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PubMed1975

Glucose- and phlorrhizin-protected thiol groups in pig intestinal brush-border membranes.

Purified brush borders, prepared fro newborn pig intestine, were incubated in the presence of 203Hg-labelled p-chloromercuribenzenesulphonic acid and the membrane proteins later separated by polyacrylamide-gel electrophoresis. The presence of either D-glucose or phlorrhizin, during a preliminary incubation in non-radioactive p-chloromercuribenzenesulphonic acid, increased the subsequent binding of the 203Hg-labelled compound to a protein of molecular weight 31500. This increase appeared to be specific for the low-molecular-weight protein, provided that the concentration of protecting agent used corresponded to that used to produce a biological response in the intact tissue. These results are discussed in relation to the known properties of other presumptive sugar carriers isolated from different membranes.

Animals↗

PubMed1988

Brain slice glucose utilization.

The metabolism of 2-deoxyglucose has been studied in 540 micron and 1,000 micron hypothalamic brain slices. Slice 2-deoxyglucose (2DG) and 2-deoxyglucose-6-phosphate (2DG6P) levels were measured after tissue homogenization and perchloric acid extraction. By analyzing the uptake and washout kinetics with nonlinear least-squares methods, we have determined the rate constants for three-, four-, or five-parameter kinetic models and obtained a value for the in vitro lumped constant (LC). The kinetic analysis reveals a small, slowly decaying, 2DG component that is not predicted by any of the models. If this component is treated as a separate, parallel compartment, then the four- and five-parameter models are essentially equivalent. To compare our data to prior in vivo data, we combined 2DG and 2DG6P to produce Ci*, the total slice radioactivity, and analyzed the first 45 min of uptake. These data were fit best by a three-parameter model and the slowly decaying pool was not identified. Calculation of glucose utilization from total tissue radioactivity, measured by whole slice homogenization and by image analysis of autoradiograms, showed excellent correlation between the two methods. Image analysis of radioactivity in the suprachiasmatic nucleus, which is present in these slices, revealed a spontaneous diurnal variation in in vitro glucose utilization in close quantitative agreement with prior in vivo measurements. The kinetic analysis of the 1,000 micron slice was qualitatively similar to that of the 540 micron slice but revealed an increase in the LC and a large decrease in k1 as well as the expected large increase in the hexokinase rate constant, k3. Overall, in vitro glucose utilization increased by about 60%. These results are consistent with our prior studies of the 1,000 micron slice and support our interpretation that the 1,000 micron slice is an excellent in vitro model for brain ischemia without infarction.

Animals↗

PubMed1988

Utilization of glucose, alanine, lactate, and glycerol as lipogenic substrates by periuterine adipose tissue in situ in fed and starved rats.

A technique previously used to study placental transfer in pregnant rats, consisting of labeled tracer infusion through the left uterine artery, was employed to determine the utilization of lipogenic substrates by periuterine adipose tissue in the fed and 48-hr starved female virgin rat. After 20 min infusion with either D[U-14C]glucose, L-[U-14C]alanine, [U-14C]glycerol or L-[U-14C]lactate, the radioactivity appearing in periuterine adipose tissue 14C-labeled lipids from the left side was always higher than that appearing in tissue from the right side. Negligible radioactivity was detected in the tissue from either side when the infusion was done with non-metabolizable derivatives such as L-[1-14C]glucose or [1-14C]alpha-aminoisobutyric acid. Simultaneous infusion of L-[U-14C]alanine and an alanine transaminase inhibitor (aminooxyacetic acid) into the left uterine artery completely blocked the conversion of the alanine transaminase inhibitor (aminooxyacetic acid) into the left uterine artery completely blocked the conversion of the alanine into periuterine adipose tissue 14C-labeled lipids. The utilization of the infused substrate for fatty acids and glyceride-glycerol synthesis by the tissue was quantified by taking into account the infused radioactivity, the difference in the amount of 14C-labeled lipids appearing in periuterine adipose tissue on the left and the right sides, the arterial plasma concentration of the studied metabolite, and the uterine horn blood flow. In fed animals, the highest fatty acid synthesis was found with lactate, followed by glucose, alanine, and glycerol. This process was intensely decreased with all the substrates in 48-hr starved rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Adipose Tissue↗

PubMed1985

Effect of glucocorticoids on hexose transport in rat adipocytes. Evidence for decreased transporters in the plasma membrane.

