Interrogating in vivo T-cell metabolism in mice using stable isotope labeling metabolomics and rapid cell sorting.

Autor: Sheldon RD; Department of Metabolism and Nutritional Programming, Van Andel Institute, Grand Rapids, MI, USA.; Metabolomics and Bioenergetics Core Facility, Van Andel Institute, Grand Rapids, MI, USA., Ma EH; Department of Metabolism and Nutritional Programming, Van Andel Institute, Grand Rapids, MI, USA., DeCamp LM; Department of Metabolism and Nutritional Programming, Van Andel Institute, Grand Rapids, MI, USA., Williams KS; Department of Metabolism and Nutritional Programming, Van Andel Institute, Grand Rapids, MI, USA., Jones RG; Department of Metabolism and Nutritional Programming, Van Andel Institute, Grand Rapids, MI, USA. russell.jones@vai.org.
Jazyk: angličtina
Zdroj: Nature protocols [Nat Protoc] 2021 Sep; Vol. 16 (9), pp. 4494-4521. Date of Electronic Publication: 2021 Aug 04.
DOI: 10.1038/s41596-021-00586-2
Abstrakt: T cells are integral players in the adaptive immune system that readily adapt their metabolism to meet their energetic and biosynthetic needs. A major hurdle to understand physiologic T-cell metabolism has been the differences between in vitro cell culture conditions and the complex in vivo milieu. To address this, we have developed a protocol that merges traditional immunology infection models with whole-body metabolite infusion and mass-spectrometry-based metabolomic profiling to assess T-cell metabolism in vivo. In this protocol, pathogen-infected mice are infused via the tail vein with an isotopically labeled metabolite (2-6 h), followed by rapid magnetic bead isolation to purify T-cell populations (<1 h) and then stable isotope labeling analysis conducted by mass spectrometry (~1-2 d). This procedure enables researchers to evaluate metabolic substrate utilization into central carbon metabolic pathways (i.e., glycolysis and the tricarboxylic acid cycle) by specific T-cell subpopulations in the context of physiological immune responses in vivo.
(© 2021. The Author(s), under exclusive licence to Springer Nature Limited.)
Databáze: MEDLINE
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