Stevioside Ameliorates Palmitic Acid-Induced Abnormal Glucose Uptake via the PDK4/AMPK/TBC1D1 Pathway in C2C12 Myotubes.

BACKGROUND: Stevioside (SV) with minimal calories is widely used as a natural sweetener in beverages due to its high sweetness and safety. However, the effects of SV on glucose uptake and the pyruvate dehydrogenase kinase isoenzyme (PDK4) as an important protein in the regulation of glucose metabolism, remain largely unexplored. In this study, we used C2C12 skeletal muscle cells that was induced by palmitic acid (PA) to assess the effects and mechanisms of SV on glucose uptake and PDK4. METHODS: The glucose uptake of C2C12 cells was determined by 2-NBDG; expression of the Pdk4 gene was measured by quantitative real-time PCR; and expression of the proteins PDK4, p-AMPK, TBC1D1 and GLUT4 was assessed by Western blotting. RESULTS: In PA-induced C2C12 myotubes, SV could significantly promote cellular glucose uptake by decreasing PDK4 levels and increasing p-AMPK and TBC1D1 levels. SV could promote the translocation of GLUT4 from the cytoplasm to the cell membrane in cells. Moreover, in Pdk4-overexpressing C2C12 myotubes, SV decreased the level of PDK4 and increased the levels of p-AMPK and TBC1D1. CONCLUSION: SV was found to ameliorate PA-induced abnormal glucose uptake via the PDK4/AMPK/TBC1D1 pathway in C2C12 myotubes. Although these results warranted further investigation for validation, they may provide some evidence of SV as a safe natural sweetener for its use in sugar-free beverages to prevent and control T2DM.

LIN28B induced PCAT5 promotes endometrial cancer progression and glycolysis via IGF2BP3 deubiquitination.

Endometrial cancer (EC) cells exhibit abnormal glucose metabolism, characterized by increased aerobic glycolysis and decreased oxidative phosphorylation. Targeting cellular glucose metabolism in these cells could be an effective therapeutic approach for EC. This study aimed to assess the roles of LIN28B, PCAT5, and IGF2BP3 in the glucose metabolism, proliferation, migration, and invasion of EC cells. LIN28B highly expressed in EC, binds and stabilizes PCAT5. PCAT5, overexpressed in EC, and its 1485-2288nt region can bind to the KH1-2 domain of IGF2BP3 to prevent MKRN2 from binding to the K294 ubiquitination site of IGF2BP3, thus stabilizing IGF2BP3. Finally, IGF2BP3 promotes the aerobic glycolysis, proliferation, migration and invasion of EC cells by stabilizing the key enzymes of glucose metabolism HK2 and PKM2. Taken together, our data reveal that the LIN28B/PCAT5/IGF2BP3 axis is critical for glucose reprogramming and malignant biological behavior in EC cells. Therefore, targeting this axis may contribute to the development of a novel therapeutic strategy for EC metabolism.

GLUT4 localisation with the plasma membrane is unaffected by an increase in plasma free fatty acid availability.

BACKGROUND: Insulin-stimulated glucose uptake into skeletal muscle occurs via translocation of GLUT4 from intracellular storage vesicles to the plasma membrane. Elevated free fatty acid (FFA) availability via a lipid infusion reduces glucose disposal, but this occurs in the absence of impaired proximal insulin signalling. Whether GLUT4 localisation to the plasma membrane is subsequently affected by elevated FFA availability is not known. METHODS: Trained (n = 11) and sedentary (n = 10) individuals, matched for age, sex and body mass index, received either a 6 h lipid or glycerol infusion in the setting of a concurrent hyperinsulinaemic-euglycaemic clamp. Sequential muscle biopsies (0, 2 and 6 h) were analysed for GLUT4 membrane localisation and microvesicle size and distribution using immunofluorescence microscopy. RESULTS: At baseline, trained individuals had more small GLUT4 spots at the plasma membrane, whereas sedentary individuals had larger GLUT4 spots. GLUT4 localisation with the plasma membrane increased at 2 h (P = 0.04) of the hyperinsulinemic-euglycemic clamp, and remained elevated until 6 h, with no differences between groups or infusion type. The number of GLUT4 spots was unchanged at 2 h of infusion. However, from 2 to 6 h there was a decrease in the number of small GLUT4 spots at the plasma membrane (P = 0.047), with no differences between groups or infusion type. CONCLUSION: GLUT4 localisation with the plasma membrane increases during a hyperinsulinemic-euglycemic clamp, but this is not altered by elevated FFA availability. GLUT4 appears to disperse from small GLUT4 clusters located at the plasma membrane to support glucose uptake during a hyperinsulinaemic-euglycaemic clamp.

