Effects of ileal glucose infusion on enteropancreatic hormone secretion in humans: relationship to glucose absorption.

BACKGROUNDS: The distal small intestine plays an important role in regulating the secretion of entero-pancreatic hormones that are critical to the control of glucose metabolism and appetite, but the quantitative contribution of a specific segment to these effects is unknown. PURPOSES: To determine the effects of 30 cm of the ileum exposed to glucose on the secretion of ghrelin, glucagon-like peptide-1 (GLP-1), glucose-dependent insulinotropic polypeptide (GIP) insulin, C-peptide and glucagon, in relation to glucose absorption in non-diabetic subjects. BASIC PROCEDURES: 10 non-diabetic subjects with a loop ileostomy after early-stage rectal cancer resection were studied on 2 days in a double-blind, randomized and crossover fashion, when a catheter was inserted retrogradely 30 cm from the ileostomy for infusion of a glucose solution containing 30 g glucose and 3 g 3-O-methylglucose (as a marker of active glucose absorption), or 0.9% saline, over 60 min. Ghrelin, GIP, GLP-1, insulin, C-peptide, glucagon and ileal glucose absorption (from concentrations of 3-O-methylglucose in serum and glucose in ileostomy effluent) were measured over 180 min. MAIN FINDINGS: 12.0 ± 1.2 g glucose was absorbed over 180 min. Compared to saline, ileal glucose resulted in minimal increases in blood glucose and plasma insulin and C-peptide, but substantial increases in plasma GLP-1, without affecting ghrelin, GIP or glucagon. The magnitude of the GLP-1 response to glucose was strongly related to the increase in serum 3-O-methylglucose. PRINCIPAL CONCLUSIONS: Stimulation of the terminal ileum by glucose, even over a short length (30 cm), induces substantial GLP-1 release, coupled primarily to active glucose absorption. CLINICAL REGISTRATION: NCT05030376 (ClinicalTrials.gov).

In vitro and in vivo comparison of MRI chemical exchange saturation transfer (CEST) properties between native glucose and 3-O-Methyl-D-glucose in a murine tumor model.

D-Glucose and 3-O-Methyl-D-glucose (3OMG) have been shown to provide contrast in magnetic resonance imaging-chemical exchange saturation transfer (MRI-CEST) images. However, a systematic comparison between these two molecules has yet to be performed. The current study deals with the assessment of the effect of pH, saturation power level (B1 ) and magnetic field strength (B0 ) on the MRI-CEST contrast with the aim of comparing the in vivo CEST contrast detectability of these two agents in the glucoCEST procedure. Phosphate-buffered solutions of D-Glucose or 3OMG (20 mM) were prepared at different pH values and Z-spectra were acquired at several B1 levels at 37°C. In vivo glucoCEST images were obtained at 3 and 7 T over a period of 30 min after injection of D-Glucose or 3OMG (at doses of 1.5 or 3 g/kg) in a murine melanoma tumor model (n = 3-5 mice for each molecule, dose and B0 field). A markedly different pH dependence of CEST response was observed in vitro for D-Glucose and 3OMG. The glucoCEST contrast enhancement in the tumor region following intravenous administration (at the 3 g/kg dose) was comparable for both molecules: 1%-2% at 3 T and 2%-3% at 7 T. The percentage change in saturation transfer that resulted was almost constant for 3OMG over the 30-min period, whereas a significant increase was detected for D-Glucose. Our results show similar CEST contrast efficiency but different temporal kinetics for the metabolizable and the nonmetabolizable glucose derivatives in a tumor murine model when administered at the same doses.

Feasibility of deuterium magnetic resonance spectroscopy of 3-O-Methylglucose at 7 Tesla.

