Glucose Transporter Function Research Articles

Regulation and biological function of a flagellar glucose transporter in Leishmania mexicana: a potential glucose sensor.

In Leishmania mexicana parasites, a unique glucose transporter, LmxGT1, is selectively targeted to the flagellar membrane, suggesting a possible sensory role that is often associated with ciliary membrane proteins. Expression of LmxGT1 is down-regulated ∼20-fold by increasing cell density but is up-regulated ∼50-fold by depleting glucose from the medium, and the permease is strongly down-regulated when flagellated insect-stage promastigotes invade mammalian macrophages and transform into intracellular amastigotes. Regulation of LmxGT1 expression by glucose and during the lifecycle operates at the level of protein stability. Significantly, a ∆lmxgt1 null mutant, grown in abundant glucose, undergoes catastrophic loss of viability when parasites deplete glucose from the medium, a property not exhibited by wild-type or add-back lines. These results suggest that LmxGT1 may function as a glucose sensor that allows parasites to enter the stationary phase when they deplete glucose and that in the absence of this sensor, parasites do not maintain viability when they run out of glucose. However, alternate roles for LmxGT1 in monitoring glucose availability are considered. The absence of known sensory receptors with defined ligands and biologic functions in Leishmania and related kinetoplastid parasites underscores the potential significance of these observations.

Expression and function of glucose transporter 1 (GLUT1) expression in activated hepatic stellate cells

Expression and function of glucose transporter 1 (GLUT1) expression in activated hepatic stellate cells

The Interrelationship between Diabetes and Depression

Both diabetes and depression are highly prevalent health problems that have a negative impact on various aspects of health, quality of life, and mortality. It has been well documented that depression is more common among people with diabetes than among the general population. Although the exact nature of the relationship between diabetes and depression is not fully understood, the bidirectional relationship of these two diseases has been suggested.1) A meta-analysis by Mezuk et al.2) showed that depression is associated with a 60% increased risk of type 2 diabetes and type 2 diabetes is associated with 15% increased risk of depression. Depression is associated with biochemical changes including increased activity of counter-regulatory hormones, altered glucose transport function, and increased production of proinflammatory cytokines, which contribute to insulin resistance, beta islet cell dysfunction, and ultimately diabetes. Depression is also associated with unhealthy behaviors including smoking, physical inactivity, and hypercaloric diets, which are risk factors for the development of diabetes. In patients with diabetes, the psychosocial burden of having a chronic illness may result in development of depression.3,4) Comorbid depression in patients with diabetes is related to poor glycemic control, higher severity of diabetic complications, increased risk of cardiovascular diseases, higher functional disability, and higher all-cause mortality.5) In the present issue, Sung et al.6) investigated the relationship between diabetes and depressive symptoms among Korean women using a nationally representative data of Korean. They obtained the data from the fifth Korea National Health and Nutrition Examination survey 2010-2011, which consisted of 6,572 Korean women aged 30 years and over. They found that diabetes has a relationship with depressive symptoms but that patients with diabetes were less likely to be treated for their depression. The result that women with diabetes have a higher likelihood of having depressive symptoms is consistent with findings from previous research. However, the authors report that there is a lack of prior research on the treatment of depression in individuals with diabetes. Gill et al.7) investigated the prescription of antidepressant medications for patients with medical comorbidities including diabetes in the primary care setting using electronic health record data. They found that primary care physicians prescribed antidepressant medication less aggressively for patients with multiple comorbidities, although there was no significant difference in the antidepressant medication prescription for diabetes alone. Patients with medical comorbidities may be already taking multiple medications for management of comorbid medical diseases so additional medications may increase their burden in relation to potential side effects and cost. Low rates of depression treatment in patients with diabetes may be related to a concern about the adverse metabolic effects of antidepressant medication.8) However, Sung et al.'s study is a cross-sectional design which does not prove causality. Although they described that diabetes is associated with an increased risk of depressive symptoms, the direction of relationship between diabetes and depressive symptoms was unclear. One limitation of this study was that depressive symptoms were assessed by the Patient Health Questionnaire-2 with low specificity, not by a formal depression diagnostic interview. The comorbidity of diabetes and depression is associated with significantly higher morbidity, disability, and mortality beyond those due to either diabetes or depression alone.9) Sung et al.6)'s study suggests that diabetes and depression frequently exist together and there is a possibility that depression treatment is suboptimal in patients with diabetes. Therefore, it should be emphasized that the coordinated care system which optimizes diagnosis and treatment of depression in patients with diabetes is necessary.