Effects of Saikosaponin-A on Insulin Resistance in Obesity: Computational and Animal Experimental Study.

Obesity is known to be associated with increased inflammation and dysregulated autophagy, both of which contribute to insulin resistance. Saikosaponin-A (SSA) has been reported to exhibit anti-inflammatory and lipid-lowering properties. In this research, we employed a combination of computational modeling and animal experiments to explore the effects of SSA. Male C57BL/6 mice were categorized into four groups: normal diet, high-fat diet (HFD), HFD + atorvastatin 10 mg/kg, and HFD + SSA 10 mg/kg. We conducted oral glucose and fat tolerance tests to assess metabolic parameters and histological changes. Furthermore, we evaluated the population of Kupffer cells (KCs) and examined gene expressions related to inflammation and autophagy. Computational analysis revealed that SSA displayed high binding affinity to tumor necrosis factor (TNF)-α, nuclear factor (NF)-κB, fibroblast growth factor 21 (FGF21), and autophagy-related 7 (ATG7). Animal study demonstrated that SSA administration improved fasting and postprandial glucose levels, homeostatic model assessment of insulin resistance (HOMA-IR) index, as well as triglyceride, free fatty acid, total cholesterol, low-density lipoprotein cholesterol (LDL-C)-cholesterol, and high-density lipoprotein cholesterol (HDL-C)-cholesterol levels in HFD-fed mice. Moreover, SSA significantly reduced liver weight and fat accumulation, while inhibiting the infiltration and M1 activation of KCs. At the mRNA level, SSA downregulated TNF-α and NF-κB expression, while upregulating FGF21 and ATG7 expression. In conclusion, our study suggests that SSA may serve as a therapeutic agent for addressing the metabolic complications associated with obesity. This potential therapeutic effect is attributed to the suppression of inflammatory cytokines and the upregulation of FGF21 and ATG7.

Gastrin: a new branch of the gastropancreatic axis that can explain the effect of sleeve gastrectomy on glucose metabolism.

BACKGROUND: Among bariatric techniques, sleeve gastrectomy (SG) stands out owing to its efficiency. The role of the stomach as a secretory organ of many substances, such as gastrin, related to insulin secretion is well known. Gastrin induces insulin release in isolated pancreatic islets, limiting somatostatin-14 intraislet release, and has been associated with blood glucose level improvement in diabetic models after SG. SG involves gastric resection along the greater curvature. This study aimed to determine the role of gastrin in glucose metabolism improvement after SG with the aid of the gastrin antagonist netazepide. METHODS: In 12 sham-operated, 12 SG-operated, and 12 SG-operated/netazepide-treated Wistar rats, we compared medium- and long-term plasma insulin, oral glucose tolerance test (OGTT) results, and plasma gastrin levels. In addition, gastrin expression was assessed in the gastric remnant, and the beta-cell mass was measured. RESULTS: SG induced a medium-term elevation of the insulin response and plasma gastrin levels without modification of the OGTT results. However, long-term depletion of the insulin response with elevated OGTT areas under the curve and plasma gastrin levels appeared after SG. Netazepide prevented the SG effect on these parameters. Gastrin tissue expression was greater in SG animals than in SG/netazepide-treated or control animals. The beta-cell mass was lower in the SG group than in the control or SG/netazepide group. CONCLUSION: Gastrin plays a central role in glucose improvement after SG. It stimulates a medium-term strong insulin response but also causes long-term beta-cell mass depletion and a loss of insulin response. These effects are prevented by gastrin antagonists such as netazepide.

Aberrant splicing of CaV1.2 calcium channel induced by decreased Rbfox1 enhances arterial constriction during diabetic hyperglycemia.