Deuterium Magnetic Resonance Spectroscopy (DMRS) is a non-invasive technique that allows the detection of deuterated compounds in vivo. DMRS has a large potential to analyze uptake, perfusion, washout or metabolism, since deuterium is a stable isotope and therefore does not decay during biologic processing of a deuterium labelled substance. Moreover, DMRS allows the distinction between different deuterated substances. In this work, we performed DMRS of deuterated 3-O-Methylglucose (OMG). OMG is a non-metabolizable glucose analog which is transported similar to D-glucose. DMRS of OMG was performed in phantom and in vivo measurements using a preclinical 7 Tesla MRI system. The chemical shift (3.51 ± 0.1 ppm) and relaxation times were determined. OMG was injected intravenously and spectra were acquired over a period of one hour to monitor the time evolution of the deuterium signal in tumor-bearing rats. The increase and washout of OMG could be observed. Three different exponential functions were compared in terms of how well they describe the OMG washout. A mono-exponential model with offset seems to describe the observed time course best with a time constant of 1910 ± 770 s and an offset of 2.5 ± 1.2 mmol/l (mean ± std, N = 3). Chemical shift imaging could be performed with a voxel size of 7.1 mm x 7.1 mm x 7.9 mm. The feasibility of DMRS with deuterium labelled OMG could be demonstrated. These data might serve as basis for future studies that aim to characterize glucose transport using DMRS.

Retinol binding protein 3 is increased in the retina of patients with diabetes resistant to diabetic retinopathy.

The Joslin Medalist Study characterized people affected with type 1 diabetes for 50 years or longer. More than 35% of these individuals exhibit no to mild diabetic retinopathy (DR), independent of glycemic control, suggesting the presence of endogenous protective factors against DR in a subpopulation of patients. Proteomic analysis of retina and vitreous identified retinol binding protein 3 (RBP3), a retinol transport protein secreted mainly by the photoreceptors, as elevated in Medalist patients protected from advanced DR. Mass spectrometry and protein expression analysis identified an inverse association between vitreous RBP3 concentration and DR severity. Intravitreal injection and photoreceptor-specific overexpression of RBP3 in rodents inhibited the detrimental effects of vascular endothelial growth factor (VEGF). Mechanistically, our results showed that recombinant RBP3 exerted the therapeutic effects by binding and inhibiting VEGF receptor tyrosine phosphorylation. In addition, by binding to glucose transporter 1 (GLUT1) and decreasing glucose uptake, RBP3 blocked the detrimental effects of hyperglycemia in inducing inflammatory cytokines in retinal endothelial and Müller cells. Elevated expression of photoreceptor-secreted RBP3 may have a role in protection against the progression of DR due to hyperglycemia by inhibiting glucose uptake via GLUT1 and decreasing the expression of inflammatory cytokines and VEGF.

CEST MRI of 3-O-methyl-D-glucose uptake and accumulation in brain tumors.

PURPOSE: 3-O-Methyl-D-glucose (3-OMG) is a nonmetabolizable structural analog of glucose that offers potential to be used as a CEST-contrast agent for tumor detection. Here, we explore it for CEST-detection of malignant brain tumors and compare it with D-glucose. METHODS: Glioma xenografts of a U87-MG cell line were implanted in five mice. Dynamic 3-OMG weighted images were collected using CEST-MRI at 11.7 T at a single offset of 1.2 ppm, showing the effect of accumulation of the contrast agent in the tumor, following an intravenous injection of 3-OMG (3 g/kg). RESULTS: Tumor regions showed higher enhancement as compared to contralateral brain. The CEST contrast enhancement in the tumor region ranged from 2.5-5.0%, while it was 1.5-3.5% in contralateral brain. Previous D-glucose studies of the same tumor model showed an enhancement of 1.5-3.0% and 0.5-1.5% in tumor and contralateral brain, respectively. The signal gradually stabilized to a value that persisted for the length of the scan. CONCLUSIONS: 3-OMG shows a CEST contrast enhancement that is approximately twice as much as that of D-glucose for a similar tumor line. In view of its suggested low toxicity and transport properties across the BBB, 3-OMG provides an option to be used as a nonmetallic contrast agent for evaluating brain tumors.

