Pentose Cycle Research Articles

Metabolic Changes in Rabbit Lens Induced by Treatment with Dexamethasone

Metabolic changes in the rabbit lens have been studied by means of nuclear magnetic resonance spectroscopy. These changes have been induced by prolonged topical treatment with dexamethasone. Our results demonstrate an increase in sorbitol, sorbitol-3-phosphate, fructose-3-phosphate, glycerol-3-phosphate and glucose-6-phosphate levels and a decrease in glutathione sulphate (GSH) and myo-inositol levels, in agreement with what was observed in lenses from streptozocin-diabetic rats before lens opacity. The hyperglycaemia can only partially explain all these observed biochemical variations. The lack of increase in the intermediates of pentose cycle, such as sedoheptulose-7-phosphate, seems to support the hypothesis of an inhibition of glucose-6-phosphate dehydrogenase by dexamethasone treatment. Finally dexamethasone treatment induces a decrease in GSH. The decreasing or the loss of GSH has been suggested as a possible pathogenic mechanism in the cataract formation.

Genistein inhibits nonoxidative ribose synthesis in MIA pancreatic adenocarcinoma cells: a new mechanism of controlling tumor growth.

Genistein is a plant isoflavonoid bearing potent tumor growth-regulating characteristics. This effect of genistein has been attributed partially to its tyrosine kinase-regulating properties, resulting in cell-cycle arrest and limited angiogenesis. Genistein has been used in chemotherapy-resistant cases of advanced leukemia with promising results. Here we demonstrate that genistein primarily affects nucleic acid synthesis and glucose oxidation in tumor cells using the [1,2-(13)C2]glucose isotope as the single tracer and gas chromatography/mass spectrometry to follow various intracellular glucose metabolites. The ribose fraction of RNA demonstrated a rapid 4.6%, 16.4%, and 46.3% decrease in isotope uptake through the nonoxidative branch of the pentose cycle and a sharp 4.8%. 24.6%, and 48% decrease in 13CO2 release from glucose after 2, 20, and 200 micromol/L genistein treatment, respectively. Fatty acid synthesis and the 13C enrichment of acetyl units were not significantly affected by genistein treatment. De novo glycogen synthesis from media glucose was not detected in cultured MIA cells. It can be concluded from these studies that genistein controls tumor growth primarily through the regulation of glucose metabolism, specifically targeting glucose carbon incorporation into nucleic acid ribose through the nonoxidative steps of the pentose cycle, which represents a new paradigm for the antiproliferative action of a plant phytochemical.

Loss of [13C]Glycerol Carbon via the Pentose Cycle: IMPLICATIONS FOR GLUCONEOGENESIS MEASUREMENT BY MASS ISOTOPER DISTRIBUTION ANALYSIS

Whereas many reports substantiated the suitability of using [2-(13)C]glycerol and Mass Isotoper Distribution Analysis for gluconeogenesis, the use of [(13)C]glycerol had been shown to give lower estimates of gluconeogenesis (GNG). The reason for the underestimation has been attributed to asymmetric isotope incorporation during gluconeogenesis as well as zonation of gluconeogenic enzymes and a [(13)C]glycerol gradient across the liver. Since the cycling of glycerol carbons through the pentose cycle pathways can introduce asymmetry in glucose labeling pattern and tracer dilution, we present here a study of the role of the pentose cycle in gluconeogenesis in Fao cells. The metabolic regulation of glucose release and gluconeogenesis by insulin was also studied. Serum-starved cells were incubated for 24 h in Dulbecco's modified Eagle's media containing 1.5 mm [U-(13)C]glycerol. Mass isotopomers of whole glucose from medium or glycogen and those of the C-1-C-4 fragment were highly asymmetrical, typical of that resulting from the cycling of glucose carbon through the pentose cycle. Substantial exchange of tracer between hexose and pentose intermediates was observed. Our results offer an alternative mechanism for the asymmetrical labeling of glucose carbon from triose phosphate. The scrambling of (13)C in hexose phosphate via the pentose phosphate cycle prior to glucose release into the medium is indistinguishable from dilution of labeled glucose by glycogen using MIDA and probably accounts for the underestimation of GNG using (13)C tracer methods.