Diabetic hyperglycemia induces dysfunctions of arterial smooth muscle, leading to diabetic vascular complications. The CaV1.2 calcium channel is one primary pathway for Ca2+ influx, which initiates vasoconstriction. However, the long-term regulation mechanism(s) for vascular CaV1.2 functions under hyperglycemic condition remains unknown. Here, Sprague-Dawley rats fed with high-fat diet in combination with low dose streptozotocin and Goto-Kakizaki (GK) rats were used as diabetic models. Isolated mesenteric arteries (MAs) and vascular smooth muscle cells (VSMCs) from rat models were used to assess K+-induced arterial constriction and CaV1.2 channel functions using vascular myograph and whole-cell patch clamp, respectively. K+-induced vasoconstriction is persistently enhanced in the MAs from diabetic rats, and CaV1.2 alternative spliced exon 9* is increased, while exon 33 is decreased in rat diabetic arteries. Furthermore, CaV1.2 channels exhibit hyperpolarized current-voltage and activation curve in VSMCs from diabetic rats, which facilitates the channel function. Unexpectedly, the application of glycated serum (GS), mimicking advanced glycation end-products (AGEs), but not glucose, downregulates the expression of the splicing factor Rbfox1 in VSMCs. Moreover, GS application or Rbfox1 knockdown dynamically regulates alternative exons 9* and 33, leading to facilitated functions of CaV1.2 channels in VSMCs and MAs. Notably, GS increases K+-induced intracellular calcium concentration of VSMCs and the vasoconstriction of MAs. These results reveal that AGEs, not glucose, long-termly regulates CaV1.2 alternative splicing events by decreasing Rbfox1 expression, thereby enhancing channel functions and increasing vasoconstriction under diabetic hyperglycemia. This study identifies the specific molecular mechanism for enhanced vasoconstriction under hyperglycemia, providing a potential target for managing diabetic vascular complications.

Oral post-exercise garlic extract supplementation enhances glycogen replenishment but does not up-regulate mitochondria biogenesis mRNA expression in human-exercised skeletal muscle.

PURPOSE: Garlic extract (GA) is purported to enhance antioxidant and anti-inflammatory activity and glucose regulation in humans. The present study investigated the effects of post-exercise GA supplementation on GLUT4 expression, glycogen replenishment, and the transcript factors involved with mitochondrial biosynthesis in exercised human skeletal muscle. METHODS: The single-blinded crossover counterbalanced study was completed by 12 participants. Participants were randomly divided into either GA (2000 mg of GA) or placebo trials immediately after completing a single bout of cycling exercise at 75% Maximal oxygen uptake (VO2max) for 60 minutes. Participants consumed either GA (2000 mg) or placebo capsules with a high glycemic index carbohydrate meal (2 g carb/body weight) immediately after exercise. Muscle samples were collected at 0-h and 3-h post-exercise. Muscle samples were used to measure glycogen levels, GLUT4 protein expression, as well as transcription factors for glucose uptake, and mitochondria biogenesis. Plasma glucose, insulin, glycerol, non-esterified fatty acid (NEFA) concentrations, and respiratory exchange ratio (RER) were also analyzed during the post-exercise recovery periods. RESULTS: Skeletal muscle glycogen replenishment was significantly elevated during the 3-h recovery period for GA concurrent with no difference in GLUT4 protein expression between the garlic and placebo trials. PGC1-α gene expression was up-regulated for both GA and placebo after exercise (p < 0.05). Transcript factors corresponding to muscle mitochondrial biosynthesis were significantly enhanced under acute garlic supplementation as demonstrated by TFAM and FIS1. However, the gene expression of SIRT1, ERRα, NFR1, NFR2, MFN1, MFN2, OPA1, Beclin-1, DRP1 were not enhanced, nor were there any improvements in GLUT4 expression, following post-exercise garlic supplementation. CONCLUSION: Acute post-exercise garlic supplementation may improve the replenishment of muscle glycogen, but this appears to be unrelated to the gene expression for glucose uptake and mitochondrial biosynthesis in exercised human skeletal muscle.

Impact of cannabidiol on brain glucose metabolism of C57Bl/6 male mice previously exposed to cocaine.

