Glucose Metabolites Research Articles

Adaptations in hepatic glucose metabolism after chronic social defeat stress in mice

Chronic stress has been shown to induce hyperglycemia in both peripheral blood and the brain, yet the detailed mechanisms of glucose metabolism under stress remain unclear. Utilizing 13C6-labeled glucose to trace metabolic pathways, our study investigated the impact of stress by chronic social defeat (CSD) on glucose metabolites in the liver and brain one week post-stress. We observed a reduction in 13C6-enrichment of glucose metabolites in the liver, contrasting with unchanged levels in the brain. Notably, hepatic glycogen levels were reduced while lactate concentrations were elevated, suggesting lactate as an alternative energy source during stress. Long-term effects were also examined, revealing normalized blood glucose levels and restored glycogen stores in the liver three weeks post-CSD, despite sustained increases in food intake. This normalization is hypothesized to result from diminished glucagon levels leading to reduced glycogen phosphorylase activity. Our findings highlight a temporal shift in glucose metabolism, with hyperglycemia and glycogen depletion in the liver early after CSD, followed by a later phase of metabolic stabilization. These results underscore the liver’s critical role in adapting to CSD and provide insights into the metabolic adjustments that maintain glucose homeostasis under prolonged stress conditions.

The effect of anti‐imbalance on glycolipid metabolic homeostasis via the liver–gut axis intervened by 3′‐sialyllactose in obese mice

AbstractSialic acid could ameliorate disorders associated with glycolipid metabolism. 3′‐Sialyllactose (3′‐SL), a type of oligosaccharide, contains sialic acid. We examined the effects of 3′‐SL on glycolipid metabolism in obese mice with high fat diet and explore the underlying mechanisms. A total of 160 male C57BL/6J mice were fed with high‐fat diet for 8 weeks to construct obese mice model. Sixty successfully constructed obese mice were randomly divided three 3′‐SL dosage‐dependent groups, one free sialic acid N‐acetylneuraminic acid (Neu5Ac) group, and one blank control group. Each group of obese mice (n = 12) was continuously supplemented by 3′‐SL or Neu5Ac for 10 weeks. Ten‐week 3′‐SL supplementation and Neu5Ac supplementation both yielded remarkedly lower body weight, reduced fat content, increased energy expenditure and active daily exercises with improved glycolipid biomarkers, and attenuated proinflammation. 3′‐SL increased the colonic abundances of Akkermansia, Lactobacillus, and Solibacillus and reduced those of Enterobacter and Enterococcus. Changes were also observed in colonic and liver transcript and metabolites, which were mainly enriched in glycolipid metabolism, ameliorated immune function, and mitigated inflammatory signals, autophagy, bile acid metabolism, and insulin resistance. Akkermansia and Solibacillus were significantly associated with changes in colonic metabolites and transcriptome genes. The liver bile component and glucose metabolites were significantly correlated with transcriptional gene expression in the aforementioned differential pathways. Therefore, 3′‐SL can enhance the gut microbiota, regulate intestinal immunity and bile acid metabolism, reduce liver oxidative stress and autophagy processes, alleviate chronic inflammation levels, and improve islet function and lower blood sugar levels by acting through the gut–liver axis.

Elucidating the Antiglycation Effect of Creatine on Methylglyoxal-Induced Carbonyl Stress In Vitro.

Advanced glycation end products (AGEs) with multiple structures are formed at the sites where carbonyl groups of reducing sugars bind to free amino groups of proteins through the Maillard reaction. In recent years, it has been highlighted that the accumulation of AGEs, which are generated when carbonyl compounds produced in the process of sugar metabolism react with proteins, is involved in various diseases. Creatine is a biocomponent that is homeostatically present throughout the body and is known to react nonenzymatically with α-dicarbonyl compounds. This study evaluated the antiglycation potential of creatine against methylglyoxal (MGO), a glucose metabolite that induces carbonyl stress with formation of AGEs in vitro. Further, to elucidate the mechanism of the cytoprotective action of creatine, its effect on the accumulation of carbonyl proteins in the cells and the MGO-induced cellular damage were investigated using neuroblastoma cells. The results revealed that creatine significantly inhibits protein carbonylation by directly reacting with MGO, and creatine added to the culture medium suppressed MGO-derived carbonylation of intracellular proteins and exerted a protective effect on MGO-induced cytotoxicity. These findings suggest that endogenous and supplemented creatine may contribute to the attenuation of carbonyl stress in vivo.

Thiamine Deficiency and its Implications on Microvascular Complications of Diabetes Mellitus

Thiamine is the first vitamin discovered and belongs to Vit B family. The main effect seen with thiamine deficiency is Beri-Beri, Wernicke’s encephalopathy, Wernicke-Korsak off syndrome and are considered as a serious condition but often can be reversed. The deficient status of thiamine can also cause varied affects and can overlap with other conditions to exacerbate its potent effects. It is seen that thiamine is necessary for the metabolism of glucose in the form of cofactors, deficiency of which leads to accumulation of toxic glucose metabolites leading to formation of free radicals and oxidative stress. Glucose is not only important for the formation of energy but its improper metabolism proves to have deleterious effects in the body. In this review, an attempt is made to correlate microvascular complications of diabetes with thiamine deficiency and can be discerned that oxidative stress is one of the important factors for the progress of microvascular complications, as well as diabetic ketoacidosis, atherosclerosis and cardiovascular damage in patients with diabetes mellitus and these can be prevented or maintained by optimizing thiamine levels in the body.