FGT-1 is the major glucose transporter in C. elegans and is central to aging pathways

Caenorhabditis elegans is widely used as a model for investigation of the relationships between aging, nutrient restriction and signalling via the DAF-2 (abnormal dauer formation 2) receptor for insulin-like peptides and AGE-1 [ageing alteration 1; orthologue of PI3K (phosphoinositide 3-kinase)], but the identity of the glucose transporters that may link these processes is unknown. We unexpectedly find that of the eight putative GLUT (glucose transporter)-like genes only the two splice variants of one gene have a glucose transport function in an oocyte expression system. We have named this gene fgt-1 (facilitated glucose transporter, isoform 1). We show that knockdown of fgt-1 RNA leads to loss of glucose transport and reduced glucose metabolism in wild-type worms. The FGT-1 glucose transporters of C. elegans thus play a key role in glucose energy supply to C. elegans. Importantly, knockdown of fgt-1 leads to an extension of lifespan equivalent, but not additive, to that observed in daf-2 and age-1 mutant worms. The results of the present study are consistent with DAF-2 and AGE-1 signalling stimulating glucose transport in C. elegans and this process being associated with the longevity phenotype in daf-2 and age-1 mutant worms. We propose that fgt-1 constitutes a common axis for the lifespan extending effects of nutrient restriction and reduced insulin-like peptide signalling.

A Small Volatile Bacterial Molecule Triggers Mitochondrial Dysfunction in Murine Skeletal Muscle

Mitochondria integrate distinct signals that reflect specific threats to the host, including infection, tissue damage, and metabolic dysfunction; and play a key role in insulin resistance. We have found that the Pseudomonas aeruginosa quorum sensing infochemical, 2-amino acetophenone (2-AA), produced during acute and chronic infection in human tissues, including in the lungs of cystic fibrosis (CF) patients, acts as an interkingdom immunomodulatory signal that facilitates pathogen persistence, and host tolerance to infection. Transcriptome results have led to the hypothesis that 2-AA causes further harm to the host by triggering mitochondrial dysfunction in skeletal muscle. As normal skeletal muscle function is essential to survival, and is compromised in many chronic illnesses, including infections and CF-associated muscle wasting, we here determine the global effects of 2-AA on skeletal muscle using high-resolution magic-angle-spinning (HRMAS), proton (1H) nuclear magnetic resonance (NMR) metabolomics, in vivo 31P NMR, whole-genome expression analysis and functional studies. Our results show that 2-AA when injected into mice, induced a biological signature of insulin resistance as determined by 1H NMR analysis-, and dramatically altered insulin signaling, glucose transport, and mitochondrial function. Genes including Glut4, IRS1, PPAR-γ, PGC1 and Sirt1 were downregulated, whereas uncoupling protein UCP3 was up-regulated, in accordance with mitochondrial dysfunction. Although 2-AA did not alter high-energy phosphates or pH by in vivo 31P NMR analysis, it significantly reduced the rate of ATP synthesis. This affect was corroborated by results demonstrating down-regulation of the expression of genes involved in energy production and muscle function, and was further validated by muscle function studies. Together, these results further demonstrate that 2-AA, acts as a mediator of interkingdom modulation, and likely effects insulin resistance associated with a molecular signature of mitochondrial dysfunction in skeletal muscle. Reduced energy production and mitochondrial dysfunctional may further favor infection, and be an important step in the establishment of chronic and persistent infections.

Estrogen: a master regulator of bioenergetic systems in the brain and body.

