Deletion of the Imprinted Phlda2 Gene Increases Placental Passive Permeability in the Mouse.

Genomic imprinting, an epigenetic phenomenon that causes the expression of a small set of genes in a parent-of-origin-specific manner, is thought to have co-evolved with placentation. Many imprinted genes are expressed in the placenta, where they play diverse roles related to development and nutrient supply function. However, only a small number of imprinted genes have been functionally tested for a role in nutrient transfer capacity in relation to the structural characteristics of the exchange labyrinthine zone. Here, we examine the transfer capacity in a mouse model deficient for the maternally expressed Phlda2 gene, which results in placental overgrowth and a transient reduction in fetal growth. Using stereology, we show that the morphology of the labyrinthine zone in Phlda2-/+ mutants is normal at E16 and E19. In vivo placental transfer of radiolabeled solutes 14C-methyl-D-glucose and 14C-MeAIB remains unaffected at both gestational time points. However, placental passive permeability, as measured using two inert hydrophilic solutes (14C-mannitol; 14C-inulin), is significantly higher in mutants. Importantly, this increase in passive permeability is associated with fetal catch-up growth. Our findings uncover a key role played by the imprinted Phlda2 gene in modifying placental passive permeability that may be important for determining fetal growth.

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.

Small intestinal epithelial permeability to water-soluble nutrients higher in passerine birds than in rodents.

In the small intestine transcellular and paracellular pathways are implicated in water-soluble nutrient absorption. In small birds the paracellular pathway is quantitatively important while transcellular pathway is much more important in terrestrial mammals. However, there is not a clear understanding of the mechanistic underpinnings of the differences among taxa. This study was aimed to test the hypothesis that paracellular permeability in perfused intestinal segments is higher in passerine birds than rodents. We performed in situ intestinal perfusions on individuals of three species of passerine birds (Passer domesticus, Taeniopygia guttata and Furnarius rufus) and two species of rodents (Mus musculus and Meriones ungiculatus). Using radio-labelled molecules, we measured the uptake of two nutrients absorbed by paracellular and transcellular pathways (L-proline and 3-O-methyl-D-glucose) and one carbohydrate that has no mediated transport (L-arabinose). Birds exhibited ~2 to ~3 times higher L-arabinose clearance per cm2 epithelium than rodents. Moreover, paracellular absorption accounted for proportionally more of 3-O-methyl-D-glucose and L-proline absorption in birds than in rodents. These differences could be explained by differences in intestinal permeability and not by other factors such as increased retention time or higher intestinal nominal surface area. Furthermore, analysis of our results and all other existing data on birds, bats and rodents shows that insectivorous species (one bird, two bats and a rodent) had only 30% of the clearance of L-arabinose of non-insectivorous species. This result may be explained by weaker natural selection for high paracellular permeability in animal- than in plant-consumers. Animal-consumers absorb less sugar and more amino acids, whose smaller molecular size allow them to traverse the paracellular pathway more extensively and faster than glucose.

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.

A microvascular compartment model validated using 11C-methylglucose liver PET in pigs.

The standard compartment model (CM) is widely used to analyse dynamic PET data. The CM is fitted to time-activity curves to estimate rate constants that describe the transport of a tracer between well-mixed compartments. The aim of this study was to develop and validate a more realistic microvascular compartment model (MCM) that includes capillary tracer concentration gradients, backflux from cells into the perfused capillaries and multiple re-uptakes during the passage through a capillary. The MCM incorporates only parameters with clear physiological meaning, it is easy to implement, and it does not require numerical solution. We compared the MCM and CM for the analysis of 3 min dynamic PET data of pig livers (N = 5) following injection of 11C-methylglucose. During PET scans, the tracer concentrations in blood were measured in the abdominal aorta, portal vein and liver vein by manual sampling. We found that the MCM outperformed the CM and that dynamic PET data include information which cannot be extracted using standard CM. The MCM fitted dynamic PET data better than the CM (Akaike values were 46 ± 4 for best MCM fits, and 82 ± 8 for best CM fits; mean ± standard deviation) and extracted physiologically reasonable parameter estimates such as blood perfusion that were in agreement with independent measurements. The difference between model-independent perfusion estimates and the best MCM perfusion estimates was -0.01 ± 0.05 ml/ml/min, whereas the difference was 0.30 ± 0.13 ml/ml/min using the CM. In addition, the MCM predicted the time course of concentrations in the liver vein, a prediction fundamentally unobtainable using the CM as it does not return tracer backflux from cells to capillary blood. The results demonstrate the benefit of using models that include more physiology and that models including concentration gradients should be preferred when analysing the blood-cell exchange of any tracer in any capillary bed.