Glucose metabolite methylglyoxal induces vascular endothelial cell pyroptosis via NLRP3 inflammasome activation and oxidative stress in vitro and in vivo

Methylglyoxal (MGO), a reactive dicarbonyl metabolite of glucose, plays a prominent role in the pathogenesis of diabetes and vascular complications. Our previous studies have shown that MGO is associated with increased oxidative stress, inflammatory responses and apoptotic cell death in endothelial cells (ECs). Pyroptosis is a novel form of inflammatory caspase-1-dependent programmed cell death that is closely associated with the activation of the NOD-like receptor 3 (NLRP3) inflammasome. Recent studies have shown that sulforaphane (SFN) can inhibit pyroptosis, but the effects and underlying mechanisms by which SFN affects MGO-induced pyroptosis in endothelial cells have not been determined. Here, we found that SFN prevented MGO-induced pyroptosis by suppressing oxidative stress and inflammation in vitro and in vivo. Our results revealed that SFN dose-dependently prevented MGO-induced HUVEC pyroptosis, inhibited pyroptosis-associated biochemical changes, and attenuated MGO-induced morphological alterations in mitochondria. SFN pretreatment significantly suppressed MGO-induced ROS production and the inflammatory response by inhibiting the NLRP3 inflammasome (NLRP3, ASC, and caspase-1) signaling pathway by activating Nrf2/HO-1 signaling. Similar results were obtained in vivo, and we demonstrated that SFN prevented MGO-induced oxidative damage, inflammation and pyroptosis by reversing the MGO-induced downregulation of the NLRP3 signaling pathway through the upregulation of Nrf2. Additionally, an Nrf2 inhibitor (ML385) noticeably attenuated the protective effects of SFN on MGO-induced pyroptosis and ROS generation by inhibiting the Nrf2/HO-1 signaling pathway, and a ROS scavenger (NAC) and a permeability transition pore inhibitor (CsA) completely reversed these effects. Moreover, NLRP3 inhibitor (MCC950) and caspase-1 inhibitor (VX765) further reduced pyroptosis in endothelial cells that were pretreated with SFN. Collectively, these findings broaden our understanding of the mechanism by which SFN inhibits pyroptosis induced by MGO and suggests important implications for the potential use of SFN in the treatment of vascular diseases.

Polyphenol-Rich Aronia melanocarpa Fruit Beneficially Impact Cholesterol, Glucose, and Serum and Gut Metabolites: A Randomized Clinical Trial.

Polyphenol-rich Aronia fruits have great potential as a functional food with anti-inflammatory, hypolipidemic, and hypoglycemic biologic activities. However, clinical intervention trials investigating the impact of Aronia fruit consumption on human health are limited. A randomized, controlled, double-blinded, parallel intervention trial was conducted using 14 human subjects who ingested either 0 mL or 100 mL of Aronia juice daily for 30 days. Anthropometric measurements, fasting, and postprandial measures of glucose and lipid metabolism and inflammation, 16S rRNA fecal microbial composition data, and mass spectrometry-acquired serum and fecal metabolomic data were collected before and after the intervention period. Data were analyzed using general linear models, ANOVA, and t-tests. Daily consumption of Aronia prevented a rise in cholesterol levels (β = -0.50, p = 0.03) and reduced postprandial glucose (β = -3.03, p < 0.01). No difference in microbial community composition by condition was identified at any taxonomic level, but a decrease (β = -18.2, p = 0.04) in microbial richness with Aronia was detected. Serum and fecal metabolomic profiles indicated shifts associated with central carbon and lipid metabolism and decreases in pro-inflammatory metabolites. Our study further informs the development of polyphenol-based dietary strategies to lower metabolic disease risk.

The reactive pyruvate metabolite dimethylglyoxal mediates neurological consequences of diabetes

Complications of diabetes are often attributed to glucose and reactive dicarbonyl metabolites derived from glycolysis or gluconeogenesis, such as methylglyoxal. However, in the CNS, neurons and endothelial cells use lactate as energy source in addition to glucose, which does not lead to the formation of methylglyoxal and has previously been considered a safer route of energy consumption than glycolysis. Nevertheless, neurons and endothelial cells are hotspots for the cellular pathology underlying neurological complications in diabetes, suggesting a cause that is distinct from other diabetes complications and independent of methylglyoxal. Here, we show that in clinical and experimental diabetes plasma concentrations of dimethylglyoxal are increased. In a mouse model of diabetes, ilvb acetolactate-synthase-like (ILVBL, HACL2) is the enzyme involved in formation of increased amounts of dimethylglyoxal from lactate-derived pyruvate. Dimethylglyoxal reacts with lysine residues, forms Nε−3-hydroxy-2-butanonelysine (HBL) as an adduct, induces oxidative stress more strongly than other dicarbonyls, causes blood-brain barrier disruption, and can mimic mild cognitive impairment in experimental diabetes. These data suggest dimethylglyoxal formation as a pathway leading to neurological complications in diabetes that is distinct from other complications. Importantly, dimethylglyoxal formation can be reduced using genetic, pharmacological and dietary interventions, offering new strategies for preventing CNS dysfunction in diabetes.

