Targeting Metabolic Dependencies Fueling the TCA Cycle to Circumvent Therapy Resistance in Acute Myeloid Leukemia.

Acute myeloid leukemia (AML) is one of the most prevalent blood cancers, characterized by a dismal survival rate. This poor outcome is largely attributed to AML cells that persist despite treatment and eventually result in relapse. Relapse-initiating cells exhibit diverse resistance mechanisms, encompassing genetic factors and, more recently discovered, nongenetic factors such as metabolic adaptations. Leukemic stem cells (LSC) rely on mitochondrial metabolism for their survival, whereas hematopoietic stem cells primarily depend on glycolysis. Furthermore, following treatments such as cytarabine, a standard in AML treatment for over four decades, drug-persisting leukemic cells exhibit an enhanced reliance on mitochondrial metabolism. In this issue of Cancer Research, two studies investigated dependencies of AML cells on two respiratory substrates, α-ketoglutarate and lactate-derived pyruvate, that support mitochondrial oxidative phosphorylation (OXPHOS) following treatment with the imipridone ONC-213 and the BET inhibitor INCB054329, respectively. Targeting lactate utilization by interfering with monocarboxylate transporter 1 (MCT1 or SLC16A1) or lactate dehydrogenase effectively sensitized cells to BET inhibition in vitro and in vivo. In addition, ONC-213 affected αKGDH, a pivotal NADH-producing enzyme of the TCA cycle, to induce a mitochondrial stress response through ATF4 activation that diminished the expression of the antiapoptotic protein MCL1, consequently promoting apoptosis of AML cells. In summary, targeting these mitochondrial dependencies might be a promising strategy to kill therapy-naïve and treatment-resistant OXPHOS-reliant LSCs and to delay or prevent relapse. See related articles by Monteith et al., p. 1101 and Su et al., p. 1084.

Wild-type IDH2 is a therapeutic target for triple-negative breast cancer.

Mutations in isocitrate dehydrogenases (IDH) are oncogenic events due to the generation of oncogenic metabolite 2-hydroxyglutarate. However, the role of wild-type IDH in cancer development remains elusive. Here we show that wild-type IDH2 is highly expressed in triple negative breast cancer (TNBC) cells and promotes their proliferation in vitro and tumor growth in vivo. Genetic silencing or pharmacological inhibition of wt-IDH2 causes a significant increase in α-ketoglutarate (α-KG), indicating a suppression of reductive tricarboxylic acid (TCA) cycle. The aberrant accumulation of α-KG due to IDH2 abrogation inhibits mitochondrial ATP synthesis and promotes HIF-1α degradation, leading to suppression of glycolysis. Such metabolic double-hit results in ATP depletion and suppression of tumor growth, and renders TNBC cells more sensitive to doxorubicin treatment. Our study reveals a metabolic property of TNBC cells with active utilization of glutamine via reductive TCA metabolism, and suggests that wild-type IDH2 plays an important role in this metabolic process and could be a potential therapeutic target for TNBC.

Functional synergy of a human-specific and an ape-specific metabolic regulator in human neocortex development.

Metabolism has recently emerged as a major target of genes implicated in the evolutionary expansion of human neocortex. One such gene is the human-specific gene ARHGAP11B. During human neocortex development, ARHGAP11B increases the abundance of basal radial glia, key progenitors for neocortex expansion, by stimulating glutaminolysis (glutamine-to-glutamate-to-alpha-ketoglutarate) in mitochondria. Here we show that the ape-specific protein GLUD2 (glutamate dehydrogenase 2), which also operates in mitochondria and converts glutamate-to-αKG, enhances ARHGAP11B's ability to increase basal radial glia abundance. ARHGAP11B + GLUD2 double-transgenic bRG show increased production of aspartate, a metabolite essential for cell proliferation, from glutamate via alpha-ketoglutarate and the TCA cycle. Hence, during human evolution, a human-specific gene exploited the existence of another gene that emerged during ape evolution, to increase, via concerted changes in metabolism, progenitor abundance and neocortex size.

Immune patterns of cuproptosis in ischemic heart failure: A transcriptome analysis.

