bims-imicid Biomed News
on Immunometabolism of infection, cancer and immune-mediated disease
Issue of 2022‒05‒15
twenty-two papers selected by
Dylan Ryan
University of Cambridge

  1. Biochim Biophys Acta Mol Basis Dis. 2022 May 06. pii: S0925-4439(22)00097-7. [Epub ahead of print]1868(9): 166427
      Macrophages undergo extensive metabolic rewiring upon activation which assist the cell in roles beyond energy production and synthesis of anabolic building blocks. So-called immunometabolites that accumulate upon immune activation can serve as co-factors for enzymes and can act as signaling molecules to modulate cellular processes. As such, the Krebs-cycle-associated metabolites succinate, itaconate and alpha-ketoglutarate (αKG) have emerged as key regulators of macrophage function. Here, we describe that 2-hydroxyglutarate (2HG), which is structurally similar to αKG and exists as two enantiomers, accumulates during later stages of LPS-induced inflammatory responses in mouse and human macrophages. D-2HG was the most abundant enantiomer in macrophages and its LPS-induced accumulation followed the induction of Hydroxyacid-Oxoacid Transhydrogenase (HOT). HOT interconverts αKG and gamma-hydroxybutyrate into D-2HG and succinic semialdehyde, and we here identified this enzyme as being immune-responsive and regulated during the course of macrophage activation. The buildup of D-2HG may be further explained by reduced expression of D-2HG Dehydrogenase (D2HGDH), which converts D-2HG back into αKG, and showed inverse kinetics with HOT and D-2HG levels. We tested the immunomodulatory effects of D-2HG during LPS-induced inflammatory responses by transcriptomic analyses and functional profiling of D-2HG-pre-treated macrophages in vitro and mice in vivo. Together, these data suggest a role for D-2HG in the negative feedback regulation of inflammatory signaling during late-stage LPS-responses in vitro and as a regulator of local and systemic inflammatory responses in vivo. Finally, we show that D-2HG likely exerts distinct anti-inflammatory effects, which are in part independent of αKG-dependent dioxygenase inhibition. Together, this study reveals an immunometabolic circuit resulting in the accumulation of the immunomodulatory metabolite D-2HG that can inhibit inflammatory macrophage responses.
    Keywords:  2-HG; 2-hydroxyglutarate; Immunometabolism; Immunometabolite; Innate immunity; Macrophage
  2. Clin Exp Immunol. 2022 May 10. pii: uxac048. [Epub ahead of print]
    Keywords:  Dendritic cells; Metabolism; antitumour immunity; vaccines
  3. FASEB J. 2022 May;36 Suppl 1
      Sepsis is a life-threatening clinical implication with increased uncontrolled host immune response to an infection, mortality, morbidity, and financial burden worldwide. Due to its critical role in metabolic reprogramming in inflammation, the Irg1/itaconate has received much attention as an immunomodulator. However, understanding of itaconate's anti-inflammatory and immunometabolic activities in response to infections is mostly limited to immune cells. By employing multi-omic approaches, here we show that in the context of sepsis, a disruption in TCA cycle flow drives hepatic itaconate accumulation, indicating potential non-immune functions. However, the functional role of itaconate in this central organ critical to maintaining systemic metabolism is yet to be elucidated. To gain more insight into its physiological role in the liver during sepsis, we subjected wild-type and whole-body Irg1 knockout mice to sepsis via cecal slurry injection. In conjunction with our previous findings, we find wild-type septic mice develop hepatic steatosis. Interestingly, global Irg1 knockout mice develop a more severe form of a hepatic steatotic phenotype and a significant increase in hepatic lipid burden compared to wild-type counterparts in response to sepsis. This data demonstrates itaconate as a negative regulator of hepatic lipid accumulation in the context of sepsis. However, the exact molecular mechanism by which itaconate interacts with regulators of hepatic lipid metabolism during sepsis is yet to be uncovered. As an anti-inflammatory metabolite, itaconate limits the glycolytic response in activated immune cells. Similarly, invitro, we find that 4-OI, an itaconate derivative, antagonizes LPS induced glycolysis in hepatocytes. Given our findings of heightened lipid accumulation in septic Irg1 knockout mice, we further hypothesize glycolysis to be elevated and fueling de novolipogenesis via the production of acetyl-CoA. Indeed, unbiased metabolomics data show heightened hepatic lactate levels indicative of hyperactivated glycolysis in knockout septic mice. Mechanistically, we find elevated gene and protein expression of lactate dehydrogenase, the enzyme that facilitates the conversion of pyruvate to lactate, driving this accumulation of lactate seen in septic Irg1 knockout mice. In summary, our preliminary findings thus far highlights itaconate as a negative modulator of hepatic glycolysis as well as de novo lipogenesis in restraining sepsis-induced hepatic steatosis.