Estrogen is a fundamental regulator of the metabolic system of the female brain and body. Within the brain, estrogen regulates glucose transport, aerobic glycolysis, and mitochondrial function to generate ATP. In the body, estrogen protects against adiposity, insulin resistance, and type II diabetes, and regulates energy intake and expenditure. During menopause, decline in circulating estrogen is coincident with decline in brain bioenergetics and shift towards a metabolically compromised phenotype. Compensatory bioenergetic adaptations, or lack thereof, to estrogen loss could determine risk of late-onset Alzheimer's disease. Estrogen coordinates brain and body metabolism, such that peripheral metabolic state can indicate bioenergetic status of the brain. By generating biomarker profiles that encompass peripheral metabolic changes occurring with menopause, individual risk profiles for decreased brain bioenergetics and cognitive decline can be created. Biomarker profiles could identify women at risk while also serving as indicators of efficacy of hormone therapy or other preventative interventions.

KHARON1 Mediates Flagellar Targeting of a Glucose Transporter in Leishmania mexicana and Is Critical for Viability of Infectious Intracellular Amastigotes

The LmxGT1 glucose transporter is selectively targeted to the flagellum of the kinetoplastid parasite Leishmania mexicana, but the mechanism for targeting this and other flagella-specific membrane proteins among the Kinetoplastida is unknown. To address the mechanism of flagellar targeting, we employed in vivo cross-linking, tandem affinity purification, and mass spectrometry to identify a novel protein, KHARON1 (KH1), which is important for the flagellar trafficking of LmxGT1. Kh1 null mutant parasites are strongly impaired in flagellar targeting of LmxGT1, and trafficking of the permease was arrested in the flagellar pocket. Immunolocalization revealed that KH1 is located at the base of the flagellum, within the flagellar pocket, where it associates with the proximal segment of the flagellar axoneme. We propose that KH1 mediates transit of LmxGT1 from the flagellar pocket into the flagellar membrane via interaction with the proximal portion of the flagellar axoneme. KH1 represents the first component involved in flagellar trafficking of integral membrane proteins among parasitic protozoa. Of considerable interest, Kh1 null mutants are strongly compromised for growth as amastigotes within host macrophages. Thus, KH1 is also important for the disease causing stage of the parasite life cycle.

Age-related alteration in cerebral blood flow and energy failure is correlated with cognitive impairment in the senescence-accelerated prone mouse strain 8 (SAMP8)

Cerebrovascular dysfunction is an early pathogenic event in Alzheimer's disease (AD) and plays a key role in the disease process. Cerebral hypoperfusion, brain glucose hypometabolism and disrupted blood-brain barrier (BBB) integrity contributed to the onset and progression of AD. However, the relationships between the age-related cognitive impairment and cerebral blood flow (CBF), energy metabolism and BBB have not been clearly explained. In this study, we investigated the cognitive function, CBF, BBB damage and expression level of glucose transporter (GLUT) 1 and 3 of senescence-accelerated mouse prone 8 (SAMP8), and the correlations between each of them were analyzed. When compared with SAMR1 (senescence-accelerated mouse resistant 1), the cognitive abilities of SAMP8 were damaged apparently even at 4 months of age, showing up a slower and more capricious acquisition in Morris water maze tasks. In both SAMP8 and SAMR1, reduced CBF and increased BBB leakage were observed with increasing age, but an earlier and more severe impairment was detected in SAMP8. In addition, alterations of GLUT1 and GLUT3 protein expression in cortex and hippocampus were more prominent in SAMP8. Correlation analysis demonstrated that the increased escape latency was correlated negatively with CBF and expression of glucose transporters; and positively with BBB permeability in the hippocampus. These results suggested that CBF, BBB integrity, the expression of GLUT1 and GLUT3 were significantly affected by age and strain, which were also closely associated with cognitive ability. The alteration in CBF and energy failure induced by aging and vascular insults resulted in cognitive decline in SAMP8.

