Search Results (188)

Global climate change is pushing insects into colder regions. Understanding their cold tolerance is important for predicting population dynamics. During overwintering, Streltzoviella insularis larvae activate the AMPK signaling pathway. This suggests that energy metabolism plays a key role under cold stress. In this study, we used enzyme activity assays, LC-MS-based targeted metabolomics, and transcriptome sequencing. We focused on six key enzymes in glycolysis and the TCA cycle. We also measured related metabolites and regulatory genes. Hexokinase (HK) and citrate synthase (CS) activities were highly sensitive to temperature. HK increased then markedly decreased; CS was significantly downregulated. Pyruvate kinase (PK), isocitrate dehydrogenase (IDH), and α-ketoglutarate dehydrogenase (KGD) showed trends that matched changes in larval cold tolerance, exhibiting an up–down–up expression trend. Glycolytic metabolites (glucose-6-phosphate, fructose-6-phosphate, 1,6-fructose-diphosphate, phosphoenolpyruvic acid) peaked at −10 °C. TCA intermediates (citrate, acetyl-CoA, α-ketoglutaric acid, and isocitrate) were more abundant at 0–4 °C. Pyruvate increased significantly. PYR content showed a significant increase followed by a decrease, peaking at 0 °C. It was converted into lactate and acetyl-CoA. ATP levels dropped and then increased, reaching their lowest level at 0 °C. These results suggest a shift from aerobic to mixed aerobic–anaerobic metabolism. Transcriptome data showed differential expression of key metabolic genes such as phosphoenolpyruvate carboxykinase, phosphoglycerate kinase, and ATP synthase subunit beta. These gene changes supported the trends in enzymes and metabolites. Our findings reveal a coordinated metabolic and transcriptional response to cold. This provides a basis for understanding the cold adaptation and potential range expansion of S. insularis. Full article

Background/Objectives: Oxidative stress constitutes a principal pathophysiological mechanism driving neurodegeneration and brain aging. α-Ketoglutarate (AKG), a key intermediate of the tricarboxylic acid (TCA) cycle, has shown potential in longevity and oxidative stress resistance. However, the role of AKG in oxidative stress-induced neuronal senescence and its interaction with the mTOR signaling pathway during neuronal aging remain poorly understood, posing a key challenge for developing senescence-targeted therapies. Methods: We investigated the neuroprotective effects of AKG using H₂O₂-induced senescence in HT22 cells and a D-galactose-induced brain aging mouse model. Assessments encompassed SA-β-gal staining, EdU incorporation, mitochondrial membrane potential (JC-1), and ROS measurement. Antioxidant markers, ATP levels, and the NAD⁺/NADH ratio were also analyzed. Proteomic profiling (DIA-MS) and KEGG/GSEA enrichment analyses were employed to identify AKG-responsive signaling pathways, and Western blotting validated changes in mTOR signaling and downstream effectors. Results: AKG significantly alleviated H₂O₂-induced senescence in HT22 cells, evidenced by enhanced cell viability, reduced ROS level, restored mitochondrial function, and downregulated p53/p21 expression. In vivo, AKG administration improved cognitive deficits and vestibulomotor dysfunction while ameliorating brain oxidative damage in aging mice. Proteomics revealed mTOR signaling pathways as key targets for AKG’s anti-aging activity. Mechanistically, AKG suppressed mTOR phosphorylation and activated ULK1, suggesting modulation of autophagy and metabolic homeostasis. These effects were accompanied by enhanced antioxidant enzyme activities and improved redox homeostasis. Conclusions: Our study demonstrates that AKG mitigates oxidative stress-induced neuronal senescence through suppression of the mTOR pathway and enhancement of mitochondrial and antioxidant function. These findings highlight AKG as a metabolic intervention candidate for age-related neurodegenerative diseases. Full article