Glyceraldehyde metabolism in mouse brain and the entry of blood-borne glyceraldehyde into the brain.

D-Glyceraldehyde, a reactive aldehyde metabolite of fructose and glucose, is neurotoxic invitro by forming advanced glycation end products (AGEs) with neuronal proteins. In Alzheimer's disease brains, glyceraldehyde-containing AGEs have been detected intracellularly and in extracellular plaques. However, little information exists on how the brain handles D-glyceraldehyde metabolically or if glyceraldehyde crosses the blood-brain barrier from the circulation into the brain. We injected [U-13C]-D-glyceraldehyde intravenously into awake mice and analyzed extracts of serum and brain by 13C nuclear magnetic resonance spectroscopy. 13C-Labeling of brain lactate and glutamate indicated passage of D-glyceraldehyde from blood to brain and glycolytic and oxidative D-glyceraldehyde metabolism in brain cells. 13C-Labeling of serum glucose and lactate through hepatic metabolism of [U-13C]-D-glyceraldehyde could not explain the formation of 13C-labeled lactate and glutamate in the brain. Cerebral glyceraldehyde dehydrogenase and reductase activities, leading to the formation of D-glycerate and glycerol, respectively, were 0.27-0.28 nmol/mg/min; triokinase, which phosphorylates D-glyceraldehyde to D-glyceraldehyde-3-phosphate, has been demonstrated previously at low levels. Thus, D-glyceraldehyde metabolism toward glycolysis could proceed both through D-glycerate, glycerol, and D-glyceraldehyde-3-phosphate. The aldehyde group of D-glyceraldehyde was overwhelmingly hydrated into a diol in aqueous solution, but the diol dehydration rate greatly exceeded glyceraldehyde metabolism and did not restrict it. We conclude that (1) D-glyceraldehyde crosses the blood-brain barrier, and so blood-borne glyceraldehyde could contribute to AGE formation in the brain, (2) glyceraldehyde is taken up and metabolized by brain cells. Metabolism thus constitutes a detoxification mechanism for this reactive aldehyde, a mechanism that may be compromised in disease states.

Altered bile acid and correlations with gut microbiome in transition dairy cows with different glucose and lipid metabolism status

The transition from pregnancy to lactation is critical in dairy cows. Among others, dairy cows experience a metabolic stress due to a large change in glucose and lipid metabolism. Recent studies revealed that bile acids (BA), besides being involved in both the emulsification and solubilization of fats during intestinal absorption, can also affect the metabolism of glucose and lipids, both directly or indirectly by affecting the gut microbiota. Thus, we used untargeted and targeted metabolomics and 16S rRNA sequencing approaches to investigate the concentration of plasma metabolites and BA, the composition of the rectum microbial community, and assess their interaction in transition dairy cows. In Experiment 1, we investigated BA and other blood parameters and gut microbiota in dairy cows without clinical diseases during the transition period, which can be seen as well adapted to the challenge of changed glucose and lipid metabolism. As expected, we detected an increased plasma concentration of β-hydroxybutyrate (BHBA) and nonesterified fatty acids (NEFA) but decreased concentration of glucose, cholesterol, and triglycerides (TG). Untargeted metabolomic analysis of the plasma revealed primary BA biosynthesis was one of the affected pathways, and was consistent with the increased concentration of BA in the plasma. A correlation approach revealed a complex association between BA and microbiota with the host plasma concentration of glucose and lipid metabolites. Among BA, chenodeoxycholic acid derivates such as glycolithocholic acid, taurolithocholic acid, lithocholic acid, taurochenodeoxycholic acid, and taurodeoxycholic acid were the main hub nodes connecting microbe and blood metabolites (such as glucose, TG, and NEFA). In Experiment 2, we investigated early postpartum dairy cows with or without hyperketonemia (HPK). As expected, HPK cows had increased concentration of NEFA and decreased concentrations of glucose and triglycerides. The untargeted metabolomic analysis of the plasma revealed that primary BA biosynthesis was also one of the affected pathways. Even though the BA concentration was similar among the 2 groups, the profiles of taurine conjugated BA changed significantly. A correlation analysis also revealed an association between BA and microbiota with the concentration in plasma of glucose and lipid metabolites (such as BHBA). Among BA, cholic acid and its derivates such as taurocholic acid, tauro α-muricholic acid, and taurodeoxycholic acid were the main hub nodes connecting microbe and blood metabolites. Our results indicated an association between BA, intestinal microbe, and glucose and lipid metabolism in transition dairy cows. These findings provide new insight into the adaptation mechanisms of dairy cows during the transition period.

