Metabolomic profiling of macrophages determines the discrete metabolomic signature and metabolomic interactome triggered by polarising immune stimuli

Abstract

Priming and activating immune stimuli have profound effects on macrophages, however, studies generally evaluate stimuli in isolation rather than in combination. In this study we have investigated the effects of pro-inflammatory and anti-inflammatory stimuli either alone or in combination on macrophage metabolism. These stimuli include host factors such as IFNγ and ovalbumin-immunoglobulin immune complexes, or pathogen factors such as LPS. Untargeted LC-MS based metabolomics provided an in-depth profile of the macrophage metabolome, and revealed specific changes in metabolite abundance upon either individual stimuli or combined stimuli. Here, by factoring in an interaction term in the linear model, we define the metabolome interactome. This approach allowed us to determine whether stimuli interact in a synergistic or antagonistic manner. In conclusion this study demonstrates a robust approach to interrogate immune-metabolism, especially systems that model host-pathogen interactions.

Metabolic characterisation of THP-1 macrophage polarisation using LC–MS-based metabolite profiling

“…Our data demonstrated a significant increase in the purines; hypoxanthine and inosine (Supplementary 3). In agreement with our results, hypoxanthine was reported to be increased in the presence of LPS (Rattigan et al 2018). Pyrimidines and purines can be used for the activated cell biosynthesis and provide NADPH which is the substrate for NADPH oxidase in order to produce reactive oxygen species (Bedard and Krause 2007) that act as a bacterial killing mechanism (West et al 2011) representing the main function of M1 macrophages.…”

“…Therefore, metabolic responses to different stimuli can be sufficiently different, contradictory and seriously complex. This in turn suggests that the studies with single or limited number of stimuli do not reflect the complexity of the picture in vivo (Rattigan et al 2018). Secondly, and most importantly, most of the previous studies discussed the metabolic signature of M1 and M2 in mice macrophages, either primary or cell line stimulated with in vitro stimuli whereas our experiment was conducted using human cell line (THP-1 cells).…”

“…GM-CSF and M-CSF can give rise to pro-inflammatory and anti-inflammatory macrophages, respectively, and so it is expected that they impact the metabolic profile of macrophage phenotypes (Akagawa 2002;Rattigan et al 2018). It was already reported that using LPS alone, IFN-γ alone or both of them together gives rise to different metabolic response of macrophages (Rattigan et al 2018). Therefore, metabolic responses to different stimuli can be sufficiently different, contradictory and seriously complex.…”

“…Using LC-MS metabolomics, Mills et al have shown that itaconate is an anti-inflammatory metabolites which has important role in regulation of macrophage functions (Mills et al 2018). However, Rattigan et al demonstrated a complex role of itaconate as an inflammatory and anti-inflammatory key metabolite (Rattigan et al 2018). Moreover, another metabolomics study of macrophages has identified succinate as a participant in innate immune signalling in response to LPS (M1 macrophages) where its increase results from impaired TCA cycle and leads to boost the production of the inflammatory cytokine, IL-1β during inflammation (Tannahill et al 2013b).…”

Metabolic reprograming of LPS-stimulated human lung macrophages involves tryptophan metabolism and the aspartate-arginosuccinate shunt

“…The methodological approach combined high-resolution LC-MS, with a split scan-range acquisition method providing improved feature detection when compared to classical methods [23], and GC-MS to provide the most extensive metabolome coverage. Metabolomics studies were previously performed with macrophages to assess the effects of drugs, plasticizers, pollutants and nanomaterials in mouse macrophage cell lines [44][45][46][47], to study the interactome in IFN-γ or/and LPS-primed murine macrophages [48] or the cellular metabolism in HIV-infected human monocyte-derived macrophages [49]. Since the availability of human primary lung macrophages is limited and prolonged culture cannot be readily performed, alternative cellular models are commonly used as surrogate to study macrophage biology.…”