The enzyme ABH2, one of nine human DNA dioxygenases of the AlkB family, belongs to the superfamily of Fe(II)/α-ketoglutarate-dependent dioxygenases and plays a crucial role in the direct reversal repair of nonbulky alkyl lesions in DNA nucleobases. ABH2 has broad substrate specificity, directly oxidizing DNA damages such as N¹-methyladenine, N³-methylcytosine, 1,N⁶-ethenoadenine, 3,N⁴-ethenocytosine, and a number of others. In our investigation, we sought to uncover the subtleties of the mechanisms governing substrate specificity in ABH2 by focusing on several critical amino acid residues situated in its active site. To gain insight into the function of this enzyme, we performed a functional mapping of its active site region, concentrating on pivotal residues, participating in forming a damaged binding pocket of the enzyme (Val99 and Ser125), as well as the residues directly involved in interactions with damaged bases, namely Arg110, Phe124, Arg172, and Glu175. To support our experimental data, we conducted a series of molecular dynamics simulations, exploring the interactions between the ABH2 mutant forms, bearing corresponding substitutions and DNA substrates, and harboring various types of methylated bases, specifically N¹-methyladenine or N³-methylcytosine. The comparative studies revealed compelling data indicating that alterations in most of the studied amino acid residues significantly influence both the binding affinity of the enzyme for DNA and its catalytic efficiency. Intriguingly, the findings suggest that the mutations impact the catalytic activity of ABH2 to a greater extent than its ability to associate with DNA strands. Collectively, these results show how changes to the active site affect molecular dynamics and reaction kinetics, improving our understanding of the substrate recognition process in this pivotal enzyme. Full article

Epigenetic modifications act as crucial regulators of gene activity and are influenced by both internal and external environmental factors, with diet being the most impactful external factor. On the other hand, cellular metabolism encompasses a complex network of biochemical reactions essential for maintaining cellular function, and it impacts every cellular process. Many metabolic cofactors are critical for the activity of chromatin-modifying enzymes, influencing methylation and the global acetylation status of the epigenome. For instance, dietary nutrients, particularly those involved in one-carbon metabolism (e.g., folate, vitamins B12 and B6, riboflavin, methionine, choline, and betaine), take part in the generation of S-adenosylmethionine (SAM), which represents the main methyl donor for DNA and histone methylation; α-ketoglutarate and ascorbic acid (vitamin C) act, respectively, as a co-substrate and cofactor for Ten-eleven Translocation (TET), which is responsible for DNA demethylation; and metabolites such as Acetyl-CoA directly impact histone acetylation, linking metabolism of the TCA cycle to epigenetic regulation. Further, bioactive compounds, such as polyphenols, modulate epigenetic patterns by affecting methylation processes or targeting epigenetic enzymes. Since diet and nutrition play a critical role in shaping epigenome functions and supporting human health, this review offers a comprehensive update on recent advancements in metabolism, epigenetics, and nutrition, providing insights into how nutrients contribute to metabolic balance, epigenome integrity maintenance and, consequently, disease prevention. Full article

Glutamate is one of the major naturally occurring non-essential amino acids. The aim of this review is to provide a comprehensive analysis of the role of glutamate as a key metabolite in the metabolism of plant and animal organisms. Its role in nutrition and neurotransmission has intrigued researchers for many years. In both plants and animals, glutamate primarily exists in a monoanionic form characterised by unique physical and chemical properties. In plants, it is involved in the glutamine synthetase/glutamate synthase (GS/GOGAT) cycle, while in animals, it plays a role in the glutamine/glutamate cycle, which is closely related to the urea cycle. Glutamate is also closely linked to the Krebs cycle in both groups of organisms through α-ketoglutarate. Glutamate is essential in both biosynthetic and catabolic pathways and participates in numerous physiological processes in plants and animals. Animals acquire glutamate from food, while plants acquire it from the soil; however, both also synthesise it de novo. Once present in the body, it is transported across cell membranes by specific transporters driven by ionic gradients (a mechanism known as secondary active transport). It is involved in cellular and systemic signalling pathways by interacting with ionotropic and metabotropic receptors. Additionally, glutamate is an important ‘building block’ of many proteins, including storage proteins. It also occurs in the form of monosodium glutamate (MSG), a flavour enhancer that is widely used but often criticised. Due to its important role in metabolism and signalling, the significance of glutamate in nutrition and its impact on human health are vital areas of research in food biochemistry. These investigations contribute to the development of nutritious food products and the design of effective pharmaceuticals. In this paper, we also address unresolved questions in glutamate research and consider its practical applications. Full article

