Metabolic reprogramming in immune response and tissue inflammation

Abstract

Innate and adaptive immunity participate in and regulate numerous human diseases. Increasing evidence implies that metabolic reprogramming mediates immune cell functional change during immune responses. In this review, we present and discuss our current understanding of metabolic regulation in different immune cells and their subsets in response to pathological stimuli. An interactive biochemical and molecular model was established to characterize metabolic reprogramming and their functional implication in anti-inflammatory, immune resolution and pro-inflammatory responses. We summarize two major features of metabolic reprogramming in inflammatory stages in innate and adaptive immune cells, 1) energy production and biosynthesis reprogramming, including increased glycolysis and decreased oxidative phosphorylation, in order to secure faster ATP production and biosynthesis for defense response and damage repair, and 2) epigenetic reprogramming, including enhanced histone acetylation and suppressed DNA methylation, due to altered accessibility of acetyl/methyl group donor and metabolite-modulated enzymatic activity. Finally, we discuss current strategies of metabolic and epigenetic therapy in cardiovascular disease and recommend cell-specific metabolic and gene-targeted site-specific epigenetic alterations for future therapies.

Graphical Abstract

Introduction

Atherosclerosis is characterized by chronic inflammation and increased infiltration of innate and adaptive immune cells in the arterial wall¹. It is well-established that immune cells undergo phenotypical and functional changes in response to various stimuli and micro-environmental changes in cardiovascular diseases^2–5. We have published updated molecular processes in innate and adaptive immune response and immune cell subsets differentiation recently^5–7. Of note, emerging evidence has shown that alteration of cellular metabolic processes can fuel and regulate functional changes in immune cells, ranging from proliferation, differentiation, to effector function, by regulating the balance between energy generation and utilization and macromolecules breaking down and biosynthesis, redox hemostasis, epigenetic landscape, and signal transduction^8–10. Therefore, unraveling the role and underling mechanisms of metabolic reprogramming during an immune response and tissue inflammation holds the promise of novel therapeutic strategies for atherosclerotic cardiovascular diseases and other chronic inflammatory diseases.

In Figure 1, we depict the general interplay between immune response and metabolic reprogramming. Innate immunity is characterized by activation of innate immune cells by pathogen/danger-associated molecular pattern or metabolite-associated danger signal via pattern recognition receptors or metabolic sensor to facilitate the downstream signaling cascade to initiate an immune response. In contrast, adaptive immunity is composed of cell-mediated and humoral-mediated immunity which are both antigen-specific processes of activating T and B cells involving in several signals (signal 1 antigen recognition, signal 2 immune checkpoint, signal 3 cytokine stimulation, signal 4 metabolite-associated danger signal recognition)⁶. Metabolic reprogramming happens during all these immune responses, in which various cellular metabolic processes in immune cells are altered to achieve an adequate function. Moreover, this process is so dynamic and intricate that 2749 metabolic pathways in 3067 different organisms (477 in human), as identified from the MetaCyc database (https://metacyc.org, Ver. 23.1), may be involved and finally lead to a metabolic optimization in which distinct metabolic signature determines specific inflammatory or anti-inflammatory immune cell subset differentiation.

Fig. 1. Metabolic reprogramming in innate and adaptive immune responses.

Innate immunity is characterized by activation of innate immune cells by pathogen/danger associated-molecular pattern (PAMP/DAMP) or metabolite-associated danger signal (MADS) via pattern recognition receptors (PRR) or metabolic sensor (MS) to facilitate the downstream signaling cascade to initiate an immune response. In contrast, adaptive immunity is composed of cell-mediated and humoral-mediated immunity, which are both antigen-specific processes of activating T and B cells involving in several signals (signal 1 antigen recognition, signal 2 immune checkpoint, signal 3 cytokine stimulation, signal 4 MADS recognition). Metabolic reprogramming participates in both innate and adaptive immune response to regulate immune cell functions, which includes up to 2749 metabolic pathways from 3067 different organisms and 477 human metabolic pathways as identified from the MetaCyc database (https://metacyc.org Ver. 23.1). Metabolic reprogramming may be a dynamic process in which glucose, fatty acid and amino acid metabolism are the most described ones, and finally lead to a metabolic optimization in which distinct metabolic signature determines specific inflammatory or anti-inflammatory immune cell subset differentiation. APC indicates antigen-presenting cell; PAMP, pathogen-associated molecular pattern; DAMP, danger-associated molecular pattern; MADS, metabolites-associated danger signal; MS, metabolic sensor; PRR, pattern recognition receptor; *, MetaCyc database (https://metacyc.org, Ver. 23.1),

In this review, we will summarize an updated understanding of metabolic reprogramming in different immune cells and their subsets during different inflammatory stages and the process of tissue inflammation and in particular explore the potential therapeutic targets for chronic inflammatory diseases, with a focus on cardiovascular disease.