Cuproptosis is a recently discovered programmed cell death pattern that affects the tricarboxylic acid (TCA) cycle by disrupting the lipoylation of pyruvate dehydrogenase (PDH) complex components. However, the role of cuproptosis in the progression of ischemic heart failure (IHF) has not been investigated. In this study, we investigated the expression of 10 cuproptosis-related genes in samples from both healthy individuals and those with IHF. Utilizing these differential gene expressions, we developed a risk prediction model that effectively distinguished healthy and IHF samples. Furthermore, we conducted a comprehensive evaluation of the association between cuproptosis and the immune microenvironment in IHF, encompassing infiltrated immunocytes, immune reaction gene-sets and human leukocyte antigen (HLA) genes. Moreover, we identified two different cuproptosis-mediated expression patterns in IHF and explored the immune characteristics associated with each pattern. In conclusion, this study elucidates the significant influence of cuproptosis on the immune microenvironment in ischemic heart failure (IHF), providing valuable insights for future mechanistic research exploring the association between cuproptosis and IHF.

Tricarboxylic Acid Cycle Intermediates and Individual Ageing.

Anti-ageing biology and medicine programmes are a focus of genetics, molecular biology, immunology, endocrinology, nutrition, and therapy. This paper discusses metabolic therapies aimed at prolonging longevity and/or health. Individual components of these effects are postulated to be related to the energy supply by tricarboxylic acid (TCA) cycle intermediates and free radical production processes. This article presents several theories of ageing and clinical descriptions of the top markers of ageing, which define ageing in different categories; additionally, their interactions with age-related changes and diseases related to α-ketoglutarate (AKG) and succinate SC formation and metabolism in pathological states are explained. This review describes convincingly the differences in the mitochondrial characteristics of energy metabolism in animals, with different levels (high and low) of physiological reactivity of functional systems related to the state of different regulatory systems providing oxygen-dependent processes. Much attention is given to the crucial role of AKG and SC in the energy metabolism in cells related to amino acid synthesis, epigenetic regulation, cell stemness, and differentiation, as well as metabolism associated with the development of pathological conditions and, in particular, cancer cells. Another goal was to address the issue of ageing in terms of individual characteristics related to physiological reactivity. This review also demonstrated the role of the Krebs cycle as a key component of cellular energy and ageing, which is closely associated with the development of various age-related pathologies, such as cancer, type 2 diabetes, and cardiovascular or neurodegenerative diseases where the mTOR pathway plays a key role. This article provides postulates of postischaemic phenomena in an ageing organism and demonstrates the dependence of accelerated ageing and age-related pathology on the levels of AKG and SC in studies on different species (roundworm Caenorhabditis elegans, Drosophila, mice, and humans used as models). The findings suggest that this approach may also be useful to show that Krebs cycle metabolites may be involved in age-related abnormalities of the mitochondrial metabolism and may thus induce epigenetic reprogramming that contributes to the senile phenotype and degenerative diseases. The metabolism of these compounds is particularly important when considering ageing mechanisms connected with different levels of initial physiological reactivity and able to initiate individual programmed ageing, depending on the intensity of oxygen consumption, metabolic peculiarities, and behavioural reactions.

Pyrimidines maintain mitochondrial pyruvate oxidation to support de novo lipogenesis.

Cellular purines, particularly adenosine 5'-triphosphate (ATP), fuel many metabolic reactions, but less is known about the direct effects of pyrimidines on cellular metabolism. We found that pyrimidines, but not purines, maintain pyruvate oxidation and the tricarboxylic citric acid (TCA) cycle by regulating pyruvate dehydrogenase (PDH) activity. PDH activity requires sufficient substrates and cofactors, including thiamine pyrophosphate (TPP). Depletion of cellular pyrimidines decreased TPP synthesis, a reaction carried out by TPP kinase 1 (TPK1), which reportedly uses ATP to phosphorylate thiamine (vitamin B1). We found that uridine 5'-triphosphate (UTP) acts as the preferred substrate for TPK1, enabling cellular TPP synthesis, PDH activity, TCA-cycle activity, lipogenesis, and adipocyte differentiation. Thus, UTP is required for vitamin B1 utilization to maintain pyruvate oxidation and lipogenesis.