3-O-Methyl-D-glucose mutarotation and proton exchange rates assessed by 13C, 1H NMR and by chemical exchange saturation transfer and spin lock measurements.

3-O-Methyl-D-glucose (3OMG) was recently suggested as an agent to image tumors using chemical exchange saturation transfer (CEST) MRI. To characterize the properties of 3OMG in solution, the anomeric equilibrium and the mutarotation rates of 3OMG were studied by 1H and 13C NMR. This information is essential in designing the in vivo CEST experiments. At room temperature, the ratio of α and ß 3OMG anomers at equilibrium was 1:1.4, and the time to reach 95% equilibrium was 6 h. The chemical exchange rates between the hydroxyl protons of 3OMG and water, measured by CEST and spin lock at pH 6.14 and a temperature of 4 °C, were in the range of 360-670 s-1.

Effects of starvation and short-term refeeding on gastric emptying and postprandial blood glucose regulation in adolescent girls with anorexia nervosa.

Postprandial glucose is reduced in malnourished patients with anorexia nervosa (AN), but the mechanisms and duration for this remain unclear. We examined blood glucose, gastric emptying, and glucoregulatory hormone changes in malnourished patients with AN and during 2 wk of acute refeeding compared with healthy controls (HCs). Twenty-two female adolescents with AN and 17 age-matched female HCs were assessed after a 4-h fast. Patients were commenced on a refeeding protocol of 2,400 kcal/day. Gastric emptying (13C-octanoate breath test), glucose absorption (3-O-methylglucose), blood glucose, plasma glucagon-like peptide-1 (GLP-1), glucose-dependent insulinotropic polypeptide (GIP), insulin, C-peptide, and glucagon responses to a mixed-nutrient test meal were measured on admission and 1 and 2 wk after refeeding. HCs were assessed once. On admission, patients had slower gastric emptying, lower postprandial glucose and insulin, and higher glucagon and GLP-1 than HCs ( P < 0.05). In patients with AN, the rise in glucose (0-30 min) correlated with gastric emptying ( P < 0.05). With refeeding, postprandial glucose and 3-O-methylglucose were higher, gastric emptying faster, and baseline insulin and C-peptide less ( P < 0.05), compared with admission. After 2 wk of refeeding, postprandial glucose remained lower, and glucagon and GLP-1 higher, in patients with AN than HCs ( P < 0.05) without differences in gastric emptying, baseline glucagon, or postprandial insulin. Delayed gastric emptying may underlie reduced postprandial glucose in starved patients with AN; however, postprandial glucose and glucoregulatory hormone changes persist after 2 wk of refeeding despite improved gastric emptying. Future research should explore whether reduced postprandial glucose in AN is related to medical risk by examining associated symptoms alongside continuous glucose monitoring during refeeding.

CEST MRI of 3-O-methyl-D-glucose on different breast cancer models.

PURPOSE: To test the ability of chemical exchange saturation transfer (CEST) MRI of 3-O-methyl-D-glucose (3OMG) to detect tumors in several breast cancer models of murine and human origin, for different routes of administration of the agent and to compare the method with glucoCEST and with 18 FDG-PET on the same animals. METHODS: In vivo CEST MRI experiments were performed with a 7T Biospec animal MRI scanner on implanted orthotopic mammary tumors of mice before and after administration of 3OMG. RESULTS: A marked 3OMG-CEST MRI contrast that was correlated with the administrated dose was obtained in different breast cancer models and by intravenous, intraperitoneal, and per os methods of administration. The most aggressive breast cancer model yielded the highest CEST contrast. 3OMG-CEST contrast reached its maximum at 20 min after administration and lasted for more than an hour, while that of glucose was lower and diminished after 20 min. 3OMG-CEST showed comparable results to that of FDG PET. CONCLUSION: The sensitivity of the 3OMG-CEST MRI method indicates its potential for the detection of tumors in the clinic. Magn Reson Med 79:1061-1069, 2018. © 2017 International Society for Magnetic Resonance in Medicine.