  4. FASEB J. 2022 May;36 Suppl 1
      Noncoding RNAs (ncRNAs) are emerging players in cell signaling and metabolism. Studies from our laboratory demonstrate that long noncoding RNA (lncRNA) HOTAIR plays critical roles in inflammation via regulation of NF-kB activation and cytokine expression. HOTAIR also regulates glucose metabolism in macrophages during inflammation. Importantly, metabolic reprogramming is critically linked to inflammation and macrophage activation. Therefore, our findings showing the HOTAIR mediated regulation of glucose metabolism in macrophages indicate potential roles of lncRNAs in macrophage activation and metabolic reprogramming. In a recent study, we explored potential involvement of other ncRNAs in inflammation and immune signaling. Based on independent RNAseq analyses in mouse and human macrophages, we have discovered a series novel mouse and human lncRNAs which are potential regulators of cytokine expression, inflammation, and immune response. Notably, there appear to be poor conservation among the lncRNAs between mouse and human. Mechanistic studies demonstrated that some of the novel inflammatory lncRNAs are regulated via NF-kB pathways while others are not, suggesting distinct modes of action of different lncRNAs during inflammatory response in macrophages. Here, we will present the discovery of novel mouse and human lncRNAs that are potential regulators of inflammation and immune signaling and discuss functional roles of selected lncRNAs in cytokine regulation, inflammation, and glucose metabolism in macrophages.
  5. Life Sci Alliance. 2022 Sep;pii: e202201405. [Epub ahead of print]5(9):
      Genome-scale metabolic models (GSMMs) can provide novel insights into metabolic reprogramming during disease progression and therapeutic interventions. We developed a context-specific system-level GSMM of people living with HIV (PLWH) using global RNA sequencing data from PBMCs with suppressive viremia either by natural (elite controllers, PLWHEC) or drug-induced (PLWHART) control. This GSMM was compared with HIV-negative controls (HC) to provide a comprehensive systems-level metabo-transcriptomic characterization. Transcriptomic analysis identified up-regulation of oxidative phosphorylation as a characteristic of PLWHART, differentiating them from PLWHEC with dysregulated complexes I, III, and IV. The flux balance analysis identified altered flux in several intermediates of glycolysis including pyruvate, α-ketoglutarate, and glutamate, among others, in PLWHART The in vitro pharmacological inhibition of OXPHOS complexes in a latent lymphocytic cell model (J-Lat 10.6) suggested a role for complex IV in latency reversal and immunosenescence. Furthermore, inhibition of complexes I/III/IV induced apoptosis, collectively indicating their contribution to reservoir dynamics.
  6. FASEB J. 2022 May;36 Suppl 1
      BACKGROUND: Significant proportion of inflammatory bowel disease (IBD) patients continue to respond inconsistently to therapies, underscoring disease complexity and the need for efficacious treatment. Interleukin 21 (IL-21), which is known to support T helper (Th) cell function, is highly expressed within inflamed intestinal tissues of IBD patients compared to healthy controls. In addition, inflammatory regulatory T cells (Tregs) have been linked to refractory human IBD. Given that healthy Tregs are critical for self-tolerance and prevention of IBD, we investigated the metabolic role of IL-21 in instigating Treg dysfunction and the therapeutic ramifications of targeting metabolism pathways during IBD pathogenesis.METHODS: Human Tregs as well as relevant control effector Th cells were generated from naïve CD4+ T cells isolated from healthy blood donors. Microarray analysis was utilized for targeted metabolic transcriptional profiling. Immune phenotyping was assessed by fluorescence-activated cell sorting. Metabolic phenotyping of cells was assessed by Seahorse flux analysis and mass spectrometry-based metabolomics. Ultrastructural analysis of mitochondria was performed by confocal and transmission electron microscopy. Intestinal inflammation was induced in Rag1-/- (T and B cell deficient) mice by the adoptive transfer of pathogenic naïve CD4+ T cells.
    RESULTS: Acute IL-21 stimulation of human Tregs induced glycolysis and fluctuations in mitochondrial respiration (i.e. oxidative phosphorylation - OXPHOS), as assessed by Seahorse flux analysis. In agreement, microarray analysis, validated by qPCR, revealed an IL-21-mediated increase in the expression of genes associated with glycolysis and pathways known to support anabolic and OXPHOS metabolism, thus resembling a hypermetabolic state. Furthermore, IL-21 stimulation rendered Tregs susceptible to inflammatory response, as evidenced by the production of effector Th cell-associated cytokines such as interferon γ, tumor necrosis factor, IL-17A, and IL-17F. Exploring the mechanisms underlying IL-21-induced effects, we found significant disruption of mitochondrial integrity with concomitant activation of glycogen synthase kinase 3 (GSK3) β, a kinase known to prevent pyruvate entry into the mitochondria. IL-21-induced GSK3β activation was accompanied by a marked increase in intracellular and extracellular metabolites such as pyruvate and lactate, as assessed by metabolomics. Importantly, GSK3 inhibition or supplementation with mitochondrial membrane-permeable methyl pyruvate broadly abrogated metabolic wiring of and inflammatory responses by IL-21-stimulated Tregs and effector Th cells. Collectively, these results suggest that impaired mitochondrial pyruvate metabolism is a feature of inflammatory CD4+ T cells. Lastly, GSK3 inhibition prevented pathogenic CD4+ T cell-induced colitis in mice as evidenced by reduced Disease Activity Index, Mouse Colon Histology Index, and serum inflammatory cytokines.
