Isotope Tracing Untargeted Metabolomics Reveals Macrophage Polarization-State-Specific Metabolic Coordination across Intracellular Compartments

Patrycja Puchalska, Xiaojing Huang, Shannon E. Martin, Xianlin Han, Gary J. Patti, Peter A. Crawford

Research output: Contribution to journalArticle

11 Scopus citations


We apply stable isotope tracing, mass-spectrometry-based untargeted metabolomics, to reveal the biochemical space labeled by 13C-substrates in bone-marrow-derived macrophages. At the pathway level, classically (lipopolysaccharide [LPS]-polarized, M1)and alternatively (interleukin [IL]-4-polarized, M2)polarized macrophages were 13C-labeled with surprising concordance. Total pools of uridine diphosphate N-acetylglucosamine (UDP-GlcNAc), an intermediate in the hexosamine biosynthetic pathway, were equally abundant in LPS- and IL-4-polarized macrophages. Informatic scrutiny of 13C-isotopologues revealed that LPS-polarized macrophages leverage the pentose phosphate pathway to generate UDP-GlcNAc, whereas IL-4-polarized macrophages rely on intact glucose and mitochondrial metabolism of glucose carbon. Labeling from [13C]glucose is competed by unlabeled fatty acids and acetoacetate, underscoring the broad roles for substrate metabolism beyond energy conversion. Finally, the LPS-polarized macrophage metabolite itaconate is imported into IL-4-polarized macrophages, in which it reprograms [13C]glucose metabolism. Thus, use of fully unsupervised isotope tracing metabolomics in macrophages reveals polarization-state-specific metabolic pathway connectivity, substrate competition, and metabolite allocation among cellular compartments.

Original languageEnglish (US)
Pages (from-to)298-313
Number of pages16
StatePublished - Nov 30 2018



  • Components of the Immune System
  • Immunology
  • Metabolomics

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