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The differentiation and suppressive functions of regulatory CD4 T cells (Tregs) are supported by a broad array of metabolic changes, providing potential therapeutic targets for immune modulation. In this study, we focused on the regulatory role of glycolytic enzymes in Tregs and identified phosphoglycerate mutase (PGAM) as being differentially overexpressed in Tregs and associated with a highly suppressive phenotype. Pharmacologic or genetic inhibition of PGAM reduced Treg differentiation and suppressive function while reciprocally inducing markers of a pro-inflammatory, T helper 17 (Th17)-like state. The regulatory role of PGAM was dependent on the contribution of 3-phosphoglycerate (3PG), the PGAM substrate, to de novo serine synthesis. Blocking de novo serine synthesis from 3PG reversed the effect of PGAM inhibition on Treg polarization, while exogenous serine directly inhibited Treg polarization. Additionally, altering serine levels in vivo with a serine/glycine-free diet increased peripheral Tregs and attenuated autoimmunity in a murine model of multiple sclerosis. Mechanistically, we found that serine limits Treg polarization by contributing to one-carbon metabolism and methylation of Treg-associated genes. Inhibiting one-carbon metabolism increased Treg polarization and suppressive function both in vitro and in vivo in a murine model of autoimmune colitis. Our study identifies a novel physiologic role for PGAM and highlights the metabolic interconnectivity between glycolysis, serine synthesis, one-carbon metabolism, and epigenetic regulation of Treg differentiation and suppressive function.

The liver plays a significant role in regulating a wide range of metabolic, homeostatic, and host-defense functions. However, the impact of liver injury on the host's ability to control bacteremia and morbidity in sepsis is not well understood. Leukocyte recruitment and activation lead to cytokine and chemokine release, which, in turn, trigger hepatocellular injury and elevate nucleotide levels in the extracellular milieu. P2Y2 purinergic receptors, G protein-coupled and activated by extracellular ATP/UTP, are expressed at the cell surface of hepatocytes and nonparenchymal cells. We sought to determine whether P2Y2 purinergic receptor function is necessary for the maladaptive host response to bacterial infection and endotoxin-mediated inflammatory liver injury and mortality in mice. We report that P2Y2 purinergic receptor knockout mice (P2Y2 -/- ) had attenuated inflammation and liver injury, with improved survival in response to LPS/galactosamine (LPS/GalN; inflammatory liver injury) and cecal ligation and puncture (CLP; polymicrobial sepsis). P2Y2 -/- livers had attenuated c-Jun NH2-terminal kinase activation, matrix metallopeptidase-9 expression, and hepatocyte apoptosis in response to LPS/GalN and attenuated inducible nitric oxide synthase and nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain containing 3 protein expression in response to CLP. Implicating liver injury in the disruption of amino acid homeostasis, CLP led to lower serum arginine and higher bacterial load and morbidity in the WT mice, whereas serum arginine levels were comparable to sham-operated controls in P2Y2 -/- mice, which had attenuated bacteremia and improved survival. Collectively, our studies highlight the pathophysiological relevance of P2Y2 purinergic receptor function in inflammatory liver injury and dysregulation of systemic amino acid homeostasis with implications for sepsis-associated immune dysfunction and morbidity in mice. NEW & NOTEWORTHY Our studies provide experimental evidence for P2Y2 purinergic receptor-mediated potentiation of inflammatory liver injury, morbidity, and mortality, in two well-established animal models of inflammatory liver injury. Our findings highlight the potential to target P2Y2 purinergic signaling to attenuate the induction of "cytokine storm" and prevent its deleterious consequences on liver function, systemic amino acid homeostasis, host response to bacterial infection, and sepsis-associated morbidity and mortality.

Background: Non-small cell lung cancer (NSCLC) comprises the majority (~85%) of all lung tumors, with lung adenocarcinoma (LUAD) and squamous cell carcinoma (LUSC) being the most frequently diagnosed histological subtypes. Multi-modal omics profiling has been carried out in NSCLC, but no studies have yet reported a unique metabolite-related gene signature and altered metabolic pathways associated with LUAD and LUSC., Methods: We integrated transcriptomics and metabolomics to analyze 30 human lung tumors and adjacent noncancerous tissues. Differential co-expression was used to identify modules of metabolites that were altered between normal and tumor., Results: We identified unique metabolite-related gene signatures specific for LUAD and LUSC and key pathways aberrantly regulated at both transcriptional and metabolic levels. Differential co-expression analysis revealed that loss of coherence between metabolites in tumors is a major characteristic in both LUAD and LUSC. We identified one metabolic onco-module gained in LUAD, characterized by nine metabolites and 57 metabolic genes. Multi-omics integrative analysis revealed a 28 metabolic gene signature associated with poor survival in LUAD, with six metabolite-related genes as individual prognostic markers., Conclusions: We demonstrated the clinical utility of this integrated metabolic gene signature in LUAD by using it to guide repurposing of AZD-6482, a PI3Kβ inhibitor which significantly inhibited three genes from the 28-gene signature. Overall, we have integrated metabolomics and transcriptomics analyses to show that LUAD and LUSC have distinct profiles, inferred gene signatures with prognostic value for patient survival, and identified therapeutic targets and repurposed drugs for potential use in NSCLC treatment., (© 2022 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)