Cytochemical and physiological studies of the energetic metabolism and osmotrophy in Sagitta friderici (chaetognath)

Different glucidic labelled compounds were used for carbohydrate metabolism exploration, tissular glycogen storage localization and amino acid uptake response to osmotic variation in the adult of Sagitta friderici.After [1-14C] glucose or [6-14C] glucose (osmotrophic) assimilation, the relative activity of the different metabolic pathways (glycolysis–citric acid cycle and pentose cycle) have been estimated by means of a micro-radiorespirometric method. The kinetics of expired 14CO2 showed that adult Sagitta mainly used the glycolytic–Krebs cycle rather than the pentose cycle in glucose catabolism.After 2-deoxy-D-[1-14C] glucose (osmotrophic) assimilation, tissular glycogen storages were localized. It was shown that glycogen deposits are abundant in several tissues except the locomotor muscles, which probably represent the main glucose consumers in these very active animals. The problem of glucose transport from the storage localizations to the sites of utilization has been discussed.The L-3H-leucine uptake variations into the Sagitta body after an osmotic shock have been estimated.

Langerhans Islet Implants in Diabetic Patients. Preliminary Results

Glycolysis, the Krebs cycle, oxidative phosphorylation and the pentose cycle are blocked in diabetes mellitus (DM). This results in alterations in the production of ATP and an increase in free radicals which damage protein lipids and DNAmt. In 1981, Cerami et al. stated that an excess of glucose is the cause of the classical complications in diabetics. The transplanting of pancreatic beta cells was proposed as a method of control for diabetes mellitus. This procedure in animals and humans has given rise to encouraging results, but only in patients receiving immunosuppressive therapy for renal transplants. On an experimental basis, we have implanted pancreatic beta islets and have obtained a reduction in glucose levels. In the present paper we evaluate the clinical and laboratory response of diabetic patients who, prior treatment with a modified form of RNAt (RNAt HAM) to induce the synthesis of IFN, received parenteral implants of Langerhans islets extracted from sheep. Immediately after, patients were given parenteral injections of thiamine pyrophosphate (X‐2). Three days after the implant blood glucose levels had reduced. Clinical improvement was recorded through the disappearance of asthenia, adynamia and depression. The administration of hypoglycemic agents or insulin was reduced or suspended according to laboratory results. We conclude that the implanting of pancreatic islets is a good option for the treatment of DM, on condition that RNAt HAM is used to avoid rejection from immunological tolerance.

Blood sugar formation from dietary carbohydrate is facilitated by the pentose phosphate pathway in an insect Manduca sexta Linnaeus