    CONCLUSIONS: IL-21 potently engages human Tregs in a hypermetabolic state that augments inflammatory cytokine production via induction of mitochondrial dysfunction. Therefore desensitizing CD4+ T cells to detrimental cues, such as IL-21, may also augment Treg function during human IBD.
  7. FASEB J. 2022 May;36 Suppl 1
      Intraepithelial lymphocytes expressing the γδ T cell receptor (γδ IEL) are located within the intestinal epithelium and serve as the first line of defense against pathogen invasion. We recently identified a novel hyperproliferative and hypermotile γδ IEL (γδHYP ) phenotype that can be transferred both horizontally and vertically to wildtype (WT) mice via the gut microbiota. Given the close relationship between metabolism and immune cell function, as well as the influence of commensals on T cell metabolism, we hypothesized that the bioenergetic potential of γδHYP IELs is increased to support this enhanced proliferation and motility. Transmission electron microscopy of sorted small intestinal γδ IELs revealed that γδHYP IELs exhibit a 70% increase in the number of mitochondria per cell (p=0.005) accompanied by a 24% increase in mitochondrial area (p=0.04) and 40% increase in aspect ratio (p=0.02) relative to WT. Since elongated mitochondria may be indicative of increased oxidative phosphorylation, we next performed Seahorse mitochondrial stress assays on sorted γδ IELs to assess the bioenergetic capacity of these lymphocytes. We find that γδHYP IELs exhibit a 50% increase in spare respiratory capacity compared to WT (p=0.014). Bulk RNA sequencing was performed on WT and γδHYP IELs and transcriptomic analysis revealed that γδHYP IELs upregulate genes involved in cell cycle (p<0.0001), as well as cholesterol and lipid metabolism (p<0.0001) compared to WT. Given the role of cholesterol and lipid metabolism in T cell activation and proliferation, we next analyzed intracellular cholesterol and lipid content within WT and γδHYP IELs using Filipin III and LipidTOX staining followed by flow cytometry. Interestingly, while there is a 3-fold increase in neutral lipids in γδHYP IELs (p<0.0001), cholesterol content is equivalent between WT and γδHYP IELs. Glucose and mitochondrial metabolism, as well as neutral lipid content, was recently shown to influence cytokine production by γδ T cells. Upon stimulation with phorbol myristate acetate (PMA) and ionomycin, the frequency of IFNγ+ γδ IELs was reduced by 59% with a 44% decrease in IFNγ mean fluorescence intensity (MFI) in γδHYP IELs relative to WT (p<0.0001). Although IFNγ production is reduced in γδHYP IELs, transcriptomic analysis revealed an increase in genes associated with antimicrobial defense including Reg3g (p<0.0001). In support of this, we find that γδHYP mice exhibit reduced bacterial burden in the spleen and liver 6 days post-infection with Salmonella Typhimurium compared to WT. In summary, our data demonstrate that the increased mitochondrial mass and oxidative phosphorylation observed in γδHYP IELs, in addition to increased neutral lipid accumulation, may contribute to this hyperproliferative and hypermotile phenotype and decreased IFNγ production. Further understanding of the metabolic mechanisms regulating γδ IEL homeostasis and effector function may ultimately allow fine tuning of mucosal surveillance to protect against intestinal injury or infection.
  8. Endocrinology. 2022 May 13. pii: bqac067. [Epub ahead of print]
      A high prevalence of metabolic syndrome (MetS) has been reported in multiple cohorts of systemic lupus erythematosus (SLE) patients, most likely as one of the consequences of autoimmune pathogenesis. Although MetS has been associated with inflammation, its consequences on the lupus immune system and on disease manifestations are largely unknown. The metabolism of immune cells is altered and overactivated in mouse models as well as in patients with SLE, and several metabolic inhibitors have shown therapeutic benefits. Here we review recent studies reporting these findings, as well as the effect of dietary interventions in clinical and pre-clinical studies of SLE. We also explore potential causal links between systemic and immunometabolism in the context of lupus, and the knowledge gap that needs to be addressed.
    Keywords:  autoimmunity; immunometabolism; lupus; systemic metabolism
  9. Int Immunopharmacol. 2022 May 10. pii: S1567-5769(22)00301-0. [Epub ahead of print]109 108817
      L-type amino acid transporter 1 (LAT1, slc7a5) supplies large neutral amino acids to highly proliferative cells. LAT1 is an attractive therapeutic target for treating overactive T cell-mediated immune disorders due to its high expression in activated T cells, but not in resting T cells. Here, we demonstrate that LAT1 plays a crucial role in T helper (Th) 17-mediated autoimmune arthritis in SKG mice, an animal model of human rheumatoid arthritis (RA). Administration of JPH203, a LAT1-specific inhibitor, suppressed mannan-induced joint swelling, synoviocyte proliferation and inflammatory cell infiltration in SKG mice. A diminished metabolic reprogramming, including a decrease in oxidative phosphorylation that regulates Hif-1α expression and subsequent control of glycolysis enzymes, was involved in the downregulation of Th17 differentiation by LAT1 inhibition. Moreover, publicly released database analysis revealed facilitated expression of LAT1 in T cells with cytotoxic features in patients with RA. Our results demonstrate the essential contribution of LAT1 to the development of RA, proposing a potential therapeutic approach targeting amino acid transporters for treating hypersensitive immune diseases.
