Lumenal Glucose Research Articles

Commentary: On the Pathophysiology of Coronary Heart Disease

This article highlights some important aspects of the pathophysiology of coronary artery disease, namely: • The distribution of lesions within the arterial tree at sites of low wall shear stress. • The potential role of low flow-mediated arterial dilatation at sites of low wall stress. • Low flow-mediated dilatation leads to low nitric oxide production by the arterial endothelium and consequent reduced protection against lesion formation. • Flow-mediated dilatation is reduced by high lumenal glucose concentration. • The role of the glycocalyx dysfunction in mediating flow-mediated dilatation and consequent reduced NO production by the arterial endothelium and cell adhesion. • Stenoses cause convective acceleration of blood velocity and a consequent increase in platelet shear stress. • Increased platelet shear stress activates platelets with release of serotonin. • Serotonin activates more platelet activation via the 5HT2A platelet receptor causing a positive feedback and thrombus growth. • Arterial thrombus growth is abolished by 5HT2A receptor antagonists, the key to improved treatment of the disease. • One 5HT2A receptor antagonist has been shown in humans to be safe and to cause no excess bleeding from wounds.

Intestinal glucose-induced calcium-calmodulin kinase signaling in the gut-brain axis in awake rats

Lumenal glucose initiates changes in gastrointestinal (GI) function, including inhibition of gastric emptying, stimulation of pancreatic exocrine and endocrine secretion, and intestinal fluid secretion. Glucose stimulates the release of GI hormones and 5-hydroxytryptamine (5-HT), and activates intrinsic and extrinsic neuronal pathways to initiate changes in GI function. The precise mechanisms involved in luminal glucose-sensing are not clear; studying gut endocrine cells is difficult due to their sparse and irregular localization within the epithelium. Here we show a technique to determine activation of gut epithelial cells and the gut-brain pathway in vivo in rats using immunohistochemical detection of the activated, phosphorylated, form of calcium-calmodulin kinase II (pCaMKII). Perfusion of the gut with glucose (60 mg) increased pCaMKII immunoreactivity in 5-HT-expressing enterochromaffin (EC) cells, cytokeratin-18 immunopositive brush cells, but not in enterocytes or cholecystokinin-expressing cells. Lumenal glucose increased pCaMKII in neurons in the myenteric plexus and nodose ganglion, nucleus of the solitary tract, dorsal motor nucleus of the vagus and the arcuate nucleus. pCaMKII expression in neurons, but not in EC cells, was significantly attenuated by pretreatment with the 5-HT(3) R antagonist ondansetron. Deoxynojirimycin, a selective agonist for the putative glucose sensor, sodium-glucose cotransporter-3 (SGLT-3), mimicked the effects of glucose with increased pCaMKII in ECs and neurons; galactose had no effect. The data suggest that native EC cells in situ respond to glucose, possibly via SGLT-3, to activate intrinsic and extrinsic neurons and thereby regulate GI function.

Solute carrier family 2, member 9 and uric acid homeostasis

The goal of this article is to review the possible physiological roles of the recently identified urate transporter, solute carrier family 2 (facilitated glucose transporter), member 9 (SLC2A9), in the renal handling of urate. Glucose transporter 9 is a high affinity hexose transporter encoded by the SLC2A9 gene found on human chromosome 4. The two splice variants SLC2A9b and SLC2A9a are expressed in the apical and basolateral membranes, respectively, of the proximal convoluted tubule. Recent reports have found significant correlations between two different sets of single nucleotide polymorphisms in SLC2A9. In one case, they are associated with increases in plasma urate levels and/or the incidence of hypertension or gout. The second set of single nucleotide polymorphisms correlate with hypouricaemia in Japanese patients. Expression of SLC2A9a and b in Xenopus laevis oocytes shows that these proteins mediate rapid urate fluxes and can exchange glucose for urate. Indirect evidence also suggests that the transporter is electrogenic. This review proposes that SLC2A9 contributes significantly in two ways to the fluxes of urate across the proximal convoluted tubule. Firstly, the apical expression of SLC2A9b secretes urate back into the urine in exchange for lumenal glucose. Secondly, the basolateral membrane SLC2A9a could be the primary route for urate movement out of the epithelium into the peritubular space.

