article  PubMed  Google Scholar 

  • Relaix F, Montarras D, Zaffran S, Gayraud-Morel B, Rocancourt D, Tajbakhsh S, Mansouri A, Cumano A, Buckingham M (2006) Pax3 and Pax7 have distinct and overlapping functions in adult muscle progenitor cells. J Cell Biol 172:91–102

    Article  PubMed  CAS  Google Scholar 

  • Renault V, Rolland E, Thornell LE, Mouly V, Butler-Browne G (2002a) Distribution of satellite cells in the human vastus lateralis muscle during aging. Exp Gerontol 37:1513–1514

    Article  PubMed  CAS  Google Scholar 

  • Renault V, Thornell LE, Eriksson PO, Butler-Browne G, Mouly V (2002b) Regenerative potential of human skeletal muscle during aging. Aging Cell 1:132–139

    Article  PubMed  CAS  Google Scholar 

  • Roth SM, Martel GF, Ivey FM, Lemmer JT, Metter EJ, Hurley BF, Rogers MA (2000) Skeletal muscle satellite cell populations in healthy young and older men and women. Anat Rec 260:351–358

    Article  PubMed  CAS  Google Scholar 

  • Sajko S, Kubinova L, Cvetko E, Kreft M, Wernig A, Erzen I (2004) Frequency of M-cadherin-stained satellite cells declines in human muscles during aging. J Histochem Cytochem 52:179–185

    PubMed  CAS  Google Scholar 

  • Schmalbruch H, Hellhammer U (1976) The number of satellite cells in normal human muscle. Anat Rec 185:279–287

    Article  PubMed  CAS  Google Scholar 

  • Schubert W, Zimmermann K, Cramer M, Starzinski-Powitz A (1989) Lymphocyte antigen Leu-19 as a molecular marker of regeneration in human skeletal muscle. Proc Natl Acad Sci U S A 86:307–311

    Article  PubMed  CAS  Google Scholar 

  • Seale P, Sabourin LA, Girgis-Gabardo A, Mansouri A, Gruss P, Rudnicki MA (2000) Pax7 is required for the specification of myogenic satellite cells. Cell 102:777–786

    Article  PubMed  CAS  Google Scholar 

  • Shefer G, Yablonka-Reuveni Z (2007) Reflections on lineage potential of skeletal muscle satellite cells: do they sometimes go MAD? Crit Rev Eukaryot Gene Expr 17:13–29

    PubMed  CAS  Google Scholar 

  • Shefer G, Van de Mark DP, Richardson JB, Yablonka-Reuveni Z (2006) Satellite-cell pool size does matter: defining the myogenic potency of aging skeletal muscle. Dev Biol 294:50–66

    Article  PubMed  CAS  Google Scholar 

  • Sinha-Hikim I, Roth SM, Lee MI, Bhasin S (2003) Testosterone-induced muscle hypertrophy is associated with an increase in satellite cell number in healthy, young men. Am J Physiol Endocrinol Metab 285:E197–E205

    PubMed  CAS  Google Scholar 

  • Sinha-Hikim I, Cornford M, Gaytan H, Lee ML, Bhasin S (2006) Effects of testosterone supplementation on skeletal muscle fiber hypertrophy and satellite cells in community-dwelling older men. J Clin Endocrinol Metab 91:3024–3033

    Article  PubMed  CAS  Google Scholar 

  • Snow MH (1978) An autoradiographic study of satellite cell differentiation into regenerating myotubes following transplantation of muscles in young rats. Cell Tissue Res 186:535–540

    Article  PubMed  CAS  Google Scholar 

  • Thornell L-E, Eriksson A, Edström L (1983) Intermediate filaments in human myophaties. Dowben RM, Shay JW (eds) Cell and muscle motility, vol 4. Plenum Press, New York, pp 85–136

    Google Scholar 

  • Thornell LE, Lindstrom M, Renault V, Mouly V, Butler-Browne GS (2003) Satellite cells and training in the elderly. Scand J Med Sci Sports 13:48–55

    Article  PubMed  Google Scholar 

  • Verdijk LB, Koopman R, Schaart G, Meijer K, Savelberg HH, van Loon LJ (2007) Satellite cell content is specifically reduced in type II skeletal muscle fibers in the elderly. Am J Physiol Endocrinol Metab 292:E151–E157

    Article  PubMed  CAS  Google Scholar 

  • Verney J, Kadi F, Charifi N, Feasson L, Saafi MA, Castells J, Piehl-Aulin K, Denis C (2008) Effects of combined lower body endurance and upper body resistance training on the satellite cell pool in elderly subjects. Muscle Nerve 38:1147–1154

    Article  PubMed  Google Scholar 

  • Zammit PS (2008) The muscle satellite cell: the story of a cell on the edge. In: Schiaffino S, Partridge T (eds) Skeletal muscle repair and regeneration. Springer, pp 45–64

  • Zammit PS, Golding JP, Nagata Y, Hudon V, Partridge TA, Beauchamp JR (2004) Muscle satellite cells adopt divergent fates: a mechanism for self-renewal? J Cell Biol 166:347–357

    Article  PubMed  CAS  Google Scholar 

  • Zammit PS, Partridge TA, Yablonka-Reuveni Z (2006a) The skeletal muscle satellite cell: the stem cell that came in from the cold. J Histochem Cytochem 54:1177–1191

    Article  PubMed  CAS  Google Scholar 

  • Zammit PS, Relaix F, Nagata Y, Perez Ruiz A, Collins CA, Partridge TA, Beauchamp JR (2006b) Pax7 and myogenic progression in skeletal muscle satellite cells. J Cell Sci 119:1824–1832

    Article  PubMed  CAS  Google Scholar 

  • Zheng B, Cao B, Crisan M, Sun B, Li G, Logar A, Yap S, Pollett JB, Drowley L, Cassino T, Gharaibeh B, Deasy BM, Huard J, Peault B (2007) Prospective identification of myogenic endothelial cells in human skeletal muscle. Nat Biotechnol 25:1025–1034

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

We wish to thank Mrs. Anna-Karin Olofsson and Margaretha Enerstedt for excellent technical assistance, Asst. profs Peter Anton and Leif Nilsson for statistical support, Ph.D Lena Carlsson and Prof. Fatima Pedrosa Domellöf for valuable comments on the manuscript and Ph.D Anders Eriksson for referring the power-lifter subjects to us. This study was supported by grants from the Association Française contre les Myopathies, MYORES Network of Excellence (contract 511978) of the European Union, the Swedish Research Council (12x3934), the Swedish National Centre for Research in Sports (108/08, 99/09) and the Medical Faculty of Umeå University, Sweden.

Author information

Authors and Affiliations

  1. Section for Anatomy, Department of Integrative Medical Biology, Umeå University, 901 87, Umeå, Sweden

    Mona Lindström & Lars-Eric Thornell

Authors
  1. Mona Lindström
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Lars-Eric Thornell
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Lars-Eric Thornell.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Lindström, M., Thornell, LE. New multiple labelling method for improved satellite cell identification in human muscle: application to a cohort of power-lifters and sedentary men. Histochem Cell Biol 132, 141–157 (2009). https://doi.org/10.1007/s00418-009-0606-0

Download citation

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00418-009-0606-0

Keywords

Search

Navigation