    Keywords:  JPH203; L-type amino acid transporter (LAT) 1; Rheumatoid arthritis; T helper (Th) 17
  10. Cell Rep. 2022 May 10. pii: S2211-1247(22)00569-1. [Epub ahead of print]39(6): 110802
      Animals must adapt their growth to fluctuations in nutrient availability to ensure proper development. These adaptations often rely on specific nutrient-sensing tissues that control whole-body physiology through inter-organ communication. While the signaling mechanisms that underlie this communication are well studied, the contributions of metabolic alterations in nutrient-sensing tissues are less clear. Here, we show how the reprogramming of adipose mitochondria controls whole-body growth in Drosophila larvae. We find that dietary nutrients alter fat-body mitochondrial morphology to lower their bioenergetic activity, leading to rewiring of fat-body glucose metabolism. Strikingly, we find that genetic reduction of mitochondrial bioenergetics just in the fat body is sufficient to accelerate body growth and development. These growth effects are caused by inhibition of the fat-derived secreted peptides ImpL2 and tumor necrosis factor alpha (TNF-α)/Eiger, leading to enhanced systemic insulin signaling. Our work reveals how reprogramming of mitochondrial metabolism in one nutrient-sensing tissue can couple nutrient availability to whole-body growth.
    Keywords:  CP: Metabolism; Drosophila; OxPhos; TFAM; TNF-α; adipose tissue; fat body; growth; insulin; metabolism; mitochondria
  11. FASEB J. 2022 May;36 Suppl 1
      BACKGROUND: Macrophages play a critical role in left ventricular (LV) remodeling after myocardial infarction (MI), and limiting macrophage-mediated inflammatory responses while enhancing reparative roles is a promising therapeutic strategy. Monocyte-derived M1 macrophages mediate the early inflammatory response, while M2 macrophages mediate the later wound healing and scar formation phase. Metabolic reprogramming from glycolysis to mitochondrial oxidative phosphorylation mediates M1 to M2 polarization. We hypothesized that metabolic reprogramming occurs in macrophages over the course of post-MI remodeling, and that blocking macrophage glycolysis attenuates post-MI inflammation.METHODS: MI was induced in adult (16-20 week old) male C57BL/6J mice by permanent ligation of the left coronary artery for 1, 3, or 7 days, and cardiac function was assessed by echocardiography. Macrophages were extracted from the LV infarct area by immunomagnetic sorting. Macrophage metabolic flux was assessed by Extracellular Flux Analysis (Seahorse); upon glucose administration, glycolysis was measured by extracellular acidification rate (ECAR) while glucose oxidation was measured by oxygen consumption (OCR). Leukocyte subtypes (neutrophils, monocytes, resident macrophages) were assessed by flow cytometry. Real-time PCR was used to assess gene expression by relative mRNA levels. To specifically block macrophage glycolysis, LysMCre mice were crossed with mice floxed for Slc2a1, the major macrophage glucose transporter, to generate macrophage-Slc2a1 null mice.
    RESULTS: MI led to LV contractile dysfunction (decreased ejection fraction; days 1, 3, and 7 versus day 0), thinning of the infarct wall (days 1, 3, and 7) and non-infarct wall (day 7), and dilation (increased internal diameter at diastole and end-diastolic volume; day 7). At day 1, increased neutrophils (Ly6G+) and monocytes (Ly6Chi ) were observed in the infarcted LV; at day 7, neutrophils and monocytes returned to baseline levels and the majority of macrophages were resident M2 subtype (Ly6Clow versus day 1). Day 1 macrophages showed elevated glycolysis (ECAR versus day 0) and increased mRNA levels of the M1 marker Il1b (versus day 3 and 7); day 3 macrophages showed increased glycolysis (ECAR versus day 0) and decreased glucose oxidation (versus day 0); day 7 macrophages showed decreased glycolysis (ECAR versus day 1 and 3) and increased glucose oxidation (OCR versus day 1 and 3), and increased Slc2a1 mRNA, Krebs cycle genes (Pdha1, Idh1/2, Sdha/bmRNA), pentose phosphate pathway genes (G6pd2/x, Pgd, Rpia, Taldo1 mRNA), and Il10 (M2 marker) mRNA. In macrophage-Slc2a1 null mice, macrophage glycolysis was decreased and glucose oxidation increased at day 3, and infarct levels of Il1b were decreased compared to floxed controls.