A citric acid cycle-deficient Escherichia coli as an efficient chassis for aerobic fermentations.

Tricarboxylic acid cycle (TCA cycle) plays an important role for aerobic growth of heterotrophic bacteria. Theoretically, eliminating TCA cycle would decrease carbon dissipation and facilitate chemicals biosynthesis. Here, we construct an E. coli strain without a functional TCA cycle that can serve as a versatile chassis for chemicals biosynthesis. We first use adaptive laboratory evolution to recover aerobic growth in minimal medium of TCA cycle-deficient E. coli. Inactivation of succinate dehydrogenase is a key event in the evolutionary trajectory. Supply of succinyl-CoA is identified as the growth limiting factor. By replacing endogenous succinyl-CoA dependent enzymes, we obtain an optimized TCA cycle-deficient E. coli strain. As a proof of concept, the strain is engineered for high-yield production of four separate products. This work enhances our understanding of the role of the TCA cycle in E. coli metabolism and demonstrates the advantages of using TCA cycle-deficient E. coli strain for biotechnological applications.

Tricarboxylic acid cycle metabolites: new players in macrophage.

Metabolic remodeling is a key feature of macrophage activation and polarization. Recent studies have demonstrated the role of tricarboxylic acid (TCA) cycle metabolites in the innate immune system. In the current review, we summarize recent advances in the metabolic reprogramming of the TCA cycle during macrophage activation and polarization and address the effects of these metabolites in modulating macrophage function. Deciphering the crosstalk between the TCA cycle and the immune response might provide novel potential targets for the intervention of immune reactions and favor the development of new strategies for the treatment of infection, inflammation, and cancer.

OGDH and Bcl-xL loss causes synthetic lethality in glioblastoma.

Glioblastoma (GBM) remains an incurable disease, requiring more effective therapies. Through interrogation of publicly available CRISPR and RNAi library screens, we identified the α-ketoglutarate dehydrogenase (OGDH) gene, which encodes an enzyme that is part of the tricarboxylic acid (TCA) cycle, as essential for GBM growth. Moreover, by combining transcriptome and metabolite screening analyses, we discovered that loss of function of OGDH by the clinically validated drug compound CPI-613 was synthetically lethal with Bcl-xL inhibition (genetically and through the clinically validated BH3 mimetic, ABT263) in patient-derived xenografts as well neurosphere GBM cultures. CPI-613-mediated energy deprivation drove an integrated stress response with an upregulation of the BH3-only domain protein, Noxa, in an ATF4-dependent manner, as demonstrated by genetic loss-of-function experiments. Consistently, silencing of Noxa attenuated cell death induced by CPI-613 in model systems of GBM. In patient-derived xenograft models of GBM in mice, the combination treatment of ABT263 and CPI-613 suppressed tumor growth and extended animal survival more potently than each compound on its own. Therefore, combined inhibition of Bcl-xL along with disruption of the TCA cycle might be a treatment strategy for GBM.

Search progress of pyruvate kinase M2 (PKM2) in organ fibrosis.

Fibrosis is characterized by abnormal deposition of the extracellular matrix (ECM), leading to organ structural remodeling and loss of function. The principal cellular effector in fibrosis is activated myofibroblasts, which serve as the main source of matrix proteins. Metabolic reprogramming, transitioning from mitochondrial oxidative phosphorylation to aerobic glycolysis, is widely observed in rapidly dividing cells such as tumor cells and activated myofibroblasts and is increasingly recognized as a fundamental pathogenic basis in organ fibrosis. Targeting metabolism represents a promising strategy to mitigate fibrosis. PKM2, a key enzyme in glycolysis, plays a pivotal role in metabolic reprogramming through allosteric regulation, impacting both metabolic and non-metabolic pathways. Therefore, metabolic reprogramming induced by PKM2 activation is involved in the occurrence and development of fibrosis in various organs. A comprehensive understanding of the role of PKM2 in fibrotic diseases is crucial for seeking new anti-fibrotic therapeutic targets. In this context, we summarize PKM2's role in glycolysis, mediating the intricate mechanisms underlying fibrosis in multiple organs, and discuss the potential value of PKM2 inhibitors and allosteric activators in future clinical treatments, aiming to identify novel therapeutic targets for proliferative fibrotic diseases.

