A matter of time: temporal structure and functional relevance of macrophage metabolic rewiring

Abstract

The response of macrophages to stimulation is a dynamic process which coordinates the orderly adoption and resolution of various immune functions. Accumulating work over the past decade has demonstrated that during the immune response macrophage metabolism is substantially rewired to support important cellular processes, including the production of bioactive molecules, intercellular communication, and the regulation of intracellular signaling and transcriptional programming. In particular, we discuss an important concept emerging from recent studies - metabolic rewiring during the immune response is temporally structured. We review the regulatory mechanisms that drive the dynamic remodeling of metabolism, and examine the functional implications of these metabolic dynamics.

Keywords: dynamics; immunometabolism; inflammatory response and resolution; macrophage; metabolic regulation.

Figure 1.. Macrophage metabolic rewiring supports the production and release of bioactive compounds.

In response to PAMP stimulation, macrophages reprogram their metabolism to support the production of bioactive and bactericidal compounds that are released into the extracellular space. Reprogramming of the TCA cycle leads to extracellular accumulation of itaconate. A shift in arginine metabolism flux to nitric oxide production leads to an increase of reactive nitrogen species (RNS). Reactive oxygen species (ROS) production also increases. Itaconate, RNS and ROS all play a direct bactericidal role either by inhibiting key metabolic enzymes in bacteria (itaconate, RNS, ROS) or due to broad oxidative damage of pathogen cell components (ROS, RNS). Other metabolically derived compounds are released and signal in an autocrine or paracrine fashion to shape the immune response. Lipid mediators, such as eicosanoids, are produced due to a reprogramming of lipid metabolism. Succinate can be released and accumulate in the extracellular space due to inhibition of the TCA cycle enzyme succinate dehydrogenase (SDH). Both eicosanoids and succinate can promote both inflammatory and anti-inflammatory functions depending on the context and target cell type. Activated macrophages increase tryptophan degradation leading to increased release of the tryptophan metabolite kynurenine and depletion of extracellular tryptophan, both of which promote an anti-inflammatory microenvironment. Additionally, macrophages and dying cells can release ATP, which acts to promote inflammatory functions by signaling to purinergic receptors. The extracellular ATP can be converted by the macrophage to its metabolite, adenosine, which generally promotes anti-inflammatory functions and contributes to resolution of the inflammatory response.

Figure 2.. Macrophage metabolism influences inflammasome activation.

Activation of the inflammasome allows for processing of the inflammatory cytokines, IL1β and IL18, and gasdermin D (GSDMD) to their mature and active forms. Inflammasome activation is influenced by changes in multiple metabolic pathways, many of which impinge on mitochondria. Release of mitochondrial DNA (mtDNA) or its oxidized form (ox-mtDNA) from mitochondria serves as an inflammasome activating signal and is regulated by multiple metabolic changes. In response to stimulation, upregulation of the mitochondrial metabolic enzyme, CMPK2, which participates in the synthesis of mitochondrial dCTP and dUTP, supports the production of mtDNA, while alteration in cholesterol and choline production influence mitochondrial membrane composition and mtDNA release. Reactive oxygen species (ROS) from both mitochondrial and cytosolic sources has also been implicated in the activation of the inflammasome. Mitochondrial ROS (mtROS) has been shown to accumulate in response to stimulation and is reported to originate either from complex III of the electron transport chain or from complex I via a phenomenon termed reverse electron transport (RET). In response to activation, there is also an accumulation of reactive nitrogen species (RNS) and the TCA cycle metabolite, itaconate, both of which have been shown to inhibit inflammasome activation.

Figure 3.. LPS induced rewiring in central carbon metabolism leads to a dynamic alteration in immunoregulatory metabolites.

During the early response (“Early”) to LPS (± IFNγ) stimulation, increased glycolysis and oxidative respiration, sustained flux through pyruvate dehydrogenase (PDH), and activation of ATP citrate lyase (ACLY) supports production of cytosolic acetyl-CoA. After some time (“Mid” stage), when reactive nitrogen species (RNS) start to accumulate, isocitrate dehydrogenase (IDH) is inhibited by nitrosation and RNS-independent transcriptional suppression. This promotes accumulation of the upstream metabolite citrate. At a similar time, increased immune responsive gene 1 (IRG1) expression causes an accumulation of itaconate, which inhibits succinate dehydrogenase (SDH), leading to a concurrent increase in succinate. This “Mid” stage is therefore characterized by accumulation of the immunomodulatory metabolites, itaconate, succinate, and citrate and corresponds with high levels of HIF-1α, TNF-α and IL6. As time continues (“Late” stage), RNS and ROS accumulate, and oxidative metabolism continues to shut down. This is driven by blockages in flux through PDH, oxoglutarate dehydrogenase (OGDH), and aconitase. Aconitase is inhibited by RNS disruption of its Fe-S cluster. PDH inhibition is mediated by multiple mechanisms targeting each of its subunits. OGDH inhibition is due to loss of catalytic lipoylation of its E2 subunit. All this leads to a decrease in itaconate, succinate, citrate, acetyl-CoA, and succinyl-CoA, which correlates with a decrease in HIF-1α, TNF-α and IL6. Inhibition of the electron transport chain also contributes to the sustained low respiration rate during the late response. The figure shows a timeline synthesized from multiple studies in recent literature, to the best of our knowledge. As there are variations among these studies, comparing exact timing across different studies can be challenging.