Glucocorticoids are known to rapidly inhibit glucose transport when added to isolated rat adipocytes. To determine whether this inhibition of transport persists following isolation of the plasma membranes, adipocytes were incubated in the absence or presence of a maximally inhibitory concentration of dexamethasone, a synthetic glucocorticoid, and plasma membrane vesicles were prepared. D-Glucose uptake into vesicles from steroid-treated cells was inhibited by an average of 40%. The ability of dexamethasone to inhibit transport depended upon pretreatment of cells with hormone prior to membrane isolation. Furthermore, the decreased rate of transport was prevented by the simultaneous addition to the cell of actinomycin D or cycloheximide with dexamethasone, indicating a requirement for RNA and protein synthesis. The effect of dexamethasone on glucose transport was further investigated using our recently developed cytochalasin B affinity-labeling protocol to identify the transporter on sodium dodecyl sulfate-polyacrylamide gels. A peak of radioactivity having Mr = 54,000 was identified which exhibited the properties expected for the glucose transporter, in that label incorporation was prevented by D-glucose and unlabeled cytochalasin B, but not by D-sorbitol or unlabeled cytochalasins A, D, or E. Dexamethasone was found to cause a significant (average 33%) decrease in the amount of labeled transporter in the plasma membrane which was prevented by the simultaneous addition of actinomycin D with dexamethasone to the cells. A similar percentage decrease was not found in a microsomal membrane fraction nor in a total cellular membrane fraction. These results suggest that glucocorticoids may decrease glucose transport in rat adipocytes by selectively decreasing the number of transporters in the plasma membrane.

Adipose Tissue↗

PubMed1979

The effect of 2-ethyl-3-(4-hydroxybenzoyl)-benzofuran (benzarone) on the metabolism of arterial tissue and cultured arterial smooth muscle cells.

Studies on the action of 2-ethyl-3-(4-hydroxybenzoyl)-benzofuran (benzarone) on the metabolism of cultured arterial smooth muscle cells and arterial tissue had the following results: 1. On incubation of calf arterial tissue in the presence of 0.03--0.1 mmol/l benzarone (8--26 micrograms/ml medium) the metabolic transformation of [14C]-glucose to [14C]-lactate and 14CO2 and the incorporation of 14C radioactivity into the total lipids is not significantly altered as compared with control values. In cultured human arterial smooth muscle cells 0.03 mmol/l benzarone stimulates the incorporation of [14C]-acetate and [3H]-palmitate into the cellular lipids while the receptor mediated uptake of homologous low-density lipoproteins (LDL) by the cells and their release are not influenced. 2. In concentrations greater than 0.2 mmol/l benzarone the glucose utilisation of arterial tissue is enhanced, while the labelling of lipids, in particular the labelling of the triglyceride fraction, is depressed. Under the same conditions the protein biosynthesis and the incorporation of [14C]-acetate and [3H]-palmitate into the total lipids of cultured arterial smooth muscle cells are decreased.

Acetates↗

PubMed1996

Comparative analysis of glucose and glutamine metabolism in transformed mammalian cell lines, insect and primary liver cells.

