Glycolytic Inhibitor 2-deoxyglucose Research Articles

Inherent metabolic preferences differentially regulate the sensitivity of Th1 and Th2 cells to ribosome-inhibiting antibiotics.

Mitochondrial translation is essential to maintain mitochondrial function and energy production. Mutations in genes associated with mitochondrial translation cause several developmental disorders, and immune dysfunction is observed in many such patients. Besides genetic mutations, several antibiotics targeting bacterial ribosomes are well-established to inhibit mitochondrial translation. However, the effect of such antibiotics on different immune cells is not fully understood. Here, we addressed the differential effect of mitochondrial translation inhibition on different subsets of helper T cells (Th) of mice and humans. Inhibition of mitochondrial translation reduced the levels of mitochondrially encoded electron transport chain subunits without affecting their nuclear-encoded counterparts. As a result, mitochondrial oxygen consumption reduced dramatically, but mitochondrial mass was unaffected. Most importantly, we show that inhibition of mitochondrial translation induced apoptosis, specifically in Th2 cells. This increase in apoptosis was associated with higher expression of Bim and Puma, two activators of the intrinsic pathway of apoptosis. We propose that this difference in the sensitivity of Th1 and Th2 cells to mitochondrial translation inhibition reflects the intrinsic metabolic demands of these subtypes. Though Th1 and Th2 cells exhibit similar levels of oxidative phosphorylation, Th1 cells exhibit higher levels of aerobic glycolysis than Th2 cells. Moreover, Th1 cells are more sensitive to the inhibition of glycolysis, while higher concentrations of glycolysis inhibitor 2-deoxyglucose are required to induce cell death in the Th2 lineage. These observations reveal that selection of metabolic pathways for substrate utilization during differentiation of Th1 and Th2 lineages is a fundamental process conserved across species.

A multiplexed high throughput screening assay using flow cytometry identifies glycolytic molecular probes in bloodstream form Trypanosoma brucei

Kinetoplastid organisms, including Trypanosoma brucei, are a significant health burden in many tropical and semitropical countries. Much of their metabolism is poorly understood. To better study kinetoplastid metabolism, chemical probes that inhibit kinetoplastid enzymes are needed. To discover chemical probes, we have developed a high-throughput flow cytometry screening assay that simultaneously measures multiple glycolysis-relevant metabolites in live T. brucei bloodstream form parasites. We transfected parasites with biosensors that measure glucose, ATP, or glycosomal pH. The glucose and ATP sensors were FRET biosensors, while the pH sensor was a GFP-based biosensor. The pH sensor exhibited a different fluorescent profile from the FRET sensors, allowing us to simultaneously measure pH and either glucose or ATP. Cell viability was measured in tandem with the biosensors using thiazole red. We pooled sensor cell lines, loaded them onto plates containing a compound library, and then analyzed them by flow cytometry. The library was analyzed twice, once with the pooled pH and glucose sensor cell lines and once with the pH and ATP sensor cell lines. Multiplexing sensors provided some internal validation of active compounds and gave potential clues for each compound's target(s). We demonstrated this using the glycolytic inhibitor 2-deoxyglucose and the alternative oxidase inhibitor salicylhydroxamic acid. Individual biosensor-based assays exhibited a Z′-factor value acceptable for high-throughput screening, including when multiplexed. We tested assay performance in a pilot screen of 14,976 compounds from the Life Chemicals Compound Library. We obtained hit rates from 0.2 to 0.4% depending on the biosensor, with many compounds impacting multiple sensors. We rescreened 44 hits, and 28 (64%) showed repeatable activity for one or more sensors. One compound exhibited EC50 values in the low micromolar range against two sensors. We expect this method will enable the discovery of glycolytic chemical probes to improve metabolic studies in kinetoplastid parasites.

N-Glycosylation of MRS2 balances aerobic and anaerobic energy production by reducing rapid mitochondrial Mg2+ influx in conditions of high glucose or impaired respiratory chain function.