Regulation of GLUT4 activity in myotubes by 3-O-methyl-d-glucose.

The rate of glucose influx to skeletal muscles is determined primarily by the number of functional units of glucose transporter-4 (GLUT4) in the myotube plasma membrane. The abundance of GLUT4 in the plasma membrane is tightly regulated by insulin or contractile activity, which employ distinct pathways to translocate GLUT4-rich vesicles from intracellular compartments. Various studies have indicated that GLUT4 intrinsic activity is also regulated by conformational changes and/or interactions with membrane components and intracellular proteins in the vicinity of the plasma membrane. Here we show that the non-metabolizable glucose analog 3-O-methyl-d-glucose (MeGlc) augmented the rate of hexose transport into myotubes by increasing GLUT4 intrinsic activity without altering the content of the transporter in the plasma membrane. This effect was not a consequence of ATP depletion or hyperosmolar stress and did not involve Akt/PKB or AMPK signal transduction pathways. MeGlc reduced the inhibitory potency (increased Ki) of indinavir, a selective inhibitor of GLUT4, in a dose-dependent manner. Kinetic analyses indicate that MeGlc induced changes in GLUT4 or GLUT4 complexes within the plasma membrane, which enhanced the hexose transport activity and reduced the potency of indinavir inhibition. Finally, we present a simple kinetic analysis for screening and discovering low molecular weight compounds that augment GLUT4 activity.

WZB117 (2-Fluoro-6-(m-hydroxybenzoyloxy) Phenyl m-Hydroxybenzoate) Inhibits GLUT1-mediated Sugar Transport by Binding Reversibly at the Exofacial Sugar Binding Site.

WZB117 (2-fluoro-6-(m-hydroxybenzoyloxy) phenyl m-hydroxybenzoate) inhibits passive sugar transport in human erythrocytes and cancer cell lines and, by limiting glycolysis, inhibits tumor growth in mice. This study explores how WZB117 inhibits the erythrocyte sugar transporter glucose transport protein 1 (GLUT1) and examines the transporter isoform specificity of inhibition. WZB117 reversibly and competitively inhibits erythrocyte 3-O-methylglucose (3MG) uptake with Ki(app) = 6 µm but is a noncompetitive inhibitor of sugar exit. Cytochalasin B (CB) is a reversible, noncompetitive inhibitor of 3MG uptake with Ki(app) = 0.3 µm but is a competitive inhibitor of sugar exit indicating that WZB117 and CB bind at exofacial and endofacial sugar binding sites, respectively. WZB117 inhibition of GLUTs expressed in HEK293 cells follows the order of potency: insulin-regulated GLUT4 ≫ GLUT1 ≈ neuronal GLUT3. This may explain WZB117-induced murine lipodystrophy. Molecular docking suggests the following. 1) The WZB117 binding envelopes of exofacial GLUT1 and GLUT4 conformers differ significantly. 2) GLUT1 and GLUT4 exofacial conformers present multiple, adjacent glucose binding sites that overlap with WZB117 binding envelopes. 3) The GLUT1 exofacial conformer lacks a CB binding site. 4) The inward GLUT1 conformer presents overlapping endofacial WZB117, d-glucose, and CB binding envelopes. Interrogating the GLUT1 mechanism using WZB117 reveals that subsaturating WZB117 and CB stimulate erythrocyte 3MG uptake. Extracellular WZB117 does not affect CB binding to GLUT1, but intracellular WZB117 inhibits CB binding. These findings are incompatible with the alternating conformer carrier for glucose transport but are consistent with either a multisubunit, allosteric transporter, or a transporter in which each subunit presents multiple, interacting ligand binding sites.

