Diabetes mellitus (DM) is a metabolic disorder that results from a dysfunction of insulin secretion (type 1) and/or sensitivity (type 2). Type 1 and in many cases type 2 diabetic patients require daily insulin injections to control blood glucose levels and retard the appearance of diabetes-related complications. The liver plays a central role in the context of whole-body glucose homoeostasis and fuel management in general. Under physiological conditions, insulin is released by the pancreas into the portal vein and reaches the liver along with the nutrient flow absorbed in the gastrointestinal tract (GIT). Under these conditions, the actions of insulin on liver metabolism play a key role in promoting glucose and lipid storage. DM is characterized by impairment of these actions resulting in abnormal levels of blood glucose and lipids that, if left untreated, provoke widespread secondary complications. With current therapies, insulin is delivered subcutaneously and this results in a predominantly peripheral uptake, in contrast with the physiological release profile. The experimental work described on this thesis addressed the pathways for glucose disposal and storage in the liver under two experimental settings representing the fasted to fed transition and the fed state. The studies were performed in nondiabetic control rats and in a rat model for type 1 DM...

Background and aims: Na,K-ATPase is an integral membrane protein responsible for generating and maintaining transmembrane ionic gradients. It has been demonstrated that, in pancreatic β-cells, Na,K-ATPase is regulated by glucose and that this phenomenon is impaired in glucose intolerant subjects. However, the mechanism underlying glucose-induced modulation of Na,K-ATPase is still unclear. The AMP-activated protein kinase (AMPK) plays a key role in energy homeostasis, providing exquisite sensitivity to small changes in intracellular AMP. Since glucose has marked effects on oxidative metabolism and intracellular ATP and AMP levels in pancreatic β-cell, the involvement of AMPK in the cascade of events regulating Na,K-ATPase was postulated. The aim of this work was to evaluate the putative role of AMPK in the glucose-evoked regulation of Na,K-ATPase activity in the pancreatic β-cell. Materials and methods: Pancreatic -cells from normal (control) or glucose-intolerant Wistar rats (GIR) were isolated and cultured (48h). After a pre-incubation (30min) with 2.1mM glucose (G2), cell batches were challenged with G2 or G8 (8.4mM glucose) for 20min, in the presence or absence of AMPK agonist (AICAR 1mM) and antagonist (compound C (CC), 10μM). Na...

Introduction: Glucose is the most important physiological insulin secretagogue. However, the mechanisms underlying glucose-induced insulin release are not fully understood. The role of electrogenic systems such as ionic pumps, to these events remains essentially uninvestigated. Na,K-ATPase, responsible for maintaining Na+ and K+ gradients across the plasma membrane and generates a net outward current, thus changes in its activity may contribute to the early ionic events regulating insulin secretion (Therien and Blostein, 2000). Objective: The aim of this work was to evaluate the regulation of Na,K-ATPase activity by glucose in intact -cells of normal and glucose intolerant (GI) rats and its putative contribution to the regulation of insulin secretion. Material and Methods: Pancreatic -cells, from normal or control or GI rats, were isolated and cultured (48h). Cell batches were pre-incubated (30min) with 2mM glucose to reach basal. Afterwards cells were challenged with glucose in the interval 0-11mM for 60min, for dose-dependence evaluation, or with 8mM glucose for 5-120min, for time-dependence evaluation. ATPase activity was assessed in intact cells by colorimetric quantification of Pi formed in 30min. Na,K-ATPase activity was calculated by the difference between the activities obtained in the absence and in presence the of 1mM ouabain (Costa et al....

Resumo: Os mecanismos que regulam a homeostase da glucose no pós-prandial são distintos dos mecanismos desencadeados em situações de jejum. Desta forma o fígado parece desempenhar um papel fundamental na acção periférica da insulina após a refeição através de um mecanismo que envolve os nervos parassimpáticos hepáticos e o óxido nítrico (NO). Esta dissertação procura evidenciar a importância de ambos na fi siologia de manutenção da glicémia pós-prandial e na fi siopatologia da resistência à insulina. Dos resultados obtidos observou-se que após a administração de uma refeição mista o perfi l glicémico foi distinto em animais com ou sem ablação dos nervos parassimpáticos hepáticos. A desnervação parassimpática hepática aumentou as excursões de glucose imediatamente após a refeição. Estas diferenças nas excursões de glucose dependentes do parassimpático ocorreram devido a uma diminuição da clearance de glucose, sem que fosse afectada a taxa de aparecimento de glucose no sangue, a produção endógena de glucose e secreção de insulina ou péptido-C. Este aumento das excursões de glucose revelou-se ser devida à diminuição da clearance de glucose pós-prandial exclusivamente no músculo-esquelético...