Metabolic reprogramming in innate and adaptive immune cells

In this section, we will elucidate the contribution of metabolic reprogramming in several immune cells and their subsets, including monocyte, macrophage, dendritic cell (DC), T cell, and B cell, to tissue inflammation (Table 1).

Table 1.

Metabolic reprogramming in immune cells

Cell type: subset	Stimuli (status)	Metabolic change	Functional change	Signal molecules	PMID
Monocyte:
CD14+	LPS	↑Glycolysis, ↓OXPHOS	↑Inflammatory cytokines (TNF-a. IL6, IL1β, IL10), ↑Phagocytosis	TLR4	27991883
	P3C	↑Glycolysis, ↑OXPHOS		TLR2	27991883
	Fungi Candida	↑Glycolysis, ↑OXPHOS, ↑Glutaminolysis	↑Inflammatory cytokine (TNFα. IL6, IL1β), ↑ROS	C-type lectin	28922415
CD14++CD16−	NA	↑Glycolysis, ↑PPP	Defense response	NA	24671955
CD14+CD16++	NA	↑OXPHOS	Anti-inflammatory	NA	24671955
CD14+CD40+	Hcy (CKD)	↑HCY, ↑SAH, ↑SAM/SAH	↓DNA methylation, ↑Inflammatory cytokine, chemokines	↓DNMT1-↑CD40	27992360
Macrophage:
M1	LPS/IFNγ	↑Glycolysis, ↑PPP	Pro-inflammatory	AKT/mTOR/HIF1α	29777212
M2	IL4	↑FAO, ↑OXPHOS↓PPP	Anti-inflammatory	STAT6-AMPK	29777212
MOX	OxPL	↓Glycolysis, ↑Glutaminolysis	Anti-oxidant activity, anti-inflammatory	TLR2-Syk	29891687
Dendritic cell:
BM-derived DC	No stimuli	↑lipid β-oxidation/OXPHOS	Antagonize DC activation	AMPK	20351312
BM-derived DC	LPS/Zymosan/curdlan	↑Glycolysis, ↓OXPHOS	↑Maturation, motility,migration	PI3K/AKT	20351312
BM-derived DC	CCL21/19	↑Glycolysis	↑Migration, trafficking to draining lymph node	CCR7-HIF1α	30824325
CD1c+ mDC	TLR agonists	↑Glycolysis, ↑Mitophagy ↓OXPHOS	Activation	TLR7/8-BNIP3	30455688
pDC	TLR agonists	↑OXPHOS,↑Glutaminolysis	Activation	TLR7/8-BNIP3	30455688
Tolerogenic DC	Dexamethason & vitamin D3	↑OXPHOS,↑FAO	Tolerogenic, ↑ROS	NA	25917094
T cell:
Naïve	NA	OXPHOS, FAO	Homestasis	TSC1-mTOR1	29677474
Th1	NA	↑Glycolysis	↑Inflammatory cytokine(IFNƔ)	LDHA, histone acetylation of Ifng	27708054
Th17	NA	↑Glycolysis	Pro-inflammatory	HIF1	21871655
Treg	NA	↑OXPHOS	Anti-inflammatory	Myc	28416194
Memory CD8+ T cell	NA	↑OXPHOS, FAO	Longevity, quick response	NA	22889213
B cell:
Mouse splenic B cell	BCR engagement	↑Glycolysis	↑Proliferation/growth	PI3K/Akt/mTOR	16449529
	IL-4	↑Glycolysis	↑Survival	STAT6	17911579
Peripheral blood B cell	anti-IgM/LPS	↑Glycolysis, ↑OXPHOS	↑Proliferation and antibody production	HIF1α,cMyc	24616478
Memory B cell	NA	↑OXPHOS	Durable antibody production	Mpc2	27396958

Metabolic changes in different immune cells and subsets in response to different stimuli obtained through literature searching. Immune cell functional changes associated with metabolic reprogramming and related signal molecules are presented. LPS indicates lipopolysaccharides; Hcy, homocysteine; OXPHOS, oxidative phosphorylation; TLR, toll-like receptor; BCR, B cell receptor; DC, dendritic cell; PPP, pentose phosphate pathway; OxPL, oxidized phospholipids; CKD, chronic kidney disease; BM, bone-marrow; FAO, fatty acid oxidation.

