Abstract
Natural killer (NK) cells are cytotoxic cells that mediate anti-tumor and anti-viral immunity. The response of NK cells to different cytokines and stimuli may involve cell survival, proliferation, and changes in their cytotoxic function. These responses will be supported by changes in cellular metabolism. Therefore, changes in NK metabolic parameters could somehow predict changes in NK cell function and cytotoxicity. In this chapter, we describe a protocol to measure NK cell metabolism in primary human NK cells by using an extracellular flux analyzer. This machine measures pH and oxygen changes in the medium and allows the study of NK cell glycolysis and mitochondrial respiration in real time with a small number of cells.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
O’Brien KL, Finlay DK (2019) Immunometabolism and natural killer cell responses. Nat Rev Immunol 19(5):282–290. https://doi.org/10.1038/s41577-019-0139-2
Brand MD, Nicholls DG (2011) Assessing mitochondrial dysfunction in cells. Biochem J 435(2):297–312. https://doi.org/10.1042/BJ20110162
Divakaruni AS, Brand MD (2011) The regulation and physiology of mitochondrial proton leak. Physiology 26(3):192–205. https://doi.org/10.1152/physiol.00046.2010
Stram AR, Payne RM (2016) Post-translational modifications in mitochondria: protein signaling in the powerhouse. Cell Mol Life Sci 73(21):4063–4073. https://doi.org/10.1007/s00018-016-2280-4
Mookerjee SA, Brand MD (2015) Measurement and analysis of extracellular acid production to determine glycolytic rate. J Vis Exp (106):e53464. https://doi.org/10.3791/53464
Mookerjee SA, Gerencser AA, Nicholls DG, Brand MD (2017) Quantifying intracellular rates of glycolytic and oxidative ATP production and consumption using extracellular flux measurements. J Biol Chem 292(17):7189–7207. https://doi.org/10.1074/jbc.M116.774471
Mookerjee SA, Nicholls DG, Brand MD (2016) Determining maximum glycolytic capacity using extracellular flux measurements. PLoS One 11(3):e0152016. https://doi.org/10.1371/journal.pone.0152016
Mookerjee SA, Goncalves RLS, Gerencser AA, Nicholls DG, Brand MD (2015) The contributions of respiration and glycolysis to extracellular acid production. Biochim Biophys Acta 1847(2):171–181. https://doi.org/10.1016/j.bbabio.2014.10.005
Tognarelli S, Jacobs B, Staiger N, Ullrich E (2016) Flow cytometry-based assay for the monitoring of NK cell functions. J Vis Exp (116). https://doi.org/10.3791/54615
Theorell J, Bryceson YT (2016) Analysis of intracellular Ca(2+) mobilization in human NK cell subsets by flow cytometry. Methods Mol Biol 1441:117–130. https://doi.org/10.1007/978-1-4939-3684-7_10
Traba J, Waldmann TA, Anton OM (2020) Analysis of human natural killer cell metabolism. J Vis Exp (160). https://doi.org/10.3791/61466
Traba J, Miozzo P, Akkaya B, Pierce SK, Akkaya M (2016) An optimized protocol to analyze glycolysis and mitochondrial respiration in lymphocytes. J Vis Exp (117). https://doi.org/10.3791/54918
Acknowledgements
We thank Michael N. Sack (National Heart, Lung and Blood Institute, NIH) and Thomas A. Waldmann (National Cancer Institute, NIH) for continuous support and discussion. This study was supported by grants RYC2018-026050-I and PID2019-105665RA-I00 of MICINN (Spain) to J.T. This study was also supported by the Intramural Research Programs of the National Institutes of Health, National Cancer Institute, and National Heart, Lung and Blood Institute.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2022 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Traba, J., Antón, O.M. (2022). Assessing Changes in Human Natural Killer Cell Metabolism Using the Seahorse Extracellular Flux Analyzer. In: Shimasaki, N. (eds) Natural Killer (NK) Cells. Methods in Molecular Biology, vol 2463. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2160-8_12
Download citation
DOI: https://doi.org/10.1007/978-1-0716-2160-8_12
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-2159-2
Online ISBN: 978-1-0716-2160-8
eBook Packages: Springer Protocols