A global analysis of SNX27–retromer assembly and cargo specificity reveals a function in glucose and metal ion transport

The PDZ-domain-containing sorting nexin 27 (SNX27) promotes recycling of internalized transmembrane proteins from endosomes to the plasma membrane by linking PDZ-dependent cargo recognition to retromer-mediated transport. Here, we employed quantitative proteomics of the SNX27 interactome and quantification of the surface proteome of SNX27- and retromer-suppressed cells to dissect the assembly of the SNX27 complex and provide an unbiased global view of SNX27-mediated sorting. Over 100 cell surface proteins, many of which interact with SNX27, including the glucose transporter GLUT1, the Menkes disease copper transporter ATP7A, various zinc and amino acid transporters, and numerous signalling receptors, require SNX27-retromer to prevent lysosomal degradation and maintain surface levels. Furthermore, we establish that direct interaction of the SNX27 PDZ domain with the retromer subunit VPS26 is necessary and sufficient to prevent lysosomal entry of SNX27 cargo. Our data identify the SNX27-retromer as a major endosomal recycling hub required to maintain cellular nutrient homeostasis.

Mechanism of insulin resistance induced by bisphenol A

Bisphenol A(BPA) is one of the environmental endocrine disruptors with estrogenic properties,and it is also one of the most widespread compounds in the manufacture of polycarbonate plastics.Many studies confirmed that BPA exposure for long-term could lead to obesity,precocious puberty,insulin resistance (IR) et al.IR was defined as the decrease sensitivity in insulin action on the target tissues,which would result in impaired glucose tolerance and diabetes.A number of experiments and clinical investigations found that BPA could induce IR by affecting the function of islet beta cell,glucose transporter and adiponectin secretion.This review aims to summarize the mechanism of IR induced by BPA. Key words: Bisphenol A; Insulin resistance ; Environmental endocrine disruptor; Metabolic syndrome

Comparative effects of single-mode vs. duration-matched concurrent exercise training on body composition, low-grade inflammation, and glucose regulation in sedentary, overweight, middle-aged men

The effect of duration-matched concurrent exercise training (CET) (50% resistance (RET) and 50% endurance (EET) training) on physiological training outcomes in untrained middle-aged men remains to be elucidated. Forty-seven men (age, 48.1 ± 6.8 years; body mass index, 30.4 ± 4.1 kg·m(-2)) were randomized into 12-weeks of EET (40-60 min of cycling), RET (10 exercises; 3-4 sets × 8-10 repetitions), CET (50% serial completion of RET and EET), or control condition. The following were determined: intervention-based changes in fitness and strength; abdominal visceral adipose tissue (VAT), total body fat (TB-FM) and fat-free (TB-FFM) mass; plasma cytokines (C-reactive protein (CRP), tumor necrosis factor-α (TNFα) interleukin-6 (IL-6)); muscle protein content of p110α and glucose transporter 4 (GLUT4); mRNA expression of GLUT4, peroxisome proliferator-activated receptor-γ coactivator-1α-β, cytochrome c oxidase, hexokinase II, citrate synthase; oral glucose tolerance; and estimated insulin sensitivity. CET promoted commensurate improvements of aerobic capacity and muscular strength and reduced VAT and TB-FM equivalently to EET and RET (p < 0.05), yet only RET increased TB-FFM (p < 0.05). Although TNFα and IL-6 were reduced after all training interventions (p < 0.05), CRP remained unchanged (p > 0.05). EET reduced area under the curve for glucose, insulin, and C-peptide, whilst CET and RET respectively reduced insulin and C-peptide, and C-peptide only (p < 0.05). Notwithstanding increased insulin sensitivity index after all training interventions (p < 0.05), no change presented for GLUT4 or p110α total protein, or chronic mRNA expression of the studied mitochondrial genes (p > 0.05). In middle-aged men, 12 weeks of duration-matched CET promoted commensurate changes in fitness and strength, abdominal VAT, plasma cytokines and insulin sensitivity, and an equidistant glucose tolerance response to EET and RET; despite no change of measured muscle mechanisms associative to insulin action, glucose transport, and mitochondrial function.