Monocyte — Monocytes are innate immune cells that monitor environmental changes and replenish the pool of tissue macrophages and dendritic cells⁷. In response to lipopolysaccharides (LPS), human CD14⁺ monocytes favor glycolysis over oxidative phosphorylation (OXPHOS) to enable a quick host defense mechanism by inflammatory cytokine (TNFα. IL6, IL1β) production and phagocytosis¹¹, which is similar to the Warburg effect observed in cancer cells^{12, 13}. In glucose deprivation condition, monocytes could perform pro-inflammatory functions by increasing fatty acid oxidation (FAO), fueling OXPHOS compensating for the Warburg effect to meet the increased energy demand during LPS stimulation¹⁴. However, unlike LPS, synthetic bacterial lipopeptide P3C or Candida albicans enhanced both glycolysis and OXPHOS with increased pro-inflammatory cytokine production and reactive oxygen species (ROS) generation through toll-like receptor 2 or C-type lectin-signaling pathways¹⁵. These findings suggested that although increased glycolysis is one of the hall marks of an activated monocyte, it may be accompanied by or compensated with oxidative processes.

It is well accepted that monocytes are highly heterogenic. At least three monocyte subsets are defined in mice (CD11b⁺Ly6C^high, CD11b⁺Ly6C^middle and CD11b⁺Ly6C^low) and human (classical CD14⁺⁺CD16⁻, intermediate CD14⁺⁺CD16⁺, and non-classical CD14⁺CD16⁺⁺)⁷. Transcription profiling confirmed distinct functions of the human monocyte subsets and demonstrated that they had distinct metabolic signatures¹⁶. Human CD14⁺⁺CD16⁻ classical monocytes were associated with defense response and expressed higher levels of genes involved in glycolysis and pentose phosphate pathway (PPP), such as hexokinase 2, glyceraldehyde 3-phosphate dehydrogenase, phosphofructokinase, lactate dehydrogenase B and glucose 6-phosphate dehydrogenase, priming them for anaerobic energy production¹⁶. In contrast, CD14⁺CD16⁺⁺ patrolling non-classical monocytes was associated with increased expression of protein metabolism genes and OXPHOS genes encoding components of complex I, II, III, and V of the mitochondrial respiratory chain¹⁶. CD40⁺ monocyte is a new monocyte subset we recently discovered, which had a stronger inflammatory feature than CD14⁺CD16⁺ inflammatory intermediate monocytes and was an effective biomarker for chronic kidney disease severity¹⁷. CD40⁺ monocytes from chronic kidney disease patients or derived by homocysteine treated splenocytes exhibited increased intracellular levels of homocysteine and S-adenosylhomocysteine (SAH), reduced S-adenosylmethionine (SAM)/SAH ratio, and suppressed DNA methylation on NF-kB consensus element in CD40 core promoter which is associated with DNA methyltransferase 1 inhibition¹⁷.

Macrophage — Macrophages play a central role in atherogenesis through their diverse roles in cholesterol metabolism and immune response¹⁸. Animal and human studies indicate that both the quantity of macrophage and the balance of inflammatory and anti-inflammatory subsets influence plaque fate^{3, 19}. LPS/interferon gamma (IFNγ)-induced M1 macrophages play an important role in host defense and tissue inflammation by inflammatory cytokine secretion, ROS generation, and Th1 response amplification^{20, 21}. M1 macrophage has enhanced glycolysis and PPP coincident with impaired OXPHOS and reduced ATP production in mitochondria via a broken tricarboxylic acid (TCA) cycle, which is controlled by Akt/mTOR/ HIF1α²². The broken TCA cycle in M1 macrophages leads to the accumulation of intermediates succinate and citrate, which are involved in the formation and elimination of ROS, epigenetic regulation and signal transduction^{22, 23}. However, IL4-induced M2 macrophages rely on FAO and OXPHOS for their survival and anti-inflammatory role in immune resolution²⁴. M2 macrophage polarization and the increased mitochondria oxidative are associated with signal transducer and activator of transcription 6 signaling²². In response to oxidized phospholipids, macrophage could differentiate to Mox and exert anti-inflammatory and anti-oxidant activities²⁵. Recent studies found that compared with M1 and M2, Mox macrophages had a different metabolic profile, which was characterized by suppressed aerobic glycolysis and increased glutathione production, via toll-like receptor 2-Syk driven mechanism²⁶.

Dendritic cell — Evidence from mice and human proves that DC, a professional antigen-presenting cell, contribute to atherosclerotic vascular disease by various pathogenic and protective mechanisms, such as early foam cell formation, lipid metabolism regulation, and adaptive immunity stimulation⁴. Recent studies suggest that metabolic reprogramming accompanies DC development, activation and differentiation^{27, 28}. Resting DC predominantly uses lipid β-oxidation and OXPHOS for energy generation²⁷. AMP-activated protein kinase is a central regulator of these metabolisms²⁷. Upon activation by different toll-like receptor agonists, DC metabolism would switch from mitochondrial oxidation to glycolytic metabolism to support their motility, migration and pro-inflammatory phenotypes which are associated with increased HIF1α activation and AKT/TBK/mTOR signaling pathways^28–30. However, although DCs displayed increased mitochondrial OXPHOS to maintain ATP production when culturing DCs without glucose or with 2-deoxy-D-glucose (glycolytic inhibitor), this metabolic compensation is not sufficient to support DC shape and motility³¹.