Alpha-ketoglutaric acid (AKG) is a key intermediate in the tricarboxylic acid cycle and plays a crucial role in energy production and amino acid metabolism. This study aimed to evaluate the effects of AKG on growth performance, nutrient digestibility, plasma biochemical parameters, and plasma amino acid profiles in weaned piglets. A total of 72 weaned piglets with an average weight of 7.33 kg (±0.50 kg) and an average age of 28 (±2 days) were randomly assigned to 3 dietary treatments with 6 replicates per group in a 42-day trial. The treatments included a basal diet (CT), a basal diet with 500 g/t AKG (AKG1), and a basal diet with 1000 g/t AKG (AKG2). Blood samples were collected on days 14 and 42, and fecal samples were collected on day 42. The results showed that diets including 500 g/t and 1000 g/t AKG significantly reduced diarrhea incidence in piglets compared to the CT group (p < 0.01). Moreover, diets including 1000 g/t AKG enhanced fecal dry matter digestibility, plasma albumin (ALB), and glucose (GLU) concentrations on day 42 compared to the AKG1 group (p < 0.05). In conclusion, diets including AKG reduced diarrhea incidence in weaned piglets, potentially through improved nutrient digestibility and enhanced systemic health. The 1000 g/t level exhibited greater beneficial effects than 500 g/t. Full article

Citric acid serves as the principal organic acid in citrus fruits, with its concentration critically determining fruit flavor and market acceptability. Isocitrate dehydrogenase (IDH), a key enzyme in citric acid metabolism, mediates the conversion of citrate to α-ketoglutarate. This study cloned six candidate genes encoding IDH from grapefruit (Citrus paradisi). Bioinformatics analysis showed that all six genes contained the typical characteristic structure of IDH. Gene expression analysis found that CpNADP-IDH1 is highly expressed in mature and low-acid varieties. Overexpression of CpNADP-IDH1 significantly increased IDH enzyme activity and decreased citric acid content in transgenic grapefruit callus. These results showed that at least six genes encoding IDH exist in grapefruit, among which CpNADP-IDH1 catalyzes the decomposition of citric acid and regulates the organic acid content in fruits at maturity. CpNADP-IDH1 can be used as a candidate gene for molecular breeding of low-acid citrus varieties and as an essential target gene for developing citrus cultivation technology for reducing acid content. Full article

N6-methyladenosine (m⁶A) methylation functions as a vital post-transcriptional and epigenetic modification in higher plants regulated by α-ketoglutarate-dependent dioxygenases (ALKBH). However, the role of ALKBH genes in oriental melon (Cucumis melo L.) fruit ripening has not been explored. Therefore, we treated oriental melon with an exogenous m⁶A demethylase inhibitor (mechlorfenamic acid) then analyzed endogenous ethylene production and ripening-related indicators to explore the effects of m⁶A methylation on ripening. Bioinformatics and real-time quantitative PCR analyses were used to determine the impact of ALKBH genes on key ethylene synthesis gene expression. Treatment effectively inhibited endogenous ethylene production, firmness changes, and soluble solid contents, thereby extending fruit ripening. Eight ALKBH gene family members belonging to five major groups were identified in the melon genome. All members were expressed in ripening fruits, with different expression patterns during ripening. CmALKBH6, CmALKBH7, and CmALKBH8 expression was inhibited by an ethylene inhibitor (1-methylcyclopropene). The transient overexpression (OE) of CmALKBH8 in oriental melon led to the increased expression of the ethylene synthesis genes CmACS1, CmACS2, and CmACO1. In summary, the ethylene-regulated gene CmALKBH8 may participate in oriental melon fruit ripening regulation by modulating the methylation levels of ethylene synthesis-related genes. These findings help us better understand how m⁶A methylation regulates melon ripening. Full article