N-linked glycoproteins function in numerous biological processes, modulating enzyme activities as well as protein folding, stability, oligomerization, and trafficking. While N-glycosylation of mitochondrial proteins has been detected by untargeted MS-analyses, the physiological existence and roles of mitochondrial protein N-linked glycosylation remain under debate. Here, we report that MRS2, a mitochondrial inner membrane protein that functions as the high flux magnesium transporter, is N-glycosylated to various extents depending on cellular bioenergetic status. Both N-glycosylated and unglycosylated isoforms were consistently detected in mitochondria isolated from mouse liver, rat and mouse liver fibroblast cells (BRL 3A and AFT024, respectively) as well as human skin fibroblast cells. Immunoblotting of MRS2 showed it was bound to, and required stringent elution conditions to remove from, lectin affinity columns with covalently bound concanavalin A or Lens culinaris agglutinin. Following peptide:N-glycosidase F (PNGase F) digestion of the stringently eluted proteins, the higher Mr MRS2 bands gel-shifted to lower Mr and loss of lectin affinity was seen. BRL 3A cells treated with two different N-linked glycosylation inhibitors, tunicamycin or 6-diazo-5-oxo-l-norleucine, resulted in decreased intensity or loss of the higher Mr MRS2 isoform. To investigate the possible functional role of MRS2 N- glycosylation, we measured rapid Mg2+ influx capacity in intact mitochondria isolated from BRL 3A cells in control media or following treatment with tunicamycin or 6-diazo-5-oxo-l-norleucine. Interestingly, rapid Mg2+ influx capacity increased in mitochondria isolated from BRL 3A cells treated with either N-glycosylation inhibitor. Forcing reliance on mitochondrial respiration by treatment with either galactose media or the glycolytic inhibitor 2-deoxyglucose or by minimizing glucose concentration similarly reduced the N-glycosylated isoform of MRS2, with a correlated concomitant increase in rapid Mg2+ influx capacity. Conversely, inhibiting mitochondrial energy production in BRL 3A cells with either rotenone or oligomycin resulted in an increased fraction of N-glycosylated MRS2, with decreased rapid Mg2+ influx capacity. Collectively, these data provide strong evidence that MRS2 N-glycosylation is directly involved in the regulation of mitochondrial matrix Mg2+, dynamically communicating relative cellular nutrient status and bioenergetic capacity by serving as a physiologic brake on the influx of mitochondrial matrix Mg2+ under conditions of glucose excess or mitochondrial bioenergetic impairment.

The troglitazone derivative EP13 disrupts energy metabolism through respiratory chain complex I inhibition in breast cancer cells and potentiates the antiproliferative effect of glycolysis inhibitors

BackgroundThe metabolism of cancer cells generally differs from that of normal cells. Indeed, most cancer cells have a high rate of glycolysis, even at normal oxygen concentrations. These metabolic properties can potentially be exploited for therapeutic intervention. In this context, we have developed troglitazone derivatives to treat hormone-sensitive and triple-negative breast cancers, which currently lack therapeutic targets, have an aggressive phenotype, and often have a worse prognosis than other subtypes. Here, we studied the metabolic impact of the EP13 compound, a desulfured derivative of Δ2-troglitazone that we synthetized and is more potent than its parent compounds.MethodsEP13 was tested on two triple-negative breast cancer cell lines, MDA-MB-231 and Hs578T, and on the luminal cell line MCF-7. The oxygen consumption rate (OCR) of the treated cell lines, Hs578T mammospheres and isolated mitochondria was measured using the XFe24 Seahorse analyser. ROS production was quantified using the MitoSOX fluorescent probe. Glycolytic activity was evaluated through measurement of the extracellular acidification rate (ECAR), glucose consumption and lactate production in extracellular medium. The synergistic effect of EP13 with glycolysis inhibitors (oxamate and 2-deoxyglucose) on cell cytotoxicity was established using the Chou-Talalay method.ResultsAfter exposure to EP13, we observed a decrease in the mitochondrial oxygen consumption rate in MCF7, MDA-MB-231 and Hs578T cells. EP13 also modified the maximal OCR of Hs578T spheroids. EP13 reduced the OCR through inhibition of respiratory chain complex I. After 24 h, ATP levels in EP13-treated cells were not altered compared with those in untreated cells, suggesting compensation by glycolysis activity, as shown by the increase in ECAR, the glucose consumption and lactate production. Finally, we performed co-treatments with EP13 and glycolysis inhibitors (oxamate and 2-DG) and observed that EP13 potentiated their cytotoxic effects.ConclusionThis study demonstrates that EP13 inhibits OXPHOS in breast cancer cells and potentiates the effect of glycolysis inhibitors.