The Role of Glucose Transporters in Brain Disease: Diabetes and Alzheimer’s Disease

The occurrence of altered brain glucose metabolism has long been suggested in both diabetes and Alzheimer’s diseases. However, the preceding mechanism to altered glucose metabolism has not been well understood. Glucose enters the brain via glucose transporters primarily present at the blood-brain barrier. Any changes in glucose transporter function and expression dramatically affects brain glucose homeostasis and function. In the brains of both diabetic and Alzheimer’s disease patients, changes in glucose transporter function and expression have been observed, but a possible link between the altered glucose transporter function and disease progress is missing. Future recognition of the role of new glucose transporter isoforms in the brain may provide a better understanding of brain glucose metabolism in normal and disease states. Elucidation of clinical pathological mechanisms related to glucose transport and metabolism may provide common links to the etiology of these two diseases. Considering these facts, in this review we provide a current understanding of the vital roles of a variety of glucose transporters in the normal, diabetic and Alzheimer’s disease brain.

Proteomic and bioinformatic analysis of hydrophobic membrane protein extracts reveals the presence of several novel CD antigens, glucose transporters and voltage gated anion channels in articular chondrocytes

Purpose: Membrane proteins are a key interface between the interior and exterior of living cells. In chondrocytes they perform many important cellular functions including cell-matrix interactions, communication, transport, differentiation and receptor signaling. Membrane proteins are known targets for many currently available pharmaceuticals. Defining the membrane proteome of chondrocytes will therefore advance our understanding of fundamental biological processes in cartilage and may reveal new targets for drug discovery efforts. However, the study of plasma membrane proteins is challenging due to their low abundance, hydrophobicity and large size. Proteomic analysis of plasma membrane proteins by conventional proteomic approaches using 2D-electrophoresis is particularly problematic. The aim of this study was to develop a detergentbased separation method to extract membrane proteins from chondrocytes and use nanoLC-MS/MS with shotgun and gel-based proteomics methodologies to identify membrane proteins and validate the expression of selected proteins using immunofluorescence microscopy. Methods: Primary cultures of chondrocytes were established from equine cartilage sourced from a local abattoir. Chondrocytes were released from cartilage using type I collagenase. To ensure preservation of the chondrocyte phenotype, only freshly dissociated, first expansion and first passage cells were used. Chondrocyte protein extracts were separated into hydrophobic and hydrophilic fractions using Triton X-114 phase separation. Trypsin digested protein fractions were analyzed by LC-MSMS using Bruker Easy-nanoLC chromatography and a Bruker amaZon ion trap instrument. The top 5 most abundant peptides from each MS scan were selected for fragmentation. MS data were submitted to the MASCOT search engine and compared to entries in the NCBInr database. The sequences of the proteins identified were analyzed by CELLO to predict their subcellular localizations. Proteins were divided into groups based on their predicted localization and further analyzed with DAVID Bioinformatics Resources. Features including function, presence in pathways and other associated proteins were uncovered using DAVID. Immunofluorescence techniques were used to confirm the expression of selected membrane proteins. Results: More than 200 proteins were identified. Analysis of the hydrophobic membrane fraction confirmed the presence of several novel membrane proteins as well as a number of previously described CD antigens, channels and glucose transporters. Table 1 summarizes the identities, MASCOT scores, locations and biological functions of the proteins identified. The immunodetection of the GLUT1 and GLUT3 proteins (Figure 1) confirms previously published reports on the expression and function of these hypoxia-responsive glucose transporters in chondrocytes. Expression of GLUT14 (Figure 1), a duplicon of GLUT3 originally cloned from testis, is a novel observation that provides further evidence for a critical role for glucose transport and metabolism in chondrocyte biology. Identification of the mitochondrial VDAC channel proteins confirms recent proteomic data obtained from human cartilage. Conclusions: In summary, the data presented support the view that the study of membrane proteins in chondrocytes is feasible using phase separation and high-throughput LC-MSMS based proteomics. Bioinformatic tools such as MASCOT, CELLO and DAVID can enhance and refine this discovery approach by revealing information about the cellular localization and functions of the proteins identified. This strategy will help us learnmore about membrane proteins in chondrocytes andmay prove to be useful for identifying new membrane antigens and elusive cell surface biomarkers.