Pathological Interplay between Inflammation and Mitochondria Aggravates Glutamate Toxicity.

Mitochondrial dysfunction and glutamate toxicity are associated with neural disorders, including brain trauma. A review of the literature suggests that toxic and transmission actions of neuronal glutamate are spatially and functionally separated. The transmission pathway utilizes synaptic GluN2A receptors, rapidly released pool of glutamate, evoked release of glutamate mediated by Synaptotagmin 1 and the amount of extracellular glutamate regulated by astrocytes. The toxic pathway utilizes extrasynaptic GluN2B receptors and a cytoplasmic pool of glutamate, which results from the spontaneous release of glutamate mediated by Synaptotagmin 7 and the neuronal 2-oxoglutarate dehydrogenase complex (OGDHC), a tricarboxylic acid (TCA) cycle enzyme. Additionally, the inhibition of OGDHC observed upon neuro-inflammation is due to an excessive release of reactive oxygen/nitrogen species by immune cells. The loss of OGDHC inhibits uptake of glutamate by mitochondria, thus facilitating its extracellular accumulation and stimulating toxic glutamate pathway without affecting transmission. High levels of extracellular glutamate lead to dysregulation of intracellular redox homeostasis and cause ferroptosis, excitotoxicity, and mitochondrial dysfunction. The latter affects the transmission pathway demanding high-energy supply and leading to cell death. Mitochondria aggravate glutamate toxicity due to impairments in the TCA cycle and become a victim of glutamate toxicity, which disrupts oxidative phosphorylation. Thus, therapies targeting the TCA cycle in neurological disorders may be more efficient than attempting to preserve mitochondrial oxidative phosphorylation.

FLVCR1a Controls Cellular Cholesterol Levels through the Regulation of Heme Biosynthesis and Tricarboxylic Acid Cycle Flux in Endothelial Cells.

Feline leukemia virus C receptor 1a (FLVCR1a), initially identified as a retroviral receptor and localized on the plasma membrane, has emerged as a crucial regulator of heme homeostasis. Functioning as a positive regulator of δ-aminolevulinic acid synthase 1 (ALAS1), the rate-limiting enzyme in the heme biosynthetic pathway, FLVCR1a influences TCA cycle cataplerosis, thus impacting TCA flux and interconnected metabolic pathways. This study reveals an unexplored link between FLVCR1a, heme synthesis, and cholesterol production in endothelial cells. Using cellular models with manipulated FLVCR1a expression and inducible endothelial-specific Flvcr1a-null mice, we demonstrate that FLVCR1a-mediated control of heme synthesis regulates citrate availability for cholesterol synthesis, thereby influencing cellular cholesterol levels. Moreover, alterations in FLVCR1a expression affect membrane cholesterol content and fluidity, supporting a role for FLVCR1a in the intricate regulation of processes crucial for vascular development and endothelial function. Our results underscore FLVCR1a as a positive regulator of heme synthesis, emphasizing its integration with metabolic pathways involved in cellular energy metabolism. Furthermore, this study suggests that the dysregulation of heme metabolism may have implications for modulating lipid metabolism. We discuss these findings in the context of FLVCR1a's potential heme-independent function as a choline importer, introducing additional complexity to the interplay between heme and lipid metabolism.

[Immunometabolism in Sepsis]. / Immunmetabolismus in der Sepsis.