MODL-05. UNDERSTANDING THE METABOLIC VULNERABILITIES IN INFILTRATING GLIOMATOSIS CEREBRI CELLS

Abstract BACKGROUND Gliomatosis cerebri (GC), a rare form of high-grade glioma, is characterized by a particularly invasive phenotype within multiple lobes of the brain and a dismal prognosis. This entity was removed in 2016 of the WHO brain tumor classification because of an absence of specific GC driving molecular signature. Its diffusive aspect on MRI is defined by an absence of cohesive tumor mass, a mild contrast enhancement and a rare neoangiogenesis. Based on those observations, our objectives were to understand how these cells are adapting to their brain environment without a physiological nutritive efflux associated to a reduced oxygen concentration and determine a metabolic signature of GC. METHODS Preliminary results in a large cohort of HGGs bearing several drivers (H3.3 G34V, NMYC amplified, BCOR positive) or a more “wildtype” phenotype with new entities (pedRTK 1 and 2) were underlining glucose and glutamine as major nutrients in cell metabolic suppliance. Therefore, epigenetic assessments, bulk metabolomics, and Seahorse technology, as well as labeled glucose and glutamine metabolites study, helped us in pediatric GC-derived cell lines (BT97 and BT139, PEDIAMODECAN program) to understand the role of both nutrients and their central role in GC in hypoxic microenvironment. RESULTS Both glucose and glutamine are involved in carbon fueling biosynthesis in those cells. Glutamine is much more involved in supplying nitrogen to fuel several specific amino acids and mitochondrial metabolisms. Meanwhile, we developed drug testing using Matrigel invasion aspects. The tested drugs were able to reduce invasiveness of GC cells and modify the intratumoral metabolism. CONCLUSION All those observations seem to explain GC cell adaptation into the hypoxic microenvironment, their plasticity and resistance through metabolic reprogramming based on glutaminolysis and glycolysis.

Isolation, screening, and characterization of the newly isolated osmotolerant yeast Wickerhamomyces anomalus BKK11-4 for the coproduction of glycerol and arabitol.

This study explored the isolation and screening of an osmotolerant yeast, Wickerhamomyces anomalus BKK11-4, which is proficient in utilizing renewable feedstocks for sugar alcohol production. In batch fermentation with high initial glucose concentrations, W. anomalus BKK11-4 exhibited notable production of glycerol and arabitol. The results of the medium optimization experiments revealed that trace elements, such as H3BO3, CuSO4, FeCl3, MnSO4, KI, H4MoNa2O4, and ZnSO4, did not increase glucose consumption or sugar alcohol production but substantially increased cell biomass. Osmotic stress, which was manipulated by varying initial glucose concentrations, influenced metabolic outcomes. Elevated glucose levels promoted glycerol and arabitol production while decreasing citric acid production. Agitation rates significantly impacted the kinetics, enhancing glucose utilization and metabolite production rates, particularly for glycerol, arabitol, and citric acid. The operational pH dictated the distribution of the end metabolites, with glycerol production slightly reduced at pH 6, while arabitol production remained unaffected. Citric acid production was observed at pH 6 and 7, and acetic acid production was observed at pH 7. Metabolomic analysis using GC/MS identified 29 metabolites, emphasizing the abundance of sugar/sugar alcohols. Heatmaps were generated to depict the variations in metabolite levels under different osmotic stress conditions, highlighting the intricate metabolic dynamics occurring post-glucose uptake, affecting pathways such as the pentose phosphate pathway and glycerolipid metabolism. These insights contribute to the optimization of W. anomalus BKK11-4 as a whole-cell factory for desirable products, demonstrating its potential applicability in sustainable sugar alcohol production from renewable feedstocks.