Abstract A023: High glucose increases DNA damage and elevates the expression of multiple DDR genes

Abstract The DNA Damage Response (DDR) pathways sense DNA damage and coordinate robust DNA repair and bypass mechanisms. A series of repair proteins are recruited depending on the type of breaks and lesions to ensure overall survival. An increase in glucose levels was shown to induce genome instability, yet the links between DDR and glucose are still not well investigated. In this study, we aimed to identify dysregulation in the transcriptome of normal and cancerous breast cell lines upon changing glucose levels. We first performed bioinformatics analysis using a microarray dataset containing the triple-negative breast cancer (TNBC) MDA-MB-231 and the normal human mammary epithelium MCF10A cell lines grown in high glucose (HG) or in the presence of the glycolysis inhibitor 2-deoxyglucose (2DG). Interestingly, multiple DDR genes were significantly upregulated in both cell lines grown in HG. In the wet lab, we remarkably found that HG results in severe DNA damage to TNBC cells as observed using the comet assay. In addition, several DDR genes were confirmed to be upregulated using qPCR analysis in the same cell line. Our results propose a strong need for DDR pathways in the presence of HG to oppose the severe DNA damage induced in cells. Citation Format: Mai A. Rahmoon, Reem A. Elghaish, Aya A. Ibrahim, Zina Alaswad, Mohamed Z. Gad, Sherif F. El-Khamisy, Menattallah Elserafy. High glucose increases DNA damage and elevates the expression of multiple DDR genes [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: DNA Damage Repair: From Basic Science to Future Clinical Application; 2024 Jan 9-11; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2024;84(1 Suppl):Abstract nr A023.

Uncoupling protein 2 modulates polarization and metabolism of human primary macrophages via glycolysis and the NF‑κB pathway.

Metabolic abnormalities, particularly the M1/M2 macrophage imbalance, play a critical role in the development of various diseases, leading to severe inflammatory responses. The present study aimed to investigate the role of uncoupling protein 2 (UCP2) in regulating macrophage polarization, glycolysis, metabolic reprogramming, reactive oxygen species (ROS) and inflammation. Primary human macrophages were first polarized into M1 and M2 subtypes, and these two subtypes were infected by lentivirus-mediated UCP2 overexpression or knockdown, followed by enzyme-linked immunosorbent assay, reverse transcription-quantitative PCR, western blotting and flow cytometry to analyze the effects of UCP2 on glycolysis, oxidative phosphorylation (OXPHOS), ROS production and cytokine secretion, respectively. The results demonstrated that UCP2 expression was suppressed in M1 macrophages and increased in M2 macrophages, suggesting its regulatory role in macrophage polarization. UCP2 overexpression decreased macrophage glycolysis, increased OXPHOS, decreased ROS production, and led to the conversion of M1 polarization to M2 polarization. This process involved NF-κB signaling to regulate the secretion profile of cytokines and chemokines and affected the expression of key enzymes of glycolysis and a key factor for maintaining mitochondrial homeostasis (nuclear respiratory factor 1). UCP2 knockdown in M2 macrophages exacerbated inflammation and oxidative stress by promoting glycolysis, which was attenuated by the glycolysis inhibitor 2-deoxyglucose. These findings highlight the critical role of UCP2 in regulating macrophage polarization, metabolism, inflammation and oxidative stress through its effects on glycolysis, providing valuable insights into potential therapeutic strategies for macrophage-driven inflammatory and metabolic diseases.