Immunometabolism is a fascinating field of research that investigates the interactions between metabolic processes and the immune response. This intricate connection plays a pivotal role in regulating inflammatory reactions and consequently exerts a significant impact on the course of sepsis. The proinflammatory response during an immune reaction is closely tied to a high energy demand in immune cells. As a result, proinflammatory immune cells rapidly require substantial amounts of energy in the form of ATP, necessitating a fundamental and swift shift in their metabolism, i.e., their means of generating energy. This entails a marked increase in glycolysis within the proinflammatory response, thereby promptly meeting the energy requirements and providing essential metabolic building blocks for the biosynthesis of macromolecules. Alongside glycolysis, there is heightened activity in the pentose phosphate pathway (PPP). The PPP significantly contributes to NADPH production within the cell, thus maintaining redox equilibrium. Elevated PPP activity consequently leads to an increased NADPH level, resulting in enhanced production of reactive oxygen species (ROS) and nitric oxide (NO). While these molecules are crucial for pathogen elimination, an excess can also induce tissue damage. Simultaneously, there are dual interruptions in the citric acid cycle. In the cellular resting state, the citric acid cycle acts as a sort of "universal processor", where metabolic byproducts of glycolysis, fatty acid breakdown, and amino acid degradation are initially transformed into NADH and FADH2, subsequently yielding ATP. While the citric acid cycle and its connected oxidative phosphorylation predominantly generate energy at rest, it becomes downregulated in the proinflammatory phase of sepsis. The two interruptions lead to an accumulation of citrate and succinate within cells, reflecting mitochondrial dysfunction. Additionally, the significantly heightened glycolysis through fermentation yields lactate, a pivotal metabolite for sepsis diagnosis and prognosis. Conversely, cells in an anti-inflammatory state revert to a metabolic profile akin to the resting state: Glycolysis is attenuated, PPP is suppressed, and the citric acid cycle is reactivated. Of particular interest is that not only does the immune reaction influence metabolic pathways, but this connection also operates in reverse. Thus, modulation of metabolic pathways also modulates the immunity of the corresponding cell and thereby the state of the immune system itself. This could potentially serve as an intriguing avenue in sepsis therapy.

Energy metabolism: a new target for gastric cancer treatment

Gastric cancer is the fifth most common malignancy worldwide having the fourth highest mortality rate. Energy metabolism is key and closely linked to tumour development. Most important in the reprogramming of cancer metabolism is the Warburg effect, which suggests that tumour cells will utilise glycolysis even with normal oxygen levels. Various molecules exert their effects by acting on enzymes in the glycolytic pathway, integral to glycolysis. Second, mitochondrial abnormalities in the reprogramming of energy metabolism, with consequences for glutamine metabolism, the tricarboxylic acid cycle and oxidative phosphorylation, abnormal fatty acid oxidation and plasma lipoprotein metabolism are important components of tumour metabolism. Third, inflammation-induced oxidative stress is a danger signal for cancer. Fourth, patterns of signalling pathways involve all aspects of metabolic transduction, and many clinical drugs exert their anticancer effects through energy metabolic signalling. This review summarises research on energy metabolism genes, enzymes and proteins and transduction pathways associated with gastric cancer, and discusses the mechanisms affecting their effects on postoperative treatment resistance and prognoses of gastric cancer. We believe that an in-depth understanding of energy metabolism reprogramming will aid the diagnosis and subsequent treatment of gastric cancer (AU)

Optimization of the quantitative protocol for organic acid in fecal samples using gas chromatography-mass spectrometry.

Organic acids (OAs) play important roles in a variety of intracellular metabolic pathways, such as the tricarboxylic acid cycle, fatty acid oxidation, glycolysis. The accurate detection of OAs in fecal samples was crucial for comprehending the metabolic changes associated with various metabolic disease. However, the analytical protocol detecting OAs profiling in feces have received scant attention. In this work, an optimized protocol based on chromatography-mass spectrometry for simultaneous quantification of 23 OAs in rat feces was developed. The optimal conditions involved using a 40-mg fecal sample mixed with isopropyl alcohol, acetonitrile, and deionized water (3:2:2 vol ratio) with a total volume of 1500 µL, followed by ultrasonic extraction and a derivatization reaction with an 80 µL derivative agent. The protocol showed an acceptable linearity (R2 ≥ 0.9906), the satisfactory precision (RSD% ≤ 14.87%), the low limits of detection (0.001 to 1 µg/mL) and the limit of quantification (0.005 to 1.5 µg/mL). Moreover, the dried residues of the extracted solution showed the better stability of OAs at -20 °C, which was more suitable for a large-scale sample analysis. Finally, the developed protocol was successfully applied to compare the difference of OAs profiling in fecal samples harvested from normal and nonalcoholic fatty liver disease rats, which was beneficial to find out the metabolic change of OAs profiling and explain the related mechanism of the disease.