Dietary carbohydrate, the principal energy source for insects, also determines the level of the blood sugar trehalose. This disaccharide, a byproduct of glycolysis, occurs at highly variable concentrations that play a key role in regulating feeding behavior and growth. Little is known of how developing insects partition the metabolism of dietary carbohydrate to meet the needs for blood trehalose, ribose sugars and NADPH, as well as energy production. This study examined the effects of varying dietary sucrose levels between 3.4 and 34 g/l in an artificial diet on growth rate, depot fat content and blood sugar formation from 13C-enriched glucose in Manduca sexta. (2- 13C)Glucose or (1,2- 13C 2)glucose were administered to larvae by injection and after 6 h blood was analyzed by nuclear magnetic resonance spectroscopy. [2- 13C]Trehalose was the principal product of [2- 13C]glucose, but trehalose was also 13C-enriched at C1 and C3, demonstrating activity of the pentose phosphate pathway. The trehalose C1/C2 13C-enrichment ratio, a measure of the substrate cycled through the pentose pathway, significantly increased with increasing dietary sugar, and reached a mean of 0.22 at the highest level. Blood trehalose concentration increased from approximately 38 mM at the lowest dietary carbohydrate level to 75 mM at the highest. Moreover, blood trehalose, growth rate and depot fat all increased in precisely the same way in relation to the level of pentose cycling. Based on the multiplet 13C-NMR signal structure of trehalose synthesized from [1,2- 13C 2]glucose by insects maintained on a high carbohydrate diet, it was established that the formation of trehalose from glucose phosphate derived directly from the administered substrate, with no involvement of the pentose pathway, was greater than that from glucose phosphate metabolized through the pentose pathway prior to trehalose synthesis. On the other hand, glucose phosphate first metabolized through the pentose pathway contributed more to pyruvate formation than did glucose phosphate formed from the labeled substrate metabolized directly to pyruvate via glycolysis; this finding based on the multiplet 13C-NMR signal structure in alanine derived from pyruvate. The results suggest that as dietary carbohydrate increases blood sugar synthesis from glucose phosphate derived directly from dietary sugar is facilitated by the pentose pathway which provides an increasing amount of substrate to pyruvate formation.

Oxythiamine and dehydroepiandrosterone induce a G 1 phase cycle arrest in Ehrlich’s tumor cells through inhibition of the pentose cycle

Transketolase (TK) reactions play a crucial role in tumor cell nucleic acid ribose synthesis utilizing glucose carbons, yet, current cancer treatments do not target this central pathway. Experimentally, a dramatic decrease in tumor cell proliferation after the administration of the TK inhibitor oxythiamine (OT) was observed in several in vitro and in vivo tumor models. Here, we demonstrate that pentose cycle (PC) inhibitors, OT and dehydroepiandrosterone (DHEA), efficiently regulate the cell cycle and tumor proliferation processes. Increasing doses of OT or DHEA were administered by daily intraperitoneal injections to Ehrlich’s ascites tumor hosting mice for 4 days. The tumor cell number and their cycle phase distribution profile were determined by DNA flow histograms. Tumors showed a dose dependent increase in their G 0-G 1 cell populations after both OT and DHEA treatment and a simultaneous decrease in cells advancing to the S and G 2-M cell cycle phases. This effect of PC inhibitors was significant, OT was more effective than DHEA, both drugs acted synergistically in combination and no signs of direct cell or host toxicity were observed. Direct inhibition of PC reactions causes a G 1 cell cycle arrest similar to that of 2-deoxyglucose treatment. However, no interference with cell energy production and cell toxicity is observed. PC inhibitors, specifically ones targeting TK, introduce a new target site for the development of future cancer therapies to inhibit glucose utilizing pathways selectively for nucleic acid production.

Blood sugar formation due to abnormally elevated gluconeogenesis: aberrant regulation in a parasitized insect, Manduca sexta Linnaeus

Alterations of carbohydrate metabolism associated with parasitism were examined in an insect, Manduca sexta L. In insect larvae maintained on a low carbohydrate diet gluconeogenesis from [3- 13C]alanine was established from the fractional 13C enrichment in trehalose, a disaccharide of glucose and the blood sugar of insects and other invertebrates. After transamination of the isotopically substituted substrate to [3- 13C]pyruvate, the latter was carboxylated to oxaloacetate ultimately leading to de novo glucose synthesis and trehalose formation. Trehalose was selectively enriched with 13C at C1 and C6 followed by C2 and C5. 13C enrichment of blood sugar in insects parasitized by Cotesia congregata (Say) was significantly greater than was observed in normal animals. The relative contributions of pyruvate carboxylation and decarboxylation to trehalose labeling were determined from the 13C distribution in glutamine, synthesized as a byproduct of the tricarboxylic acid cycle. The relative contribution of carboxylation was significantly greater in parasitized larvae than in normal insects providing additional evidence of elevated gluconeogenesis due to parasitism. Despite the increased gluconeogenesis in parasitized insects the level of blood sugar was the same in all animals. Because de novo glucose synthesis does not normally maintain blood sugar level in insects maintained under these dietary conditions the findings suggest an aberrant regulation over gluconeogenesis. The 13C labeling in trehalose was nearly symmetric in all insects but the mean C1/C6 13C ratio was higher in parasitized animals suggesting a lower activity of the pentose phosphate pathway that brings about a redistribution of 13C in trehalose following de novo glucose synthesis. Additional studies with insects maintained on a high carbohydrate diet and administered [1,2- 13C 2]glucose confirmed a decreased level of pentose cycling during parasitism consistent with a lower level of lipogenesis. It is suggested, however, that the pentose pathway may facilitate the synthesis of trehalose from dietary carbohydrate by directing hexose phosphate cycled through the pathway to the production of energy.