297 Effects of plasma zinc concentration on longissimus thoracis metabolic status of beef steers

Abstract The objective of this study was to examine the role of Zn status on muscle glucose and other metabolites. Angus steers [n = 144; body weight (BW) = 525 ± 30 kg] with varying plasma Zn concentration and implant status served as the initial pool for this secondary experiment. Steers were assigned to one of two implant (IMP) treatments: no implant (NO) or Component TE-200 (TE-200; Elanco, Greenfield, IN) on d 0. Zinc was supplemented as ZnSO4 at 0 mg Zn/kg dry matter (DM; analyzed 54 mg Zn/kg DM), 30 mg Zn/kg DM, or 100 mg Zn/kg DM, starting 60 d before implant day. Steers were fed via GrowSafe bunks (GrowSafe Systems Ltd., Airdie, AB, Canada), and steer was experimental unit. Steers were fed in two blocks with a 14-d stagger for sample logistics. Blood and muscle biopsies of the longissimus thoracis were collected on d 40 after implant, a minimum of 2 h post-feed delivery. Plasma Zn was quantified via ICP-OES and 144 samples were stratified into quantiles by plasma Zn concentration. Samples (n = 12 low and 12 high from NO and TE-200 groups) were identified and designated as low plasma Zn (LOW, 1.1 mg Zn/L average) and high plasma Zn (HI, 1.6 mg Zn/L average) treatments (TRT). Muscle samples from these 48 steers were homogenized and derivatized for subsequent analysis via gas chromatography-mass spectrometry (GCMS) for non-targeted metabolomic analysis. Data were analyzed as a complete randomized design using the MIXED procedure of SAS 9.4 (SAS Inst. Inc., Cary, NC) with fixed effects of TRT, IMP, BLOCK, and TRT×IMP. A TRT×IMP effect was observed for muscle glycine (P = 0.02) where LOW/IMP was similar to HI/NO but had greater glycine concentrations than LOW/NO and HI/IMP. There was a tendency for a TRT×IMP (P = 0.07) effect where LOW/IMP and HI/NO tended to have greater concentrations of hydroxyisovaleric acid. Given the role of glycine in protein synthesis and hydroxyisovaleric acid in leucine metabolism these data suggest differential protein responses to implants in cattle of lesser vs. greater plasma Zn concentrations. Hydroxybutyric acid, a ketone body which can be utilized for energy, tended to be greater in muscle of steers receiving an IMP (P = 0.09). Contrary to our hypothesis that greater plasma Zn would support differential muscle glucose metabolism, no effects of plasma Zn TRT or IMP were noted for glucose, lactate, and other detected metabolites. Both LOW and HI plasma Zn concentrations were still considered adequate, and future research should explore the impact of distinctly deficient and adequate plasma Zn concentration on glucose metabolism pathways in muscle.

Metabolic dysfunction in the myocardium three months after exertional heat stroke in mice

In humans, exposure to heat stroke has proven to be a risk factor for development of cardiovascular disease later in life. We hypothesized that exposure to exertional heat stroke (EHS) would result in the development of similar long-term cardiac disease in mice. In this study we followed mice for three months after EHS, using ultrasound imaging of the heart, metabolomics and histological analysis of ventricular tissue and biomarker analysis after 4 weeks of recovery. Methods: Mice were instrumented with temperature telemetry devices and were trained to run on forced running wheels using an incremental exercise protocol. EHS mice ran in an temperature elevated environmental chamber; females 37.5°C; males 34°C; whereas, matched exercise control mice (EXC) ran in 22-23°C. Running stopped when EHS mice reached symptom limitation (unconsciousness). The mice were followed for 12 wks post EHS, with two ultrasound measures at 2 and 10 wks of recovery. At the end of the study, mice were anesthetized, a blood sample drawn, and ventricular tissue collected for metabolomics. RESULTS: Male but not female EHS mice gained ~10% greater body mass compared to EXC over the course of recovery ( P&lt;0.01). Hearts from male but not female EHS mice exhibited elevations in heart mass per tibia length ( P&lt;0.01). At three months, metabolomics in male EHS hearts revealed significant elevations in glucose and glucose metabolites and significant decreases in acylcarnitines consistent with a metabolic shift toward glucose and away from lipid metabolism. In contrast, female hearts exhibited fewer abnormalities but with significant elevations in lipid and arachidonate metabolites. At two weeks post, there were no differences in male ultrasound measures between EHS and EXC mice but in females there was a reduced posterior wall velocity ( P=0.012) and a trend towards narrower posterior wall thickness ( P= 0.06) . Between 2 and 12 wks post EHS, there were significant elevations in fractional shortening in males ( P&lt;0.05) and elevations in posterior wall velocity ( P&lt;0.01). In contrast, females exhibited significant reductions posterior wall velocity. From blood samples, male EHS mice had increased HDL and females exhibited decreased HDL (p&lt;0.01 in both). LDL increased only in males P&lt;0.05. Vascular adhesion molecules sICAM ( P&lt;0.01) and Pecam1 ( P&lt;0.01) were significantly decreased in EHS males and females and e-Selectin was decreased in EHS males only ( P&lt;0.001). Total serum protein was significantly reduced in both male and females ( P&lt;0.01). Male EHS mice also exhibited significant elevations in white blood cell count and total lymphocytes ( P=0.01) suggesting ongoing activation of the immune system . CONCLUSIONS: In this controlled preclinical model of EHS, both male and female mice exhibited long-lasting cardiovascular and systemic disorders that could account for the lingering incidence of cardiovascular disease and poor overall health in heat stroke victims. Supported by US Army Med. Research and Devel. Command BA180078 and the BK and Betty Stevens Endowment. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Analysis of Lysine Metabolism in a Rat Model of Type 2 Diabetes Reveals Modulation of Endocytosis and Glucose Metabolism