Glucose and glutamine metabolism in several cultured mammalian cell lines (BHK, CHO, and hybridoma cell lines) were investigated by correlating specific utilization and formation rates with specific maximum activities of regulatory enzymes involved in glycolysis and glutaminolysis. Results were compared with data from two insect cell lines and primary liver cells. Flux distribution was measured in a representative mammalian (BHK) and an insect (Spodoptera frugiperda) cell line using radioactive substrates. A high degree of similarity in many aspects of glucose and glutamine metabolism was observed among the cultured mammalian cell lines examined. Specific glucose utilization rates were always close to specific hexokinase activities, indicating that formation of glucose-6-phosphate from glucose (catalyzed by hexokinase) is the rate limiting step of glycolysis. No activity of the key enzymes connecting glycolysis with the tricarboxylic acid cycle, such as pyruvate dehydrogenase, pyruvate carboxylase, and phosphoenolpyruvate carboxykinase, could be detected. Flux distribution in BHK cells showed glycolytic rates very similar to lactate formation rates. No glucose- or pyruvate-derived carbon entered the tricarboxylic acid cycle, indicating that glucose is mainly metabolized via glycolysis and lactate formation. About 8% of utilized glucose was metabolized via the pentose phosphate shunt, while 20 to 30% of utilized glucose followed pathways other than glycolysis, the tricarboxylic acid cycle, or the pentose phosphate shunt. About 18% of utilized glutamine was oxidized, consistent with the notion that glutamine is the major energy source for mammalian cell lines. Mammalian cells cultured in serum-free low-protein medium showed higher utilization rates, flux rates, and enzyme activities than the same cells cultured in serum-supplemented medium. Insect cells oxidized glucose and pyruvate in addition to glutamine. Furthermore, insect cells produced little or no lactate and were able to channel glycolytic intermediates into the tricarboxylic acid cycle. Metabolic profiles of the type presented here for a variety of cell lines may eventually enable one to interfere with the metabolic patterns of cells relevant to biotechnology, with the hope of improving growth rate and/or productivity.

Animals↗

PubMed1996

Partitioning of CO2 production between glucose and lactate in excised sympathetic ganglia, with implications for brain.

Chains of lumbar sympathetic ganglia from 15-day-old chicken embryos were incubated for 4 h at 36 degrees C in a bicarbonate-buffered salt solution equilibrated with 5% CO2-95% O2. Glucose (1-10 mM), lactate (1-10 mM), [U-14C]glucose, [1(-14)C]glucose, [6(-14)C]glucose, and [U-14C]lactate were added as needed. 14CO2 output was measured continuously by counting the radioactivity in gas that had passed through the incubation chamber. Lactate reduced the output of CO2 from [U(-14)C]glucose, and glucose reduced that from [U(-14)C]lactate. When using uniformly labeled substrates in the presence of 5.5 mM glucose, the output of CO2 from lactate exceeded that from glucose when the lactate concentration was > 2 mM. The combined outputs at each concentration tested were greater than those from either substrate alone. The 14CO2 output from [1(-14)C]glucose always exceeded that from [6(-14)C]glucose, indicating activity of the hexose monophosphate shunt. Lactate reduced both of these outputs, with the maximum difference between them during incubation remaining constant as the lactate concentration was increased, suggesting that lactate may not affect the shunt. Modeling revealed many details of lactate metabolism as a function of its concentration. Addition of a blood-brain barrier to the model suggested that lactate can be a significant metabolite for brain during hyperlactemia, especially at the high levels reached physiologically during exercise.

Acetates↗

PubMed1988

Effects of Electroconvulsive Therapy on Brain Glucose Metabolism: A Preliminary Study.

We measured (15)O water uptake and [(18)F]fluorodeoxyglucose (FDG) uptake in four depressed individuals before electroconvulsive therapy (ECT) and 24 h after completion of a series of six to 11 bilateral ECT treatments. Studies of radioactive uptake were done using positron emission tomography, with FDG as a measure for glucose metabolism and with oxygen-15-labeled water as a measure for cerebral blood flow. The four patients showed decreased uptake of FDG in the frontal cortex after ECT treatment. Two of the three patients in whom (15)O water was measured showed decreased uptake in the frontal cortex after ECT.