Several DC subsets have been identified, including conventional/myeloid DC, plasmacytoid DC, regulatory DC, and monocyte-derived DC^{32, 33}. It was found that CD1c⁺ myeloid DCs increased glycolysis and mitophagy in response to a toll-like receptor agonist, whereas plasmacytoid DCs relied on glutaminolysis and OXPHOS for energy production³⁴. Cellular metabolism also determines tolerogenic DC differentiation, which promotes immune tolerance through the secretion of immunosuppressive cytokines IL-10 and regulation of Treg polarization³⁵. Compared to activate immunogenic DC, tolerogenic DC displayed higher mitochondrial OXPHOS activity and more abundant electron transport chain complexes with increased ROS production⁴.

T cell — It is appreciated that metabolic reprogramming regulates lymphocyte differentiation, activation and effector functions^{36, 37}. Peripheral naïve T cells use OXPHOS as a major source of ATP production to maintain weak T cell receptor interactions with self-peptides presented on MHC molecules for survival and homeostatic proliferation^21,37. T cells are activated when encountering antigens on an antigen-presenting cell along with costimulatory signals and cytokine stimulation⁶. Recent studies suggested that anaerobic glycolysis is an important metabolic hallmark of activated effector T cell even though sufficient oxygen is present to support glucose catabolism via the TCA cycle and OXPHOS³⁸. In activated Th1 cells, lactate dehydrogenase A was induced to support aerobic glycolysis and promote expression of IFNγ via maintaining high levels of acetyl-CoA to enhance histone acetylation and transcription of Ifnγ³⁹. T cell-specific ablation of Ldha could protect mice from immunopathology triggered by excessive IFNγ expression³⁹. Another study pointed out that glycolysis is specifically required for the effector function of activated T cells, such as pro-inflammatory cytokine IFNγ production. However, OXPHOS, but not glycolysis, was essential for the proliferation and survival of these cells⁴⁰. By contrast, Treg had suppressed glycolysis and enhanced OXPHOS, which might allow a metabolic advantage in low glucose, lactate rich environment by resisting lactate mediated suppression of T cell function and proliferation^{41, 42}.

B cell — Increased glycolysis after activation, like in activated T cell, was also found in B cells following B cell receptor engagement or cytokine stimulation^{43, 44}. In these cases, glycolysis supports B cell growth through PI3K signaling or protect B cell from apoptosis via signal transducer and activator of transcription 6 signaling^{43, 44}. A recent study examining the metabolic reprogramming of purified B and T cells upon antigen challenging found that activated B cells increased both oxygen consumption rate as well as the rate of glycolysis reflected by the extracellular acidification rate, and maintained a balanced oxygen consumption rate/ extracellular acidification rate ratio⁴⁵. In contrast, activated T cells shifted to a more glycolytic metabolism⁴⁵. This study also suggested that increased glycolysis is essential for B cell proliferation and antibody production, whereas glycolysis inhibition by Glut1-depletion or dichloroacetate (a pyruvate dehydrogenase kinase inhibitor) sharply suppressed B cell proliferation and antibody secretion⁴⁵. We know that short-lived plasma cells would differentiate to long-lived memory B cells for durable antibody production. Metabolic studies revealed that long-lived memory B cells rely on increased mitochondrial respiratory capacity, which was attribute to increased respiratory substrates such as pyruvate, although they uptake more glucose than short-lived plasma cells⁴⁶. Because a large amount of the cellular glucose is used for antibody glycosylation, inhibition of pyruvate import by knocking down Mpc1 or Mpc2 (pyruvate transporter) would significantly reduce long-lived memory B cells numbers and antibody secreting^{46, 47}.

In general, in response to the stimulation of various danger signals, dynamic balance of pro- and anti-inflammatory metabolic reprograming determines the direction of immune cell responses. Anti-inflammatory immune cell subsets, such as CD14⁺CD16⁺⁺ monocyte subsets, M2 macrophage and Treg, favor oxidative metabolism (FAO and OXPHOS) for efficient energy production to maintain metabolic/cellular function homeostasis and achieve immune resolution or anti-inflammatory function (Figure 2A, left panel). However, in a pro-inflammatory immune response, most of the immune cells, including CD14⁺⁺CD16⁻ monocyte, M1 macrophage and Th1/Th2/Th17 cells, switch toward enhanced glycolysis and PPP activity to prompt an activation and proliferation status and adopt the pro-inflammatory effector functions (Figure 2A, right panel).