Lysine is an essential amino acid that serves as a building block for proteins and plays a role in metabolic processes. While a recent study provided evidence that lysine is beneficial for hypertension-associated kidney disease, possibly through inhibiting renal tubular reabsorption and forming new lysine conjugates, knowledge about the role of lysine in diabetic nephropathy (DN) is rudimentary. Therefore, the aim of the study was to investigate the role of lysine in kidney function in the setting of DN and its underlying mechanisms. We hypothesized that lysine may impact the kidneys in the following ways: 1) by mediating protein endocytosis in proximal tubules; 2) by altering protein abundance in the kidneys and other organs; and 3) by disrupting metabolic pathways through the formation of conjugates or degradation products. We investigated the effects of lysine on proximal tubule endocytosis and other organs using proteomic approaches and studied changes in lysine conjugates and metabolic products using targeted metabolomics. The results of urinary proteomics suggested that lysine’s impact on endocytosis may be associated with the inhibition of megalin and cubilin. Furthermore, lysine affected proteins related to glucose metabolism. Subsequently, we proceeded to investigate the influence of lysine on various organs in Type 2 Diabetic Nephropathy (T2DN) rats. Gene Ontology (GO) enrichment revealed that lysine exerts tissue-specific effects on different organs, with pathways enriched in the kidney including phagocytosis and enzyme inhibitor activity. Additionally, within the kidney, the lysine-treated group showed downregulation of glycolysis-related enzymes. The metabolomics results showed that lysine leads to a decrease in glucose and an increase in fructose 1,6-bisphosphate, while also resulting in an upregulation of related metabolites in the tricarboxylic acid cycle (TCA cycle). Our findings showed changes in the protein composition involved in lysine-mediated endocytosis and the impact of lysine on glucose metabolism enzymes and metabolites. The specific molecular mechanisms are still under investigation. R01 DK135644 and I01 BX004024 (to AS). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Citrate, a TCA cycle metabolite, plays a role as a hydroxyl radical scavenger in vitro

Reactive oxygen species, such as superoxide anions, hydrogen peroxide, and hydroxyl radicals, are strongly associated with diseases and aging. Superoxide anions and hydrogen peroxide are scavenged by superoxide dismutase and catalase, respectively. These scavenging reactions are supported by enzymes. In contrast, hydroxyl radicals are scavenged by non-enzymatic compounds, such as ascorbic acid, glutathione, and phenolic compounds. Recently, metabolites of glucose and ethanol have been shown to act as hydroxyl radical scavengers. In this study, we evaluated hydroxyl radical-scavenging activity of metabolites of the TCA cycle. We found that these metabolites have a scavenging activity against hydroxyl radicals. In particular, citrate exhibited significant scavenging activity at the cellular level. Additionally, citrate protected superoxide dismutase from hydroxyl radicals. We also confirmed the protective role of citrate against hydroxyl radical-induced DNA strand breakage. The previously reported rate constants and intracellular concentrations did not rule out a protective role in mitochondria. Collectively, our results indicate that citrate acts as a hydroxyl radical scavenger.

Effects of Ganjianglingzhu Decoction on Lean Non-Alcoholic Fatty Liver Disease in Mice Based on Untargeted Metabolomics.

Non-alcoholic fatty liver disease (NAFLD) is usually associated with obesity. However, it is crucial to recognize that NAFLD can also occur in lean individuals, which is frequently overlooked. Without an approved pharmacological therapy for lean NAFLD, we aimed to investigate whether the Ganjianglingzhu (GJLZ) decoction, a representative traditional Chinese medicine (TCM), protects against lean NAFLD and explore the potential mechanism underlying these protective effects. The mouse model of lean NAFLD was established with a methionine-choline-deficient (MCD) diet in male C57BL/6 mice to be compared with the control group fed the methionine-choline-sufficient (MCS) diet. After four weeks, physiological saline, a low dose of GJLZ decoction (GL), or a high dose of GJLZ decoction (GH) was administered daily by gavage to the MCD group; the MCS group was given physiological saline by gavage. Untargeted metabolomics techniques were used to explore further the potential mechanism of the effects of GJLZ on lean NAFLD. Different doses of GJLZ decoction were able to ameliorate steatosis, inflammation, fibrosis, and oxidative stress in the liver; GL performed a better effect on lean NAFLD. In addition, 78 candidate differential metabolites were screened and identified. Combined with metabolite pathway enrichment analysis, GL was capable of regulating the glucose and lipid metabolite pathway in lean NAFLD and regulating the glycerophospholipid metabolism by altering the levels of sn-3-O-(geranylgeranyl)glycerol 1-phosphate and lysoPC(P-18:0/0:0). GJLZ may protect against the development of lean NAFLD by regulating glucose and lipid metabolism, inhibiting the levels of sn-3-O-(geranylgeranyl)glycerol 1-phosphate and lysoPC(P-18:0/0:0) in glycerophospholipid metabolism.