Journal Article↗

PubMed1988

Leukotriene C4 metabolism during its action on glucose and lactate balance and flow in perfused rat liver.

Rat livers were perfused in a non-recirculating mode at constant pressure via the portal vein with media containing 5 mM glucose, 2 mM lactate, and 0.2 mM pyruvate. [3H]LTC4 was infused for a period of 5 min to a final concentration of 20 nM; it increased glucose and lactate output and reduced perfusion flow. 1) Leukotriene radioactivity was recovered 10 min after the onset of [3H]LTC4 infusion to about 40% in the effluent, to 20% in the bile, and to 40% in the liver. 2) Radioactivity in the effluent increased to a maximum 4-5 min after the onset and decreased again to essentially zero 3 min after completion of [3H]LTC4 infusion. [3H]LTC4 and [3H]LTD4 were the major labeled components in the effluent accounting for 45% and 38%, respectively, of the effluent radioactivity. 3) [3H]LTC4 and [3H]LTD4 were also the major components in bile; they accounted for 50% and 30%, respectively, of the radioactivity excreted, while more polar [3H]leukotriene metabolites accounted for the remainder. 4) In the liver, [3H]LTC4 and [3H]LTD4 were the major and [3H]LTE4, N-acetyl-[3H]LTE4 as well as omega-hydroxy-N-acetyl-[3H]LTE4 and omega-carboxy-N-acetyl-[3H]LTE4 were minor components detected 5 min after completion of [3H]LTC4 infusion. It is concluded from the present findings that during a 5 min infusion period about one third each of the infused LTC4 remained unchanged, was converted to LTD4, and was further degraded to LTE4 and polar metabolites including omega-oxidation products of N-acetyl-LTE4.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed2000

Kinetic differences and similarities among 3 tracers of myocardial glucose uptake.

UNLABELLED: Two glucose tracer analogs, uniformly labeled [14C]2-deoxyglucose ([U-14C]2DG) and FDG, are widely used to assess myocardial glucose uptake. Despite the similar electron configuration of the fluorine and hydrogen atoms, uptake of the 2 tracer analogs may not be the same because of their different electronegativity. METHODS: To test this hypothesis, we determined glucose uptake in isolated rat hearts simultaneously from the accumulation of [U-14C]2DG radioactivity in the tissue, by continuous monitoring of FDG accumulation with a pair of coincidence detectors and by cumulative release of 3HOH from [2-3H]glucose. A first group of hearts was perfused at physiologic workload with Krebs-Henseleit buffer containing 10 mmol/L glucose; a second group, with the buffer containing 5 mmol/L glucose plus 0.4 mmol/L oleate and 1 mU/mL insulin. Third and fourth groups were subjected to ischemia (i.e., a 75% reduction in coronary flow) and reperfused. For the third group, the buffer contained 5 mmol/L glucose; for the fourth, 5 mmol/L glucose plus 0.4 mmol/L oleate. RESULTS: No difference in the total amount of tracer accumulation in any group was seen between the 2 tracer analogs. The ratio [+/-SD] of [U-14C]2DG to FDG ranged from 0.93+/-0.09 to 1.31+/-0.11. However, both tracer analogs paralleled glucose uptake in the absence of insulin but underestimated glucose uptake significantly in the presence of insulin. Changes in 2DG uptake with ischemia and reperfusion could be detected only with FDG. CONCLUSION: Although uptake of [U-14C]2DG equals uptake of FDG quantitatively, acute changes in 2DG uptake (and, thus, in the tracer-tracee relationship) are detectable only with the fluorine-labeled tracer.

Animals↗

PubMed1974

The accumulation of adenosine in rabbit intestinal muscle.

