Primary Human M2 Macrophage Subtypes Are Distinguishable by Aqueous Metabolite Profiles

The complexity of macrophage (MΦ) plasticity and polarization states, which include classically activated pro-inflammatory (M1) and alternatively activated anti-inflammatory (M2) MΦ phenotypes, is becoming increasingly appreciated. Within the M2 MΦ polarization state, M2a, M2b, M2c, and M2d MΦ subcategories have been defined based on their expression of specific cell surface receptors, secreted cytokines, and specialized immune effector functions. The importance of immunometabolic networks in mediating the function and regulation of MΦ immune responses is also being increasingly recognized, although the exact mechanisms and extent of metabolic modulation of MΦ subtype phenotypes and functions remain incompletely understood. In this study, proton (¹H) nuclear magnetic resonance (NMR) metabolomics was employed to determine the polar metabolomes of M2 MΦ subtypes and to investigate the relationship between aqueous metabolite profiles and M2 MΦ functional phenotypes. Results from this study demonstrate that M2a MΦs are most distinct from M2b, M2c, and M2d MΦ subtypes, and that M2b MΦs display several metabolic traits associated with an M1-like MΦ phenotype. The significance of metabolome differences for metabolites implicated in glycolysis, the tricarboxylic acid (TCA) cycle, phospholipid metabolism, and creatine-phosphocreatine cycling is discussed. Altogether, this study provides biochemical insights into the role of metabolism in mediating the specialized effector functions of distinct M2 MΦ subtypes and supports the concept of a continuum of macrophage activation states rather than two well-separated and functionally distinct M1/M2 MΦ classes, as originally proposed within a classical M1/M2 MΦ framework.