Tissue localization of GM-CSF receptor in bovine ovarian follicles and its role on glucose uptake by mural granulosa cells.

The granulocyte-macrophage colony stimulating factor (GM-CSF) is a multifunctional cytokine implicated in proliferation, differentiation, and activation of several cell types including those involved in hematopoiesis and reproduction. In the present study, the expression of the α- and ß-subunit genes of GM-CSF receptor during follicular development in cattle was assessed. The spatial association of α- and ß-subunits of GM-CSF with follicle stimulating hormone receptor (FSHR) and 3ß-hydroxysteroid dehydrogenase (3ß-HSD), and the temporal associations with gene expression of hexose transporters (GLUTs) in granulosa cells of cattle were also evaluated. The effect of GM-CSF on the functionality of hexose transporters was also determined in an in vitro primary culture of granulosa cells. The spatial association of subunits of the GM-CSF receptor with 3ß-HSD and FSHR suggests a potential steroidogenic regulation of GM-CSF in granulosa cells. Immunodetection of GLUTs and uptake kinetic assays confirmed expression and functionality of these genes for hexose transporters in granulosa cells of cattle. Treatment of granulosa cells with GM-CSF, FSH or insulin- like growth factor-I (IGF-I) alone increased 2-deoxyglucose (DOG) or 3-0-methylglucose (OMG) uptake; however, when cells were treated with various combination of these factors there were no additive effect. Unexpectedly, the combination of GM-CSF and FSH decreased DOG uptake compared to FSH treatment alone. Thus, the expression pattern of GM-CSF receptor subunit genes during follicle development in cattle and promotion of DOG and OMG uptake in granulosa cells indicate a role for GM-CSF, FSH and/or IGF-I alone in regulating granulosa cell metabolic activity, specifically by promoting glucose uptake.

Effects of intraduodenal hydroxycitrate on glucose absorption, incretin release, and glycemia in response to intraduodenal glucose infusion in health and type 2 diabetes: A randomised controlled trial.

OBJECTIVE: Hydroxycitric acid (HCA), derived from the fruit Garcinia cambogia, reduces the rate of glucose absorption and lowers postprandial glycemia in rodents, but its effect in humans is unknown. The aim of this study was to investigate the effects of small intestinal perfusion with HCA on glucose absorption, as well as the incretin and glycemic responses to a subsequent intraduodenal glucose infusion, in both healthy individuals and patients with type 2 diabetes. METHODS: Twelve healthy participants and 8 patients with type 2 diabetes received an intraduodenal infusion of HCA (2800 mg in water) or control (water) over 60 min, followed by an intraduodenal infusion of 60 g glucose over 120 min, in a double-blind, randomized crossover design. In healthy individuals, 5 g 3-O-methylglucose (3-OMG) was co-infused with glucose as a marker of glucose absorption. Blood was sampled frequently. RESULTS: In healthy individuals, blood glucose was lower with HCA than control, both before and during the intraduodenal glucose infusion (P < 0.05 for each). Plasma glucose-dependent insulinotropic polypeptide (GIP; P = 0.01) and glucagon (P = 0.06) were higher with HCA, but there were no differences in plasma glucagon-like peptide (GLP)-1, insulin, or serum 3-OMG concentrations. In patients with type 2 diabetes, blood glucose, and plasma GIP, GLP-1, and insulin did not differ between HCA and control either before or after intraduodenal glucose, but during glucose infusion, plasma glucagon was higher with HCA (P = 0.04). CONCLUSION: In healthy individuals, small intestinal exposure to HCA resulted in a modest reduction in glycemia and stimulation of plasma GIP and glucagon, but no effect on plasma GLP-1 or insulin, or on glucose absorption. HCA had no effect on glycemia in patients with type 2 diabetes.

Measurement of Glucose Uptake in Cultured Cells.