Intestinal paracellular absorption is necessary to support the sugar oxidation cascade in nectarivorous bats.

We made the first measurements of the capacity for paracellular nutrient absorption in intact nectarivorous bats. Leptonycteris yerbabuenae (20 g mass) were injected with or fed inert carbohydrate probes L-rhamnose and D(+)-cellobiose, which are absorbed exclusively by the paracellular route, and 3-O-methyl-D-glucose (3OMD-glucose), which is absorbed both paracellularly and transcellularly. Using a standard pharmacokinetic technique, we collected blood samples for 2 h after probe administration. As predicted, fractional absorption (f) of paracellular probes declined with increasing Mr in the order of rhamnose (f=0.71)>cellobiose (f=0.23). Absorption of 3OMD-glucose was complete (f=0.85; not different from unity). Integrating our data with those for glucose absorption and oxidation in another nectarivorous bat, we conclude that passive paracellular absorption of glucose is extensive in nectarivorous bat species, as in other bats and small birds, and necessary to support high glucose fluxes hypothesized for the sugar oxidation cascade.

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.

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.

Measurement of gastric emptying in the critically ill.

BACKGROUND & AIMS: Enteral nutrition is important in critically ill patients and is usually administered via a nasogastric tube. As gastric emptying is frequently delayed, and this compromises the delivery of nutrient, it is important that the emptying rate can be quantified. METHODS: A comprehensive search of MEDLINE/PubMed, of English articles, from inception to 1 July 2014. References of included manuscripts were also examined for additional studies. RESULTS: A number of methods are available to measure gastric emptying and these broadly can be categorised as direct- or indirect-test and surrogate assessments. Direct tests necessitate visualisation of the stomach contents during emptying and are unaffected by liver or kidney metabolism. The most frequently used direct modality is scintigraphy, which remains the 'gold standard'. Indirect tests use a marker that is absorbed in the proximal small intestine, so that measurements of the marker, or its metabolite measured in plasma or breath, correlates with gastric emptying. These tests include drug and carbohydrate absorption and isotope breath tests. Gastric residual volumes (GRVs) are used frequently to quantify gastric emptying during nasogastric feeding, but these measurements may be inaccurate and should be regarded as a surrogate measurement. While the inherent limitations of GRVs make them less suitable for research purposes they are often the only technique that is available for clinicians at the bedside. CONCLUSIONS: Each of the available techniques has its strength and limitations. Accordingly, the choice of gastric emptying test is dictated by the particular requirement(s) and expertise of the investigator or clinician.

Interactions among glucose delivery, transport, and phosphorylation that underlie skeletal muscle insulin resistance in obesity and type 2 Diabetes: studies with dynamic PET imaging.

Dynamic positron emission tomography (PET) imaging was performed using sequential tracer injections ([(15)O]H2O, [(11)C]3-O-methylglucose [3-OMG], and [(18)F]fluorodeoxyglucose [FDG]) to quantify, respectively, skeletal muscle tissue perfusion (glucose delivery), kinetics of bidirectional glucose transport, and glucose phosphorylation to interrogate the individual contribution and interaction among these steps in muscle insulin resistance (IR) in type 2 diabetes (T2D). PET imaging was performed in normal weight nondiabetic subjects (NW) (n = 5), obese nondiabetic subjects (OB) (n = 6), and obese subjects with T2D (n = 7) during fasting conditions and separately during a 6-h euglycemic insulin infusion at 40 mU · m(-2) · min(-1). Tissue tracer activities were derived specifically within the soleus muscle with PET images and magnetic resonance imaging. During fasting, NW, OB, and T2D subjects had similar [(11)C]3-OMG and [(18)F]FDG uptake despite group differences for tissue perfusion. During insulin-stimulated conditions, IR was clearly evident in T2D (P < 0.01), and [(18)F]FDG uptake by muscle was inversely correlated with systemic IR (P < 0.001). The increase in insulin-stimulated glucose transport was less (P < 0.01) in T2D (twofold) than in NW (sevenfold) or OB (sixfold) subjects. The fractional phosphorylation of [(18)F]FDG during insulin infusion was also significantly lower in T2D (P < 0.01). Dynamic triple-tracer PET imaging indicates that skeletal muscle IR in T2D involves a severe impairment of glucose transport and additional impairment in the efficiency of glucose phosphorylation.