Fig. 2. Models of metabolic reprogramming in immune response.

A. Metabolic reprogramming in anti- and pro-inflammatory immune response. In response to the stimulation of various danger signals, dynamic balance of pro- and anti- inflammatory metabolic reprograming determines the direction of immune cell responses. In physiological condition or anti-inflammatory response (left panel), immune cells favor oxidative metabolism, including fatty acid oxidation (FAO) and mitochondrial oxidative phosphorylation (OXPHOS), for energy production and homeostasis of metabolic/cellular function. During pro-inflammatory response (right panel), immune cells switch to active glycolysis and biosynthesis (fatty acid and nucleotide synthesis) for quick energy production and regeneration, whereas, FAO and OXPHOS are suppressed. This metabolic reprogramming produces high levels of cytosolic ROS and mitochondrial ROS. Words and arrows in red indicate activate intermediates or process, whereas the blue color depicts the suppressed ones. B. Metabolic switch in innate and adaptive immune cells according to the inflammatory stage. Innate and adaptive immune cells undergo similar metabolic reprogramming in the early inflammatory stage but switch to different metabolic statuses during the memory stage. In general, upon activation, immune cells switch from OXPHOS to glycolysis for quick ATP production. In the memory stage, innate immune cell relies on glycolytic activity, but adaptive immune cell can switch back to more efficient mitochondrial OXPHOS for long-term survival. Biosynthetic and proliferative activities are increased in the activate stage and memory stage in innate immune cell, whereas may be reduced in memory stage in adaptive immune cell. Methylation status is suppressed in the active stage and may maintain as such in the memory stage of innate and adaptive immune cells. Acetylation-mediated epigenetic reprogramming is potentially upregulated due to increased acetyl CoA production and metabolite-modulated enzymatic regulation. Solid lines represent metabolic changes established. Dashed lines represent metabolic changes that remain to be confirmed. PAMP indicates pathogen-associated molecular pattern; DAMP, danger-associated molecular pattern; MADS, metabolite-associated danger signal; G6P, glucose 6-phosphate; R5P, ribose 5-phosphate; PPP, pentose phosphate pathway; NAD, nicotinamide adenine dinucleotide; NADPH, nicotinamide adenine dinucleotide phosphate; ROS, reactive oxygen species; mtROS, mitochondrial ROS; CH, cholesterol; TG, triacylglycerol; FA, fatty acid; FAO, fatty acid oxidation; FAS, fatty acid synthesis; α-KG, alpha-ketoglutarate; TCA cycle, citric acid cycle; OXPHOS, oxidative phosphorylation; SAM, S-adenosylmethionine; SAH, S-adenosylhomocysteine; Met, methione; Hcy, homocysteine.

Model of metabolic reprogramming in immune response — connection between metabolic change and inflammatory feature

In hemostasis situation (Figure 2A, left panel), immune cells most likely perform anti-inflammatory and immune resolution functions. Glucose enters the cells via glucose transporters (such as Glut1) and is catabolized to pyruvate along with ATP and nicotinamide adenine dinucleotide (NADH) production in the cytosol via glycolysis⁴⁸. Pyruvate is primarily transported to the mitochondria and decarboxylated to acetyl-CoA, which acts as fuel for the TCA cycle⁴⁹. Additionally, acetyl-CoA from increased FAO and alpha-ketoglutarate from increased glutaminolysis also replenishes the TCA cycle to generate the reducing equivalent NADH and Flavin adenine dinucleotide (FADH₂)⁵⁰. In further reactions, NADH and FADH₂ donate electrons to O₂ via the electron transport chain and results in an electron chemical gradient across the mitochondrial inner membrane to drive increased ATP synthesis in the OXPHOS process.

In contrast, in an activated situation (Figure 2A, left panel) immune cells exhibit pro-inflammatory functions. Glucose uptake and glycolytic rate are significantly increased partially by inducing the expression of the glucose transporter and multiple glycolytic enzymes in an inflammatory response^{51, 52}. Pyruvate is preferentially converted to lactate in the cytosol instead of fully oxidized in the mitochondrial, even in normal O₂ environments. Since glycolysis (2 ATP per molecule glucose) is a less efficient but more rapid way for ATP production than OXPHOS (30–32 ATP per molecule glucose), these pro-inflammatory cells rely on glycolysis to meet their demand for quick ATP generation and increased lipid, protein and nucleotide biosynthesis, which is critical for rapid cell proliferation and effector molecular production⁵³. PPP, a branch of glycolysis starting from glucose 6-phosphate, is also increased in pro-inflammatory immune cells and supports immune cell survival and proliferation by providing ribulose 5-phosphate for nucleotide synthesis and NADPH^{54, 55}, which is further used for fatty acid synthesis (FAS) and redox homeostasis. The PPP is a principal source of cytosolic ROS, which is a powerful antimicrobial weapon in innate immunity. However, excessive ROS production also results in oxidative stress, which may cause damage to lipids, proteins and DNA and contribute to various diseases, including atherosclerosis.