1 Strips of longitudinal muscle from rabbit intestine accumulated radioactivity when exposed to [(3)H]-adenosine.2 Accumulation of radioactivity was not sodium-dependent or ouabain-sensitive, but was reduced by cooling, zero glucose plus bubbling with N(2), 2,4,dinitrophenol, dipyridamole, hexobendine and lidoflazine.3 After 7 min exposure to [(3)H]-adenosine, the tissue was found to contain radioactive adenosine triphosphate (ATP), adenosine diphosphate (ADP), adenosine monophosphate (AMP) and adenosine itself in the approximate ratio 13: 6: 4: 1.4 In the presence of dipyridamole, hexobendine or lidoflazine (each 1 muM), the amounts of radioactive ATP, ADP, AMP and adenosine were reduced with the concentration of adenosine not significantly different from controls.5 It is concluded that energy-dependent uptake of adenosine does not occur in the longitudinal muscle of rabbit intestine. Adenosine enters the tissue by a passive process and rapidly becomes phosphorylated giving rise to apparently high tissue: medium ratios.6 The drugs dipyridamole, hexobendine and lidoflazine appear to reduce the accumulation of radioactivity by preventing the formation of adenosine phosphate derivatives.

Adenosine↗

PubMed2005

Biosynthetic (14)C-labelling of xanthohumol in hop cones.

Xanthohumol (Xn) has well-established chemopreventive potential in vitro. In order to carry out in vivo bioavailability and tissue distribution studies,( 14)C-labelled Xn was produced by biolabelling. Supplying hop sprouts with 5 mCi [U-(14)C]glucose led to incorporation of (14)C into Xn. Delivering the radioactive precursor at once resulted in Xn with a specific activity of 318 microCi.mmol(-1); if however the amount was supplied in aliquots over 4 days a specific activity of only 53.1 microCi.mmol(-1) Xn was obtained.

Carbon Radioisotopes↗

PubMed1987

A glycoprotein rapidly labelled with [14C]glucose present in a murine melanoma cell line.

If a murine melanoma cell line PG19 is metabolically labelled with [14C]glucose for a short time, one particular membrane component incorporates more radioactivity than the rest. This is a glycoprotein of about Mr 100,000, which is not preferentially labelled in non-malignant variants of the melanoma or in suppressed hybrid cells formed between PG19 and mouse fibroblasts.

Absorptiometry, Photon↗

PubMed1990

Standardization of serum fructosamine assays.

We have calibrated a secondary serum protein standard by use (as primary standards) of samples of albumin and polylysine glycated with [14C]glucose in vitro, the glycation of which was assessed by radioactivity measurements and by elementary analysis for C and N. Using this standard for calibration in our improved fructosamine assay, one obtains an average fructosamine value of 247 mumol/L for nondiabetic individuals (or, in terms of total serum protein, 3.2 mumol/g)--about a tenth the value we obtained when we used the fructosamine assay of Johnson et al. (Clin Chim Acta 1983;127:87-95), standardized with desoxymorpholinofructose. In contrast, results corresponded well with the value for mean glycation of serum proteins, 3 mumol/g, determined by a furosine/HPLC method. Evidently the proposed procedure, in which a standard sharing the binding characteristics of endogenous glycated proteins is used together with our modified new fructosamine assay, leads to more realistic values for the concentrations of glycated serum proteins.

Carbon↗

PubMed1977

[Physiology and biochemistry of streptomycetes. XI. Different incorporation of D-glucose-u-14C into the paromomycin isomers and the precursors of paromomycin I].

During application of D-glucose-u-14C paromomycine II is higher labelled and shows a different dependence on the application time than paromomycine I, which is isomer at the paromose part. For the two paromose isomeres different rates of synthesis are supposed that change nonproportionally to each other. The distribution of radioactivity in paromomycine I shows that there is no fragmentation of the glucose chain during the biosynthesis of glucosamine, ribose, and paromose I. As to the 2-deoxystreptamine the result has not been ascertained.

Glucose↗

PubMed1999

Diffusive transfer of water and glucose across the chorionic plate of the isolated human term placenta.