K+-ATP-channel-related protein complexes: potential transducers in the regulation of epithelial tight junction permeability

K(+)-ATP channels are composed of an inwardly rectifying Kir6 subunit and an auxiliary sulfonylurea receptor (SUR) protein. The SUR subunits of Kir6 channels have been recognized as an ATPase, which appears to work as a mechanochemical device like other members of the ABC protein family. Thus, in spite of just gating ions, Kir6/Sur might, in addition, regulate completely different cellular systems. However, so far no model system was available to directly investigate this possibility. Using highly specific antibodies against Kir6.1-SUR2A and an in vitro model system of the rat small intestine, we describe a new function of the Kir6.1-SUR2A complex, namely the regulation of paracellular permeability. The Kir6.1-SUR2A complex localizes to regulated tight junctions in a variety of gastrointestinal, renal and liver tissues of rat, pig and human, whereas it is absent in the urothelium. Changes in paracellular permeability following food intake was investigated by incubating the lumen of morphological well-defined segments of rat small intestine with various amounts of glucose. Variations in the lumenal glucose concentrations and regulators of Kir6.1/SUR2A activity, such as tolbutamide or diazoxide, specifically modulate paracellular permeability. The data presented here shed new light on the physiological and pathophysiological role K(+)-ATP channels might have for the regulation of tight junctions.

Glucose sensing in the intestinal epithelium

Dietary sugars regulate expression of the intestinal Na+/glucose cotransporter, SGLT1, in many species. Using sheep intestine as a model, we showed that lumenal monosaccharides, both metabolisable and nonmetabolisable, regulate SGLT1 expression. This regulation occurs not only at the level of transcription, but also at the post-transcriptional level. Introduction of d-glucose and some d-glucose analogues into ruminant sheep intestine resulted in > 50-fold enhancement of SGLT1 expression. We aimed to determine if transport of sugar into the enterocytes is required for SGLT1 induction, and delineate the signal-transduction pathways involved. A membrane impermeable d-glucose analogue, di(glucos-6-yl)poly(ethylene glycol) 600, was synthesized and infused into the intestines of ruminant sheep. SGLT1 expression was determined using transport studies, Northern and Western blotting, and immunohistochemistry. An intestinal cell line, STC-1, was used to investigate the signalling pathways. Intestinal infusion with di(glucos-6-yl)poly(ethylene glycol) 600 led to induction of functional SGLT1, but the compound did not inhibit Na+/glucose transport into intestinal brush-border membrane vesicles. Studies using cells showed that increased medium glucose up-regulated SGLT1 abundance and SGLT1 promoter activity, and increased intracellular cAMP levels. Glucose-induced activation of the SGLT1 promoter was mimicked by the protein kinase A (PKA) agonist, 8Br-cAMP, and was inhibited by H-89, a PKA inhibitor. Pertussis toxin, a G-protein (Gi)-specific inhibitor, enhanced SGLT1 protein abundance to levels observed in response to glucose or 8Br-cAMP. We conclude that lumenal glucose is sensed by a glucose sensor, distinct from SGLT1, residing on the external face of the lumenal membrane. The glucose sensor initiates a signalling pathway, involving a G-protein-coupled receptor linked to a cAMP-PKA pathway resulting in enhancement of SGLT1 expression.

Provision of phosphorylatable substrate during hypoxia decreases jejunal barrier function

There is an emerging consensus that early enteral nutrition benefits the high-risk surgical patient. However, in patients with inadequate gastrointestinal perfusion, food in the intestine may increase the oxygen demand beyond that which can be satisfied by the available delivery, potentially leading to intestinal malfunction. The effect of metabolic substrate on gastrointestinal function during various oxygenation states was investigated. Jejunal samples obtained from 32 male Sprague-Dawley rats (263 +/- Q15 g) were stripped of the muscularis, mounted in modified Ussing chambers, and randomized to be incubated in media equilibrated with one of four gas mixtures (95%, 75%, 50%, and 25% oxygen). After equilibration, fluorescent probes (4400 and 17 200 molecular weight [MW]) were added to the incubation media on the mucosal side. The rate of probe accumulation on the serosal side was determined before and after the addition of one of four substrates to the mucosal medium: mannitol (an osmotic control), glucose (which is transported, phosphorylated, and metabolized), 2-deoxyglucose (a glucose analog that is transported and phosphorylated but not metabolized), or 3-O-methylglucose (a glucose analog that is transported but not phosphorylated or metabolized). Lumenal glucose, 2-deoxyglucose, and 3-O-methylglucose increased permeation of the 4400-MW probe at all oxygen levels, whereas mannitol did not alter permeation in the 95% and 75% oxygen groups. Lumenal glucose increased (P < 0.05) the permeation rate of the 17 200-MW probe at all oxygen levels, whereas 2-deoxyglucose and 3-O-methylglucose did not increase the permeation rate of the 17 200-MW probe until oxygen was lowered to 75% and 50%, respectively. Regardless of substrate treatment, jejunal permeation of the 4400-MW (P < 0.001) and 17 200-MW (P < 0.05) probes increased in the 25% and 50% oxygen groups compared with the 75% and 95% oxygen groups. These initial results suggest that the provision of lumenal nutrients exacerbates the loss of gastrointestinal barrier function during hypoxia. Although the early provision of nutrients is an important intervention in acutely injured patients, care must be taken to ensure that gastrointestinal perfusion is adequate to allow substrate metabolism and prevent further compromise in gastrointestinal function.