Facilitative glucose uptake transport systems are ubiquitous in animal cells and are responsible for transporting glucose across cell surface membranes. Evaluation of glucose uptake is crucial in the study of numerous diseases and metabolic disorders such as myocardial ischemia, diabetes mellitus, and cancer. Detailed in this unit are laboratory methods for assessing glucose uptake into mammalian cells. The unit is divided into five sections: (1) a brief overview of glucose uptake assays in cultured cells; (2) a method for measuring glucose uptake using radiolabeled 3-O-methylglucose; (3) a method for measuring glucose uptake using radiolabeled 2-deoxyglucose (2DG); (4) a microplate method for measuring 2DG-uptake using an enzymatic, fluorometric assay; and (5) a microplate-based method using a fluorescent analog of 2DG.

Glucose-Sensing Receptor T1R3: A New Signaling Receptor Activated by Glucose in Pancreatic ß-Cells.

Subunits of the sweet taste receptors T1R2 and T1R3 are expressed in pancreatic ß-cells. Compared with T1R3, mRNA expression of T1R2 is considerably lower. At the protein level, expression of T1R2 is undetectable in ß-cells. Accordingly, a major component of the sweet taste-sensing receptor in ß-cells may be a homodimer of T1R3 rather than a heterodimer of T1R2/T1R3. Inhibition of this receptor by gurmarin or deletion of the T1R3 gene attenuates glucose-induced insulin secretion from ß-cells. Hence the T1R3 homodimer functions as a glucose-sensing receptor (GSR) in pancreatic ß-cells. When GSR is activated by the T1R3 agonist sucralose, elevation of intracellular ATP concentration ([ATP]i) is observed. Sucralose increases [ATP]i even in the absence of ambient glucose, indicating that sucralose increases [ATP]i not simply by activating glucokinase, a rate-limiting enzyme in the glycolytic pathway. In addition, sucralose augments elevation of [ATP]i induced by methylsuccinate, suggesting that sucralose activates mitochondrial metabolism. Nonmetabolizable 3-O-methylglucose also increases [ATP]i and knockdown of T1R3 attenuates elevation of [ATP]i induced by high concentration of glucose. Collectively, these results indicate that the T1R3 homodimer functions as a GSR; this receptor is involved in glucose-induced insulin secretion by activating glucose metabolism probably in mitochondria.

Selective sodium-dependent glucose transporter 1 inhibitors block glucose absorption and impair glucose-dependent insulinotropic peptide release.

GSK-1614235 and KGA-2727 are potent, selective inhibitors of the SGLT1 sodium-dependent glucose transporter. Nonclinical (KGA-2727) and clinical (GSK-1614235) trials assessed translation of SGLT1 inhibitor effects from rats to normal human physiology. In rats, KGA-2727 (0.1 mg/kg) or vehicle was given before oral administration of 3-O-methyl-α-d-glucopyranose (3-O-methylglucose, 3-OMG) containing 3-[3H]OMG tracer. Tracer absorption and distribution were assessed from plasma, urine, and fecal samples. SGLT1 inhibition reduced urine 3-OMG recovery and increased fecal excretion. SGLT1 inhibitor effects on plasma glucose, insulin, gastric inhibitory peptide (GIP), and glucagon-like peptide-1 (GLP-1) concentrations were also measured during a standard meal. Incremental glucose, insulin, and GIP concentrations were decreased, indicating downregulation of ß-cell and K cell secretion. Minimal effects were observed in the secretion of the L cell product, GLP-1. With the use of a three-way, crossover design, 12 healthy human subjects received placebo or 20 mg GSK-1614235 immediately before or after a meal. Five minutes into the meal, 3-OMG was ingested. Postmeal dosing had little impact, yet premeal dosing delayed and reduced 3-OMG absorption, with an AUC0-10 of 231±31 vs. 446±31 µg·h(-1)·ml(-1), for placebo. Recovery of tracer in urine was 1.2±0.7 g for premeal dosing and 2.2±0.1 g for placebo. Incremental concentrations of insulin, C-peptide, and GIP were reduced for 2 h with premeal GSK-1614235. Total GLP-1 concentrations were significantly increased, and a trend for increased peptide YY (PYY) was noted. SGLT1 inhibitors block intestinal glucose absorption and reduce GIP secretion in rats and humans, suggesting SGLT1 glucose transport is critical for GIP release. Conversely, GLP-1 and PYY secretion are enhanced by SGLT1 inhibition in humans.