Les résultats présentés dans cette thèse précisent certains aspects de la fonction du cotransporteur Na+/glucose (SGLT1), une protéine transmembranaire qui utilise le gradient électrochimique favorable des ions Na+ afin d’accumuler le glucose à l’intérieur des cellules épithéliales de l’intestin grêle et du rein. Nous avons tout d’abord utilisé l’électrophysiologie à deux microélectrodes sur des ovocytes de xénope afin d’identifier les ions qui constituaient le courant de fuite de SGLT1, un courant mesuré en absence de glucose qui est découplé de la stoechiométrie stricte de 2 Na+/1 glucose caractérisant le cotransport. Nos résultats ont démontré que des cations comme le Li+, le K+ et le Cs+, qui n’interagissent que faiblement avec les sites de liaison de SGLT1 et ne permettent pas les conformations engendrées par la liaison du Na+, pouvaient néanmoins générer un courant de fuite d’amplitude comparable à celui mesuré en présence de Na+. Ceci suggère que le courant de fuite traverse SGLT1 en utilisant une voie de perméation différente de celle définie par les changements de conformation propres au cotransport Na+/glucose, possiblement similaire à celle empruntée par la perméabilité à l’eau passive. Dans un deuxième temps...

Les oxydants infusés avec la nutrition parentéral (NP) néonatale induisent une modification du métabolisme des lipides et du glucose, donnant lieu à l’âge adulte à un phénotype de carence énergétique (faible poids, baisse de l’activité physique). L’hypothèse qu’une diète précoce riche en glucose prévient ces symptômes plus tard dans la vie, fut évalué chez le cobaye par un ANOVA en plan factoriel complet à deux facteurs (p < 0:05) : NP du jour 3 à 7, suivit d’une nourriture régulière (chow) (NP+) vs. chow à partir du 3ième jour (NP-), combiné avec une eau de consommation enrichie en glucose (G+) ou non (G-) à partir de la 3ième semaine. Les paramètres suivant ont été mesurés à l’âge de 9 semaine: taux de croissance, activité physique, activité de phosphofructokinase-1 et glucokinase (GK), niveau hépatique de glucose-6-phosphate (G6P), glycogène, pyruvate et potentiel redox du glutathion, poids du foie, glycémie, tolérance au glucose, concentrations hépatiques et plasmatiques en triacylglycérides (TG) et cholestérol. Le groupe G+ (vs. G-) avait un taux de croissance plus bas, une activité de GK et une concentration en G6P plus élevée, et un potentiel redox plus bas (moins oxydé). Le niveau plasmatique de TG était moins élevé dans le groupe NP+ (vs. NP-). Les traitements n’eurent aucun effet sur les autres paramètres. Ces résultats suggèrent qu’indépendamment de la NP...

Recent observations indicate that gastric emptying may be influenced by patterns of previous nutrient intake. The aims of this study were to determine the effects of a high glucose diet on gastric emptying of glucose and fructose, and the impact of any changes in gastric emptying on plasma concentrations of glucose, insulin and gastric inhibitory polypeptide in response to glucose and fructose loads. Gastric emptying of glucose and fructose (both 75 g dissolved in 350 ml water) were measured in seven normal volunteers on separate days while each was on a ‘standard’ diet and an identical diet supplemented with 440 g/day of glucose for 4–7 days. Venous blood samples for measurement of plasma glucose, insulin and gastric inhibitory polypeptide levels were taken immediately before and for 180 min after ingestion of glucose and fructose loads. Dietary glucose supplementation accelerated gastric emptying of glucose (50% emptying time 82±8 vs 106±10 min, p=0.004) and fructose (73±9 vs 106±9 min, p=0.001). After ingestion of glucose, plasma concentrations of insulin (p<0.05) and gastric inhibitory polypeptide (p<0.05) were higher during the glucose-supplemented diet. In contrast, plasma glucose concentrations at 60 min and 75 min were lower (p<0.05) on the glucose-supplemented diet. We conclude that short-term supplementation of the diet with glucose accelerates gastric emptying of glucose and fructose...