Despite evidence of the beneficial effects of cannabidiol (CBD) in animal models of cocaine use disorder (CUD), CBD neuronal mechanisms remain poorly understood. This study investigated the effects of CBD treatment on brain glucose metabolism, in a CUD animal model, using [18F]FDG positron emission tomography (PET). Male C57Bl/6 mice were injected with cocaine (20 mg/kg, i.p.) every other day for 9 days, followed by 8 days of CBD administration (30 mg/kg, i.p.). After 48 h, animals were challenged with cocaine. Control animals received saline/vehicle. [18F]FDG PET was performed at four time points: baseline, last day of sensitization, last day of withdrawal/CBD treatment, and challenge. Subsequently, the animals were euthanized and immunohistochemistry was performed on the hippocampus and amygdala to assess the CB1 receptors, neuronal nuclear protein, microglia (Iba1), and astrocytes (GFAP). Results showed that cocaine administration increased [18F]FDG uptake following sensitization. CBD treatment also increased [18F]FDG uptake in both saline and cocaine groups. However, animals that were sensitized and challenged with cocaine, and those receiving only an acute cocaine injection during the challenge phase, did not exhibit increased [18F]FDG uptake when treated with CBD. Furthermore, CBD induced modifications in the integrated density of NeuN, Iba, GFAP, and CB1R in the hippocampus and amygdala. This is the first study addressing the impact of CBD on brain glucose metabolism in a preclinical model of CUD using PET. Our findings suggest that CBD disrupts cocaine-induced changes in brain energy consumption and activity, which might be correlated with alterations in neuronal and glial function.

Decoding the role of oxidative stress resistance and alternative carbon substrate assimilation in the mature biofilm growth mode of Candida glabrata.

BACKGROUND: Biofilm formation is viewed as a vital mechanism in C. glabrata pathogenesis. Although, it plays a significant role in virulence but transcriptomic architecture and metabolic pathways governing the biofilm growth mode of C. glabrata remain elusive. The present study intended to investigate the genes implicated in biofilm growth phase of C. glabrata through global transcriptomic approach. RESULTS: Functional analysis of Differentially expressed genes (DEGs) using gene ontology and pathways analysis revealed that upregulated genes are involved in the glyoxylate cycle, carbon-carbon lyase activity, pre-autophagosomal structure membrane and vacuolar parts whereas, down- regulated genes appear to be associated with glycolysis, ribonucleoside biosynthetic process, ribosomal and translation process in the biofilm growth condition. The RNA-Seq expression of eight selected DEGs (CgICL1, CgMLS1, CgPEP1, and CgNTH1, CgERG9, CgERG11, CgTEF3, and CgCOF1) was performed with quantitative real-time PCR (RT-qPCR). The gene expression profile of selected DEGs with RT-qPCR displayed a similar pattern of expression as observed in RNA-Seq. Phenotype screening of mutant strains generated for genes CgPCK1 and CgPEP1, showed that Cgpck1∆ failed to grow on alternative carbon substrate (Glycerol, Ethanol, Oleic acid) and similarly, Cgpep1∆ unable to grow on YPD medium supplemented with hydrogen peroxide. Our results suggest that in the absence of glucose, C. glabrata assimilate glycerol, oleic acid and generate acetyl coenzyme-A (acetyl-CoA) which is a central and connecting metabolite between catabolic and anabolic pathways (glyoxylate and gluconeogenesis) to produce glucose and fulfil energy requirements. CONCLUSIONS: The study was executed using various approaches (transcriptomics, functional genomics and gene deletion) and it revealed that metabolic plasticity of C. glabrata (NCCPF-100,037) in biofilm stage modulates its virulence and survival ability to counter the stress and may promote its transition from commensal to opportunistic pathogen. The observations deduced from the present study along with future work on characterization of the proteins involved in this intricate process may prove to be beneficial for designing novel antifungal strategies.

Breast cancer therapy: from the perspective of glucose metabolism and glycosylation.

Breast cancer is a leading cause of mortality and the most prevalent form of malignant tumor among women worldwide. Breast cancer cells exhibit an elevated glycolysis and altered glucose metabolism. Moreover, these cells display abnormal glycosylation patterns, influencing invasion, proliferation, metastasis, and drug resistance. Consequently, targeting glycolysis and mitigating abnormal glycosylation represent key therapeutic strategies for breast cancer. This review underscores the importance of protein glycosylation and glucose metabolism alterations in breast cancer. The current research efforts in developing effective interventions targeting glycolysis and glycosylation are further discussed.

Endocytic vesicles act as vehicles for glucose uptake in response to growth factor stimulation.