Quantitative determination of the main glucose metabolic fluxes in human erythrocytes by 13C- and 1H-MR spectroscopy

Information displayed by homonuclear and heteronuclear spin-coupling patterns in 1 3C- and 1H-MR spectra allowed us to identify the major lactate isotopomers produced either from [1- 1 3C]-glucose or from [2- 1 3C]-glucose by human erythrocytes. Relative concentrations of detectable isotopomers were determined by integrating the corresponding MR signals. The interpretation of these data in terms of the fractional glucose metabolised through glycolysis and pentose phosphate pathway was performed by a computer simulation of the metabolism that took into account metabolic schemes pertaining to glycolysis and to the F-type of pentose phosphate pathway. The simulation was organised in a way to anticipate the populations of the isotopomers produced from any precursor at a priori established metabolic steady state. By the simulation, isotopomer populations were determined according to different values of pentose cycle, defined as the flux of glyceraldehyde 3-phosphate originating from pentose phosphate pathway at unitary glucose uptake. The populations of the isotopomers originating from [2- 1 3C]-glucose were described by polynomials, and ratios between the polynomials were used in conjunction with 13C- and 1H-MR data to determine pentose cycle values. The knowledge of glucose uptake and of pentose cycle value allowed us to perform accurate measurement of the pentose phosphate pathway flux, of the hexokinase and phosphofructokinase fluxes as well as, indirectly, of the carbon dioxide production.

Methotrexate: pentose cycle and oxidative stress.

The effect of methotrexate (MTX) and leucovorin (LCV) on pentose cycle enzymes and the activity of enzymes involved in enzyme defence mechanisms against ROS in HeLa cells, were studied. The effect of MTX was also investigated on the cellular levels of glutathione. MTX inhibited the activity of glucose-6-phosphate and 6-phosphogluconate dehydrogenases. The activities of glutathione reductase and gamma-glutamylcysteine synthetase were also inhibited by the drug. No effect was observed on the activities of catalase, superoxide dismutase or transketolase. LCV had no effect on any of the enzymes studied. MTX decreased the cellular levels of glutathione (70 per cent), while the presence of LCV and glutamine did not interfere with the effect of MTX. The net results appear to show that the biological situation resulting from treatment with MTX leads to a reduction of effectiveness of the antioxidant enzyme defence system.