BACKGROUND: Dietary interventions represent a promising therapeutic strategy to optimize postprandial glycemia. The addition of protein to oral glucose has been reported to improve the glycemic profile. OBJECTIVE: The aim of the current study was to evaluate the mechanisms by which protein supplementation lowers the blood glucose response to oral glucose. DESIGN: Nine healthy men were studied on 3 d each in a random order. Subjects consumed 300-mL drinks containing either 50 g glucose (Glucose), 30 g gelatin (Protein), or 50 g glucose with 30 g gelatin (Glucose + Protein) in water labeled with 150 mg [(13)C]acetate. Blood and breath samples were subsequently collected for 3 h to measure blood glucose and plasma insulin, glucagon-like peptide 1 (GLP-1), and glucose-dependent insulinotropic polypeptide (GIP) concentrations and gastric half-emptying time, which was calculated from (13)CO(2) excretion. RESULTS: The blood glucose response was less after Glucose + Protein than after Glucose (P < 0.005); GIP was lower (P < 0.005), and there were no significant differences in plasma insulin or GLP-1. Protein alone stimulated insulin, GLP-1, and GIP (P < 0.05 for each) without elevating blood glucose. The gastric half-emptying time was greater after Glucose + Protein than after Glucose (P < 0.05) and tended to be greater for Glucose than for Protein (P = 0.06). CONCLUSIONS: In healthy humans...

The human digestive tract is a complex system that, in addition to the digestion and absorption of nutrients, serves an important neuroendocrine role. The focus of this thesis is to examine how changes in the motor function of the gastroduodenal region influence glucose absorption, gut hormone secretion, and postprandial blood glucose regulation, in different human populations, including the healthy young and those with cystic fibrosis. The studies included utilise a mix of established and novel techniques to evaluate gastroduodenal motor function and glucose absorption, and provide insights into the function of the human gut. Strict overall glycaemic control dramatically reduces the incidence and progression of micro-, and probably macrovascular, complications associated with type 1 and type 2 diabetes. Postprandial glycaemia is now recognised as an important determinant of overall glycaemia, as indicated by the glycated haemoglobin (HbA1c). The rate of glucose absorption after a meal has a major influence on postprandial glycaemia and has, therefore, been a focus of increasing research interest in recent years. Postprandial blood glucose concentrations are a poor indicator of glucose absorption due to peripheral glucose uptake and hepatic glucose release. The glucose analogue 3-O-methylglucose (3-OMG) is absorbed in the small intestine by the same mechanism as glucose...

3-O-methyl-D-glucose has been extensively used as a proxy for d-glucose uptake. This nonmetabolizable analog has lower affinity for transporters, potentially leading to underestimates of glucose absorption rates as well as overestimates of the nutritional significance of passive uptake. Here we sought to precisely quantify the bias, if any, incurred when using 3-O-methyl-D-glucose by comparing relative absorption rates with D-glucose in vivo in a seasonally frugivorous bird, the American robin. By simultaneously administering these D-glucose probes with L-glucose--the latter absorbed only via nonmediated mechanisms and the former absorbed by both mediated and nonmediated mechanisms--using common pharmacokinetic procedures, we were able to estimate the nutritional significance of paracellular uptake in this species. The relative absorption rate of 3-O-methyl-D-glucose calculated over the initial absorptive phase was not significantly different from that of D-glucose, indicating that the former provides reasonable estimates of glucose absorption rates in vivo. The ratio of L-glucose to D-glucose cumulative fractional absorption indicates that around 60% of total glucose absorption in American robins is paracellular and showed no apparent bias in using 3-O-methyl-D-glucose when averaged over the entire initial absorptive phase. Although the absorption and elimination kinetics of radiolabeled D-glucose were appropriate for pharmacokinetic analysis in this study...