Mitochondria play a key regulatory role during an immune response to coordinate the production of energy and mitochondrial ROS, fatty acid biosynthesis, acetylation and methylation^{56, 57}. It has been shown that pro-inflammatory immune cells repurpose their mitochondria to increased ROS generation instead of ATP production, which can further shutdown mitochondrial energy production by inducing mitochondrial DNA mutations and mitochondrial respiratory chain damage⁵⁸. Besides, although mitochondrial oxidation, FAO and OXPHOS, is impaired in pro-inflammatory cells⁵⁹, the TCA cycle is not entirely shut down and it is reported that several intermediates, such as citrate, succinate and itaconate, in the TCA cycle are increased in the M1 macrophage due to the inactivating of enzymes catalyzing the corresponding reactions^{60, 61}. The accumulation of these metabolites either supply biosynthesis pathways to support immune cell proliferation or serve as signaling molecules affecting other metabolic and functional pathways. Moreover, the increased citrate can be catabolized to acetyl-CoA, which is an important precursor for both FAS and acetylation after exported from the mitochondria to the cytosol via citrate-carriers.

In summary, anti-inflammatory immune cells rely on OXPHOS for more prolonged survival and homeostasis. Whereas pro-inflammatory immune cells show increased glycolysis that is connected with fast ATP generation and activated PPP that is related to increased nucleotide synthesis and NADPH production. These metabolic changes support rapid cell proliferation fulfilling the demand for defense response and damage repair. In addition, cytosolic and mitochondrial ROS are increased in pro-inflammatory immune cells due to NADPH production and remodeled mitochondrial respiratory chain. Although mitochondrial oxidation (FAO and OXPHOS) are impaired, TCA cycle intermediates are increased to support immune cell proliferation or serve as signaling molecules affecting other metabolic and functional pathways in pro-inflammatory immune cells.

Metabolic reprogramming in immune memory cells

After activation, most of the effector T cell and plasma B cell are eliminated under apoptosis, and only a small number of them become memory cells which stay in a resting state and respond immediately to re-challenge from the same antigen^{62, 63}. Current knowledge supports the view that memory T and B cells are more resistant to apoptosis than the activated cells and switch back to a catabolic metabolism breaking down nutrients to fuel TCA cycle and efficiently generate ATP via OXPHOS for long-term survival^{47, 64}.

In the past few years, emerging evidence has suggested that innate immune cells also display immunological memory function by the observation that stimulation of innate immune cells with certain pathogens or stimuli enables them an augmented response to a re-stimulation^{65, 66, 67}. Compared to adaptive immune memory, the process of trained immunity is a less specific and short-lived phenomenon (last from 1 week to several months)⁶⁷. Importantly, atherogenic stimuli, like oxidized low-density lipoprotein, has been shown to induce trained immunity in innate immune cells^{68, 69}, and the prolonged activated phenotype of trained innate immune cells was proposed as a novel mechanism linking non-resolving inflammation and atherosclerosis⁷⁰.

Similar to immune cell activation, immune cells rely on elevated glycolysis for trained immunity as well⁷¹, and Akt-mTOR-HIF-1α pathway was responsible for the metabolic shift from OXPHOS to glycolysis. Increased glutaminolysis⁷² and mevalonate pathway^{72, 73} are also critical for trained immunity. Subsequently, the cellular metabolic pathway changes can in part regulate gene expression and contribute to a trained phenotype by modulating epigenetic landscapes, especially at the level of histone methylation and acetylation^{72, 74, 75}.

Metabolic switch according to inflammatory stage

Innate and adaptive inflammatory responses are described as three stages, quiescent/naive, activation and trained immunity/immune memory (Figure 2B). Collectively, innate and adaptive immune cells undergo a metabolic switch between glycolysis and OXPHOS and change their biosynthesis status. Upon activation, glycolysis and macromolecular biosynthesis are increased, whereas OXPHOS is impaired for rapid cell proliferation in both innate and adaptive immune cells. However, switching from OXPHOS to glycolysis does not always happen in an immune response. For example, B cells displayed a balanced increase in both lactate production (glycolysis) and oxygen consumption rate (OXPHOS) following LPS or B cell receptor stimulation⁴⁵. During the memory stage, innate immune cells still rely on active glycolysis, whereas the adaptive immune cells switch back to more efficient mitochondrial OXPHOS for long-term survival. Moreover, biosynthetic pathways return to a less active status in adaptive memory immune cells, which is likely to be induced and associated with the induction of trained immunity in innate immune cells⁷².

