Use of integrated metabolomics, transcriptomics, and signal protein profile to characterize the effector function and associated metabotype of polarized macrophage phenotypes

Abstract

MΦs display remarkable plasticity and the ability to activate diverse responses to a host of intracellular and external stimuli. Despite extensive characterization of M1 MΦs and a broad set of M2 MΦs, comprehensive characterization of functional phenotype and associated metabotype driving this diverse MΦ activation remains. Herein, an ex vivo model was utilized to produce 6 MΦ functional phenotypes. Isolated CD14+ PBMCs were differentiated into resting M0 MΦs, and then polarized into M1 (IFN‐γ/LPS), M2a (IL‐4/IL‐13), M2b (IC/LPS), M2c (IL‐10), and M2d (IL‐6/LIF) MΦs. The MΦs were profiled using a bioanalyte matrix of 4 cell surface markers, ∼50 secreted proteins, ∼800 expressed myeloid genes, and ∼450 identified metabolites relative to M0 MΦs. Signal protein and expressed gene profiles grouped the MΦs into inflammatory (M1 and M2b) and wound resolution (M2a, M2c, and M2d) phenotypes; however, each had a unique metabolic profile. While both M1 and M2b MΦs shared metabotype profiles consistent with an inflammatory signature; key differences were observed in the TCA cycle, FAO, and OXPHOS. Additionally, M2a, M2c, and M2d MΦs all profiled as tissue repair MΦs; however, metabotype differences were observed in multiple pathways including hexosamine, polyamine, and fatty acid metabolism. These metabolic and other key functional distinctions suggest phagocytic and proliferative functions for M2a MΦs, and angiogenesis and ECM assembly capabilities for M2b, M2c, and M2d MΦs. By integrating metabolomics into a systems analysis of MΦ phenotypes, we provide the most comprehensive map of MΦ diversity to date, along with the global metabolic shifts that correlate to MΦ functional plasticity in these phenotypes.

Quantitative Proteomics of Polarised Macrophages Derived from Induced Pluripotent Stem Cells

“…Principal component analysis (PCA) of the proteins identified by MS showed that M0 and M2 iPSDM are more closely related to each other compared to M1 iPSDM ( Figure 2 A). These findings are in line with published transcriptomics and nCounter gene expression analyses of polarised MDM [ 18 , 19 ]. In addition, it has been shown previously [ 20 ] that M-CSF alone polarises M Φ more to a M2-like phenotype.…”

Cyclic Attractors Are Critical for Macrophage Differentiation, Heterogeneity, and Plasticity

“…They express high levels of IL-10, which induces the phosphorylation of STAT3, thus negatively regulating the production of pro-inflammatory cytokines ( Hutchins et al, 2013 ; Ma et al, 2015 ; Wang et al, 2019 ). M2d macrophages are induced by the costimulation of the adenosine A2 receptor and TLR, expressing levels of IL-10 as well as IL-12, and characterized by presenting properties of tumor-associated macrophages that carry out angiogenesis and tumor progression ( Leibovich et al, 2002 ; Grinberg et al, 2009 ; Park et al, 2009 ; Colin et al, 2014 ; Arora et al, 2018 ; Wang et al, 2019 ; Anders et al, 2021 ). However, these are not the only possible expression patterns, as variations have been found in vitro and in vivo .…”

Advances in the Regulation of Macrophage Polarization by Mesenchymal Stem Cells and Implications for ALI/ARDS Treatment

“…Glucocorticoids, IL-10, and TGF-b are involved in the induction of M2c macrophages. Activated M2c macrophages negatively regulate the production of anti-inflammatory cytokines involved in tissue remodeling and matrix deposition (43,44,(54)(55)(56). M2d macrophages are synergistically induced by TLRs and adenosine A2a receptor agonists or by IL-6.…”