Glycolysis is a fundamental cellular process, yet its regulatory mechanisms remain incompletely understood. Here, we show that a subset of glucose transporter 1 (GLUT1/SLC2A1) co-endocytoses with platelet-derived growth factor (PDGF) receptor (PDGFR) upon PDGF-stimulation. Furthermore, multiple glycolytic enzymes localize to these endocytosed PDGFR/GLUT1-containing vesicles adjacent to mitochondria. Contrary to current models, which emphasize the importance of glucose transporters on the cell surface, we find that PDGF-stimulated glucose uptake depends on receptor/transporter endocytosis. Our results suggest that growth factors generate glucose-loaded endocytic vesicles that deliver glucose to the glycolytic machinery in proximity to mitochondria, and argue for a new layer of regulation for glycolytic control governed by cellular membrane dynamics.

International Roadshow: New Advances in Endocrinology and Metabolic Diseases.

Dear Readers,Currently, there is a myriad of new developments in the field of endocrinology. In particular, significant strides have been made in the development of poly-agonists for the treatment of type 2 diabetes and obesity 1 2. Poly-agonists represent a novel therapeutic approach by combining multiple actions within a single molecule, targeting multiple receptors simultaneously to achieve enhanced efficacy. These innovative compounds aim to address the complex interplay of hormonal pathways involved in glucose regulation and metabolism, offering potential breakthroughs in the management of diabetes and obesity.

Unraveling the significance of PPP1R1A gene in pancreatic ß-cell function: A study in INS-1 cells and human pancreatic islets.

BACKGROUND AND AIMS: The protein phosphatase 1 regulatory inhibitor subunit 1A (PPP1R1A) has been linked with insulin secretion and diabetes mellitus. Yet, its full significance in pancreatic ß-cell function remains unclear. This study aims to elucidate the role of the PPP1R1A gene in ß-cell biology using human pancreatic islets and rat INS-1 (832/13) cells. RESULTS: Disruption of Ppp1r1a in INS-1 cells was associated with reduced insulin secretion and impaired glucose uptake; however, cell viability, ROS, apoptosis or proliferation were intact. A significant downregulation of crucial ß-cell function genes such as Ins1, Ins2, Pcsk1, Cpe, Pdx1, Mafa, Isl1, Glut2, Snap25, Vamp2, Syt5, Cacna1a, Cacna1d and Cacnb3, was observed upon Ppp1r1a disruption. Furthermore, silencing Pdx1 in INS-1 cells altered PPP1R1A expression, indicating that PPP1R1A is a target gene for PDX1. Treatment with rosiglitazone increased Ppp1r1a expression, while metformin and insulin showed no effect. RNA-seq analysis of human islets revealed high PPP1R1A expression, with α-cells showing the highest levels compared to other endocrine cells. Muscle tissues exhibited greater PPP1R1A expression than pancreatic islets, liver, or adipose tissues. Co-expression analysis revealed significant correlations between PPP1R1A and genes associated with insulin biosynthesis, exocytosis machinery, and intracellular calcium transport. Overexpression of PPP1R1A in human islets augmented insulin secretion and upregulated protein expression of Insulin, MAFA, PDX1, and GLUT1, while silencing of PPP1R1A reduced Insulin, MAFA, and GLUT1 protein levels. CONCLUSION: This study provides valuable insights into the role of PPP1R1A in regulating ß-cell function and glucose homeostasis. PPP1R1A presents a promising opportunity for future therapeutic interventions.

Addition of low sodium does not increase sensitivity to glucose in wild-type mice, or lead to partial glucose taste detection in T1R3 knock-out mice.

The sodium glucose cotransporter 1 (SGLT1) has been proposed as a non-T1R glucosensor contributing to glucose taste. Studies have shown that the addition of NaCl at very weak concentrations to a glucose stimulus can enhance signaling in the gustatory nerves of mice and significantly lower glucose detection thresholds in humans. Here, we trained mice with (wild-type; WT) and without (knockout; KO) a functioning T1R3 subunit on a two-response operant detection task to differentially respond to the presence or absence of a taste stimulus immediately after sampling. After extensive training (∼40 sessions), KO mice were unable to reliably discriminate 2 M glucose+0.01 M NaCl from 0.01 M NaCl alone, but all WT mice could. We then tested WT mice on a descending array of glucose concentrations (2.0-0.03 M) with the addition of 0.01 M NaCl vs. 0.01 M NaCl alone. The concentration series was then repeated with glucose alone vs. water. We found no psychophysical evidence of a non-T1R taste transduction pathway involved in the detection of glucose. The addition of NaCl to glucose did not lower taste detection thresholds in WT mice, nor did it render the stimulus detectable to KO mice, even at 2 M. The proposed pathway must contribute to functions other than sensory-discriminative detection, at least when tested under these conditions. Detection thresholds were also derived for fructose and found to be 1/3 log10 lower than for glucose, but highly correlated (r = 0.88) between the two sugars, suggesting that sensitivity to these stimuli in this task was based on a similar neural process.