This study investigated systematically the diffusive transfer of water and glucose across the chorionic plate of the human placenta. Isolated sections of human term placentae were perfused at the fetal side (open loop) with modified Ringer's solution (n=31). An artificial amniotic compartment was created on top of the chorionic plate. 3H- and 14C-labelled tracer pairs were added (donor side) to the fetal perfusion fluid or to the 'amniotic' fluid. Transfer fractions (TF, ratio of acceptor side to donor side radioactivity) were calculated as percentages. TF of water and L-glucose from perfusion fluid into the 'amniotic' fluid were 3.9+/-0.5 per cent (mean+/-SEM) and 1.2+/-0.3 per cent after 60 min and significantly different (n=6). In each sample of the following experiments the transfer fraction of the D-hexose was larger than that of the L-isomer. At 60 min, the TF were 1.6+/-0.2 and 1.1+/-0.2 per cent (D-glucose/L-glucose; fetal to amniotic compartment, n=8), from amniotic compartment to fetal perfusate 0.6+/-0.1 and 0.4+/-0.1 per cent (D-glucose/L-glucose, n=11), and 0.8+/-0.1 and 0.6+/-0.1 per cent (3-O-methyl-D-glucose/L-glucose, n=6). The difference between the latter TF lost its significance after cytochalasin B (0.1-0.2 mmol/l) had been added to the amniotic compartment. It is concluded that a limited diffusive pathway across the chorionic plate of the human placenta exists and that the transfer of D-glucose depends in part on facilitated diffusion.

3-O-Methylglucose↗

PubMedDate not supplied

Relationship between glucose transporter, hexokinase and FDG-PET in esophageal cancer.

BACKGROUND/AIMS: 18F-fluorodeoxyglucose positron emission tomography (FDG-PET) has been established as a powerful diagnosing modality in clinical oncology. FDG accumulation has been demonstrated to correlate with hexokinase activity. However, recent reports suggest that glucose transporters participate in FDG accumulation. The aim of this study is to evaluate glucose transporter and hexokinase expression and clarify the relationship between them and FDG accumulation. METHODOLOGY: FDG-PET was performed in 72 preoperative patients with esophageal cancer. The ratios of tumor radioactivity to plasma radioactivity (Ci/Cp values) were obtained 60 minutes after administration. We studied the expressions of glucose transporter 1 (Glut1) and type-II hexokinase (HK-II) by immunohistochemical analysis of the resected specimen. The percentages of cells expressing Glut1 and HK-II were scored on a 5-point scale (1=0-20%, 2=20-40%, 3=40-60%, 4=60-80%, 5=80-100%). Then the 3 scores obtained from 3 counting trials were averaged to give the Glut-index and HK-index. RESULTS: All esophageal cancers showed marked FDG accumulation. All 72 cancers expressed Glut1 and 71 of 72 cancers expressed HK-II. The Glut-index had a weak correlation with the Ci/Cp value (not significant). The HK-index had a close positive correlation with the Ci/Cp value (p<0.005). CONCLUSIONS: FDG accumulation correlates more with type-II hexokinase expression than with glucose transporter 1 expression.

Adult↗

PubMed1978

Glucosyl transferase activity of bovine galactosyl transferase.

Bovine galactosyl transferase was found to utilize UDPglucose as a substrate and elicit disaccharide biosynthesis with glucose and N-acetylglucosamine as acceptors. The relative rate of glucosyl transferase with N-acetylglucosamine as acceptor was 0.3%, the rate for N-acetyllactosamine biosynthesis. This activity was also evidenced indirectly from NMR water proton relaxation experiments, and from Mn(II) ESR experiments. In direct experiments with radioactive UDPglucose, paper chromatography showed a product which migrated with cellobiose when glucose was the acceptor and a new, glucose-containing product which resulted when GlcNAc was the acceptor. Despite this marginally expanded specificity of the donor site, spin-label experiments with a covalently bound UDPgalactose analog reaffirmed the restrictive nature of the donor site against this non-glycosyl-like analog.

Animals↗