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Myogenesis pp 213-228 | Cite as

LC-MS Analyses of Lipid Species in Skeletal Muscle Cells and Tissue

  • Marta Moreno-Torres
  • Jesper F. Havelund
  • Nils J. FaergemanEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1889)

Abstract

Liquid chromatography–mass spectrometry (LC-MS) is a widely used methodology for measuring lipids at a global level. Combined with an optimal extraction method LC-MS enables the detection and characterization of a wide range of lipid species even of low abundance. Here, we describe two extraction- and LC-MS-based quantitative analytical methods for lipid, acyl-CoA, and acyl-carnitine analyses from either mouse C2C12 myotubes or mouse skeletal tissue. We also describe the use of 13C16-palmitate and its incorporation into acyl-carnitines to show how stable isotope tracers are metabolized within cells and therefore can be implemented for lipidomic flux analysis.

Key words

Lipidomics Acyl-CoA Acyl-carnitine Liquid chromatography Mass spectrometry Stable isotope tracers C2C12 myotubes Skeletal muscle tissue Labeling Palmitate 

Notes

Acknowledgments

This work was supported by The Danish Council for Independent Research, Natural Sciences. Marta Moreno-Torres was supported by a postdoctoral grant from The Danish Diabetes Academy.

References

  1. 1.
    Coen PM, Goodpaster BH (2012) Role of intramyocelluar lipids in human health. Trends Endocrinol Metab 23(8):391–398.  https://doi.org/10.1016/j.tem.2012.05.009CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Gault CR, Obeid LM, Hannun YA (2010) An overview of sphingolipid metabolism: from synthesis to breakdown. Adv Exp Med Biol 688:1–23CrossRefGoogle Scholar
  3. 3.
    Turner N, Kowalski GM, Leslie SJ, Risis S, Yang C, Lee-Young RS, Babb JR, Meikle PJ, Lancaster GI, Henstridge DC, White PJ, Kraegen EW, Marette A, Cooney GJ, Febbraio MA, Bruce CR (2013) Distinct patterns of tissue-specific lipid accumulation during the induction of insulin resistance in mice by high-fat feeding. Diabetologia 56(7):1638–1648.  https://doi.org/10.1007/s00125-013-2913-1CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Bergman BC, Hunerdosse DM, Kerege A, Playdon MC, Perreault L (2012) Localisation and composition of skeletal muscle diacylglycerol predicts insulin resistance in humans. Diabetologia 55(4):1140–1150.  https://doi.org/10.1007/s00125-011-2419-7CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Coen PM, Dube JJ, Amati F, Stefanovic-Racic M, Ferrell RE, Toledo FG, Goodpaster BH (2010) Insulin resistance is associated with higher intramyocellular triglycerides in type I but not type II myocytes concomitant with higher ceramide content. Diabetes 59(1):80–88.  https://doi.org/10.2337/db09-0988CrossRefPubMedGoogle Scholar
  6. 6.
    Bozic J, Markotic A, Cikes-Culic V, Novak A, Borovac JA, Vucemilovic H, Trgo G, Ticinovic Kurir T (2018) Ganglioside GM3 content in skeletal muscles is increased in type 2 but decreased in type 1 diabetes rat models: implications of glycosphingolipid metabolism in pathophysiology of diabetes. J Diabetes 10(2):130–139.  https://doi.org/10.1111/1753-0407.12569CrossRefPubMedGoogle Scholar
  7. 7.
    Albarracin I, Lassaga FE, Caputto R (1974) Changes of gangliosides and other lipids in skeletal muscle from rabbits with experimental dystrophy. J Lipid Res 15(1):89–93PubMedGoogle Scholar
  8. 8.
    Higatsberger MR, Auff E (1984) Gangliosides in rabbit and human skeletal muscle with denervation atrophy. J Neurol 231(2):79–82CrossRefGoogle Scholar
  9. 9.