α-ketoglutarate dehydrogenase complex (KGDHc) is a crucial enzyme in the tricarboxylic acid (TCA) cycle that intersects monosaccharides, amino acids, and fatty acid catabolism with oxidative phosphorylation (OxPhos). A key feature of KGDHc is its ability to sense changes in the redox environment through the reversible oxidation of the vicinal lipoic acid thiols of its dihydrolipoamide succinyltransferase (DLST; E2) subunit, which controls its activity and, by extension, OxPhos. This characteristic inculcates KGDHc with redox regulatory properties for the modulation of metabolism and mediating of intra- and intercellular signals. The innate capacity of KGDHc to participate in the regulation of cell redox homeodynamics also occurs through the production of mitochondrial hydrogen peroxide (mtH₂O₂), which is generated by the dihydrolipoamide dehydrogenase (DLD; E3) downstream from the E2 subunit. Reversible covalent redox modification of the E2 subunit controls this mtH₂O₂ production by KGDHc, which not only protects from oxidative distress but also modulates oxidative eustress pathways. The importance of KGDHc in modulating redox homeodynamics is underscored by the pathogenesis of neurological and metabolic disorders that occur due to the hyper-generation of mtH₂O₂ by this enzyme complex. This also implies that the targeted redox modification of the E2 subunit could be a potential therapeutic strategy for limiting the oxidative distress triggered by KGDHc mtH₂O₂ hyper-generation. In this short article, I will discuss recent findings demonstrating KGDHc is a potent mtH₂O₂ source that can trigger the manifestation of several neurological and metabolic diseases, including non-alcoholic fatty liver disease (NAFLD), inflammation, and cancer, and the targeted redox modification of the E2 subunit could alleviate these syndromes. Full article

Background/Objectives: Gestational diabetes mellitus (GDM) is a condition characterized by hyperglycemia and is associated with increased risk of obesity and diabetes in exposed children. Differences in human milk composition between women with (GDM+) and without GDM (GDM-) suggest that GDM could impact milk production and composition, potentially influencing infant growth. However, this association remains poorly understood. The objective was to study the association between GDM and human milk composition and its influence on infant growth, focusing on metabolites and bioactive molecules involved in energy metabolism. Methods: Using a cross-sectional design, 24 metabolites were measured by GC-MS in human milk obtained at 2 months postpartum from 20 GDM+ women and 29 GDM- women. Anthropometric measures, as well as lipid and glycemic profiles, were collected. Infant weight and length data were obtained from health records. Results: Human milk metabolites significantly differ between GDM+ and GDM- mothers, with higher levels of myristic acid, glycerol, uracil, arachidonic acid, and cholesterol in GDM+ milk (p < 0.05). Specific human milk metabolites showed distinct correlations with maternal glycemic as well as infant growth, depending on GDM status. While maternal glycemia was associated with succinate and malate in all groups, maternal glycemia was specifically correlated with valine and glutamate in GDM+ mothers. Additionally, in GDM+ women, α-ketoglutarate and glycine were negatively correlated with infant growth. Conclusions: The results of this study suggest that GDM can influence the mother’s health beyond delivery, impacting the mammary gland biology with effects on the human milk composition. Further, correlations with infant growth suggest that GDM-dependent variations in milk composition potentially influence infant growth and metabolism. Full article