Caffeine inhibits glucose transport by binding at the GLUT1 nucleotide-binding site.

Glucose transporter 1 (GLUT1) is the primary glucose transport protein of the cardiovascular system and astroglia. A recent study proposes that caffeine uncompetitive inhibition of GLUT1 results from interactions at an exofacial GLUT1 site. Intracellular ATP is also an uncompetitive GLUT1 inhibitor and shares structural similarities with caffeine, suggesting that caffeine acts at the previously characterized endofacial GLUT1 nucleotide-binding site. We tested this by confirming that caffeine uncompetitively inhibits GLUT1-mediated 3-O-methylglucose uptake in human erythrocytes [Vmax and Km for transport are reduced fourfold; Ki(app) = 3.5 mM caffeine]. ATP and AMP antagonize caffeine inhibition of 3-O-methylglucose uptake in erythrocyte ghosts by increasing Ki(app) for caffeine inhibition of transport from 0.9 ± 0.3 mM in the absence of intracellular nucleotides to 2.6 ± 0.6 and 2.4 ± 0.5 mM in the presence of 5 mM intracellular ATP or AMP, respectively. Extracellular ATP has no effect on sugar uptake or its inhibition by caffeine. Caffeine and ATP displace the fluorescent ATP derivative, trinitrophenyl-ATP, from the GLUT1 nucleotide-binding site, but d-glucose and the transport inhibitor cytochalasin B do not. Caffeine, but not ATP, inhibits cytochalasin B binding to GLUT1. Like ATP, caffeine renders the GLUT1 carboxy-terminus less accessible to peptide-directed antibodies, but cytochalasin B and d-glucose do not. These results suggest that the caffeine-binding site bridges two nonoverlapping GLUT1 endofacial sites-the regulatory, nucleotide-binding site and the cytochalasin B-binding site. Caffeine binding to GLUT1 mimics the action of ATP but not cytochalasin B on sugar transport. Molecular docking studies support this hypothesis.

Acid-catalysed glucose dehydration in the gas phase: a mass spectrometric approach.

Understanding on a molecular level the acid-catalysed decomposition of the sugar monomers from hemicellulose and cellulose (e.g. glucose, xylose), the main constituent of lignocellulosic biomass is very important to increase selectivity and reaction yields in solution, key steps for the development of a sustainable renewable industry. In this work we reported a gas-phase study performed by electrospray triple quadrupole mass spectrometry on the dehydration mechanism of D-glucose. In the gas phase, reactant ions corresponding to protonated D-glucose were obtained in the ESI source and were allowed to undergo collisionally activated decomposition (CAD) into the quadrupole collision cell. The CAD mass spectrum of protonated D-glucose is characterized by the presence of ionic dehydrated daughter ion (ionic intermediates and products), which were structurally characterized by their fragmentation patterns. In the gas phase D-glucose dehydration does not lead to the formation of protonated 5-hydroxymethyl-2-furaldehyde, but to a mixed population of m/z 127 isomeric ions. To elucidate the D-glucose dehydration mechanism, 3-O-methyl-D-glucose was also submitted to the mass spectrometric study; the results suggest that the C3 hydroxyl group plays a key role in the reaction mechanism. Furthermore, protonated levulinic acid was found to be formed from the monodehydrated D-glucose ionic intermediate, an alternative pathway other than the known route consisting of 5-hydroxymethyl-2-furaldehyde double hydration.