Abstract 2874: Cancer metabolism in radiation resistance: Complementary roles of O-GlcNAc transferase and PARP1

Abstract The efficacy of image-guided radiotherapy (RT) has been linked to directing increased chromosomal double strand breaks (DSBs) to tumor cells while sparing surrounding normal tissue. DSBs can block cell cycle progression and promote mitotic catastrophe, cell death and senescence. End processing of persistent DSBs releases single strand DNA (ssDNA), driving cGAS/STING signaling and anti-tumor immune response. DSB repair, whether by non-homologous end-joining (NHEJ) or homologous recombination (HR), is a resistance mechanism, supporting cell survival, repopulation and immune evasion. Decades of effort to block DSB repair to enhance benefits of RT has yet to produce approved agents. Toxicity due to lack of cancer specificity may prevent the current generation of targeted agents from reaching the clinic. An answer may come from examining cancer metabolic reprogramming as a determinant of intrinsic radiation resistance. Along with other glucose metabolites, the Warburg effect increases N-acetyl glucosamine (GlcNAc) biosynthesis, which not only supports N-glycosylation but also modification of nucleocytoplasmic proteins by O-GlcNAc transferase (OGT). OGT-dependent O-GlcNAcylation confers radiation resistance, in part by accelerating DSB repair. OGT regulates γH2AX, 53BP1 and BRCA1 foci kinetics and suppresses ssDNA formation at DSB ends, limiting HR but favoring NHEJ for S/G2 DSB repair. Blocking OGT induces radiation sensitization in vitro and in vivo. In cells, OGT inhibition results in hyperresection, persistent RPA and RAD51 foci and accumulation of cytosolic DNA. One mediator may be the histone methyltransferase EZH2, which is rapidly recruited to deposit H3K27me3 at DSBs, promoting NHEJ repair. Irradiation of EZH2 KO cells results in hyperresection that cannot be suppressed by O-GlcNAcylation. Cancer cells may also accumulate NAD+, a substrate for PARP1 incorporated into poly-ADP-ribose to initiate DSB repair. Much like OGT inhibition, blocking PARP1 induces hyperresection and shifts repair toward HR and away from NHEJ. Importantly, increasing O-GlcNAcylation can suppress ssDNA formation even in PARP1 KO cells. In turn, inhibiting PARP1 enhances hyperesection in EZH2 KO cells, leading to a marked increase in cytosolic ssDNA. Our data assign complementary roles for EZH2 and PARP1 in DSB repair pathway choice by independently limiting 5' end resection. This directs DSBs to rapid, imprecise repair by NHEJ over slow and precise repair by HR. An implication is that PARP inhibitor resistance due to the Warburg effect may be mediated by EZH2. Driving hyperresection by targeting O-GlcNAcylation or H3K27 trimethylation along with PARP inhibition may saturate the capacity of HR and produce irreversible damage in cancer cells. Targeting impacts of metabolic reprogramming on DSB repair may not only enhance the therapeutic index of radiation but also sharpen synthetic lethality with HR deficiency. Citation Format: Elena E. Efimova, Sera Averbek, Donald J. Wolfgeher, Ding Wu, Yue Liu, Stephen J. Kron. Cancer metabolism in radiation resistance: Complementary roles of O-GlcNAc transferase and PARP1 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2874.

Abstract 1784: Glucose metabolism and de novo palmitate synthesis under normoxia and hypoxia in breast cancer cells that preferentially metastasize to lung and liver

Abstract Cancer cells are exposed to variable oxygen and nutrient availability during the metastatic process which can influence metabolic adaptations of cancer cells to metastasize to distant sites. Thus, we investigated the metabolic adaptations of breast cancer cells that preferentially metastasize to lung (metM-WntLung cells; MLg) and liver (metM-WntLiver cells; MLr) in normoxia and hypoxia and at different glucose concentrations. In normoxia, 14C-glucose uptake is similar, but mRNA abundance of hexokinase, the initial rate-limiting step in glycolysis, was 22% higher in MLg cells. This is consistent with higher 13C6-glucose flux into glycolytic metabolites pyruvate (M+3) and lactate (M+3) in MLg compared to MLr. Also, de novo palmitate synthesis from 13C6-glucose was 7.6% higher in MLg and was accompanied by higher mRNA expression of ATP-citrate lyase (ACLY), which converts citrate to acetyl-CoA for fatty acid synthesis. These suggest greater glucose metabolism in MLg compared to MLr in normoxia, consistent with reduced viability of MLr by 39% in high glucose (25 mM) concentrations. Further, mRNA abundance of CPT1A, a rate-limiting step in fatty acid oxidation, is 23% higher in MLg. Moreover, the fatty acid oxidation inhibition (etomoxir) reduced viability of MLg by 26.7% compared to MLr, suggesting that fatty acid synthesis and oxidation are critical for MLg. Protein expression of HIF1α, a transcription factor that induces glycolysis during hypoxia, is 1.5-fold higher in MLr than MLg in normoxia, consistent with higher MLr mRNA abundance of pyruvate dehydrogenase kinase (PDK1), a target of HIF1α that inhibits pyruvate dehydrogenase (PDH) activity. Surprisingly, mRNA abundance of pyruvate carboxylase (PC), the enzyme that converts pyruvate to oxaloacetate, is also higher in MLr (39%) than MLg, thus supporting a potential alternative mechanism for glucose metabolites to enter the TCA cycle in normoxia in MLr. In hypoxia, viability of MLr, relative to MLg, increased by 11% at high glucose concentration. This was accompanied by higher hexokinase (33%) and PDK1 (83%) mRNA abundance suggesting MLr’s shift towards a glycolytic phenotype. However, de novo palmitate synthesis from 13C6-glucose is higher in MLg, suggesting that palmitate synthesis is maintained in MLg even in hypoxia. Altogether, results demonstrate that glucose utilization shifts in breast cancer cells that preferentially metastasize to lung vs liver, but de novo fatty acid synthesis is higher in MLg in different oxygen conditions. Thus, targeting breast cancer cells’ adaptation to glucose and de novo fatty acid synthesis may be a potential strategy to prevent breast cancer progression to distant organs. Citation Format: Marjorie Anne A. Layosa, Morgan Conrad, Stephen Hursting, Dorothy Teegarden. Glucose metabolism and de novo palmitate synthesis under normoxia and hypoxia in breast cancer cells that preferentially metastasize to lung and liver [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1784.