Hormonal effects on glucose and ketone metabolism in a perfused liver of an elasmobranch, the North Pacific spiny dogfish, Squalus suckleyi.

Hormonal influence on hepatic function is a critical aspect of whole-body energy balance in vertebrates. Catecholamines and corticosteroids both influence hepatic energy balance via metabolite mobilization through glycogenolysis and gluconeogenesis. Elasmobranchs have a metabolic organization that appears to prioritize the mobilization of hepatic lipid as ketone bodies (e.g. 3-hydroxybutyrate [3-HB]), which adds complexity in determining the hormonal impact on hepatic energy balance in this taxon. Here, a liver perfusion was used to investigate catecholamine (epinephrine [E]) and corticosteroid (corticosterone [B] and 11-deoxycorticosterone [DOC]) effects on the regulation of hepatic glucose and 3-HB balance in the North Pacific Spiny dogfish, Squalus suckleyi. Further, hepatic enzyme activity involved in ketogenesis (3-hydroxybutyrate dehydrogenase), glycogenolysis (glycogen phosphorylase), and gluconeogenesis (phosphoenolpyruvate carboxykinase) were assessed in perfused liver tissue following hormonal application to discern effects on hepatic energy flux. mRNA transcript abundance key transporters of glucose (glut1 and glut4) and ketones (mct1 and mct2) and glucocorticoid function (gr, pepck, fkbp5, and 11ßhsd2) were also measured to investigate putative cellular components involved in hepatic responses. There were no changes in the arterial-venous difference of either metabolite in all hormone perfusions. However, perfusion with DOC increased gr transcript abundance and decreased flow rate of perfusions, suggesting a regulatory role for this corticosteroid. Phosphoenolpyruvate carboxykinase activity increased following all hormone treatments, which may suggest gluconeogenic function; E also increased 3-hydroxybutyrate dehydrogenase activity, suggesting a function in ketogenesis, and decreased pepck and fkbp5 transcript abundance, potentially showing some metabolic regulation. Overall, we demonstrate hormonal control of hepatic energy balance using liver perfusions at various levels of biological organization in an elasmobranch.

Two hexokinases of the shrimp Penaeus (Litopenaeus) vannamei are differentially expressed during oxygen limited conditions.

The white shrimp Penaeus (Litopenaeus) vannamei is the most cultivated shrimp worldwide. Compared to other shrimp species, it has higher resistance to adverse conditions. During hypoxia, the shrimp reduces oxygen consumption and adjusts energy metabolism via anaerobic glycolysis, among other strategies. Hexokinase (HK) is the first enzyme of glycolysis and a key regulation point. In mammals and other vertebrates, there are several tissue-specific HK isoforms with differences in expression and enzyme activity. In contrast, crustacean HKs have been relatively little studied. We studied the P. vannamei HK isoforms during hypoxia and reoxygenation. We cloned two HK1 sequences named HK1-long (1455 bp) and HK1-short (1302 bp), and one HK2 (1344 bp). In normoxia, total HK1 expression is higher in hepatopancreas, while HK2 is higher in gills. Severe hypoxia (1 mg/L of DO) after 12 h exposure and 1 h of reoxygenation increased HK1 expression in both organs, but HK2 expression changed differentially. In hepatopancreas, HK2 expression increased in 6 and 12 h of hypoxia but diminished to normoxia levels after reoxygenation. In gills, HK2 expression decreased after 12 h of hypoxia. HK activity increased in hepatopancreas after 12 h hypoxia, opposite to gills. These results indicate that shrimp HK isoforms respond to hypoxia and reoxygenation in a tissue-specific manner. Intracellular glucose levels did not change in any case, showing the shrimp ability to maintain glucose homeostasis during hypoxia.

Exercise increases TCA intermediate concentrations during low-calorie diet independent of insulin resistance among women with obesity.

