Abstract
Skeletal muscles contain stem cells called satellite cells, which are essential for muscle regeneration. The population of satellite cells declines with aging and the incidence of pathological conditions such as muscular dystrophy. There is increasing evidence that metabolic switches and mitochondrial function are critical regulators of cell fate decision (quiescence, activation, differentiation, and self-renewal) during myogenesis. Thus, monitoring and identifying the metabolic profile in live cells using the Seahorse XF Bioanalyzer could provide new insights on the molecular mechanisms governing stem cell dynamics during regeneration and tissue maintenance. Here we described a method to assess mitochondrial respiration (oxygen consumption rate) and glycolysis (ECAR) in primary murine satellite cells, multinucleated myotubes, and C2C12 myoblasts.
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References
Fujita R, Yoshioka K, Seko D et al (2018) Zmynd17 controls muscle mitochondrial quality and whole-body metabolism. FASEB J 32:5012–5025
Yoshioka K, Fujita R, Seko D et al (2019) Distinct roles of Zmynd17 and PGC1α in mitochondrial quality control and biogenesis in skeletal muscle. Front Cell Dev Biol 7:330
Chen H, Vermulst M, Wang YE et al (2010) Mitochondrial fusion is required for mtDNA stability in skeletal muscle and tolerance of mtDNA mutations. Cell 141:280–289
Koves TR, Ussher JR, Noland RC et al (2008) Mitochondrial overload and incomplete fatty acid oxidation contribute to skeletal muscle insulin resistance. Cell Metab 7:45–56
Mootha VK, Lindgren CM, Eriksson K-F et al (2003) PGC-1α-responsive genes involved in oxidative phosphorylation are coordinately downregulated in human diabetes. Nat Genet 34:267–273
Petersen KF (2003) Mitochondrial dysfunction in the elderly: possible role in insulin resistance. Science 300:1140–1142
Petersen KF, Dufour S, Befroy D et al (2004) Impaired mitochondrial activity in the insulin-resistant offspring of patients with type 2 diabetes. N Engl J Med 350:664–671
Amati-Bonneau P, Valentino ML, Reynier P et al (2008) OPA1 mutations induce mitochondrial DNA instability and optic atrophy “plus” phenotypes. Brain 131:338–351
Mauro A (1961) Satellite cell of skeletal muscle fibers. J Biophys Biochem Cytol 9:493–495
Yin H, Price F, Rudnicki MA (2013) Satellite cells and the muscle stem cell niche. Physiol Rev 93:23–67
Brack AS, Rando TA (2012) Tissue-specific stem cells: lessons from the skeletal muscle satellite cell. Cell Stem Cell 10:504–514
Dumont NA, Wang YX, Rudnicki MA (2015) Intrinsic and extrinsic mechanisms regulating satellite cell function. Dev Camb Engl 142:1572–1581
Rodgers JT, King KY, Brett JO et al (2014) mTORC1 controls the adaptive transition of quiescent stem cells from G0 to GAlert. Nature 510:393–396
Tang AH, Rando TA (2014) Induction of autophagy supports the bioenergetic demands of quiescent muscle stem cell activation. EMBO J 33:2782–2797
Zismanov V, Chichkov V, Colangelo V et al (2016) Phosphorylation of eIF2α is a translational control mechanism regulating muscle stem cell quiescence and self-renewal. Cell Stem Cell 18:79–90
Fujita R, Crist C (2018) Translational control of the myogenic program in developing, regenerating, and diseased skeletal muscle. In: Current topics in developmental biology. Elsevier, pp 67–98
Crist CG, Montarras D, Buckingham M (2012) Muscle satellite cells are primed for myogenesis but maintain quiescence with sequestration of Myf5 mRNA targeted by microRNA-31 in mRNP granules. Cell Stem Cell 11:118–126
Ryall JG, Dell’Orso S, Derfoul A et al (2015) The NAD+-dependent SIRT1 deacetylase translates a metabolic switch into regulatory epigenetics in skeletal muscle stem cells. Cell Stem Cell 16:171–183
Ryall JG, Cliff T, Dalton S et al (2015) Metabolic reprogramming of stem cell epigenetics. Cell Stem Cell 17:651–662
Sala D, Cunningham TJ, Stec MJ et al (2019) The Stat3-Fam3a axis promotes muscle stem cell myogenic lineage progression by inducing mitochondrial respiration. Nat Commun 10:1796
Sin J, Andres AM, Taylor DJR et al (2016) Mitophagy is required for mitochondrial biogenesis and myogenic differentiation of C2C12 myoblasts. Autophagy 12:369–380
Khacho M, Clark A, Svoboda DS et al (2016) Mitochondrial dynamics impacts stem cell identity and fate decisions by regulating a nuclear transcriptional program. Cell Stem Cell 19:232–247
Acknowledgments
The authors thank Dr. Kazuo Yamamoto (Nagasaki University, School of Medicine, Nagasaki, Japan) and Dr. Yusuke Ono (Institute of Molecular Embryology and Genetics, Kumamoto University) for technical support. This work was supported by a MEXT Leading Initiative for Excellent Young Researchers.
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Takeda, K., Takemasa, T., Fujita, R. (2023). High Throughput Screening of Mitochondrial Bioenergetics in Myoblasts and Differentiated Myotubes. In: Asakura, A. (eds) Skeletal Muscle Stem Cells. Methods in Molecular Biology, vol 2640. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3036-5_7
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DOI: https://doi.org/10.1007/978-1-0716-3036-5_7
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Publisher Name: Humana, New York, NY
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Online ISBN: 978-1-0716-3036-5
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