Prospective isolation of skeletal muscle stem cells with a Pax7 reporter

Darko Bosnakovski, Zhaohui Xu, Wei Li, Suwannee Thet, Ondine Cleaver, Rita C R Perlingeiro, Michael Kyba

Research output: Contribution to journalArticle

103 Citations (Scopus)

Abstract

Muscle regeneration occurs through activation of quiescent satellite cells whose progeny proliferate, differentiate, and fuse to make new myofibers. We used a transgenic Pax7-ZsGreen reporter mouse to prospectively isolate stem cells of skeletal muscle by flow cytometry. We show that Pax7-expressing cells (satellite cells) in the limb, head, and diaphragm muscles are homogeneous in size and granularity and uniformly labeled by certain cell surface markers, including CD34 and CD29. The frequency of the satellite cells varies between muscle types and with age. Clonal analysis demonstrated that all colonies arising from single cells within the Pax7-sorted fraction have myogenic potential. In response to injury, Pax7+ cells reduce CD34, CD29, and CXCR4 expression, increase in size, and acquire Sca-1. When directly isolated and cultured in vitro, Pax7+ cells display the hallmarks of activation and proliferate, initially as suspension aggregates and later distributed between suspension and adherence. During in vitro expansion, Pax7 (ZsGreen) and CD34 expression decline, whereas expression of PSA-NCAM is acquired. The nonmyogenic, Pax7neg cells expand as Sca1+ PDGRα+ PSA-NCAMneg cells. Satellite cells expanded exclusively in suspension can engraft and produce dystrophin+ fibers in mdx-/- mice. These results establish a novel animal model for the study of muscle stem cell physiology and a culture system for expansion of engraftable muscle progenitors.

Original languageEnglish (US)
Pages (from-to)3194-3204
Number of pages11
JournalStem Cells
Volume26
Issue number12
DOIs
StatePublished - Dec 2008

Fingerprint

Muscle Cells
Skeletal Muscle
Stem Cells
Muscles
Suspensions
Inbred mdx Mouse
Cell Physiological Phenomena
Dystrophin
Diaphragm
Regeneration
Flow Cytometry
Extremities
Animal Models
Head
Wounds and Injuries

Keywords

  • Experimental models
  • Muscle stem cells
  • Muscular dystrophy
  • Pax7

ASJC Scopus subject areas

  • Cell Biology
  • Developmental Biology
  • Molecular Medicine

Cite this

Bosnakovski, D., Xu, Z., Li, W., Thet, S., Cleaver, O., Perlingeiro, R. C. R., & Kyba, M. (2008). Prospective isolation of skeletal muscle stem cells with a Pax7 reporter. Stem Cells, 26(12), 3194-3204. https://doi.org/10.1634/stemcells.2007-1017

Prospective isolation of skeletal muscle stem cells with a Pax7 reporter. / Bosnakovski, Darko; Xu, Zhaohui; Li, Wei; Thet, Suwannee; Cleaver, Ondine; Perlingeiro, Rita C R; Kyba, Michael.

In: Stem Cells, Vol. 26, No. 12, 12.2008, p. 3194-3204.

Research output: Contribution to journalArticle

Bosnakovski, D, Xu, Z, Li, W, Thet, S, Cleaver, O, Perlingeiro, RCR & Kyba, M 2008, 'Prospective isolation of skeletal muscle stem cells with a Pax7 reporter', Stem Cells, vol. 26, no. 12, pp. 3194-3204. https://doi.org/10.1634/stemcells.2007-1017
Bosnakovski, Darko ; Xu, Zhaohui ; Li, Wei ; Thet, Suwannee ; Cleaver, Ondine ; Perlingeiro, Rita C R ; Kyba, Michael. / Prospective isolation of skeletal muscle stem cells with a Pax7 reporter. In: Stem Cells. 2008 ; Vol. 26, No. 12. pp. 3194-3204.
@article{ddb37b8445aa42db868c87fc961562d2,
title = "Prospective isolation of skeletal muscle stem cells with a Pax7 reporter",
abstract = "Muscle regeneration occurs through activation of quiescent satellite cells whose progeny proliferate, differentiate, and fuse to make new myofibers. We used a transgenic Pax7-ZsGreen reporter mouse to prospectively isolate stem cells of skeletal muscle by flow cytometry. We show that Pax7-expressing cells (satellite cells) in the limb, head, and diaphragm muscles are homogeneous in size and granularity and uniformly labeled by certain cell surface markers, including CD34 and CD29. The frequency of the satellite cells varies between muscle types and with age. Clonal analysis demonstrated that all colonies arising from single cells within the Pax7-sorted fraction have myogenic potential. In response to injury, Pax7+ cells reduce CD34, CD29, and CXCR4 expression, increase in size, and acquire Sca-1. When directly isolated and cultured in vitro, Pax7+ cells display the hallmarks of activation and proliferate, initially as suspension aggregates and later distributed between suspension and adherence. During in vitro expansion, Pax7 (ZsGreen) and CD34 expression decline, whereas expression of PSA-NCAM is acquired. The nonmyogenic, Pax7neg cells expand as Sca1+ PDGRα+ PSA-NCAMneg cells. Satellite cells expanded exclusively in suspension can engraft and produce dystrophin+ fibers in mdx-/- mice. These results establish a novel animal model for the study of muscle stem cell physiology and a culture system for expansion of engraftable muscle progenitors.",
keywords = "Experimental models, Muscle stem cells, Muscular dystrophy, Pax7",
author = "Darko Bosnakovski and Zhaohui Xu and Wei Li and Suwannee Thet and Ondine Cleaver and Perlingeiro, {Rita C R} and Michael Kyba",
year = "2008",
month = "12",
doi = "10.1634/stemcells.2007-1017",
language = "English (US)",
volume = "26",
pages = "3194--3204",
journal = "Stem Cells",
issn = "1066-5099",
publisher = "AlphaMed Press",
number = "12",