Objectives: Providing effective enteral nutrition is important during critical illness. In health, glucose is absorbed from the small intestine via sodium-dependent glucose transporter-1 and glucose transporter-2, which may both be regulated by intestinal sweet taste receptors. We evaluated the effect of critical illness on glucose absorption and expression of intestinal sodium-dependent glucose transporter-1, glucose transporter-2, and sweet taste receptors in humans and mice. Design: Prospective observational study in humans and mice. Setting: ICU and university-affiliated research laboratory. Subjects: Human subjects were 12 critically ill patients and 12 healthy controls. In the laboratory 16-week-old mice were studied. Interventions: Human subjects underwent endoscopy. Glucose (30 g) and 3-O-methylglucose (3 g), used to estimate glucose absorption, were infused intraduodenally over 30 minutes. Duodenal mucosa was biopsied before and after infusion. Mice were randomized to cecal ligation and puncture to model critical illness (n = 16) or sham laparotomy (control) (n = 8). At day 5, mice received glucose (100 mg) and 3-O-methylglucose (10 mg) infused intraduodenally prior to mucosal tissue collection. Measurements and Main Results: Quantitative polymerase chain reaction was performed to measure absolute (human) and relative levels of sodium-dependent glucose transporter-1...

Excess salt intake increases blood pressure particularly during states of hyperinsulinism and insulin resistance. Insulin is presumably effective through activation of ENaC. Excess salt intake further decreases peripheral glucose uptake thus impairing glucose tolerance. Stimulation of both, the epithelial Na+ channel ENaC and of cellular glucose uptake involves phosphatidylinositide 3-kinase (PI-3K) which signals through protein kinase B (Akt/PKB) and all three members of the serum and glucocorticoid inducible kinase (SGK) family of kinases SGK1, SGK2 and SGK3. All three kinases have been previously shown to modify a variety of transporters including ENaC and the glucose transporter SGLT1. To explore the role of SGK1 in salt sensitive hypertension and peripheral glucose uptake, experiments were performed in male or female SGK1 knockout mice (sgk1-/-) and their wild type littermates (sgk1+/+) which were subjected to standard diet, high-fat diet, high fructose diet or dexamethasone treatment and allowed free access to either tap water (control-salt) or 1% saline (high-salt). Under control diet fluid intake, blood pressure, urinary flow rate and urinary Na+, K+, Cl- excretion were similar in sgk1-/- and sgk1+/+mice, plasma aldosterone concentration was however significantly higher in sgk1-/- (1.22 ± 0.18 ng/ml) than in sgk1+/+mice (0.57 ± 0.11 ng/ml). Under standard diet...

A patofisiologia de doenças metabólicas como a diabetes mellitus tipo II (T2DM) é caracterizada pela perda de sincronia entre a síntese e catabolismo de triacilgliceróis (TAG). A lipase adiposa de triacilglicerídeos (ATGL), uma enzima envolvida na hidrólise de TAGs, tem sido alvo de vários estudos de forma a compreender o seu papel na resistência à insulina. A razão para este aumento de interesse é o fato de os ATGL knock‐outs (ATGL‐/‐) terem demonstrado maior sensibilidade à insulina e tolerancia à glucose em comparação com ratinhos sem a delecção (WT). Apesar de eles não recorrerem a ácidos gordos armazenados como fonte de energia, estas características fazem dos ratinhos ATGL‐/‐ um modelo único para a melhor compreensão da relação entre o metabolismo de TAGs e a resistência à insulina. A taxa do turnover de glucose e reciclagem pelo ciclo de Cori é altamente suscetível tanto à sensibilidade a insulina como à disponibilidade de nutrientes. Sob condições normais de jejum, a maioria da glucose é reciclada pelo ciclo de Cori. No entanto, a maior dependência de glucose por parte dos ratinhos ATGL‐/‐, levou‐nos a colocar a hipótese de que estes ratinhos seriam mais dependentes da produção de glucose endógena (EGP) para fazer face a esta necessidade e que a glucose seria eliminada principalmente através da oxidação...