    CONCLUSIONS: Our results indicate that time-dependent macrophage metabolic reprogramming occurs over the MI remodeling process, and that blocking macrophage glycolysis may be a promising strategy for limiting post-MI inflammation and adverse remodeling.
  12. Redox Biol. 2022 Apr 30. pii: S2213-2317(22)00098-2. [Epub ahead of print]53 102326
      The nonunion following a fracture is associated with severe patient morbidity and economic consequences. Currently, accumulating studies are focusing on the importance of macrophages during fracture repair. However, details regarding the process by which macrophages facilitate endochondral ossification (EO) are largely unknown. In this study, we present evidence that apoptotic chondrocytes (ACs) are not inert corpses awaiting removal, but positively modulate the osteoinductive ability of macrophages. In vivo experiments revealed that fatty acid (FA) metabolic processes up-regulated following EO. In vitro studies further uncovered that FAs derived from ACs are taken up by macrophages mainly through macrophage scavenger receptor 1 (MSR1). Then, our functional experiments confirmed that these exogenous FAs subsequently activate peroxisome proliferator-activated receptor α (PPARα), which further facilitates lipid droplets generation and fatty acid oxidation (FAO). Mechanistically, elevated FAO is involved in up-regulating the osteoinductive effect by generating BMP7 and NAD+/SIRT1/EZH2 axis epigenetically controls BMP7 expression in macrophages cultured with ACs culture medium. Our findings advanced the concept that ACs could promote bone regeneration by regulating metabolic and function reprogram in macrophages and identified macrophage MSR1 represents a valuable target for fracture treatments.
    Keywords:  Apoptotic chondrocyte; Fatty acid oxidation; MSR1; Macrophage; Osteogenic differentiation
  13. Immunometabolism. 2022 ;pii: e220011. [Epub ahead of print]4(2):
      Hematopoietic stem cells (HSC) directly initiate a response to bacterial infections by rapidly entering the cell cycle in order to produce mature blood cells. An important issue in the field of HSC biology is to understand how metabolic activities of HSC are fueled during specific condition that require HSC activation. In their paper, Mistry et al. provide evidence that bacterial infections trigger an increased in free fatty acid uptake by HSC that fuel fatty acid oxidation and mitochondrial respiration activities. This increased fatty acid uptake is exclusively dependent on the upregulation of the fatty acid transporter CD36. This study shed important light into the metabolic needs of HSC during septic conditions.
    Keywords:  CD36; fatty acid; hematopoietic stem cells; infection; mitochondria
  14. FASEB J. 2022 May;36 Suppl 1
      Otto Warburg discovered cancer cells favor increased glycolysis with pyruvate being converted to lactate rather than acetyl-CoA for the tricarboxylic acid cycle, and this is termed the Warburg effect. Warburg metabolism has been found to occur in mammalian rapidly proliferating cancer and immune cells. Anopheles species mosquitoes, the vectors for malaria, rely on their metabolic system to provide energy and intermediates for their innate immune system, so called immunometabolism. We posited Warburg metabolism is involved in Anopheles sp. mosquito's immune response. We designed an anti-bacterial immunity model using intrathoracic inoculations of Escherichia coli K12, avirulent bacterium, and Enterobacter sp. Ag1, virulent bacterium isolated from the Anopheles sp. midgut, to test the immunometabolism response. We found a statistically significant doubling in lactic acid upon Ent sp. immune challenge as compared to injection injury controls using a lactate assay (t-test, p = 0.0016). This provides evidence to support increased Warburg metabolism in immune challenged mosquitoes. We then fed Anopheles sp. mosquitoes dimethyl fumarate (DMF), a GAPDH inhibitor, and challenged the mosquitoes with E. coli. We found a significant mortality was observed in DMF treated mosquitoes compared to control (Mantel-Cox, p < 0.001). By inhibiting the utilization of glycolysis and Warburg metabolism, decreased survival related to immune challenge is observed, showing Warburg metabolism is crucial for mosquito immunity. We believe current cancer therapeutics targeting Warburg metabolism can be an effective vector control strategy to prevent mosquito-borne diseases.