P53-responsive CMBL reprograms glucose metabolism and suppresses cancer development by destabilizing phosphofructokinase PFKP

Aerobic glycolysis is critical for cancer progression and can be exploited in cancer therapy. Here, we report that the human carboxymethylenebutenolidase homolog (carboxymethylenebutenolidase-like [CMBL]) acts as a tumor suppressor by reprogramming glycolysis in colorectal cancer (CRC). The anti-cancer action of CMBL is mediated through its interactions with the E3 ubiquitin ligase TRIM25 and the glycolytic enzyme phosphofructokinase-1 platelet type (PFKP). Ectopic CMBL enhances TRIM25 binding to PFKP, leading to the ubiquitination and proteasomal degradation of PFKP. Interestingly, CMBL is transcriptionally activated by p53 in response to genotoxic stress, and p53 activation represses glycolysis by promoting PFKP degradation. Remarkably, CMBL deficiency, which impairs p53's ability to inhibit glycolysis, makes tumors more sensitive to a combination therapy involving the glycolysis inhibitor 2-deoxyglucose. Taken together, our study demonstrates that CMBL suppresses CRC growth by inhibiting glycolysis and suggests a potential combination strategy for the treatment of CMBL-deficient CRC.

Inhibition of thioredoxin-1 enhances the toxicity of glycolysis inhibitor 2-deoxyglucose by downregulating SLC1A5 expression in colorectal cancer cells.

Targeting glycolysis in cancer is an attractive approach for therapeutic intervention. 2-Deoxyglucose (2DG) is a synthetic glucose analog that inhibits glycolysis. However, its efficacy is limited by the systemic toxicity at high doses. Understanding the mechanism of 2DG resistance is important for further use of this drug in cancer treatment. The expression of thioredoxin-1 (Trx-1) in colorectal cancer (CRC) cells treated with 2DG was detected by Western blotting. The effect of Trx-1 on the cytotoxicity of 2DG in CRC cells was examined in vitro and in vivo. The molecular mechanism involved in Trx-1-mediated activation of the SLC1A5 gene promoter activity was elucidated using in vitro models. Inhibition glycolysis with 2DG increased the expression of Trx-1 in CRC cells. Overexpression of Trx-1 decreased the cytotoxicity of 2DG, whereas knockdown of Trx-1 by shRNA significantly increased the cytotoxicity of 2DG in CRC cells. The Trx-1 inhibitor PX-12 increased the cytotoxicity of 2DG on CRC cells both in vitro and in vivo. In addition, Trx-1 promoted SLC1A5 expression by increasing the promoter activity of the SLC1A5 gene by binding to SP1. We also found that the SLC1A5 expression was upregulated in CRC tissues, and inhibition of SLC1A5 significantly enhanced the inhibitory effect of 2DG on the growth of CRC cells in vitro and in vivo. Overexpression of SLC1A5 reduced the cytotoxicity of 2DG in combination with PX-12 treatment in CRC cells. Our results demonstrate a novel adaptive mechanism of glycolytic inhibition in which Trx-1 increases GSH levels by regulating SLC1A5 to rescue cytotoxicity induced by 2DG in CRC cells. Inhibition of glycolysis in combination with inhibition of Trx-1 or SLC1A5 may be a promising strategy for the treatment of CRC.

Modulation of Cellular Levels of Adenosine Phosphates and Creatine Phosphate in Cultured Primary Astrocytes

Adenosine triphosphate (ATP) is the main energy currency of all cells, while creatine phosphate (CrP) is considered as a buffer of high energy-bond phosphate that facilitates rapid regeneration of ATP from adenosine diphosphate (ADP). Astrocyte-rich primary cultures contain ATP, ADP and adenosine monophosphate (AMP) in average specific contents of 36.0 ± 6.4 nmol/mg, 2.9 ± 2.1 nmol/mg and 1.7 ± 2.1 nmol/mg, respectively, which establish an adenylate energy charge of 0.92 ± 0.04. The average specific cellular CrP level was found to be 25.9 ± 10.8 nmol/mg and the CrP/ATP ratio was 0.74 ± 0.28. The specific cellular CrP content, but not the ATP content, declined with the age of the culture. Absence of fetal calf serum for 24 h caused a partial loss in the cellular contents of both CrP and ATP, while application of creatine for 24 h doubled the cellular CrP content and the CrP/ATP ratio, but did not affect ATP levels. In glucose-deprived astrocytes, the high cellular ATP and CrP contents were rapidly depleted within minutes after application of the glycolysis inhibitor 2-deoxyglucose and the respiratory chain inhibitor antimycin A. For those conditions, the decline in CrP levels always preceded that of ATP contents. In contrast, incubation of glucose-fed astrocytes for up to 30 min with antimycin A had little effect on the high cellular ATP content, while the CrP level was significantly lowered. These data demonstrate the importance of cellular CrP for maintaining a high cellular ATP content in astrocytes during episodes of impaired ATP regeneration.