Connection between epigenetic reprogramming, metabolic reprogramming and inflammatory stage

Epigenetic reprogramming, predominantly methylation and acetylation, is crucial for immune cell development and differentiation by regulating gene transcription through altering DNA accessibility and chromatin structure.

Metabolic reprogramming mostly impacts epigenetic modifications by modulating the production and availability of methyl group donor SAM and acetyl group donor acetyl-CoA. SAM is generated in the homocysteine-methionine cycle and is converted to SAH, a competitive methyltransferase inhibitor, after the donation of its methyl group^76,77 (Fig. 2A). Through database mining, we have established that homocysteine-methionine cycle is a metabolic sensor system, and that its suppression determines histone hypomethylation, most frequently on H3K4 and H3K9, in autoimmune and cardiovascular diseases⁷⁶. In chronic kidney disease patients, suppressed homocysteine-methionine cycle-mediated homocysteine and SAH elevation mediates inflammatory CD40⁺ monocyte differentiation and is associated with DNA hypomethylation on NF-kB consensus element in CD40 core promoter¹⁷. Similarly, DNA hypomethylation is also coupled to B cell activation, proliferation, and memory cell differentiation⁷⁸. However, during effector CD8⁺ T cell differentiation in acute infection, DNA methylation is globally dynamic that hypermethylation is responsible for the repression of naïve T cell and hypomethylation is to drive differentiation and effector functions⁷⁹. Although the activation of CD4⁺ T cell has been shown to require the supply of methionine for the production of SAM to provide methyl group for DNA and histone methylation⁸⁰, genome-wide DNA methylation profiles revealed that most of the differentially methylated regions were hypermethylated in the naive phase, intermediately methylated in the effector phase, and hypomethylated in the memory phase during the process of T cell differentiation⁸¹.

Since acetyl-CoA cannot cross the mitochondria membrane, the acetyl-CoA accessible for histone modification is generated in the nucleo-cytoplasmic compartment from citrate and pyruvate (Fig. 2A). Citrate exported from the mitochondria utilizing fatty acid (FAO) and glucose (TCA cycle) has long been recognized as the predominant source for nuclear acetyl-CoA generation⁸². Moreover, it has been shown that all the subunits of the mitochondrial pyruvate dehydrogenase complex that convert pyruvate to acetyl-CoA are present and functional in the nucleus of mammalian cells and contribute to the de novo synthesis of acetyl-CoA and histones acetylation⁸³. Enhanced histone acetylation has been found during both innate and adaptive immune cell activation partially mediated by the increased availability of acetyl-CoA through increased glucose uptake and citrate accumulation^{84, 85}. For instance, increased glucose uptake and aerobic glycolysis are hallmarks of activated T cells. A recent study demonstrated that this metabolic change could promote Th1 activation via enhanced histone acetylation on H3K9³⁹.

In addition, several metabolites serve as activators or inhibitors for epigenetic enzymes and modulate epigenetic modifications. For instance, alpha-ketoglutarate is a cofactor for Jumonji C-terminal domain containing lysine demethylases. Increased alpha-ketoglutarate via glutaminolysis has been shown to support M2 macrophage activation through the Jmjd3-dependent demethylation of H3K27⁸⁶. However, fumarate and succinate are competitive antagonists of this reaction⁸⁷. The class III histone deacetylases, Sirtuin, are linked to decreased glycolysis by suppressing the expression of multiple glycolytic genes via deacetylation of H3K9 and require NAD⁺ as a cofactor to support their deacetylation activities⁸⁸. It was demonstrated that NAD⁺ levels were decreased in aging and metabolic diseases, including obesity, diabetes and cardiovascular diseases⁸⁹.

Taken together, metabolic reprogramming in immune cells modulate epigenetic reprogramming in response to inflammatory stimulation by three major mechanisms: 1) Increased glucose uptake and glycolysis lead enhanced histone acetylation by elevating the accessibility of acetyl group donor acetyl CoA, 2) impaired homocysteine-methionine cycle, a metabolic sensor system, results in suppressed SAM/SAH-dependent DNA/histone hypomethylation due to decreased methyl group donor SAM, increased methyltransferases inhibitor SAH and reduced SAM/SAH ratio, 3) accumulation of intermediate metabolites from altered TCA cycle regulate the activity of epigenetic enzymes, including methyltransferases, demethylases, acetyltransferase, and deacetylases. Current evidence supports a model that methylation is suppressed due to impaired homocysteine-methionine cycle and metabolite-modulated methylation enzyme activity, and acetylation is elevated due to increased acetyl CoA and metabolite-modulated enzymatic regulation upon immune cell activation during innate and adaptive immune responses (Fig. 2B).