Paracellular absorption in laboratory mice: Molecule size-dependent but low capacity.

Water-soluble nutrients are absorbed by the small intestine via transcellular and paracellular processes. The capacity for paracellular absorption seems lower in nonfliers than in fliers, although that conclusion rests largely on a comparison of relatively larger nonflying mammals (>155g) and relatively smaller flying birds (<155g). We report on paracellular absorption in laboratory mice, the smallest nonflying mammal species studied to date. Using a standard pharmacokinetic technique, we measured the extent of absorption (fractional absorption=f) of inert carbohydrate probes: L-arabinose (M(r)=150.13Da) and cellobiose (342.3) that are absorbed exclusively by the paracellular route, and 3-O-methyl D-glucose (3OMD-glucose) (M(r)=194) absorbed both paracellularly and transcellularly. f was measured accurately in urine collection trials of 5-10h duration. Absorption of 3OMD-glucose by mice was essentially complete (f=0.95±0.07) and much higher than that for L-arabinose (f=0.21±0.02), indicating that in mice, like other nonflying mammals, >80% of glucose is absorbed by mediated process(es) rather than the passive, paracellular route. As in all other vertebrates, absorption of cellobiose (f=0.13±0.02) was even lower than that for L-arabinose, suggesting an equivalent molecular size cut-off for flying and nonflying animals and thus a comparable effective TJ aperture. An important ecological implication is that smaller water-soluble plant secondary metabolites that have been shown to be absorbed by the paracellular path in cell culture, such as phenolics and alkaloids, might be absorbed in substantial amounts by bats and small birds relative to nonflying mammals such as mice.

AMP kinase regulation of sugar transport in brain capillary endothelial cells during acute metabolic stress.

AMP-dependent kinase (AMPK) and GLUT1-mediated sugar transport in blood-brain barrier endothelial cells are activated during acute cellular metabolic stress. Using murine brain microvasculature endothelium bEnd.3 cells, we show that AMPK phosphorylation and stimulation of 3-O-methylglucose transport by the AMPK agonist AICAR are inhibited in a dose-dependent manner by the AMPK antagonist Compound C. AMPK α1- or AMPK α2-knockdown by RNA interference or AMPK inhibition by Compound C reduces AMPK phosphorylation and 3-O-methylglucose transport stimulation induced by cellular glucose-depletion, by potassium cyanide (KCN), or by carbonyl cyanide-p-trifluoromethoxy-phenylhydrazone (FCCP). Cell surface biotinylation studies reveal that plasma membrane GLUT1 levels are increased two- to threefold by cellular glucose depletion, AICAR or KCN treatment, and that these increases are prevented by Compound C and by AMPK α1- or α2-knockdown. These results support the hypothesis that AMPK activation in blood-brain barrier-derived endothelial cells directs the trafficking of GLUT1 intracellular pools to the plasma membrane, thereby increasing endothelial sugar transport capacity.

Measurement of glucose uptake in cultured cells.

Facilitative glucose uptake transport systems are ubiquitous in animal cells and responsible for transporting glucose across the cell surface membrane. Evaluation of glucose uptake is crucial in the study of numerous diseases and metabolic disorders, such as myocardial ischemia, diabetes mellitus, and cancer. Methods for assessing glucose uptake into mammalian cells are detailed in this unit. The work is divided into four 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); and (4) an improved method for measuring 2DG-uptake using an enzymatic, fluorometric assay, eliminating the need for radiolabeled glucose analogs.

Flavonoids have differential effects on glucose absorption in rats (Rattus norvegicus) and American robins (Turdis migratorius).