The role of promoter methylation of the genes encoding the enzymes metabolizing di- and tricarboxylic acids in the regulation of plant respiration by light.

We discuss the role of epigenetic changes at the level of promoter methylation of the key enzymes of carbon metabolism in the regulation of respiration by light. While the direct regulation of enzymes via modulation of their activity and post-translational modifications is fast and readily reversible, the role of cytosine methylation is important for providing a prolonged response to environmental changes. In addition, adenine methylation can play a role in the regulation of transcription of genes. The mitochondrial and extramitochondrial forms of several enzymes participating in the tricarboxylic acid cycle and associated reactions are regulated via promoter methylation in opposite ways. The mitochondrial forms of citrate synthase, aconitase, fumarase, NAD-malate dehydrogenase are inhibited while the cytosolic forms of aconitase, fumarase, NAD-malate dehydrogenase, and the peroxisomal form of citrate synthase are activated. It is concluded that promoter methylation represents a universal mechanism of the regulation of activity of respiratory enzymes in plant cells by light. The role of the regulation of the mitochondrial and cytosolic forms of respiratory enzymes in the operation of malate and citrate valves and in controlling the redox state and balancing the energy level of photosynthesizing plant cells is discussed.

Cananga oil inhibits Salmonella infection by mediating the homeostasis of purine metabolism and the TCA cycle.

ETHNOPHARMACOLOGY RELEVANCE: Cananga oil (CO) is derived from the flowers of the traditional medicinal plant, the ylang-ylang tree. As a traditional antidepressant, CO is commonly utilized in the treatment of various mental disorders including depression, anxiety, and autism. It is also recognized as an efficient antibacterial insecticide, and has been traditionally utilized to combat malaria and acute inflammatory responses resulting from bacterial infections both in vitro and in vivo. AIM OF THE STUDY: The objective of this study is to comprehensively investigate the anti-Salmonella activity and mechanism of CO both in vitro and in vivo, with the expectation of providing feasible strategies for exploring new antimicrobial strategies and developing novel drugs. METHODS: The in vitro antibacterial activity of CO was comprehensively analyzed by measuring MIC, MBC, growth curve, time-killing curve, surface motility, biofilm, and Live/dead bacterial staining. The analysis of the chemistry and active ingredients of CO was conducted using GC-MS. To examine the influence of CO on the membrane homeostasis of Salmonella, we conducted utilizing diverse techniques, including ANS, PI, NPN, ONPG, BCECF-AM, DiSC3(5), and scanning electron microscopy (SEM) analysis. In addition, the antibacterial mechanism of CO was analyzed and validated through metabolomics analysis. Finally, a mouse infection model of Salmonella typhimurium was established to evaluate the toxic side effects and therapeutic effects of CO. RESULTS: The antibacterial effect of CO is the result of the combined action of the main chemical components within its six (palmitic acid, α-linolenic acid, stearic acid, benzyl benzoate, benzyl acetate, and myristic acid). Furthermore, CO disrupts the balance of purine metabolism and the tricarboxylic acid cycle (TCA cycle) in Salmonella, interfering with redox processes. This leads to energy metabolic disorders and oxidative stress damage within the bacteria, resulting in bacterial shock, enhanced membrane damage, and ultimately bacterial death. It is worth emphasizing that CO exerts an effective protective influence on Salmonella infection in vivo within a non-toxic concentration range. CONCLUSION: The outcomes indicate that CO displays remarkable anti-Salmonella activity both in vitro and in vivo. It triggers bacterial death by disrupting the balance of purine metabolism and the TCA cycle, interfering with the redox process, making it a promising anti-Salmonella medication.

Preparation of [1'-13 C]citric acid as a probe in a breath test to evaluate tricarboxylic acid cycle flux.

[1'-13 C]Citric acid (1) was efficiently prepared from dimethyl 1,3-acetonedicarboxylate in two steps as a probe for a breath test. The synthetic method was selected because of the yield and reproducibility. Compound 1 was orally administrated to rats, and the time course of the increase of 13 CO2 /12 CO2 ratios (Δ13 CO2 ) in their breath was successfully followed, indicating the metabolism of 1. Thus, the 13 C-breath test using 1 is a promising method to evaluate tricarboxylic acid (TCA) cycle flux.