Metabolic reprogramming in cardiovascular disease

Inflammation has long been characterized as a key factor of atherosclerotic cardiovascular disease in which a wide range of infiltrated and residential immune cells are activated, including macrophages, monocytes, dendritic cells, T cells, and B cells. A recent study performing serum metabolic profiling of over 3000 patients with subclinical atherosclerosis in carotid and coronary arteries demonstrated a strong association between metabolites and incidence of atherosclerosis and coronary artery disease. Energy metabolism, TCA cycle and glycolysis were identified as central pathways associated with atherosclerosis⁹⁰.

Extensive evidence demonstrates that monocyte and macrophage are the major populations in atherosclerotic lesion in metabolic disorders^{7, 17, 19, 91, 92}. It has been shown that monocyte and macrophage from patients with coronary artery disease were hyper-inflammatory with increased production of IL-6, IL-1β and TNFα. These inflammatory features were associated with increased glucose uptake, glycolysis, ROS production and histone demethylation^93,94. The metabolic switch from FAO and OXPHOS to glycolysis was also found in the pro-inflammatory macrophages stimulated by oxidized low-density lipoprotein⁹⁵. Of note, a recent study found that lipoprotein(a) activated aortic endothelial cells by enhancing PFKFB3-mediated glycolysis and lead to increased trans-endothelial migration of monocyte⁹⁶. Inhibition of glycolysis could reverse this pro-inflammatory and pro-adhesive state of aortic endothelial cells⁹⁶. In summary, immunometabolism plays important roles in accelerating tissue inflammation and cardiovascular diseases.

Potential metabolic therapeutic strategies for cardiovascular disease

With the increasing understanding of the crosstalk between metabolic reprogramming and immune cell activation in cardiovascular diseases, new therapeutic targets by altering the intracellular metabolism of immune cells are emerging. Some metabolic-targeting drugs have already been used in the treatment of inflammatory and autoimmune diseases, such as metformin, dimethyl fumarate, methotrexate and rapamycin, by regulating glucose, fatty acid and nucleotide metabolism⁵⁵. For instance, metformin, an oral antidiabetic medicine, has been shown to display an anti-inflammatory role by inhibiting ROS generation through reverse electron transport at Complex I⁹⁷ and inducing FAO in an AMP-activated protein kinase-dependent manner⁹⁸.

As outlined above, pro- and anti-inflammatory immune cells have different metabolic signatures. Therefore, manipulating specific metabolic targets would be beneficial to control inflammation in cardiovascular disease. For example, limiting the rate of glycolysis by inhibiting glucose transporter and key glycolytic enzymes, such as Glut1, HK2, PKM2 and GAPDH, can shift cellular metabolism in pro-inflammatory immune cells and repolarize them to an anti-inflammatory phenotype. Moreover, the transcription factor HIF-1α has been known to be one of the master regulators of the metabolic shift from OXPHOS to glycolysis in hypoxic or normorxic cells in inflammation^{99, 100} and also associated with initiation and progression of human and mouse atherosclerotic plaques^100–102. Recently, a study showed that HIF-1α activation by LPS/IFNγ in inflammatory macrophages was accompanied by upregulation of miR-210 and downregulation of miR-388 which contributed to necrotic core formation in atherosclerosis by reducing OXPHOS and energy depletion^{100, 103}. This indicates that microRNA may also be a promising approach of metabolic therapy for diseases.

In addition, metabolism-mediated epigenetic reprogramming is another promising target for cardiovascular diseases. DNA methyltransferase inhibitors and histone deacetylase inhibitors have been studied in several clinical trials. However, these inhibitors modified gene expression in a non-specific manner. If we take the hypomethylation status described above and in Fig. 2B in activated immune cells into consideration, DNA methyltransferase inhibitor therapy may not be a logical therapeutic strategy. Future studies should identify cell-specific metabolic alteration and gene-targeted site-specific epigenetic therapies.

Conclusion

This review discussed current understanding of metabolic reprogramming during the innate and adaptive immune cell responses, and established models of metabolic interplay in anti- and pro-inflammatory responses. Further studies to explore molecular and biochemical basis governing the metabolic reprogramming in immune responses would lead to the identification of novel therapies for human disease.

Nonstandard Abbreviations and Acronyms

DC

Dendritic cell

FADH₂

Flavin adenine dinucleotide

FAO

Fatty acid oxidation

FAS

Fatty acid synthesis

IFNγ

Interferon gamma

LPS

Lipopolysaccharides

NADH

Nicotinamide adenine dinucleotide

OXPHOS

Oxidative phosphorylation

PPP

Pentose phosphate pathway

ROS

Reactive oxygen species

SAH

S-adenosylhomocysteine

SAM

S-adenosylmethionine

TCA Cycle

Tricarboxylic acid cycle