Tricarboxylic acid cycle intermediates (TCAi) have been proposed to act as myokines that influence energy metabolism. We determined if 2-weeks of low-calorie diet with interval exercise (LCD + INT) would increase TCAi more than a low-calorie diet (LCD). Twenty-three women were randomized to 2-weeks of LCD (n = 12, 48.4 ± 2.5 years, 37.8 ± 1.5 kg/m2, ~1200 kcal/d) or LCD + INT (n = 11, 47.6 ± 4.3 years, 37.9 ± 2.3 kg/m2; 60 min/d supervised INT of 3 min 90% & 50% HRpeak). TCAi and amino acids (AA) were measured at 0 min of a 75 g OGTT, while glucose, insulin, and FFA were obtained at 0, 30, 60, 90, 120, and 180 min to assess total area under the curve (tAUC180min) and insulin resistance (IR; tAUC180min of Glucose × Insulin). Fuel use (indirect calorimetry) was also collected at 0, 60, 120, and 180 min as was fitness (VO2peak) and body composition (BodPod). Treatments reduced weight (p < 0.001), fasting RER (p = 0.01), and IR (p = 0.03), although LCD + INT increased VO2peak (p = 0.02) and maintained RER tAUC180min (p = 0.05) versus LCD. Treatments increased FFA tAUC180min (p = 0.005), cis-aconitate, isocitrate, and succinate (p ≤ 0.02), as well as reduced phenylalanine and tryptophan, cysteine (p ≤ 0.005). However, LCD + INT increased malate, citrate, α-ketoglutarate, and alanine more than LCD (p ≤ 0.04). Thus, INT enhanced LCD effects on some TCAi in women with obesity independent of IR.

Transcriptomics aids in uncovering the metabolic shifts and molecular machinery of Schizochytrium limacinum during biotransformation of hydrophobic substrates to docosahexaenoic acid.

BACKGROUND: Biotransformation of waste oil into value-added nutraceuticals provides a sustainable strategy. Thraustochytrids are heterotrophic marine protists and promising producers of omega (ω) fatty acids. Although the metabolic routes for the assimilation of hydrophilic carbon substrates such as glucose are known for these microbes, the mechanisms employed for the conversion of hydrophobic substrates are not well established. Here, thraustochytrid Schizochytrium limacinum SR21 was investigated for its ability to convert oils (commercial oils with varying fatty acid composition and waste cooking oil) into ω-3 fatty acid; docosahexaenoic acid (DHA). RESULTS: Within 72 h SR21 consumed ~ 90% of the oils resulting in enhanced biomass (7.5 g L- 1) which was 2-fold higher as compared to glucose. Statistical analysis highlights C16 fatty acids as important precursors of DHA biosynthesis. Transcriptomic data indicated the upregulation of multiple lipases, predicted to possess signal peptides for secretory, membrane-anchored and cytoplasmic localization. Additionally, transcripts encoding for mitochondrial and peroxisomal ß-oxidation along with acyl-carnitine transporters were abundant for oil substrates that allowed complete degradation of fatty acids to acetyl CoA. Further, low levels of oxidative biomarkers (H2O2, malondialdehyde) and antioxidants were determined for hydrophobic substrates, suggesting that SR21 efficiently mitigates the metabolic load and diverts the acetyl CoA towards energy generation and DHA accumulation. CONCLUSIONS: The findings of this study contribute to uncovering the route of assimilation of oil substrates by SR21. The thraustochytrid employs an intricate crosstalk among the extracellular and intracellular molecular machinery favoring energy generation. The conversion of hydrophobic substrates to DHA can be further improved using synthetic biology tools, thereby providing a unique platform for the sustainable recycling of waste oil substrates.

The long-chain flavodoxin FldX1 improves the biodegradation of 4-hydroxyphenylacetate and 3-hydroxyphenylacetate and counteracts the oxidative stress associated to aromatic catabolism in Paraburkholderia xenovorans.