}

TY - JOUR

T1 - Prospective isolation of skeletal muscle stem cells with a Pax7 reporter

AU - Bosnakovski, Darko

AU - Xu, Zhaohui

AU - Li, Wei

AU - Thet, Suwannee

AU - Cleaver, Ondine

AU - Perlingeiro, Rita C R

AU - Kyba, Michael

PY - 2008/12

Y1 - 2008/12

N2 - Muscle regeneration occurs through activation of quiescent satellite cells whose progeny proliferate, differentiate, and fuse to make new myofibers. We used a transgenic Pax7-ZsGreen reporter mouse to prospectively isolate stem cells of skeletal muscle by flow cytometry. We show that Pax7-expressing cells (satellite cells) in the limb, head, and diaphragm muscles are homogeneous in size and granularity and uniformly labeled by certain cell surface markers, including CD34 and CD29. The frequency of the satellite cells varies between muscle types and with age. Clonal analysis demonstrated that all colonies arising from single cells within the Pax7-sorted fraction have myogenic potential. In response to injury, Pax7+ cells reduce CD34, CD29, and CXCR4 expression, increase in size, and acquire Sca-1. When directly isolated and cultured in vitro, Pax7+ cells display the hallmarks of activation and proliferate, initially as suspension aggregates and later distributed between suspension and adherence. During in vitro expansion, Pax7 (ZsGreen) and CD34 expression decline, whereas expression of PSA-NCAM is acquired. The nonmyogenic, Pax7neg cells expand as Sca1+ PDGRα+ PSA-NCAMneg cells. Satellite cells expanded exclusively in suspension can engraft and produce dystrophin+ fibers in mdx-/- mice. These results establish a novel animal model for the study of muscle stem cell physiology and a culture system for expansion of engraftable muscle progenitors.

AB - Muscle regeneration occurs through activation of quiescent satellite cells whose progeny proliferate, differentiate, and fuse to make new myofibers. We used a transgenic Pax7-ZsGreen reporter mouse to prospectively isolate stem cells of skeletal muscle by flow cytometry. We show that Pax7-expressing cells (satellite cells) in the limb, head, and diaphragm muscles are homogeneous in size and granularity and uniformly labeled by certain cell surface markers, including CD34 and CD29. The frequency of the satellite cells varies between muscle types and with age. Clonal analysis demonstrated that all colonies arising from single cells within the Pax7-sorted fraction have myogenic potential. In response to injury, Pax7+ cells reduce CD34, CD29, and CXCR4 expression, increase in size, and acquire Sca-1. When directly isolated and cultured in vitro, Pax7+ cells display the hallmarks of activation and proliferate, initially as suspension aggregates and later distributed between suspension and adherence. During in vitro expansion, Pax7 (ZsGreen) and CD34 expression decline, whereas expression of PSA-NCAM is acquired. The nonmyogenic, Pax7neg cells expand as Sca1+ PDGRα+ PSA-NCAMneg cells. Satellite cells expanded exclusively in suspension can engraft and produce dystrophin+ fibers in mdx-/- mice. These results establish a novel animal model for the study of muscle stem cell physiology and a culture system for expansion of engraftable muscle progenitors.

KW - Experimental models

KW - Muscle stem cells

KW - Muscular dystrophy

KW - Pax7

UR - http://www.scopus.com/inward/record.url?scp=58049195427&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=58049195427&partnerID=8YFLogxK

U2 - 10.1634/stemcells.2007-1017

DO - 10.1634/stemcells.2007-1017

M3 - Article

VL - 26

SP - 3194

EP - 3204

JO - Stem Cells

JF - Stem Cells

SN - 1066-5099

IS - 12

ER -