Evidence for organic cation transporter-mediated metformin transport and 5'-adenosine monophosphate-activated protein kinase activation in rat skeletal muscles.

OBJECTIVE: 5'-Adenosine monophosphate-activated protein kinase (AMPK) is a key molecule of metabolic enhancement in skeletal muscle. We investigated whether metformin (MET) acts directly on skeletal muscle, is transported into skeletal muscle via organic cation transporters (OCTs), and activates AMPK. MATERIALS/METHODS: Isolated rat epitrochlearis and soleus muscles were incubated in vitro either in the absence or in the presence of MET. The activation status of AMPK, the intracellular energy status, and glucose and MET transport activity were then evaluated. The effect of cimetidine, which is an OCT inhibitor, on AMPK activation was also examined. RESULTS: MET (10 mmol/L, ≥60 min) increased the phosphorylation of Thr¹7² at the catalytic α subunit of AMPK in both muscles. AMPK activity assays showed that both AMPKα1 and AMPKα2 activity increased significantly. The AMPK activation was associated with energy deprivation, which was estimated from the ATP, phosphocreatine (PCr), and glycogen content, and with increased rates of 3-O-methyl-D-glucose (3MG) transport. MET did not change the basal phosphorylation status of insulin receptor signaling molecules. MET was transported into the cytoplasm in a time-dependent manner, and cimetidine suppressed MET-induced AMPK phosphorylation and 3MG transport. CONCLUSION: These results suggest that MET is acutely transported into skeletal muscle by OCTs, and stimulates AMPKα1 and α2 activity in both fast- and slow-twitch muscle types, at least in part by reducing the energy state.

Accelerated intestinal glucose absorption in morbidly obese humans: relationship to glucose transporters, incretin hormones, and glycemia.

CONTEXT: Intestinal glucose absorption is mediated by sodium-dependent glucose transporter 1 (SGLT-1) and glucose transporter 2 (GLUT2), which are linked to sweet taste receptor (STR) signaling and incretin responses. OBJECTIVE: This study aimed to examine intestinal glucose absorption in morbidly obese humans and its relationship to the expression of STR and glucose transporters, glycemia, and incretin responses. DESIGN/SETTING/PARTICIPANTS: Seventeen nondiabetic, morbidly obese subjects (body mass index [BMI], 48 ± 4 kg/m(2)) and 11 lean controls (BMI, 25 ± 1 kg/m(2)) underwent endoscopic duodenal biopsies before and after a 30-minute intraduodenal glucose infusion (30 g glucose and 3 g 3-O-methylglucose [3-OMG]). MAIN OUTCOME MEASURES: Blood glucose and plasma concentrations of 3-OMG, glucose-dependent insulinotropic polypeptide (GIP), glucagon-like peptide 1 (GLP-1), insulin, and glucagon were measured over 270 minutes. Expression of duodenal SGLT-1, GLUT2, and STR (T1R2) was quantified by PCR. RESULTS: The increase in plasma 3-OMG (P < .001) and blood glucose (P < .0001) were greater in obese than lean subjects. Plasma 3-OMG correlated directly with blood glucose (r = 0.78, P < .01). In response to intraduodenal glucose, plasma GIP (P < .001), glucagon (P < .001), and insulin (P < .001) were higher, but GLP-1 (P < .001) was less in the obese compared with lean. Expression of SGLT-1 (P = .035), but not GLUT2 or T1R2, was higher in the obese, and related to peak plasma 3-OMG (r = 0.60, P = .01), GIP (r = 0.67, P = .003), and insulin (r = 0.58, P = .02). CONCLUSIONS: In morbid obesity, proximal intestine glucose absorption is accelerated and related to increased SGLT-1 expression, leading to an incretin-glucagon profile promoting hyperinsulinemia and hyperglycemia. These findings are consistent with the concept that accelerated glucose absorption in the proximal gut underlies the foregut theory of obesity and type 2 diabetes.