    Stanley CA, Hale DE (1994) Genetic disorders of mitochondrial fatty acid oxidation. Curr Opin Pediatr 6(4):476–481CrossRefGoogle Scholar
  10. 10.
    Folch J, Lees M, Sloane Stanley GH (1957) A simple method for the isolation and purification of total lipides from animal tissues. J Biol Chem 226(1):497–509Google Scholar
  11. 11.
    Bligh EG, Dyer WJ (1959) A rapid method of total lipid extraction and purification. Can J Biochem Physiol 37(8):911–917.  https://doi.org/10.1139/o59-099CrossRefGoogle Scholar
  12. 12.
    Svennerholm L, Fredman P (1980) A procedure for the quantitative isolation of brain gangliosides. Biochim Biophys Acta 617(1):97–109CrossRefGoogle Scholar
  13. 13.
    Schiller J, Arnold K (2002) Application of high resolution 31P NMR spectroscopy to the characterization of the phospholipid composition of tissues and body fluids - a methodological review. Med Sci Monit 8(11):MT205–MT222PubMedGoogle Scholar
  14. 14.
    Iverson SJ, Lang SL, Cooper MH (2001) Comparison of the Bligh and dyer and Folch methods for total lipid determination in a broad range of marine tissue. Lipids 36(11):1283–1287CrossRefGoogle Scholar
  15. 15.
    Fong B, Norris C, Lowe E, McJarrow P (2009) Liquid chromatography-high-resolution mass spectrometry for quantitative analysis of gangliosides. Lipids 44(9):867–874.  https://doi.org/10.1007/s11745-009-3327-1CrossRefPubMedGoogle Scholar
  16. 16.
    Neess D, Bek S, Engelsby H, Gallego SF, Faergeman NJ (2015) Long-chain acyl-CoA esters in metabolism and signaling: role of acyl-CoA binding proteins. Prog Lipid Res 59:1–25.  https://doi.org/10.1016/j.plipres.2015.04.001CrossRefPubMedGoogle Scholar
  17. 17.
    McCoin CS, Knotts TA, Adams SH (2015) Acylcarnitines--old actors auditioning for new roles in metabolic physiology. Nat Rev Endocrinol 11(10):617–625.  https://doi.org/10.1038/nrendo.2015.129CrossRefPubMedPubMedCentralGoogle Scholar
  18. 18.
    Haynes CA (2011) Analysis of mammalian fatty acyl-coenzyme A species by mass spectrometry and tandem mass spectrometry. Biochim Biophys Acta 1811(11):663–668.  https://doi.org/10.1016/j.bbalip.2011.05.010CrossRefPubMedGoogle Scholar
  19. 19.
    Mackay GM, Zheng L, van den Broek NJ, Gottlieb E (2015) Analysis of cell metabolism using LC-MS and isotope tracers. Methods Enzymol 561:171–196.  https://doi.org/10.1016/bs.mie.2015.05.016CrossRefPubMedGoogle Scholar
  20. 20.
    Dettmer K, Aronov PA, Hammock BD (2007) Mass spectrometry-based metabolomics. Mass Spectrom Rev 26(1):51–78.  https://doi.org/10.1002/mas.20108CrossRefPubMedPubMedCentralGoogle Scholar
  21. 21.
    Pauling JK, Hermansson M, Hartler J, Christiansen K, Gallego SF, Peng B, Ahrends R, Ejsing CS (2017) Proposal for a common nomenclature for fragment ions in mass spectra of lipids. PLoS One 12(11):e0188394.  https://doi.org/10.1371/journal.pone.0188394CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Xia J, Wishart DS (2016) Using MetaboAnalyst 3.0 for comprehensive metabolomics data analysis. Curr Protoc Bioinformatics 55:14 10 11-14 10 91.  https://doi.org/10.1002/cpbi.11CrossRefGoogle Scholar
  23. 23.
    Dietmair S, Timmins NE, Gray PP, Nielsen LK, Kromer JO (2010) Towards quantitative metabolomics of mammalian cells: development of a metabolite extraction protocol. Anal Biochem 404(2):155–164.  https://doi.org/10.1016/j.ab.2010.04.031CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Marta Moreno-Torres
    • 1
  • Jesper F. Havelund
    • 1
  • Nils J. Faergeman
    • 1
    Email author
  1. 1.Department of Biochemistry and Molecular Biology, Villum Center for Bioanalytical SciencesUniversity of Southern DenmarkOdense MDenmark

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