Dexamethasone inhibits IL-8 via glycolysis and mitochondria-related pathway to regulate inflammatory pain

BackgroundDexamethasone (Dexa) has been recently found to exert an analgesic effect, whose action is closely related to IL-8. However, whether dexamethasone induces antinociception via glycolysis and mitochondria-related pathways is still unclear.MethodsRight hind paw inflammatory pain in mice was induced by intraplantar injection of Freund’s Complete Adjuvant (FCA). Von Frey test was then used to measure the paw withdrawal threshold. The detection of glycolysis and mitochondrial pathway-related proteins and IL-8 were determined by Western blot and ELISA. The potential interaction between Dexa and fructose-1,6-bisphosphate (FBP, a PKM2 activator) was examined by simulation predictions using molecular docking.ResultsIntrathecal administration of Dexa (20 µg/20 µL) had an obvious analgesic effect in FCA-treated mice, which was counteracted by the glycolysis inhibitor 2-deoxyglucose (2-DG, 5 mg/20 µL) or the mitochondria-related pathway inhibitor oligomycin complex (Oligo, 5 µg/20 µL). In the glycolysis pathway, Dexa decreased GLUT3 and had no impact on HIF-1α expression during FCA-induced inflammation. Additionally, Dexa further increased the PKM2 level, accompanied by the formation of hydrogen bonds between Dexa and the PKM2 activator fructose-1,6-bisphosphate (FBP). In the mitochondrial pathway, Dexa downregulated the expression of Mfn2 protein but not the PGC-1α and SIRT-1 levels in the spinal cord. Moreover, both 2-DG and Oligo decreased Mfn2 expression. Finally, IL-8 level was reduced by the single or combined administration of Dexa, 2-DG, and Oligo.ConclusionDexa attenuated IL-8 expression via glycolysis and mitochondrial pathway-related proteins, thus mediating the analgesic effect during inflammatory pain.

DNA Aptamer Beacon Probe (ABP) for Monitoring of Adenosine Triphosphate Level in SW480 Cancer Cells Treated with Glycolysis Inhibitor 2-Deoxyglucose.

Early cancer screening enables timely detection of carcinogenesis, and aids in prompt clinical intervention. Herein, we report on the development of a simple, sensitive, and rapid fluorometric assay based on the aptamer probe (aptamer beacon probe, ABP) for monitoring the energy-demand biomarker adenosine triphosphate (ATP), an essential energy source that is released into the tumor microenvironment. Its level plays a significant role in risk assessment of malignancies. The operation of the ABP for ATP was examined using solutions of ATP and other nucleotides (UTP, GTP, CTP), followed by monitoring of ATP production in SW480 cancer cells. Then, the effect of a glycolysis inhibitor, 2-deoxyglucose (2-DG), on SW480 cells was investigated. The stability of predominant ABP conformations in the temperature range of 23-91 °C and the effects of temperature on ABP interactions with ATP, UTP, GTP, and CTP were evaluated based on quenching efficiencies (QE) and Stern-Volmer constants (KSV). The optimized temperature for best selectivity of ABP toward ATP was 40 °C (KSV = 1093 M-1, QE = 42%). We have found that the inhibition of glycolysis in SW480 cancer cells by 2-deoxyglucose resulted in lowering of ATP production by 31.7%. Therefore, monitoring and modulation of ATP concentration may aid in future cancer treatment.