Mounting evidence suggests that small birds rely largely on non-mediated intestinal absorption of glucose through the paracellular pathway, while non-flying mammals rely on mediated absorption across the enterocyte membranes by using glucose transporters SGLT-1 and GLUT-2. Relying on non-mediated transport of glucose may decrease its absorption rate at low glucose concentrations but may release small birds from the effects of glucose transport inhibitors. We evaluated transport by using flavonoids known to inhibit glucose transport in vitro. Quercetin, isoquercetrin, and phloridzin were tested in rats (Rattus norvegicus) and robins (Turdis migratirius), and naringenin, naringenin-7-glucoside, genistein, epigallocatechin gallate (EGCG), and phloretin were used only in rats. By using a pharmacokinetic approach that involves serial blood collection and area under the curve calculations, we determined the bioavailability of 3-0-methyl D-glucose, the non-metabolized analogue of D-glucose. Six of the eight flavonoids tested in rats significantly decreased the absorption of 3-0-methyl D-glucose, while none of the flavonoids tested in robins significantly decreased the bioavailability of 3-0-methyl D-glucose. We conclude that flavonoids effectively decrease glucose absorption in rats, which rely on mediated absorption of glucose, but that flavonoids do not have an effect in robins, which rely on non-mediated absorption of glucose.

Assessment of radiolabeled D-glucose and the nonmetabolizable analog 3-O-methyl-D-glucose as tools for in vivo absorption studies.

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, because of the potential for interspecific differences in loss to catabolism, it should be used in vivo with caution and with independent verification of absorption efficiency.

The digestive adaptation of flying vertebrates: high intestinal paracellular absorption compensates for smaller guts.

Anecdotal evidence suggests that birds have smaller intestines than mammals. In the present analysis, we show that small birds and bats have significantly shorter small intestines and less small intestine nominal (smooth bore tube) surface area than similarly sized nonflying mammals. The corresponding >50% reduction in intestinal volume and hence mass of digesta carried is advantageous because the energetic costs of flight increase with load carried. But, a central dilemma is how birds and bats satisfy relatively high energy needs with less absorptive surface area. Here, we further show that an enhanced paracellular pathway for intestinal absorption of water-soluble nutrients such as glucose and amino acids may compensate for reduced small intestines in volant vertebrates. The evidence is that l-rhamnose and other similarly sized, metabolically inert, nonactively transported monosaccharides are absorbed significantly more in small birds and bats than in nonflying mammals. To broaden our comparison and test the veracity of our finding we surveyed the literature for other similar studies of paracellular absorption. The patterns found in our focal species held up when we included other species surveyed in our analysis. Significantly greater amplification of digestive surface area by villi in small birds, also uncovered by our analysis, may provide one mechanistic explanation for the observation of higher paracellular absorption relative to nonflying mammals. It appears that reduced intestinal size and relatively enhanced intestinal paracellular absorption can be added to the suite of adaptations that have evolved in actively flying vertebrates.

Prior exercise increases phosphorylation of Akt substrate of 160 kDa (AS160) in rat skeletal muscle.

The main purpose of this study was to determine whether the increased glucose transport (GT) found immediately postexercise (IPEX) or 4 h postexercise (4hPEX) is accompanied by increased phosphorylation of Akt substrate of 160 kDa (AS160, a protein regulator of GLUT4 translocation). Paired epitrochlearis muscles were dissected from rats (sedentary or IPEX, 2-h swim) and used to measure protein phosphorylation and insulin-independent GT. IPEX values exceeded sedentary values for GT and phosphorylations of AS160, AMP-activated protein kinase (pAMPK) and acetyl-CoA carboxylase (pACC) but not for AS160 abundance or phosphorylation of Akt serine (pSerAkt), Akt threonine (pThrAkt), or glycogen synthase kinase-3 (pGSK3). AS160 phosphorylation was significantly correlated with GT (R=0.801, P<0.01) and pAMPK (R=0.655, P<0.05). Muscles from other rats were studied 4hPEX along with sedentary controls. One muscle per rat was incubated without insulin, and the contralateral muscle was incubated with insulin. 4hPEX values exceeded sedentary values for insulin-stimulated GT. The elevated pAMPK and pACC found IPEX had reversed by 4hPEX. Insulin caused a significant increase in pSerAkt, pThrAkt, pGSK3, and AS160 phosphorylation with or without exercise. Exercise significantly increased AS160 phosphorylation, regardless of insulin, with unchanged AS160 abundance. Among the signaling proteins studied, insulin-stimulated GT was significantly correlated only with insulin-stimulated pThrAkt (R=0.720, P<0.0005). The results are consistent with a role for increased AS160 phosphorylation in the increased insulin-independent GT IPEX, and the exercise effects on AS160 phosphorylation and/or pThrAkt at 4hPEX are potentially relevant to the increased insulin-stimulated glucose transport at this time.

