Abstract
Satellite cells are mononucleated cells of the skeletal muscle lineage that exist beneath the basal lamina juxtaposed to the sarcolemma of skeletal muscle fibers. It is widely accepted that satellite cells mediate skeletal muscle regeneration. Within the satellite cell pool of adult muscle are skeletal muscle stem cells (MuSCs), also called satellite stem cells, which fulfill criteria of tissue stem cells: They proliferate and their progeny either occupies the adult MuSC niche during self-renewal or differentiates to regenerate mature muscle fibers. Here, we describe robust methods for the isolation of enriched populations of human satellite cells containing MuSCs from fresh human muscle, utilizing mechanical and enzymatic dissociation and purification by fluorescence-activated cell sorting. We also describe a process for xenotransplantation of human satellite cells into mouse muscle by injection into irradiated, immunodeficient, mouse leg muscle with concurrent notexin or bupivacaine muscle injury to increase engraftment efficiency. The engraftment of human MuSCs and the formation of human muscle can then be analyzed by histological and immunofluorescence staining, or subjected to in vivo experimentation.
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References
Collins CA, Olsen I, Zammit PS, Heslop L, Petrie A, Partridge TA, Morgan JE (2005) Stem cell function, self-renewal, and behavioral heterogeneity of cells from the adult muscle satellite cell niche. Cell 122:289–301
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
Montarras D, Morgan J, Collins C, Relaix F, Zaffran S, Cumano A, Partridge T, Buckingham M (2005) Direct isolation of satellite cells for skeletal muscle regeneration. Science 309:2064–2067
Sacco A, Doyonnas R, Kraft P, Vitorovic S, Blau HM (2008) Self-renewal and expansion of single transplanted muscle stem cells. Nature 456:502–506
Sherwood RI, Christensen JL, Conboy IM, Conboy MJ, Rando TA, Weissman IL, Wagers AJ (2004) Isolation of adult mouse myogenic progenitors: functional heterogeneity of cells within and engrafting skeletal muscle. Cell 119:543–554
Dumont NA, Bentzinger CF, Sincennes MC, Rudnicki MA (2015) Satellite cells and skeletal muscle regeneration. Compr Physiol 5:1027–1059
Yin H, Price F, Rudnicki MA (2013) Satellite cells and the muscle stem cell niche. Physiol Rev 93:23–67
Bareja A, Holt JA, Luo G, Chang C, Lin J, Hinken AC, Freudenberg JM, Kraus WE, Evans WJ, Billin AN (2014) Human and mouse skeletal muscle stem cells: convergent and divergent mechanisms of myogenesis. PLoS One 9:e90398
Boldrin L, Morgan JE (2012) Human satellite cells: identification on human muscle fibres. PLoS Curr 3:RRN1294
Castiglioni A, Hettmer S, Lynes MD, Rao TN, Tchessalova D, Sinha I, Lee BT, Tseng YH, Wagers AJ (2014) Isolation of progenitors that exhibit myogenic/osteogenic bipotency in vitro by fluorescence-activated cell sorting from human fetal muscle. Stem Cell Reports 2:92–106
Darabi R, Arpke RW, Irion S, Dimos JT, Grskovic M, Kyba M, Perlingeiro RC (2012) Human ES- and iPS-derived myogenic progenitors restore DYSTROPHIN and improve contractility upon transplantation in dystrophic mice. Cell Stem Cell 10:610–619
Ehrhardt J, Brimah K, Adkin C, Partridge T, Morgan J (2007) Human muscle precursor cells give rise to functional satellite cells in vivo. Neuromuscul Disord 17:631–638
Lindstrom M, Thornell LE (2009) 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
Marg A, Escobar H, Gloy S, Kufeld M, Zacher J, Spuler A, Birchmeier C, Izsva´k Z, Spuler S (2014) Human satellite cells have regenerative capacity and are genetically manipulable. J Clin Invest 124:4257–4265
Meng J, Chun S, Asfahani R, Lochmuller H, Muntoni F, Morgan J (2014) Human skeletal muscle-derived CD133(+) cells form functional satellite cells after intramuscular transplantation in immunodeficient host mice. Mol Ther 22:1008–1017
Miller RG, Sharma KR, Pavlath GK, Gussoni E, Mynhier M, Lanctot AM, Greco CM, Steinman L, Blau HM (1997) Myoblast implantation in Duchenne muscular dystrophy: the San Francisco study. Muscle Nerve 20:469–478
Partridge T (2002) Myoblast transplantation. Neuromuscul Disord 12(Suppl 1):S3–S6
Pisani DF, Dechesne CA, Sacconi S, Delplace S, Belmonte N, Cochet O, Clement N, Wdziekonski B, Villageois AP, Butori C, Bagins C, Di Santo JP, Kurzenne JY, Desnuelle C, Dani C (2010) Isolation of a highly myogenic CD34-negative subset of human skeletal muscle cells free of adipogenic potential. Stem Cells 28:753–764
Silva-Barbosa SD, Butler-Browne GS, de Mello W, Riederer I, Di Santo JP, Savino W, Mouly V (2008) Human myoblast engraftment is improved in laminin-enriched microenvironment. Transplantation 85:566–575
Skuk D, Paradis M, Goulet M, Chapdelaine P, Rothstein DM, Tremblay JP (2010) Intramuscular transplantation of human postnatal myoblasts generates functional donor-derived satellite cells. Mol Ther 18:1689–1697
Charville GW, Cheung TH, Yoo B, Santos PJ, Lee GK, Shrager JB, Rando TA (2015) Ex vivo expansion and in vivo self-renewal of human muscle. Stem Cell Reports 5:621–632
Xu X, Wilschut KJ, Kouklis G, Tian H, Hesse R, Garland C, Sbitany H, Hansen S, Seth R, Knott PD, Hoffman WY, Pomerantz JH (2015) Human satellite cell transplantation and regeneration from diverse skeletal muscles. Stem Cell Reports 5:419–434
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 86:307–311
Boldrin L, Neal A, Zammit PS, Muntoni F, Morgan JE (2012) Donor satellite cell engraftment is significantly augmented when the host niche is preserved and endogenous satellite cells are incapacitated. Stem Cells 30:1971–1984
Acknowledgments
This work was funded by CIRM New Faculty Physician Scientist Award RN3-06504 to JHP and the UCSF PROF-PATH program through R25MD006832 from the National Institute on Minority Health and Health Disparities fellowship to SMG.
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Garcia, S.M., Tamaki, S., Xu, X., Pomerantz, J.H. (2017). Human Satellite Cell Isolation and Xenotransplantation. In: Ryall, J. (eds) Skeletal Muscle Development. Methods in Molecular Biology, vol 1668. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7283-8_8
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DOI: https://doi.org/10.1007/978-1-4939-7283-8_8
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