C/EBPβ-LIP mediated activation of the malate-aspartate shuttle sensitizes cells to glycolysis inhibition

ObjectiveCancer cells use glycolysis for generation of metabolic intermediates and ATP needed for cell growth and proliferation. The transcription factor C/EBPβ-LIP stimulates glycolysis and mitochondrial respiration in cancer cells. We initially observed that high expression of C/EBPβ-LIP makes cells vulnerable to treatment with the glycolysis inhibitor 2-deoxyglucose. The aim of the study was to uncover the involved mechanisms of C/EBPβ-LIP induced sensitivity to glycolysis inhibition. MethodsWe used genetically engineered cell lines to examine the effect of C/EBPβ-LIP and -LAP protein isoforms on glycolysis and NADH/NAD+ metabolism in mouse embryonic fibroblasts (MEFs), and triple negative breast cancer (TNBC) cells that endogenously express high levels of C/EBPβ-LIP. Analyses included assays of cell proliferation, cell survival and metabolic flux (OCR and ECAR by Seahorse XF96). Small molecule inhibitors were used to identify underlying metabolic pathways that mediate sensitivity to glycolysis inhibition induced by C/EBPβ-LIP. ResultsThe transcription factor C/EBPβ-LIP stimulates both glycolysis and the malate-aspartate shuttle (MAS) and increases the sensitivity to glycolysis inhibition (2-deoxyglucose) in fibroblasts and breast cancer cells. Inhibition of glycolysis with ongoing C/EBPβ-LIP-induced MAS activity results in NADH depletion and apoptosis that can be rescued by inhibiting either the MAS or other NAD+-regenerating processes. ConclusionThis study indicates that a low NADH/NAD+ ratio is an essential mediator of 2-deoxyglucose toxicity in cells with high cytoplasmic NAD+-regeneration capacity and that simultaneous inhibition of glycolysis and lowering of the NADH/NAD+ ratio may be considered to treat cancer.

Correction to: Glycolytic inhibitors 2-deoxyglucose and 3-bromopyruvate synergize with photodynamic therapy respectively to inhibit cell migration

Correction to: Glycolytic inhibitors 2-deoxyglucose and 3-bromopyruvate synergize with photodynamic therapy respectively to inhibit cell migration

High Glucose Increases DNA Damage and Elevates the Expression of Multiple DDR Genes.

The DNA Damage Response (DDR) pathways sense DNA damage and coordinate robust DNA repair and bypass mechanisms. A series of repair proteins are recruited depending on the type of breaks and lesions to ensure overall survival. An increase in glucose levels was shown to induce genome instability, yet the links between DDR and glucose are still not well investigated. In this study, we aimed to identify dysregulation in the transcriptome of normal and cancerous breast cell lines upon changing glucose levels. We first performed bioinformatics analysis using a microarray dataset containing the triple-negative breast cancer (TNBC) MDA-MB-231 and the normal human mammary epithelium MCF10A cell lines grown in high glucose (HG) or in the presence of the glycolysis inhibitor 2-deoxyglucose (2DG). Interestingly, multiple DDR genes were significantly upregulated in both cell lines grown in HG. In the wet lab, we remarkably found that HG results in severe DNA damage to TNBC cells as observed using the comet assay. In addition, several DDR genes were confirmed to be upregulated using qPCR analysis in the same cell line. Our results propose a strong need for DDR pathways in the presence of HG to oppose the severe DNA damage induced in cells.

Chronic stress promotes colorectal cancer progression by enhancing glycolysis through β2-AR/CREB1 signal pathway.

Colorectal cancer (CRC) is a common malignancy worldwide, and chronic stress has been considered as a significant risk factor for CRC. However, the role of chronic stress in CRC progression is unclear. The present study showed that pre-exposure to chronic stress facilitated CRC tumor growth in mice, and epinephrine promoted CRC cell proliferation in vitro. Metabolomics analysis revealed that chronic stress reshaped metabolic pathways to enhance glycolysis. Additional studies have shown that stress enhanced the expression levels of glycolytic-associated enzymes, including GLUT1, HK2 and PFKP. Mechanistically, chronic stress activated the β2-AR/PKA/CREB1 pathway, as a result, phosphorylated CREB1 transcriptional induced glycolytic enzymes expression. Furthermore, stress-induced cell proliferation and tumor growth could be reversed by administration of glycolysis inhibitor 2-deoxyglucose (2-DG) and β2-AR antagonist ICI118,551, respectively. Altogether, these findings define novel insights into the stress-induced epinephrine-mediated CRC progression from the point of view of tumor energy metabolism reprogramming and provide a perspective on targeting glycolysis as a potential approach in stress-associated CRC treatment.