The acute neurotoxicity of inorganic mercury in Mactra chinensis philippi

Inorganic mercury (IHg) is hazardous to marine organisms especially resulting in neurotoxicity, bivalves are sensitive to pollutants as “ocean sentinel”, but data on the neurotoxicity of IHg in bivalves are sparse. So we chosed M. chinensis philippi with typical neural structures in bivalves to investigate the neurotoxicity of IHg, which could be helpful to understand the specificity of neural regulation and the response characteristics of bivalves. After acute exposed to IHg (HgCl2) for 24 h, the metabolites of ganglion tissues in M. chinensis philippi were evaluated using 1H-nuclear magnetic resonance based metabolomics; Ca2+, neurotransmitters (nitric oxide, glutamate, acetylcholine) and related enzymes (calcineurin, nitric oxide synthase and acetylcholinesterase) were measured using biochemical detection. Compared to the control group, the levels of the nitric oxide (81.04 ± 12.84 μmol/g prot) and acetylcholine (30.93 ± 12.57 μg/mg prot) in M. chinensis philippi of IHg-treated were decreased, while glutamate (2.11 ± 0.61 mmol/L) increased significantly; the activity of nitric oxide synthase (679.34 ± 135.33 U/mg prot) was increased, while acetylcholinesterase (1.39 ± 0.44 U/mg prot) decreased significantly, and the activity of calcineurin (0.52 ± 0.02 U/mg prot) had a statistically insignificant increasing tendency. The concentration of Ca2+ (0.92 ± 0.46 mmol/g prot) in the IHg-treated group was significantly higher than that in the control group. OPLS-DA was performed to reveal the difference in metabolites between the control and IHg-challenged groups, the metabolites of glucose, glutamine, inosine, succinate, glutamate, homarine, and alanine were sensitive to IHg, subsequently metabolic pathways that were affected including glucose metabolism, glutamine metabolism, nucleotide metabolism, Krebs cycle, amino acid metabolism and osmotic regulation. In our study, IHg interfered with metabolites in M. chinensis philippi, thus the corresponding metabolic pathways were changed, which influenced the neurotransmitters subsequently. Furthermore, Ca2+overload affected the synthesis or degradation of the neurotransmitters, and then the altered neurotransmitters involved in changes in metabolic pathways again. Overall, we hypothesized that the neurotoxic effects of IHg on bivalve were in close contact with metabolism, neurotransmitters, related enzymes and Ca2+, which could be effective neurotoxic biomarkers for marine environmental quality assessment, and also provide effective data for the study of the regulatory mechanism of the nervous system in response to IHg in bivalves.

Role of miRNAs in glucose metabolism of mouse cumulus cells†.

It is known that the oocyte has a limited capacity to acquire and metabolize glucose, and it must rely on cumulus cells (CCs) to take up glucose and produce pyruvate for use to produce ATP through oxidative phosphorylation. We therefore propose that miRNAs might regulate glucose metabolism (GM) in CCs and might be used as markers for oocyte quality assessment. Here, mouse CC models with impaired glycolysis or pentose phosphate pathway (PPP) were established, and miRNAs targeting the key enzymes in glycolysis/PPP were predicted using the miRNA target prediction databases. Expression of the predicted miRNAs was compared between CCs with normal and impaired glycolysis/PPP to identify candidate miRNAs. Function of the candidate miRNAs was validated by transfecting CCs or cumulus-oocyte-complexes (COCs) with miRNA inhibitors and observing effects on glucose metabolites of CCs and on competence of oocytes. The results validated that miR-23b-3p, let-7b-5p, 34b-5p and 145a-5p inhibited glycolysis, and miR-24-3p, 3078-3p,183-5p and 7001-5p inhibited PPP of CCs. Our observation using a more physiologically relevant model (intact cultured COCs) further validated the four glycolysis-targeting miRNAs we identified. Furthermore, miR-let-7b-5p, 34b-5p and 145a-5p may also inhibit PPP, as they decreased the production of glucose-6-phosphate. In conclusion, miRNAs play critical roles in GM of CCs and may be used as markers for oocyte quality assessment. Summary sentence: We identified and validated eight new miRNAs that inhibit glycolysis and/or pentose phosphate pathways in cumulus cells (CCs) suggesting that miRNAs play critical roles in glucose metabolism of CCs and may be used for oocyte quality markers.