Increased bacterial permeation in long-lasting ileoanal pouches.

BACKGROUND AND AIMS: Bacterial overgrowth appears to play an important role in the pathogenesis of ileoanal pouches. Therefore, the capability of bacterial permeation and its determinants is of great interest. The aim of this study was to examine bacterial permeation in the ileoanal pouch and to correlate the results with the degree of inflammation, the epithelial resistance, the mucosal transport function, and the age of the ileoanal pouches. MATERIALS AND METHODS: Biopsies were taken from 54 patients before colectomy (n = 13; preileal pouch-anal anastomosis [IPAA]), and closure of ileostomy (n = 7; deviation), <1 year after closure of ileostomy (n = 8; intact pouch I), >1 year after closure of ileostomy (n = 16; intact pouch II), in the case of pouchitis (n = 11), and in 11 controls. Tissues were mounted in a miniaturized Ussing chamber. Escherichia coli was added to the mucosal side of the Ussing chamber, and the permeation was proven by serosal presence of E. coli. Epithelial and subepithelial resistance was determined by transmural impedance analysis. Active Na-glucose cotransport and active Cl secretion were measured. Specimens were analyzed by fluorescent in situ hybridization with oligonucleotide probes targeting the bacterial 16s ribosomal RNA. The bacteria in and on the tissue were enumerated. RESULTS: Bacterial permeation occurred in 2 of 13 pre-IPAA, 2 of 7 deviations, 0 of 8 intact pouch I, 9 of 16 intact pouch II, 5 of 11 pouchitis specimens, and 0 of 11 ileum controls. The frequency of bacterial permeation in the intact pouch II group is higher than in the intact pouch I group (P < 0.001). Epithelial resistance, mannitol fluxes, electrogenic chloride secretion, sodium-glucose cotransport of the bacterially permeated specimens versus nonpermeated of the intact pouch II group, and the pouchitis group and subepithelial resistance remained unchanged. Intramural bacteria could be detected by fluorescence in situ hybridization mainly in long-lasting pouches, but there was no correlation with bacterial permeation. CONCLUSIONS: The long-lasting ileoanal pouch is associated with increased bacterial permeability. This is not correlated with a disturbed function of the pouch mucosa but could be a precursor of pouchitis.

Seizures, ataxia, developmental delay and the general paediatrician: glucose transporter 1 deficiency syndrome.

AIM: Glucose transporter 1 deficiency syndrome (GLUT1-DS) is an important condition for the general paediatrician's differential armamentarium. We describe a case series of eight patients in order to raise awareness of this treatable neurometabolic condition. The diagnosis of GLUT1-DS is suggested by a decreased absolute cerebrospinal fluid (CSF) glucose value (<2.2 mmol/L) or lowered CSF: plasma glucose ratio (<0.4). METHODS: This is a review of eight Queensland patients with GLUT1-DS. The clinical presentation, clinical course, laboratory investigations and treatment outcomes are discussed. RESULTS: The clinical features noted in our patient cohort include combinations of ataxia, developmental delay and a severe seizure disorder that is refractory to anticonvulsant medications. Seizures are the most common clinical manifestation and may be exacerbated by phenobarbitone. The paired CSF: plasma glucose results ranged from 0.2 to 0.39 (normal <0.6) with an average of 0.33. 3-O-Methyl-D-Glucose uptake and GLUT1 Genotyping analysis have been performed on five patients thus far. Rapid and impressive seizure control was observed in 100% of our patients once the ketogenic diet was instituted, with half of the cohort being able to wean completely from anticonvulsants. CONCLUSION: Children presenting with a clinical phenotype consisting of a refractory seizure disorder, ataxia and developmental delay should prompt the consideration of Glucose transporter 1 deficiency syndrome. While the diagnostic test of lumbar puncture is an invasive manoeuvre, the diagnosis provides a viable treatment option, the ketogenic diet. GLUT1-DS displays clinical heterogeneity, but the value of early diagnosis and treatment is demonstrated by our patient cohort.