BACKGROUND: Bacterial aromatic degradation may cause oxidative stress. The long-chain flavodoxin FldX1 of Paraburkholderia xenovorans LB400 counteracts reactive oxygen species (ROS). The aim of this study was to evaluate the protective role of FldX1 in P. xenovorans LB400 during the degradation of 4-hydroxyphenylacetate (4-HPA) and 3-hydroxyphenylacetate (3-HPA). METHODS: The functionality of FldX1 was evaluated in P. xenovorans p2-fldX1 that overexpresses FldX1. The effects of FldX1 on P. xenovorans were studied measuring growth on hydroxyphenylacetates, degradation of 4-HPA and 3-HPA, and ROS formation. The effects of hydroxyphenylacetates (HPAs) on the proteome (LC-MS/MS) and gene expression (qRT-PCR) were quantified. Bioaugmentation with strain p2-fldX1 of 4-HPA-polluted soil was assessed, measuring aromatic degradation (HPLC), 4-HPA-degrading bacteria, and plasmid stability. RESULTS: The exposure of P. xenovorans to 4-HPA increased the formation of ROS compared to 3-HPA or glucose. P. xenovorans p2-fldX1 showed an increased growth on 4-HPA and 3-HPA compared to the control strain WT-p2. Strain p2-fldX1 degraded faster 4-HPA and 3-HPA than strain WT-p2. Both WT-p2 and p2-fldX1 cells grown on 4-HPA displayed more changes in the proteome than cells grown on 3-HPA in comparison to glucose-grown cells. Several enzymes involved in ROS detoxification, including AhpC2, AhpF, AhpD3, KatA, Bcp, CpoF1, Prx1 and Prx2, were upregulated by hydroxyphenylacetates. Downregulation of organic hydroperoxide resistance (Ohr) and DpsA proteins was observed. A downregulation of the genes encoding scavenging enzymes (katE and sodB), and gstA and trxB was observed in p2-fldX1 cells, suggesting that FldX1 prevents the antioxidant response. More than 20 membrane proteins, including porins and transporters, showed changes in expression during the growth of both strains on hydroxyphenylacetates. An increased 4-HPA degradation by recombinant strain p2-fldX1 in soil microcosms was observed. In soil, the strain overexpressing the flavodoxin FldX1 showed a lower plasmid loss, compared to WT-p2 strain, suggesting that FldX1 contributes to bacterial fitness. Overall, these results suggest that recombinant strain p2-fldX1 is an attractive bacterium for its application in bioremediation processes of aromatic compounds. CONCLUSIONS: The long-chain flavodoxin FldX1 improved the capability of P. xenovorans to degrade 4-HPA in liquid culture and soil microcosms by protecting cells against the degradation-associated oxidative stress.

Evaluating the effect of rapamycin treatment in Alzheimer's disease and aging using in vivo imaging: the ERAP phase IIa clinical study protocol.

BACKGROUND: Rapamycin is an inhibitor of the mechanistic target of rapamycin (mTOR) protein kinase, and preclinical data demonstrate that it is a promising candidate for a general gero- and neuroprotective treatment in humans. Results from mouse models of Alzheimer's disease have shown beneficial effects of rapamycin, including preventing or reversing cognitive deficits, reducing amyloid oligomers and tauopathies and normalizing synaptic plasticity and cerebral glucose uptake. The "Evaluating Rapamycin Treatment in Alzheimer's Disease using Positron Emission Tomography" (ERAP) trial aims to test if these results translate to humans through evaluating the change in cerebral glucose uptake following six months of rapamycin treatment in participants with early-stage Alzheimer's disease. METHODS: ERAP is a six-month-long, single-arm, open-label, phase IIa biomarker-driven study evaluating if the drug rapamycin can be repurposed to treat Alzheimer's disease. Fifteen patients will be included and treated with a weekly dose of 7 mg rapamycin for six months. The primary endpoint will be change in cerebral glucose uptake, measured using [18F]FDG positron emission tomography. Secondary endpoints include changes in cognitive measures, markers in cerebrospinal fluid as well as cerebral blood flow measured using magnetic resonance imaging. As exploratory outcomes, the study will assess change in multiple age-related pathological processes, such as periodontal inflammation, retinal degeneration, bone mineral density loss, atherosclerosis and decreased cardiac function. DISCUSSION: The ERAP study is a clinical trial using in vivo imaging biomarkers to assess the repurposing of rapamycin for the treatment of Alzheimer's disease. If successful, the study would provide a strong rationale for large-scale evaluation of mTOR-inhibitors as a potential disease-modifying treatment in Alzheimer's disease. TRIAL REGISTRATION: ClinicalTrials.gov ID NCT06022068, date of registration 2023-08-30.