Glucose is the primary metabolic signal reflecting the current energy state of the body. Glucose influences the excitability of neurons in the area postrema (AP), a circumventricular organ (CVO), prompting my interest in investigating whether the subfornical organ (SFO), another sensory CVO can also detect glucose. Using patch-clamp electrophysiology, we investigated the influence of changing glucose concentrations on the excitability of SFO neurons. In dissociated SFO neurons, altering the bath concentration of glucose (1mM, 5mM, 10mM) influenced the excitability of 49% of neurons tested (n=67). Glucose-inhibited (GI, hyperpolarized by increased glucose or depolarized by decreased glucose) and glucose-excited (GE, depolarized by increased glucose or hyperpolarized by decreased glucose) neurons were observed. GI neurons (27%, n=18) depolarized in response to decreased glucose (n=10, mean 4.6 ± 1.0 mV) or hyperpolarized in response to increased glucose (n=8, mean -4.4 ± 0.8 mV). In contrast, GE neurons (22%, n=15) depolarized in response to increased glucose (n=9, mean 6.4 ± 0.4) or hyperpolarized in response to decreased glucose (n=6, mean -4.8 ± 0.6 mV). These data show that glucose acts on a subpopulation of SFO neurons to produce both excitatory and inhibitory actions. Using voltage-clamp recordings two groups of SFO neurons were identified: those producing an outward current (GI) and those producing an inward current (GE) in response to increasing concentrations of glucose from 1 to 10 mM (n=23). The mean glucose-induced inward current had a reversal potential of -24 ± 12 mV (mean input resistance 2.0 ± 0.4 GΩ...

Le diabète est une maladie métabolique qui se caractérise par une résistance à l’insuline des tissus périphériques et par une incapacité des cellules β pancréatiques à sécréter les niveaux d’insuline appropriés afin de compenser pour cette résistance. Pour mieux comprendre les mécanismes déficients dans les cellules β des patients diabétiques, il est nécessaire de comprendre et de définir les mécanismes impliqués dans le contrôle de la sécrétion d’insuline en réponse au glucose. Dans les cellules β pancréatiques, le métabolisme du glucose conduit à la production de facteurs de couplage métabolique, comme l’ATP, nécessaires à la régulation de l’exocytose des vésicules d’insuline. Le mécanisme par lequel la production de l’ATP par le métabolisme oxydatif du glucose déclenche l’exocytose des vésicules d’insuline est bien décrit dans la littérature. Cependant, il ne peut à lui seul réguler adéquatement la sécrétion d’insuline. Le malonyl-CoA et le NADPH sont deux autres facteurs de couplage métaboliques qui ont été suggérés afin de relier le métabolisme du glucose à la régulation de la sécrétion d’insuline. Les mécanismes impliqués demeurent cependant à être caractérisés. Le but de la présente thèse était de déterminer l’implication des navettes du pyruvate...

Plasma glucose, insulin and glucose tolerance were quantified in diabetic Goto–Kakizaki (GK) rats (342 ± 45 g, n = 5) and compared with weight-matched non-diabetic Wistars (307 ± 30 g, n = 8). Compared to Wistars, GK rats had higher fasting plasma insulin (219 ± 50 versus 44 ± 14 pmol/l, P < 0.002) and glucose (9.2 ± 2.3 versus 5.5 ± 0.5 mmol/l, P < 0.025). GK rats showed impaired glucose tolerance (IPGTT 2 h plasma glucose = 14 ± 1.5 versus 6.4 ± 0.1 mmol/l, P < 0.001). Endogenous glucose production (EGP) from glycogenolysis, phosphoenolpyruvate (PEP) and glycerol after 6 hours of fasting was quantified by a primed infusion of [U–13C]glucose and 2H2O tracers and 2H/13C NMR analysis of plasma glucose. EGP was higher in GK compared to Wistar rats (191 ± 16 versus 104 ± 27 μmol/kg per min, P < 0.005). This was sustained by increased gluconeogenesis from PEP (85 ± 12 versus 35 ± 4 μmol/kg per min, P < 0.02). Gluconeogenesis from glycerol was not different (20 ± 3 in Wistar versus 30 ± 6 μmol/kg per min for GK), and glycogenolysis fluxes were also not significantly different (76 ± 23 μmol/kg per min for GK versus 52 ± 19 μmol/kg per min for Wistar). The Cori cycle accounted for most of PEP gluconeogenesis in both Wistar and GK rats (85 ± 15% and 77 ± 10%...