The measurement of bioreductive capacity of tumor cells using methylene blue

Purpose: Methylene blue (MB) can be used as an intracellular electron acceptor. The purpose of this study was to demonstrate the usefulness of MB for the determination of total bioreductive capacity of cell suspensions. Methods and Materials: We measured oxygen consumption by Clark electrode and pentose cycle activity by release of 14CO 2 from 1- 14C-glucose. Results: Methylene blue catalyzes the reaction of intracellular reductants NADPH, NADH, and reduced glutathione (GSH) with oxygen, causing the production of hydrogen peroxide. The reaction rate correlates with the negative charge of molecule (NADPH −4 > NADH −2 > GSH −1), suggesting that reaction with positively charged oxidized MB is the limiting step of the reaction. In a cellular system MB causes the electron flow from cellular endogenous substrates to oxygen. It is activated by the disruption of the NADP +/NADPH ratio due to several processes. These are direct oxidation of NADPH and GSH, the GSH peroxidase catalyzed reaction of GSH with H 2O 2, followed by NADPH oxidation by oxidized glutathione (GSSG). This results in increased cellular oxygen consumption and stimulation of the oxidative limb of pentose cycle (PC) in the presence of MB. The cellular effect of MB differs from other electron accepting drugs. Diamide and tert-butylhydroperoxide act as direct oxidants, while MB is an electron carrier to oxygen. Accordingly, MB shows the highest effect on PC activation and oxygen consumption. Conclusions: Our results indicate that MB may be used for the determination of the total bioreductive capacity of the cells, measured by oxygen consumption and PC activation.

Metabolic energy status of diapausing nervous system in Pieris brassicae: Comparison with the relative utilization of metabolic pathways in the whole insect body

The temporal evolution of changes in energy metabolism of nervous tissues during postembryonic development in Pieris brassicae has been studied either in nondiapausing or diapausing development. We showed that the nervous system, during the diapause of insects, was characterized by a very large reduction in the quantitative level of metabolism, but the metabolic activity of the nervous system was never completely abolished. We also observed a typical diapausing metabolic pattern in the relative utilization of the catabolic pathways of glucose. The oxidative routes were 95% depressed (from 3-day-old to 30-day-old pupae), but the pentose cycle retained a relatively high degree (C6/C1≤ 0.25) of activity as compared to the Krebs cycle, clearly showing that synthetic processes were not fully abolished during the diapause period. Thus an apparently static morphological brain condition corresponds to a relatively dynamic state of metabolic activity. This phenomenon remained rather constant during all the time the insects were undergoing diapause and during wintering conditions (short daylight, low temperature for a minimum 3 months) but nevertheless appeared to be necessary to achieve diapause breakdown. After this long wintering period, when insects were replaced into summer conditions, most of them were able to exhibit a large increase of cerebral energy metabolism in both catabolic pathways of glucose. This led to adult emergence and enabled imaginal life. If diapausing insects did not undergo wintering conditions of a sufficiently long period prior to the emergence of the adult, they could not attain threshold levels of nervous system metabolic reactivation and thus would compromise the possibilities of emergence, resulting in the death (>95%) of the most of insects after diapause. J. Exp. Zool. 281:260–269, 1998. © 1998 Wiley-Liss, Inc.

Metabolic energy status of diapausing nervous system inPieris brassicae: Comparison with the relative utilization of metabolic pathways in the whole insect body

The temporal evolution of changes in energy metabolism of nervous tissues during postembryonic development in Pieris brassicae has been studied either in nondiapausing or diapausing development. We showed that the nervous system, during the diapause of insects, was characterized by a very large reduction in the quantitative level of metabolism, but the metabolic activity of the nervous system was never completely abolished. We also observed a typical diapausing metabolic pattern in the relative utilization of the catabolic pathways of glucose. The oxidative routes were 95% depressed (from 3-day-old to 30-day-old pupae), but the pentose cycle retained a relatively high degree (C6/C1≤ 0.25) of activity as compared to the Krebs cycle, clearly showing that synthetic processes were not fully abolished during the diapause period. Thus an apparently static morphological brain condition corresponds to a relatively dynamic state of metabolic activity. This phenomenon remained rather constant during all the time the insects were undergoing diapause and during wintering conditions (short daylight, low temperature for a minimum 3 months) but nevertheless appeared to be necessary to achieve diapause breakdown. After this long wintering period, when insects were replaced into summer conditions, most of them were able to exhibit a large increase of cerebral energy metabolism in both catabolic pathways of glucose. This led to adult emergence and enabled imaginal life. If diapausing insects did not undergo wintering conditions of a sufficiently long period prior to the emergence of the adult, they could not attain threshold levels of nervous system metabolic reactivation and thus would compromise the possibilities of emergence, resulting in the death (>95%) of the most of insects after diapause. J. Exp. Zool. 281:260–269, 1998. © 1998 Wiley-Liss, Inc.