GLUT1 expression in malignant tumors and its use as an immunodiagnostic marker

OBJECTIVE:To analyze glucose transporter 1 expression patterns in malignant tumors of various cell types and evaluate their diagnostic value by immunohistochemistry.INTRODUCTION:Glucose is the major source of energy for cells, and glucose transporter 1 is the most common glucose transporter in humans. Glucose transporter 1 is aberrantly expressed in several tumor types. Studies have implicated glucose transporter 1 expression as a prognostic and diagnostic marker in tumors, primarily in conjunction with positron emission tomography scan data.METHODS:Immunohistochemistry for glucose transporter 1 was performed in tissue microarray slides, comprising 1955 samples of malignant neoplasm from different cell types.RESULTS:Sarcomas, lymphomas, melanomas and hepatoblastomas did not express glucose transporter 1. Forty-seven per cent of prostate adenocarcinomas were positive, as were 29% of thyroid, 10% of gastric and 5% of breast adenocarcinomas. Thirty-six per cent of squamous cell carcinomas of the head and neck were positive, as were 42% of uterine cervix squamous cell carcinomas. Glioblastomas and retinoblastomas showed membranous glucose transporter 1 staining in 18.6% and 9.4% of all cases, respectively. Squamous cell carcinomas displayed membranous expression, whereas adenocarcinomas showed cytoplasmic glucose transporter 1 expression.CONCLUSION:Glucose transporter 1 showed variable expression in various tumor types. Its absence in sarcomas, melanomas, hepatoblastomas and lymphomas suggests that other glucose transporters mediate the glycolytic pathway in these tumors. The data suggest that glucose transporter 1 is a valuable immunohistochemical marker that can be used to identify patients for evaluation by positron emission tomography scan. The function of cytoplasmic glucose transporter 1 in adenocarcinomas must be further examined.

Glucose transporter and hypoxia-associated gene expression in the mammary gland of transition dairy cattle

Glucose is an important energy substrate, especially needed by dairy cows postpartum to support the onset of lactation. The prioritization and regulation of glucose uptake is accomplished, in part, by changes in expression of cellular glucose transport molecules (GLUT) within the mammary gland. The objectives of this study were to (1) evaluate the expression and cell-type specific localization of GLUT and hypoxia-associated genes that may regulate GLUT expression over the transition period and through lactation in bovine mammary tissue and (2) determine functionality of GLUT on primary bovine mammary endothelial cells (BMEC). Mammary tissue biopsies were taken from cows at 15 d before calving and again at 1, 15, 30, 60, 120, and 240 d post-parturition for quantitative real-time PCR analysis of GLUT and hypoxia-associated genes. Additional mammary tissue samples were used to localize GLUT within the cells of the lobulo-alveolar system via fluorescence microscopy. Cultures of primary bovine mammary endothelial cells were used to confirm the functionality of GLUT with a fluorescent glucose analog uptake assay. Significant increases in GLUT1 gene expression were observed during early lactation, whereas both GLUT3 and GLUT4 gene expression increased during late lactation. The gene expression for 2 receptors of vascular endothelial growth factor increased significantly during early lactation and remained increased throughout lactation when compared with gene expression during the transition period. All GLUT were detected on cultured BMEC and were capable of internalizing glucose through GLUT-mediated mechanisms. These data suggest mammary vascular tissues express GLUT during lactation and BMEC express functional glucose transporters. A better understanding of glucose uptake at the endothelial level may prove to be critical to improve glucose absorption from the blood for utilization by mammary epithelial cells.