Background/Objectives: Exposure to high altitudes often results in gastrointestinal disorders. This study aimed to identify probiotic strains that can alleviate such disorders. Methods: We conducted a microbiome analysis to investigate the differences in gut microbiota among volunteers during the acute response and acclimatization phases at high altitudes. Subsequently, we established a mouse model of intestinal barrier damage induced by high-altitude exposure to further investigate the roles of probiotic strains and 2-ketoglutaric acid. Additionally, we performed untargeted metabolomics and transcriptomic analyses to elucidate the underlying mechanisms. Results: The microbiome analysis revealed a significant increase in the abundance of Faecalibacterium prausnitzii during the acclimatization phase. Faecalibacterium duncaniae (F. duncaniae) significantly mitigated damage to the intestinal barrier and the reduction of 2-ketoglutaric acid levels in the cecal contents induced by high-altitude exposure in mice. Immunohistochemistry and TUNEL staining demonstrated that high-altitude exposure significantly decreased the expression of ZO-1 and occludin while increasing apoptosis in ileal tissues. In contrast, treatment with F. duncaniae alleviated the loss of ZO-1 and occludin, as well as the apoptosis induced by high-altitude exposure. Furthermore, 2-ketoglutaric acid also mitigated this damage, reducing the loss of occludin and apoptosis in mice. Transcriptomic analysis indicated that high-altitude exposure significantly affects the calcium signaling pathway; conversely, the administration of F. duncaniae significantly influenced the PPAR signaling pathway, mineral absorption, and the regulation of lipolysis in adipocytes. Additionally, the expression of the FBJ osteosarcoma oncogene (Fos) was markedly reduced following the administration of F. duncaniae. Conclusions: F. duncaniae mitigates hypoxia-induced intestinal barrier damage by increasing levels of 2-ketoglutaric acid and shows promise as a probiotic, ultimately aiding travelers in adapting to high-altitude environments. Full article

Background/Objectives: Cancer cells often display altered energy metabolism. In particular, expression levels and activity of the tricarboxylic acid cycle (TCA cycle) enzymes may change in cancer, and dysregulation of the TCA cycle is a frequent hallmark of cancer cell metabolism. MEMO1, a modulator of cancer metastasis, has been shown to bind iron and regulate iron homeostasis in the cells. MEMO1 knockout changed mitochondrial morphology and iron content in breast cancer cells. Our previous genome-wide analysis of MEMO1 genetic interactions across multiple cancer cell lines revealed that gene sets involved in mitochondrial respiration and the TCA cycle are enriched among the gain-of-function interaction partners of MEMO1. Based on these findings, we measured the TCA cycle metabolite levels in breast cancer cells with varying levels of MEMO1 expression. Methods: ShRNA knockdown assay was performed to test essentiality of key TCA cycle enzymes. TCA metabolites were quantified using liquid chromatography-tandem mass spectrometry (LC-MS/MS) in MDA-MB-231 (high MEMO1), M67-2 (MEMO1 knockdown), and M67-9 (MEMO1 knockout) cells under iron-depleted, basal iron, and iron-supplemented conditions. Results:ACO2 and OGDH knockdowns inhibit cell proliferation, indicating an essential role of the TCA cycle in MDA-MB-231 metabolism. α-Ketoglutarate and citrate levels exhibited an inverse relationship with MEMO1 expression, increasing significantly in MEMO1 knockout cells regardless of iron availability. In contrast, fumarate, malate, and glutamate levels were elevated in MEMO1 knockout cells specifically under low iron conditions, suggesting an iron-dependent effect. Conclusions: Overall, our results indicate that MEMO1 plays a role in regulating the TCA in cancer cells in an iron-dependent manner. Full article