  15. FASEB J. 2022 May;36 Suppl 1
      Free fatty acid receptor 4 (FFAR4), also known as G-protein coupled receptor 120 (GPR120), is a long-chain unsaturated fatty acid receptor expressed in adipocytes, endothelial cells, and macrophages. Activation of FFAR4 helps maintain metabolic homeostasis by regulating adipogenesis, insulin sensitivity, and inflammation. While FFAR4 is best known for its role its role in preventing obesity and diabetes, recent studies have demonstrated that FFAR4 may also play an important role in the development of atherosclerosis and cardiovascular disease (CVD). Given FFAR4's importance in anti-inflammatory signaling and high expression levels in macrophages, we designed experiments to test the hypothesis that FFAR4 prevents the development of atherosclerosis by reversing macrophage foam cell formation, a hallmark of early atherogenesis. In these studies, we isolated peritoneal macrophages from wild-type C57/BL6J mice and incubated them with oxidized low-density lipoprotein (oxLDL) to generate foam cells. We then investigated the effects of FFAR4 activation by GW9508 (a synthetic agonist) on lipid accumulation, cytokine secretion, and cholesterol efflux. Activation of FFAR4 by GW9508 decreased macrophage secretion of pro-inflammatory cytokines. We also found that activation of FFAR4 with GW9508 reduced lipid accumulation in macrophages as observed by decreased Oil Red O staining and reduced cellular cholesterol content. Additionally, activation of FFAR4 by GW9508 increased [3 H]-cholesterol efflux to high-density lipoprotein (HDL). Interestingly, the increased efflux was accompanied by decreased scavenger receptor CD36 expression (that mediates oxLDL uptake) and increased expression of ATP binding cassette transporters, ABCA1 and ABCG1 (that mediate cholesterol efflux). Taken together, our results support an exciting and novel role for FFAR4 in the reversal of foam cell formation and could emerge this receptor as a new target for treating CVD by preventing accumulation of atherosclerotic plaque.
  16. FASEB J. 2022 May;36 Suppl 1
      While inflammatory dysregulation is a major contributor to cardiac injury and worsened functional recovery, current anti-inflammatory therapies have shown limited success, necessitating studies which identify gaps in our mechanistic understanding of inflammation in the heart. Essential dietary N-3 and N-6 polyunsaturated fatty acids (PUFAs) are metabolized into a family of PUFA epoxides and downstream diols with contrasting effects in cardiac inflammatory disease, suggesting metabolite- and cell-specific roles for these molecules. Previous studies have demonstrated 12,13-dihydroxyoctadecenoic acid (12,13-DiHOME), a linoleic acid-derived diol, mediates mitochondrial damage and inflammation in cardiomyocytes, though its roles in other cardiac cell types are unknown. We hypothesized that 12,13-DiHOME enhances macrophage inflammation by promoting M1 polarization and NLRP3 inflammasome activation. To study macrophage polarization, PMA-differentiated THP1 "M0" macrophages were incubated with vehicle, 12,13-DiHOME (0.5 µM), M1-polarization stimuli (10 ng/mL LPS and 20 ng/mL IFN-gamma) or M1-polarization stimuli + 12,13-DiHOME for 24 hours. Using quantitative real-time PCR we observed that 12,13-DiHOME alone did not induce M1 polarization but M1-associated cytokine TNF gene expression was enhanced in macrophages treated with 12,13-DiHOME alone or M1-polarization stimuli + 12,13-DiHOME. The NLRP3 inflammasome response was activated in THP1 M0 macrophages by first priming with LPS (10 ng/mL) for 4.5h followed by treatment with nigericin (10 µM) for 30min. NLRP3 inflammasome activation was assessed by measuring key markers (NLRP3, caspase-1, interleukin-1β). We observed a concentration-dependent increase in NLRP3 inflammasome activation when 12,13-DiHOME was co-treated with LPS-priming. Next, we used epifluorescence microscopy to assess mitochondrial dysfunction, which is known to modulate macrophage NLRP3 inflammasome activation, by measuring alterations in membrane potential with TMRE (50nM). Time course analyses demonstrated no differences in mitochondrial membrane potential between LPS-primed and LPS+12,13-DiHOME-primed macrophages prior to nigericin-activation. However, LPS+12,13-DiHOME-primed macrophages activated with nigericin had a more rapid decline in membrane potential compared to nigericin-activated macrophages primed with LPS alone, suggesting exaggerated mitochondrial damage. Overall, our data demonstrates 12,13-DiHOME does not trigger macrophage polarization alone but enhances macrophage NLRP3 inflammasome activation potentially by causing mitochondrial injury.
  17. Theranostics. 2022 ;12(7): 3251-3272
      Rationale: Inflammatory macrophages and osteoclasts (OCs) play critical roles in joint inflammation, which feature the excessive production of reactive oxygen species (ROS), resulting in synovial inflammation and bone erosion. Scavenging ROS, especially by modulating mitochondrial metabolic activity, could be a desirable strategy for the management of inflammatory joints. This study aimed to develop a mitochondria-targeted supramolecular drug delivery system with exogenous and endogenous ROS-scavenging activities for the treatment of joint inflammation. Methods: In this study, we utilized a zinc-based metal-organic supercontainer (MOSC) as a proton sponge and electron reservoir with outstanding proton binding capacity, extracellular ROS-scavenging ability, and biocompatibility to establish an efficient supramolecular nanocarrier for endo/lysosomal escape and mitochondrial targeting. 4-Octyl itaconate (4-OI), an itaconate derivative, served as the loaded guest for the construction of a synergistic therapeutic system for inflammatory macrophages and OCs. Results: After the effective encapsulation of 4-OI, 4-OI@Zn-NH-pyr not only exhibited potent ROS-scavenging capacity, but also reduced ROS production by mediating mitochondrial respiration in inflammatory macrophages. Regarding its anti-inflammatory efficacy, 4-OI@Zn-NH-pyr ameliorated the inflammatory reaction by activating nuclear factor erythroid 2-related factor 2 (Nrf2), thus increasing the production of antioxidants, apart from the inhibition of NF-κB pathways. Additionally, receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast differentiation and function was remarkably suppressed by 4-OI@Zn-NH-pyr. Consistent with in vitro observations, 4-OI@Zn-NH-pyr efficiently inhibited synovial inflammation and subchondral bone destruction in an acute arthritis model. Conclusion: By using MOSCs that are highly reactive to ROS as drug-loaded matrices for the first time, this study provides an avenue for the management of severe joint inflammation by designing synergistic supramolecular drug-delivery systems with subcellular targeting and ROS-scavenging capacity.