The role of SH groups in the regulation of Gardos channels in glucose deficiency

Background. Disruption of the energy balance of erythrocytes under conditions of a decrease in the glycolysis level can cause a change in the ion permeability of their membrane. The aim. To study Ca2+-dependent potassium permeability of the erythrocytes membrane in the presence of SH group modifiers under conditions of glucose deficiency. Materials and methods. The study used precipitated erythrocytes obtained from the blood of 20 male Wistar rats. The change in the Ca2+-dependent potassium conductivity of the erythrocyte membrane was determined using the potentiometric method. The A23187-and redox-induced hyperpolarization responses of erythrocytes were evaluated. Results. Glucose deficiency in the medium, as well as the use of the glycolysis inhibitor 2-deoxyglucose, led to an increase in the amplitude of A23187-stimulated membrane hyperpolarization by the opening of the Gardos channels. At the same time, the redox-dependent hyperpolarization of the erythrocyte membrane turned out to be insensitive to a decrease in the glucose content in the medium and to the glycolysis inhibition. The effects of SH group modifiers in the normal incubation medium and under glucose deficiency turned out to be multidirectional and depended on the method of stimulation of Gardos channels. Conclusion. The results obtained indicate that metabolic disorders in erythrocytes under conditions of glucose deficiency lead to a change in the mechanisms of control of Gardos channels with the participation of SH groups of the proteins of these channels or their regulatory proteins.

GM3 synthase deficiency increases brain glucose metabolism in mice.

GM3 synthase (GM3S) deficiency is a rare neurodevelopmental disorder caused by an inability to synthesize gangliosides, for which there is currently no treatment. Gangliosides are brain-enriched, plasma membrane glycosphingolipids with poorly understood biological functions related to cell adhesion, growth, and receptor-mediated signal transduction. Here, we investigated the effects of GM3S deficiency on metabolism and mitochondrial function in a mouse model. By indirect calorimetry, GM3S knockout mice exhibited increased whole-body respiration and an increased reliance upon carbohydrate as an energy source. 18F-FDG PET confirmed higher brain glucose uptake in knockout mice, and GM3S deficient N41 neuronal cells showed higher glucose utilization in vitro. Brain mitochondria from knockout mice respired at a higher rate on Complex I substrates including pyruvate. This appeared to be due to higher expression of pyruvate dehydrogenase (PDH) and lower phosphorylation of PDH, which would favor pyruvate entry into the mitochondrial TCA cycle. Finally, it was observed that blocking glucose metabolism with the glycolysis inhibitor 2-deoxyglucose reduced seizure intensity in GM3S knockout mice following administration of kainate. In conclusion, GM3S deficiency may be associated with a hypermetabolic phenotype that could promote seizure activity.

Glycolytic Inhibitors Potentiated the Activity of Paclitaxel and Their Nanoencapsulation Increased Their Delivery in a Lung Cancer Model.

Antiglycolytic agents inhibit cell metabolism and modify the tumor’s microenvironment, affecting chemotherapy resistance mechanisms. In this work, we studied the effect of the glycolytic inhibitors 3-bromopyruvate (3BP), dichloroacetate (DCA) and 2-deoxyglucose (2DG) on cancer cell properties and on the multidrug resistance phenotype, using lung cancer cells as a model. All compounds led to the loss of cell viability, with different effects on the cell metabolism, migration and proliferation, depending on the drug and cell line assayed. DCA was the most promising compound, presenting the highest inhibitory effect on cell metabolism and proliferation. DCA treatment led to decreased glucose consumption and ATP and lactate production in both A549 and NCI-H460 cell lines. Furthermore, the DCA pretreatment sensitized the cancer cells to Paclitaxel (PTX), a conventional chemotherapeutic drug, with a 2.7-fold and a 10-fold decrease in PTX IC50 values in A549 and NCI-H460 cell lines, respectively. To increase the intracellular concentration of DCA, thereby potentiating its effect, DCA-loaded poly(lactic-co-glycolic acid) nanoparticles were produced. At higher DCA concentrations, encapsulation was found to increase its toxicity. These results may help find a new treatment strategy through combined therapy, which could open doors to new treatment approaches.