Background: Respiratory tract microbial dysbiosis can exacerbate inflammation and conversely inflammation may cause dysbiosis. Dysbiotic microbiome metabolites may lead to bronchopulmonary dysplasia (BPD). Hyperoxia and lipopolysaccharide (LPS) interaction alters lung microbiome and metabolome, mediating BPD lung injury sequence., Methods: C57BL6/J mice were exposed to 21% (normoxia) or 70% (hyperoxia) oxygen during postnatal days (PND) 1-14. Pups were injected with LPS (6 mg/kg) or equal PBS volume, intraperitoneally on PND 3, 5, and 7. At PND14, the lungs were collected for microbiome and metabolomic analyses (n = 5/group)., Results: Microbiome alpha and beta diversity were similar between groups. Metabolic changes included hyperoxia 31 up/18 down, LPS 7 up/4 down, exposure interaction 8. Hyperoxia increased Intestinimonas abundance, whereas LPS decreased Clostridiales, Dorea, and Intestinimonas; exposure interaction affected Blautia. Differential co-expression analysis on multi-omics data identified exposure-altered modules. Hyperoxia metabolomics response was integrated with a published matching transcriptome, identifying four induced genes (ALDOA, GAA, NEU1, RENBP), which positively correlated with BPD severity in a published human newborn cohort., Conclusions: We report hyperoxia and LPS lung microbiome and metabolome signatures in a clinically relevant BPD model. We identified four genes correlating with BPD status in preterm infants that are promising targets for therapy and prevention., Impact: Using multi-omics, we identified and correlated key biomarkers of hyperoxia and LPS on murine lung micro-landscape and examined their potential clinical implication, which shows strong clinical relevance for future research. Using a double-hit model of clinical relevance to bronchopulmonary dysplasia, we are the first to report integrated metabolomic/microbiome landscape changes and identify novel disease biomarker candidates., (© 2022. The Author(s), under exclusive licence to the International Pediatric Research Foundation, Inc.)

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Our early-life environment has a profound influence on developing organs that impacts metabolic function and determines disease susceptibility across the life-course. Using a rat model for exposure to an endocrine disrupting chemical (EDC), we show that early-life chemical exposure causes metabolic dysfunction in adulthood and reprograms histone marks in the developing liver to accelerate acquisition of an adult epigenomic signature. This epigenomic reprogramming persists long after the initial exposure, but many reprogrammed genes remain transcriptionally silent with their impact on metabolism not revealed until a later life exposure to a Western-style diet. Diet-dependent metabolic disruption was largely driven by reprogramming of the Early Growth Response 1 (EGR1) transcriptome and production of metabolites in pathways linked to cholesterol, lipid and one-carbon metabolism. These findings demonstrate the importance of epigenome:environment interactions, which early in life accelerate epigenomic aging, and later in adulthood unlock metabolically restricted epigenetic reprogramming to drive metabolic dysfunction.

Cells are internally organized into compartmentalized organelles that execute specialized functions. To understand the functions of individual organelles and their regulations, it is critical to resolve the compositions of individual organelles, which relies on a rapid and efficient isolation method for specific organellar populations. Here, we introduce a robust affinity purification method for rapid isolation of intracellular organelles (e.g. lysosomes, mitochondria and peroxisomes) by taking advantage of the extraordinarily high affinity between the twin strep tag and streptavidin variants. With this method, we can isolate desired organelles with high purity and yield in 3 min from the post-nuclear supernatant of mammalian cells or less than 8 min for the whole purification process. Using lysosomes as an example, we show that the rapid procedure is especially useful for studying transient and fast cellular activities, such as organelle-initiated signaling and organellar contents of small-molecular metabolites. Therefore, our method offers a powerful tool to dissect spatiotemporal regulation and functions of intracellular organelles., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2019. Published by The Company of Biologists Ltd.)