Basolateral D-glucose transport activity along the crypt-villus axis in rat jejunum and upregulation induced by gastric inhibitory peptide and glucagon-like peptide-2.

Phloridzin-insensitive D-glucose uptake into enterocytes isolated sequentially from along the crypt-villus axis showed the majority of transport activity to reside in cells from the upper third of the villus. In contrast, total postnuclear glucose transporter 2 (GLUT2) protein content of the cells was high even close to the crypt and was almost constant for the upper 80% of the villi. A 4 h lumenal perfusion in vivo with 100 mM D-glucose prior to harvesting the enterocytes produced a 2- to 3-fold increase in phloridzin-insensitive D-glucose uptake which extended down 70% of the villus. Vascular infusion in vivo with either 800 pM gastric inhibitory polypeptide (GIP) or glucagon-like peptide-2 (GLP-2) prior to harvesting enterocytes produced the same response as lumenal glucose, while glucagon like peptide-1 (GLP-1) had no effect. Inclusion of 30 microM brefeldin A (BFA), an inhibitor of protein trafficking, in the lumenal perfusate produced a small, but not significant, increase in the control uptake profile along the villus in isolated enterocytes. However, BFA significantly reduced the upregulation induced by lumenal glucose and vascular GIP and blocked the stimulation produced by vascular GLP-2. Biotinylation of surface proteins and isolation with protein A indicated that there was no change in the membrane abundance of GLUT2 after GLP-2 treatment. These results are discussed in relation to the role of gastrointestinal peptide hormones in controlling intestinal hexose transport and the possibility of protein trafficking being involved in mediating the upregulation of GLUT2 activity in the enterocyte basolateral membrane.

Steady-state cerebral glucose concentrations and transport in the human brain.

Understanding the mechanism of brain glucose transport across the blood-brain barrier is of importance to understanding brain energy metabolism. The specific kinetics of glucose transport have been generally described using standard Michaelis-Menten kinetics. These models predict that the steady-state glucose concentration approaches an upper limit in the human brain when the plasma glucose level is well above the Michaelis-Menten constant for half-maximal transport, Kt. In experiments where steady-state plasma glucose content was varied from 4 to 30 mM, the brain glucose level was a linear function of plasma glucose concentration. At plasma concentrations nearing 30 mM, the brain glucose level approached 9 mM, which was significantly higher than predicted from the previously reported Kt of approximately 4 mM (p < 0.05). The high brain glucose concentration measured in the human brain suggests that ablumenal brain glucose may compete with lumenal glucose for transport. We developed a model based on a reversible Michaelis-Menten kinetic formulation of unidirectional transport rates. Fitting this model to brain glucose level as a function of plasma glucose level gave a substantially lower Kt of 0.6 +/- 2.0 mM, which was consistent with the previously reported millimolar Km of GLUT-1 in erythrocyte model systems. Previously reported and reanalyzed quantification provided consistent kinetic parameters. We conclude that cerebral glucose transport is most consistently described when using reversible Michaelis-Menten kinetics.

Rapid enhancement of brush border glucose uptake after exposure of rat jejunal mucosa to glucose.