Helicobacter pylori-mediated gastric pathogenesis is attenuated by treatment of 2-deoxyglucose and metformin.

Helicobacter pylori infection causes chronic inflammation in the stomach, which is linked to the development of gastric cancer. The anti-inflammatory and anticancer effects of a glycolysis inhibitor 2-deoxyglucose (2DG) and an antidiabetic medication metformin (Met) have gotten attention. Using a Mongolian gerbil animal model, we investigated H. pylori-mediated gastric pathogenesis and how this pathogenesis is influenced by 2DG and Met. Five-week-old male gerbils were infected with H. pylori strain 7.13. After 2 weeks of infection, gerbils were fed 2DG-containing food (0.03% w/w), Met-containing water (0.5% w/v), or both (Combi) for 2 (short-term) or 10 weeks (long-term). Gastric pathogenesis and host response to H. pylori infection were examined by macroscopic and histopathologic analysis of gerbils' stomach. As a result, indicators of gastric pathogenesis by H. pylori infection including infiltration of polymorphonuclear neutrophils and lymphocytes, intestinal metaplasia, atrophy, and proliferation of gastric epithelial cells were attenuated by short-term administration of 2DG, Met, or Combi. When the infection was sustained for long-term, gastric pathogenesis in drug-treated gerbils was equivalent to that in untreated gerbils, with the exception that the infiltration of neutrophil was reduced by 2DG. Colonization of H. pylori in stomach was unaffected by both short- and long-term treatments. Our findings demonstrate that the progression of gastric pathogenesis induced by H. pylori infection can be attenuated by the short-term individual or combinational treatment of 2DG and Met, implying that 2DG or Met could be considered as a treatment option for gastric diseases in the early stages of infection.

Alcohol Impairs Immunometabolism and Promotes Naïve T Cell Differentiation to Pro-Inflammatory Th1 CD4+ T Cells.

CD4+ T cell differentiation to pro-inflammatory and immunosuppressive subsets depends on immunometabolism. Pro-inflammatory CD4+ subsets rely on glycolysis, while immunosuppressive Treg cells require functional mitochondria for their differentiation and function. Previous pre-clinical studies have shown that ethanol (EtOH) administration increases pro-inflammatory CD4+ T cell subsets; whether this shift in immunophenotype is linked to alterations in CD4+ T cell metabolism had not been previously examined. The objective of this study was to determine whether ethanol alters CD4+ immunometabolism, and whether this affects CD4+ T cell differentiation. Naïve human CD4+ T cells were plated on anti-CD3 coated plates with soluble anti-CD28, and differentiated with IL-12 in the presence of ethanol (0 and 50 mM) for 3 days. Both Tbet-expressing (Th1) and FOXP3-expressing (Treg) CD4+ T cells increased after differentiation. Ethanol dysregulated CD4+ T cell differentiation by increasing Th1 and decreasing Treg CD4+ T cell subsets. Ethanol increased glycolysis and impaired oxidative phosphorylation in differentiated CD4+ T cells. Moreover, the glycolytic inhibitor 2-deoxyglucose (2-DG) prevented the ethanol-mediated increase in Tbet-expressing CD4+ T cells but did not attenuate the decrease in FOXP3 expression in differentiated CD4+ T cells. Ethanol increased Treg mitochondrial volume and altered expression of genes implicated in mitophagy and autophagosome formation (PINK1 and ATG7). These results suggest that ethanol impairs CD4+ T cell immunometabolism and disrupts mitochondrial repair processes as it promotes CD4+ T cell differentiation to a pro-inflammatory phenotype.