Small Ubiquitin-related Modifier (SUMO)-1 Promotes Glycolysis in Hypoxia

Under conditions of hypoxia, most eukaryotic cells undergo a shift in metabolic strategy, which involves increased flux through the glycolytic pathway. Although this is critical for bioenergetic homeostasis, the underlying mechanisms have remained incompletely understood. Here, we report that the induction of hypoxia-induced glycolysis is retained in cells when gene transcription or protein synthesis are inhibited suggesting the involvement of additional post-translational mechanisms. Post-translational protein modification by the small ubiquitin related modifier-1 (SUMO-1) is induced in hypoxia and mass spectrometric analysis using yeast cells expressing tap-tagged Smt3 (the yeast homolog of mammalian SUMO) revealed hypoxia-dependent modification of a number of key glycolytic enzymes. Overexpression of SUMO-1 in mammalian cancer cells resulted in increased hypoxia-induced glycolysis and resistance to hypoxia-dependent ATP depletion. Supporting this, non-transformed cells also demonstrated increased glucose uptake upon SUMO-1 overexpression. Conversely, cells overexpressing the de-SUMOylating enzyme SENP-2 failed to demonstrate hypoxia-induced glycolysis. SUMO-1 overexpressing cells demonstrated focal clustering of glycolytic enzymes in response to hypoxia leading us to hypothesize a role for SUMOylation in promoting spatial re-organization of the glycolytic pathway. In summary, we hypothesize that SUMO modification of key metabolic enzymes plays an important role in shifting cellular metabolic strategies toward increased flux through the glycolytic pathway during periods of hypoxic stress.

Relationship between erythrocyte GLUT1 function and membrane glycation in type 2 diabetes

This paper investigates the effect of glycation on glucose transport in erythrocytes. Glucose transporter function, numbers and erythrocyte phosphorylation rates are simultaneously studied using 30 Caucasian patients with diabetes and 30 Caucasian control volunteers (mean±SD where P≤0.05; age 48±8 vs. 45±8 years [ns]; body mass index [BMI] 31±7 vs. 27±5 [P=0.035]; blood glucose 12±7 vs. 5±0.6 mmol/L [P=0.001]; HbA1c 8±2 vs. 5±0.3% [P=0.0001]; fructosamine 336±64 vs. 237±16 µmol/L [P=0.0001]; disease duration 13±11 years, respectively). Significant differences were found for glucose transporter function, with 3-O-methylglucose uptake rates (108±49 vs. 146±55 µmol/L/sec/1012 cells [P=0.010]); D-glucose influx (64±30 vs. 117±45 µmol/L/sec/1012 cells [P=0.0001]); and D-glucose net transport (31±22 vs. 74±55 µmol/L/sec/ 1012 cells [P = 0.0001]). No differences were found for phosphorylation rates using 2-deoxyglucose (33±17 vs. 38±12 µmol/L/sec/1012 cells [P=0.194]). The number of functional transporters using cytochalasin B studies measured via Bmax, was not found to be significantly different between the groups (195±139 vs. 264±174 [P=0.206]). However, Kd was lower for those with diabetes, suggesting higher binding affinity (12±11 vs. 32±25 nmol/L [P=0.006]). A negative correlation between HbA1c and D-glucose influx involving both groups was found (r=–0.670, P=0.0001). Glucose transport is shown to be decreased in people who have diabetes compared to normoglycaemic volunteers, whereas the number of glucose transporters is apparently unchanged; however, affinity for binding is increased.

Expression of Na+/glucose co-transporter 1 (SGLT1) is enhanced by supplementation of the diet of weaning piglets with artificial sweeteners

In an intensive livestock production, a shorter suckling period allows more piglets to be born. However, this practice leads to a number of disorders including nutrient malabsorption, resulting in diarrhoea, malnutrition and dehydration. A number of strategies have been proposed to overcome weaning problems. Artificial sweeteners, routinely included in piglets' diet, were thought to enhance feed palatability. However, it is shown in rodent models that when included in the diet, they enhance the expression of Na+/glucose co-transporter (SGLT1) and the capacity of the gut to absorb glucose. Here, we show that supplementation of piglets' feed with a combination of artificial sweeteners saccharin and neohesperidin dihydrochalcone enhances the expression of SGLT1 and intestinal glucose transport function. Artificial sweeteners are known to act on the intestinal sweet taste receptor T1R2/T1R3 and its partner G-protein, gustducin, to activate pathways leading to SGLT1 up-regulation. Here, we demonstrate that T1R2, T1R3 and gustducin are expressed together in the enteroendocrine cells of piglet intestine. Furthermore, gut hormones secreted by the endocrine cells in response to dietary carbohydrates, glucagon-like peptides (GLP)-1, GLP-2 and glucose-dependent insulinotrophic peptide (GIP), are co-expressed with type 1 G-protein-coupled receptors (T1R) and gustducin, indicating that L- and K-enteroendocrine cells express these taste elements. In a fewer endocrine cells, T1R are also co-expressed with serotonin. Lactisole, an inhibitor of human T1R3, had no inhibitory effect on sweetener-induced SGLT1 up-regulation in piglet intestine. A better understanding of the mechanism(s) involved in sweetener up-regulation of SGLT1 will allow the identification of nutritional targets with implications for the prevention of weaning-related malabsorption.