INTRODUCTION: Disorders that affect glucose metabolism, namely diabetes mellitus (DM), may favor the development and/or progression of osteoarthritis (OA). Thus far, little is known regarding the ability of chondrocytes to adjust to variations in the extracellular glucose concentration, resulting from hypoglycemia and hyperglycemia episodes, and so, to avoid deleterious effects resulting from deprivation or intracellular accumulation of glucose. The aim of this study was to compare the ability of normal and OA chondrocytes to regulate their glucose transport capacity in conditions of insufficient or excessive extracellular glucose and to identify the mechanisms involved and eventual deleterious consequences, namely the production of reactive oxygen species (ROS). METHODS: Chondrocytes, isolated from normal and OA human cartilage, were maintained in high-density monolayer cultures, in media without or with 10 or 30 mM glucose. Glucose transport was measured as the uptake of 2-deoxy-D-glucose (2-DG). Glucose transporter-1 (GLUT-1) mRNA and protein content were evaluated by real-time RT-PCR and western blot, respectively. ROS production was measured with 2',7'-dichlorodihydrofluorescein diacetate. RESULTS: Basal and IL-1beta-induced 2-DG uptake...

This is the accepted manuscript version. The final version is available from the American Association for Clinical Chemistry at http://www.clinchem.org/content/60/12/1500.long.; Background: Accuracy and frequency of glucose measurement is essential to achieve safe and efficacious glucose control in the ICU. Emerging continuous glucose monitors provide frequent measurements, trending information and alarms. The objective of this study was to establish the level of accuracy of continuous glucose monitoring (CGM) associated with safe and efficacious glucose control in the intensive care unit. Methods: Three established glucose control protocols (Yale, University of Washington, and NICE-SUGAR) underwent evaluation using computer simulations. Insulin delivery was informed by intermittent blood glucose (BG) measurements or CGM levels with an increasing level of the measurement error. Measures of glucose control included mean glucose, glucose variability, time glucose was in target range, and hypoglycemia episodes. Results: Apart from Washington protocol, CGM with mean absolute relative deviation (MARD) up to 15% resulted in similar mean glucose as with the use of intermittent BG measurements. Glucose variability was also similar between CGM and BG-informed protocols. Frequency and duration of hypoglycemia were not worse using CGM with MARD at or below 10%. Measures of glucose control varied more between protocols than at different levels of the CGM error. Conclusions: The efficacy of CGM-informed and BG-informed commonly used glucose protocols is similar but the risk of hypoglycemia may be reduced using CGM with MARD at or below 10%. Protocol choice has greater influence on glucose control measures than the glucose measurement method.; Edwards Lifesciences provided educational grant to conduct the study but did not play any role in data analysis or interpretation of study results.

BACKGROUND: The bioluminescence technique was used to quantify the local glucose concentration in the tissue surrounding subcutaneously implanted polyurethane material and surrounding glucose sensors. In addition, some implants were coated with a single layer of adipose-derived stromal cells (ASCs) because these cells improve the wound-healing response around biomaterials. METHODS: Control and ASC-coated implants were implanted subcutaneously in rats for 1 or 8 weeks (polyurethane) or for 1 week only (glucose sensors). Tissue biopsies adjacent to the implant were immediately frozen at the time of explant. Cryosections were assayed for glucose concentration profile using the bioluminescence technique. RESULTS: For the polyurethane samples, no significant differences in glucose concentration within 100 μm of the implant surface were found between bare and ASC-coated implants at 1 or 8 weeks. A glucose concentration gradient was demonstrated around the glucose sensors. For all sensors, the minimum glucose concentration of approximately 4 mM was found at the implant surface and increased with distance from the sensor surface until the glucose concentration peaked at approximately 7 mM at 100 μm. Then the glucose concentration decreased to 5.5-6.5 mM more than 100 μmm from the surface. CONCLUSIONS: The ASC attachment to polyurethane and to glucose sensors did not change the glucose profiles in the tissue surrounding the implants. Although most glucose sensors incorporate a diffusion barrier to reduce the gradient of glucose and oxygen in the tissue...