Inhibition of the oxidative and nonoxidative pentose phosphate pathways by somatostatin: a possible mechanism of antitumor action

Long-acting somatostatin analogs have recently become supplemental drugs in the treatment of neurofibroma because of their marked tumor growth inhibitory effect. Somatostatin is currently under extended evaluation in other cancers as a possible supplemental drug to the treatment protocols in use. The mode of action is not known. Somatostatin has been shown to cause glucose intolerance by inhibiting glucose-6-phosphate dehydrogenase (G6PD) in fish liver. Recent data generated in our laboratory indicate that it is this pathway and the transketolase reactions of the pentose cycle (PC) which are directly involved in the ribose synthesis process of pancreatic adenocarcinoma cells. In cell culture, somatostatin alone inhibited glucose carbon recycling through the PC by 5.7%, which was increased to 19.8% in combination with oxythiamine, a competitive inhibitor of transketolase. Oxythiamine produced strong apoptosis in in-vitro hosted tumor cells. We hypothesize that somatostatin- and oxythiamine-induced anti proliferative action is mediated by the inhibition of G6PD, transketolase, or both.

Mass isotopomer study of the nonoxidative pathways of the pentose cycle with [1,2-13C2]glucose.

We present a single-tracer method for the study of the pentose phosphate pathway (PPP) using [1,2-13C2]glucose and mass isotopomer analysis. The metabolism of [1,2-13C2]glucose by the glucose-6-phosphate dehydrogenase, transketolase (TK), and transaldolase (TA) reactions results in unique pentose and lactate isotopomers with either one or two 13C substitutions. The distribution of these isotopomers was used to estimate parameters of the PPP using the model of Katz and Rognstad (J. Katz and R. Rognstad. Biochemistry 6: 2227-2247, 1967). Mass and position isotopomers of ribose, and lactate and palmitate (products from triose phosphate) from human hepatoma cells (Hep G2) incubated with 30% enriched [1,2-13C2]glucose were determined using gas chromatography-mass spectrometry. After 24-72 h incubation, 1.9% of lactate molecules in the medium contained one 13C substitution (m1) and 10% contained two 13C substitutions (m2). A similar m1-to-m2 ratio was found in palmitate as expected. Pentose cycle (PC) activity determined from incubation with [1,2-13C2]glucose was 5.73 +/- 0.52% of the glucose flux, which was identical to the value of PC (5.55 +/- 0.73%) determined by separate incubations with [1-13C] and [6-13C]glucose, 13C was found to be distributed in four ribose isotopomers ([1-13C]-, [5-13C]-, [1,2-13C2]-, and [4,5-13C2]ribose). The observed ribose isotopomer distribution was best matched with that provided from simulation by substituting 0.032 for TK and 0.85 for TA activity relative to glucose uptake into the model of Katz and Rognstad. The use of [1,2-13C2]glucose not only permits the determination of PC but also allows estimation of relative rates through the TK and TA reactions.

Endotoxemia, pentose cycle, and the oxidant/antioxidant balance in the hepatic sinusoid.