Interactive changes between macrophages and adipocytes.

Obesity is associated with a proinflammatory state, with macrophage infiltration into adipose tissue. We tested the hypothesis that communication between macrophages and adipocytes affects insulin resistance by disrupting insulin-stimulated glucose transport, adipocyte differentiation, and macrophage function. To test this hypothesis, we cocultured 3T3-L1 adipocytes with C2D macrophages or primary peritoneal mouse macrophages and examined the impacts of macrophages and adipocytes on each other. Adipocytes and preadipocytes did not affect C2D macrophage TNF-alpha, IL-6, or IL-1beta transcript concentrations relative to those obtained when C2D macrophages were incubated alone. However, preadipocytes and adipocytes increased PEC-C2D macrophage IL-6 transcript levels, while preadipocytes inhibited IL-1beta transcript levels compared to those obtained when PEC-C2D macrophages were incubated in medium alone. We found that adipocyte coculture increased macrophage consumption of tumor necrosis factor alpha (TNF-alpha), interleukin 1beta (IL-1beta), and, in some cases, IL-6. C2D macrophages increasingly downregulated GLUT4 transcript levels in differentiated adipocytes. Recombinant TNF-alpha, IL-1beta, and IL-6 also downregulated GLUT4 transcript levels relative to those for the control. However, only IL-6 was inhibitory at concentrations detected in macrophage-adipocyte cocultures. IL-6 and TNF-alpha, but not IL-1beta, inhibited Akt phosphorylation within 15 min of insulin stimulation, but only IL-6 was inhibitory 30 min after stimulation. Lastly, we found that adipocyte differentiation was inhibited by macrophages or by recombinant TNF-alpha, IL-6, and IL-1beta, with IL-6 having the most impact. These data suggest that the interaction between macrophages and adipocytes is a complex process, and they support the hypothesis that the macrophage-adipocyte interaction affects insulin resistance by disrupting insulin-stimulated glucose transport, adipocyte differentiation, and macrophage function.

Metabolic evolution of energy-conserving pathways for succinate production in Escherichia coli

During metabolic evolution to improve succinate production in Escherichia coli strains, significant changes in cellular metabolism were acquired that increased energy efficiency in two respects. The energy-conserving phosphoenolpyruvate (PEP) carboxykinase (pck), which normally functions in the reverse direction (gluconeogenesis; glucose repressed) during the oxidative metabolism of organic acids, evolved to become the major carboxylation pathway for succinate production. Both PCK enzyme activity and gene expression levels increased significantly in two stages because of several mutations during the metabolic evolution process. High-level expression of this enzyme-dominated CO(2) fixation and increased ATP yield (1 ATP per oxaloacetate). In addition, the native PEP-dependent phosphotransferase system for glucose uptake was inactivated by a mutation in ptsI. This glucose transport function was replaced by increased expression of the GalP permease (galP) and glucokinase (glk). Results of deleting individual transport genes confirmed that GalP served as the dominant glucose transporter in evolved strains. Using this alternative transport system would increase the pool of PEP available for redox balance. This change would also increase energy efficiency by eliminating the need to produce additional PEP from pyruvate, a reaction that requires two ATP equivalents. Together, these changes converted the wild-type E. coli fermentation pathway for succinate into a functional equivalent of the native pathway that nature evolved in succinate-producing rumen bacteria.