Tricarboxylic Acid Cycle Metabolite-Coordinated Biohydrogels Augment Cranial Bone Regeneration Through Neutrophil-Stimulated Mesenchymal Stem Cell Recruitment and Histone Acetylation-Mediated Osteogenesis.

Cranial bone defects remain a major clinical challenge, increasing patients' life burdens. Tricarboxylic acid (TCA) cycle metabolites play crucial roles in facilitating bone tissue regeneration. However, the development of TCA cycle metabolite-modified biomimetic grafts for skull bone regeneration still needs to be improved. The mechanism underlying the release of TCA cycle metabolites from biomaterials in regulating immune responses and mesenchymal stem cell (MSC) fate (migration and differentiation) remains unknown. Herein, this work constructs biomimetic hydrogels composed of gelatin and chitosan networks covalently cross-linked by genipin (CGG hydrogels). A series of TCA cycle metabolite-coordinated CGG hydrogels with strong mechanical and antiswelling performances are subsequently developed. Remarkably, the citrate (Na3Cit, Cit)-coordinated CGG hydrogels (CGG-Cit hydrogels) with the highest mechanical modulus and strength significantly promote skull bone regeneration in rat and murine cranial defects. Mechanistically, using a transgenic mouse model, bulk RNA sequencing, and single-cell RNA sequencing, this work demonstrates that CGG-Cit hydrogels promote Gli1+ MSC migration via neutrophil-secreted oncostatin M. Results also indicate that citrate improves osteogenesis via enhanced histone H3K9 acetylation on osteogenic master genes. Taken together, the immune microenvironment- and MSC fate-regulated CGG-Cit hydrogels represent a highly efficient and facile approach toward skull bone tissue regeneration with great potential for bench-to-bedside translation.

LINC00571 drives tricarboxylic acid cycle metabolism in triple-negative breast cancer through HNRNPK/ILF2/IDH2 axis.

BACKGROUND: Triple-negative breast cancer is a complex breast malignancy subtype characterized by poor prognosis. The pursuit of effective therapeutic approaches for this subtype is considerably challenging. Notably, recent research has illuminated the key role of the tricarboxylic acid cycle in cancer metabolism and the complex landscape of tumor development. Concurrently, an emerging body of evidence underscores the noteworthy role that long non-coding RNAs play in the trajectory of breast cancer development. Despite this growing recognition, the exploration of whether long non-coding RNAs can influence breast cancer progression by modulating the tricarboxylic acid cycle has been limited. Moreover, the underlying mechanisms orchestrating these interactions have not been identified. METHODS: The expression levels of LINC00571 and IDH2 were determined through the analysis of the public TCGA dataset, transcriptome sequencing, qRT‒PCR, and Western blotting. The distribution of LINC00571 was assessed using RNA fluorescence in situ hybridization. Alterations in biological effects were evaluated using CCK-8, colony formation, EdU, cell cycle, and apoptosis assays and a tumor xenograft model. To elucidate the interaction between LINC00571, HNRNPK, and ILF2, RNA pull-down, mass spectrometry, coimmunoprecipitation, and RNA immunoprecipitation assays were performed. The impacts of LINC00571 and IDH2 on tricarboxylic acid cycle metabolites were investigated through measurements of the oxygen consumption rate and metabolite levels. RESULTS: This study revealed the complex interactions between a novel long non-coding RNA (LINC00571) and tricarboxylic acid cycle metabolism. We validated the tumor-promoting role of LINC00571. Mechanistically, LINC00571 facilitated the interaction between HNRNPK and ILF2, leading to reduced ubiquitination and degradation of ILF2, thereby stabilizing its expression. Furthermore, ILF2 acted as a transcription factor to enhance the expression of its downstream target gene IDH2. CONCLUSIONS: Our study revealed that the LINC00571/HNRNPK/ILF2/IDH2 axis promoted the progression of triple-negative breast cancer by regulating tricarboxylic acid cycle metabolites. This discovery provides a novel theoretical foundation and new potential targets for the clinical treatment of triple-negative breast cancer.