    Keywords:  Coordination containers; Exogenous itaconate; Joint inflammation; ROS scavenging; Supermolecule
  18. FASEB J. 2022 May;36 Suppl 1
      CD4 T cell differentiation to pro-inflammatory and immunosuppressive subsets requires distinct metabolic pathways. Pro-inflammatory CD4 subsets rely on glycolysis, while immunosuppressive (Treg cells) subsets, require functional mitochondria for their differentiation and function. Previous studies have shown that binge alcohol (ethanol, EtOH) administration increased Tbet-expressing (Th1) and decreased FOXP3-expressing (Treg) CD4 T cells in the colons of mice. We tested the hypothesis that EtOH dysregulates normal CD4 T cell differentiation, after stimulation, by impairing mitochondrial homeostasis. Human naïve CD4 T cells were isolated from buffy coats from blood bank donors (N = 6) using MACS sorting. Cells were stimulated using anti-CD3-coated dishes in the presence of anti-CD28 and IL-12, and exposed to EtOH (0 and 50 mM) for 3 days. Mitochondrial content was measured with Mitotracker Deep Red. Gene expression indicative of: autophagosome formation (ATG5, ATG7, ATG13, MAP1LC3B, BECN1, BNIP3L, ULK1), mitophagy (PINK1, PRKN),mitochondrial fusion (MFN1, MFN2, OPA1), mitochondrial fission (MFFand FIS1), and mitochondrial biogenesis (PPARC1A, PPARC1B, TFAM) was determined by RT2 profiler arrays. EtOH-treated CD4 T cells had increased mitochondrial content (p = 0.0008) with Tregs accounting for the greatest increase in mitochondria (p = 0.04). There was a main effect of stimulation (p < 0.05) to increase ATG5, ATG13, MAP1LC3B, BECN1, BNIP3L, ULK1, MFF, PPARC1B, and TFAM, and a main effect of EtOH (p < 0.05) to increase PINK1 and decrease ATG7. There was a main effect of both EtOH and stimulation (p < 0.05) to increase MFN2, and OPA1.Taken together, these results indicate that EtOH increases mitochondrial content in Treg cells and dysregulates mitochondrial gene expression important for mitochondrial repair and mitophagy. These EtOH-mediated alterations in gene expression could result in an inability of CD4 T cells to maintain mitochondrial homeostasis and remove damaged mitochondria that is required for normal differentiation and function of anti-inflammatory Treg cells.
  19. Antioxid Redox Signal. 2022 May 11.
      SIGNIFICANCE: Immune cell therapy, involves the administration of immune cells into patients, has emerged as one of the most common type of immunotherapy for cancer treatment. Knowledge on the biology and metabolism of the adoptively transferred immune cells and the metabolic requirements of different cell types in the tumour is fundamental for the development of immune cell therapy with higher efficacy.RECENT ADVANCES: Adoptive T cell therapy has shown to be effective in limited types of cancer. Different types and generations of adoptive T cell therapies have evolved in the recent decade. This review covers the basic principles and development of these therapies in cancer treatment.
    CRITICAL ISSUES: Our review provides an overview on the basic concepts on T cell metabolism and highlights the metabolic requirements of T and adoptively transferred T cells.
    FUTURE DIRECTIONS: Integrating the knowledge above will facilitate the development of strategies to maximize the expansion of adoptively transferred T cells ex vivo and in vivo and promote their durability and anti-tumour effects.
  20. Mol Immunol. 2022 May 06. pii: S0161-5890(22)00195-X. [Epub ahead of print]147 101-114
      TRAF-associated NF-κB activator (TANK)-binding kinase 1 (TBK1), a nonclassical IκB kinase (IKK), and its effect on inflammation have not been entirely clarified. Here, we identified that TBK1 participates in the catabolism of glutamine by mediating the phosphorylation of receptor-interacting protein kinase 3 (RIPK3) and promoting macrophage endotoxin tolerance (ET). We found that the TBK1 protein directly interacts with the RIPK3 protein and mediates the phosphorylation of RIPK3 in macrophages. Activated RIPK3 can directly bind to glutamate dehydrogenase 1 (GLUD1), which is known to be a critical enzyme for catalyzing glutamine decomposition, to improve its catalytic activity and increase the production of α-ketoglutarate (α-KG) in macrophages. α-KG generated from glutaminolysis can promote M2 activation and restrict M1 polarization, which plays a crucial role in promoting lipopolysaccharide (LPS)-induced ET. As a result of TBK1 regulating the phosphorylation level of RIPK3, overexpressed TBK1 could enhance the tolerance of macrophages to endotoxin through glutaminolysis. Overall, these findings reveal a novel mechanism for the metabolic control of inflammation and for the induction of ET by modulating glutamine metabolism.