Increased jejunal glucose transport after ingestion of carbohydrate rich diets may reflect higher concentrations of lumenal glucose. Normal processing of carbohydrate causes wide fluctuations in glucose concentration in the jejunal lumen and this raises the question of whether the high lumenal concentrations seen at peak digestion affect glucose uptake. To study the effects of 30 minute exposure of rat jejunal mucosa to glucose on sodium-glucose transporter (SGLT1) mediated glucose transport across the brush border membrane. Jejunal mucosa was exposed in vitro or in vivo to 25 mM glucose or 25 mM mannitol for 30 minutes. In addition, isolated villus enterocytes were incubated with mannitol or glucose for the same time. Brush border membrane vesicles were isolated from these preparations and phlorizin sensitive 3H-D-glucose accumulation was measured. Lumenal glucose in vivo significantly enhanced SGLT1 mediated glucose uptake by 49.2-57.2%. For jejunal loops in vitro, the increase was 32.0-85.2%. Kinetic analysis disclosed a 50% greater Vmax for glucose uptake in each preparation. The facilitated and passive components of uptake were, however, unaffected by prior exposure to glucose. Incubation of villus enterocytes with 25 mM glucose did not influence glucose uptake by brush border membranes. Finally, exposure of intact mucosa to 20 mM galactose, a nonmetabolised sugar also transported by SGLT1, did not alter glucose transport. Lumenal glucose promotes glucose transport by brush border membrane within 30 minutes. An intact mucosa is necessary for upregulation and evidence suggests that the response is mediated by locally acting mechanisms.

Ontogenesis of intestine morphology and intestinal disaccharidases in chickens (Gallus gallus) fed contrasting purified diets.

Two groups of growing posthatching Cornish x Rock cross chickens were fed with either a carbohydrate-containing (52.5%) or a carbohydrate-free diet. At 36 days after hatching some of the chicks in each group were shifted to the opposite diet. Chickens fed on a carbohydrate-containing diet grew faster and achieved higher asymptotic masses than chickens fed on a carbohydrate-free diet. Chickens fed on a carbohydrate-free diet had longer intestines and larger intestinal areas than chickens of the same mass fed on a carbohydrate-containing diet. In both groups sucrase and maltase activity (standardized by either intestinal area or mass) increased from day 1 to approximately day 17. After day 17, chickens fed on a carbohydrate-containing diet exhibited 1.8 and 1.9 times higher sucrase and maltase activities per unit intestinal area, respectively, than chickens fed on a carbohydrate-free diet. Analysis of covariance was used to estimate the contribution of sucrase and the sucrase-independent maltases to maltase activity, and to estimate the effect of diet on the sucrase-independent maltases. Sucrase contributed 80% and 75% of the maltase activity in carbohydrate and carbohydrate-free fed chickens, respectively. Chickens shifted from a carbohydrate-free to a carbohydrate diet converged in gross intestinal morphology and intestinal sucrase and maltase levels with carbohydrate-fed chickens within 8 days. Chickens shifted from carbohydrate to carbohydrate-free diets, in contrast, did not show appreciable changes in intestinal length and after 8 days had not reduced levels of sucrase and maltase to those of chickens fed on the carbohydrate-free diet. A comparison of integrated maltase intestinal activity with published data on glucose uptake showed that the ratio of maltose hydrolysis to glucose uptake seemed to be about 7 and to remain relatively invariant during ontogeny. Because so little is known about the interaction between hydrolysis and uptake in vivo, it is difficult to determine if this relatively high ratio represents excess hydrolytic capacity or if it is needed to provide high lumenal glucose concentrations that maximize uptake.

In vitro perfusion of hybrid artificial pancreas devices at low flow rates.

Type I diabetes is characterized by insulin insufficiency due to lack of functional beta cells. To replace injection therapy, schemes such as the Hybrid Artificial Pancreas (HAP) were developed. This consists of an acrylic housing enclosing a semipermeable hollow fiber membrane. Donor islets can be seeded in the annular space through a port in the housing, and thus are separated from the recipient's bloodstream or perfusate. Before scaling the HAP to human size, the dynamics of its insulin response to a perfusion glucose challenge must be better understood. In this study, the HAP's insulin response after a step increase in the lumenal glucose concentration was determined as a function of the radial thickness of the annular space (0.173-0.973 mm) and islet distribution at a flow rate of 1 ml/min. Devices containing a single, 65 mm long fiber were used. Rat islets were isolated using standard collagenase digestion techniques. In unseeded HAP perfusions, the washout time for glucose and insulin from the annular space was dependent on flow rate and radial thickness. Both solutes were removed in < 3 min from the smallest devices when perfused at 10 ml/min. Thus, solute transport within the HAP is very fast. In the seeded HAP perfusions, the devices were subjected to a step increase in the lumenal glucose concentration. Sequential samples of the HAP effluent were collected and assayed for glucose and insulin. The spatial distribution of the islets in the annular space was one of the most important factors in determining the HAP's insulin response.(ABSTRACT TRUNCATED AT 250 WORDS)