As an endogenous metabolite, α-ketoglutarate (AKG) exhibits potent antioxidant properties, yet its molecular mechanisms remain unclear. Dermal Papilla Cells (DPCs), functioning as the regulatory hub of hair follicle morphogenesis, serve as a pivotal model system for deciphering follicular functionality and regeneration mechanisms through their orchestration of signaling networks. Using a hydrogen peroxide (H₂O₂)-induced oxidative stress model in DPCs, we investigated AKG’s protective effects. AKG attenuated H₂O₂-triggered reactive oxygen species (ROS) overproduction, restored mitochondrial membrane potential, and suppressed apoptosis-related protein dysregulation. It enhanced cellular stress resistance by increasing the Bcl-2/Bax ratio, boosting antioxidant levels, and inhibiting inflammation. Mechanistically, H₂O₂ activated the Nrf2 pathway, while AKG amplified Nrf2 nuclear translocation and expression. Crucially, ERK inhibition abrogated AKG-mediated Nrf2 regulation, intensifying ROS accumulation and cell death. These results identify the ERK/Nrf2 axis as central to AKG’s antioxidative cytoprotection. This study advances AKG’s therapeutic potential and deepens insights into its multifunctional roles. Full article

Water scarcity can negatively affect crop yield, posing a significant threat to global food security, such as drought. Plant growth-promoting rhizobacteria (PGPR), either as single strains or synthetic communities (SynComs), has shown promise in alleviating drought stress in various plant species. In this study, we examined the effects of water limitation on Salvia officinalis and the potential of a SynCom composed of five phosphate-solubilizing, auxin-producing, and/or nitrogen-fixing Gram-negative bacteria to enhance plant growth and drought tolerance. Plant growth, morphology, physiology, and leaf metabolomic profiles were assessed using a combination of physiological measurements and LC-MS untargeted metabolomics. Mild water stress induced a conservative water-use strategy in S. officinalis, characterized by increased root-to-shoot ratio and altered leaf morphology, without compromising photosynthetic performance. SynCom inoculation under well-watered conditions elicited drought-like responses, including transient reductions in stomatal conductance. Leaf metabolomic analysis revealed that inoculation influenced the abundance of several metabolites, including biogenic amines and dipeptides, under both irrigation regimes. Notably, drought stress and SynCom inoculation increased histamine and α-ketoglutaric acid levels, highlighting potential impacts on food quality. Under reduced irrigation, inoculation further modulated leaf morphology and biomass allocation, promoting thicker leaves and increased root biomass allocation. These results demonstrate the ability of the SynCom to modulate plant physiology and metabolism in response to both optimal and reduced irrigation, potentially enhancing drought resilience without directly improving growth. The study also highlights the complex interactions among microbial inoculation, plant stress responses, and leaf metabolite profiles, emphasizing the importance of considering the effects on the production of bioactive compounds when developing microbial inoculants for edible plants. Full article

A divergent selection for resilience was carried out in rabbits over 12 generations. The selection criterion was increased (the HO line) and decreased litter size variability at birth (the HE line). The HO line (more resilient) shows higher litter size than the HE line (less resilient). The HO line sows higher litter size and embryo development than the HE line. The aim of this work is to investigate the plasma organic acid profile in both lines at mating and early gestation in order to analyze the effect of selection by resilience in the ovulation rate and early gestation. A total of 19 and 18 nonlactating multiparous females from the HE and HO lines were used. The ovulation rate, normal embryos, and percentage of compacted morulae at 72 h post-coitum (hpc) were studied, and blood samples were obtained at mating and 72 hpc. The organic profile was determined by HPLC. Bayesian methodology was used for statistical analysis. The HE line had 1.5% fewer normal embryos and 12.3% fewer compacted morulae than the HO line. The ovulation rate was similar in both lines. α-ketoglutaric acid and cis-aconitic acid were higher in the HE line than in the HO line. Citric acid, lactic acid, and pyruvic acid were higher at mating than at early gestation. In conclusion, the lower efficiency in the utilization of energy sources in the HE line could explain the reduced embryo production observed. The organic profile varies depending on the reproductive state in the female. Full article