    Keywords:  Endotoxin tolerance; GLUD1; RIPK3; Sepsis; TBK1; α-KG
  21. Nat Aging. 2022 Mar;2(3): 231-242
      Impaired T cell immunity with aging increases mortality from infectious disease. The branching of Asparagine-linked glycans is a critical negative regulator of T cell immunity. Here we show that branching increases with age in females more than males, in naïve more than memory T cells, and in CD4+ more than CD8+ T cells. Female sex hormones and thymic output of naïve T cells (TN) decrease with age, however neither thymectomy nor ovariectomy altered branching. Interleukin-7 (IL-7) signaling was increased in old female more than male mouse TN cells, and triggered increased branching. N-acetylglucosamine, a rate-limiting metabolite for branching, increased with age in humans and synergized with IL-7 to raise branching. Reversing elevated branching rejuvenated T cell function and reduced severity of Salmonella infection in old female mice. These data suggest sex-dimorphic antagonistic pleiotropy, where IL-7 initially benefits immunity through TN maintenance but inhibits TN function by raising branching synergistically with age-dependent increases in N-acetylglucosamine.
    Keywords:  Immunosenescence; N-acetyglucosamine; N-glycan branching; N-glycosylation; T cell, infection; aging; immunity; interleukin-7
  22. FASEB J. 2022 May;36 Suppl 1
      OBJECTIVE: The study aimed to characterize immunometabolic signatures of porcine alveolar macrophage (PAM) associated with cellular iron deficiency (ID) and iron overload (IE). We hypothesized that cellular iron imbalance modulates LPS-induced inflammatory and metabolic responses in PAM cells.METHODS: Alveolar macrophage cells were collected from 6-week old donor piglets (N= 5-6) and were cultured in complete media at 37 ˚C with 5% CO2 until attachment. Cells were treated with complete media (control, CON) or complete media supplemented with 500 µM deferiprone (DFP) or 200 µM ferric ammonium citrate (FAC) to induce iron deficiency (ID) or iron excess (IE), respectively. At the end of the 36-hour treatment, cells were challenged with 100 ng/mL LPS (L) or sterile saline (S) for another 6 hours. Cells were tested for viability using XTT assay or harvested for analysis of gene, protein expression, and untargeted metabolome using RT-qPCR, western blot, and GC time-of-flight MS, respectively. Cell culture media were collected for analysis of inflammatory cytokines (TNFα, IL-6, and IL-10) using ELISA kits. Data were analyzed for two-way ANOVA using PROC MIXED of SAS (v.9.4). Statistical significance was declared when P-value or FDR is less than 0.05 or 0.1, respectively.
    RESULTS: LPS challenge reduced cell viability by 10 - 20% regardless of the cellular iron status (P < 0.001). Ferritin H was markedly increased in IE cells compared to CON or ID cells (P < 0.001), suggesting cellular iron overload. The mRNA expression of transferrin receptor 1 was the highest in ID-S cells and was significantly higher in ID and CON compared to IE cells, irrespective of the LPS challenge (P < 0.001). LPS significantly increased mRNA expression of divalent metal transporter 1 (DMT1)and zinc transporter (ZIP14) by 10-300 fold (P< 0.001), while iron treatment had no significant effect. Unsurprisingly, LPS drastically induced (P < 0.001) genes encoding pro- (TNFA and IL1B) and anti-(TGFB1 and IL10) inflammatory cytokines. Unexpectedly, both ID and IE diminished the induction of those genes caused by the LPS challenge (P < 0.05). The response of TNFA gene was mirrored by changes in TNFα concentration in culture media. Twenty-nine, 14, and 5 metabolites were altered by cellular iron status, LPS challenge, and their interaction (FDR < 0.1), respectively. A total of 28 altered metabolites were mapped in the user database. Interestingly, metabolites involved in the glycolysis and TCA cycle (e.g. glucose-6-phosphate, citric acid, and aconitic acid) were most abundant in ID cells, whereas, in comparison with CON, both ID and IE decreased several amino acids in PAM (e.g. glycine, alanine, leucine, and lysine). LPS increased metabolites involved in glycolysis (fructose-1-phosphate) and pentose phosphate pathway (ribose-5-phosphate and fructose-6-phosphate). Cellular iron status modulated changes in itaconic acid, glucose, fumaric acid, and putrescine in response to LPS challenge; ID enhanced the LPS-induced increase in itaconic acid, a metabolite marker of macrophage activation.
    CONCLUSION: Presence of iron chelator and iron overexposure altered cellular iron metabolism of PAM cells and its inflammatory and metabolic responses to the invasion of bacterial pathogens.