During the innate immune response, excessive release of reactive oxygen species (ROS) from sequestered phagocytes and activated resident macrophages represents the predominant component of oxidative stress in the liver and other tissues. The consequence of oxidative stress is determined by the status and adaptive changes of antioxidant pathways. In this review, we present evidence that the synchronized response of hepatic sinusoidal endothelial cells, the primary sites of phagocyte attachment, plays an important role in defense against phagocyte-derived ROS. An essential component of the metabolic adaptation of hepatic sinusoidal cells to lipopolysaccharide (LPS)-induced oxidative stress is the stimulated expression of glucose-6-phosphate dehydrogenase (G6PD), the key enzyme of the pentose cycle (hexose monophosphate shunt, HMS). All major ROS-metabolic enzymes, i.e., glutathione peroxidase, glutathione reductase, catalase, superoxide dismutases, NADPH oxidase, and nitric oxide synthase, directly or indirectly depend on NADPH, which is produced in the HMS in these cells. The functional significance of up-regulated HMS within a particular cell type depends on the accompanying adaptive changes in ROS-metabolizing enzymes. In LPS-activated Kupffer cells, the elevated expression of glucose transporter GLUT1 and G6PD mainly serves primed production of superoxide anion, hydrogen peroxide, and nitric oxide. In sinusoidal endothelial cells, the LPS-induced response pattern of glucose- and ROS-metabolizing enzymes results in elevated ROS detoxifying capacity. The described studies also suggest the existence of an intercellular oxidant balance between pro-oxidant Kupffer cells and antioxidant endothelial cells in the hepatic micro-environment. Maintenance of the intercellular oxidant/antioxidant balance between phagocytes and endothelial cells may represent an important mechanism protecting the hepatic parenchyma against exogenous oxidative stress during the inflammatory response.

Biosynthesis of 13C-labeled branched polysaccharides by pestalotiopsis from 13C-labeled glucoses and the mechanism of formation

Biosynthesis of branched glucan by Pestalotiopsis from media containing D-(1- 13C)glucose, D-(2- 13C)glucose, D-(4- 13C)glucose, D-(6- 13C)glucose or a mixture of D-(1- 13C)glucose and D-(2- 13C)glucose was carried out to elucidate biosynthetic mechanism of branched polysaccharides. 13C NMR spectra of the labeled polysaccharides were determined and assigned. Analysis of 13C NMR spectra of glucitol acetates obtained from hydrolysates of the labeled branched polysaccharides indicated that transfer of labeling from C-1 to C-3 and C-6 carbons, from C-2 to C-1, C-3 and C-5 carbons, and from C-6 to C-1 carbon. From the results the percentages of routes via which the polysaccharide is biosynthesized are estimated. They show that the biosynthesis of the polysaccharide via the Embden-Meyerhof pathway and that from lipids and proteins are more active, and the pentose cycle is less active, than in the biosynthesis of cellulose and curdlan. As for the results, labeling at C-6 carbon in the branched polysaccharide cultured from D-(6- 13C)glucose was low, compared to that of cellulose and curdlan.

Effects of Hypoxia on Glucose Metabolic Pathways in the Mealworm Larval Fat Body In Vitro: Alterations of Hormonal Actions

Glucose metabolism of mealworm larval fat body tissue in vitro was studied using a highly sensitive micro-radiorespirometric method. Respiratory O2 partial pressure variations were used to measure catabolic rates of [1-14C]glucose or [6-14C]glucose molecules as substrates when introduced separately into the fat body incubation medium. Expired volumes of CO2 and 14CO2 radioactivity were determined when fat body was submitted to gradual hypoxia. The modifications in the specific radioactivity kinetics and in the cumulative yields of total 14CO2 derived from [1-14C]glucose and [6-14C]glucose molecules under gradual hypoxia, were measured. Catabolic corpora cardiaca hormonal extracts from Tenebrio strongly inhibited the utilization of the pentose cycle in the in vitro degradation of fat body glucose, when the respiratory gaseous mixture contained 30% O2. This effect was drastically reduced when fat body underwent hypoxia (5% O2). The anabolic insulin-like peptide (ILP) from Tenebrio strongly stimulated the utilization of the pentose cycle in the in vitro degradation of fat body glucose when the respiratory gaseous mixture contained 30% O2; this dose-dependent effect was increased when fat body underwent hypoxia (5% O2).

Effects of Deltamethrin on Glucose Catabolic Pathways in the Isolated Fat Body of Adult Male Locusta migratoria

The effects of various doses (non-lethal, LD 15, LD 25, LD 50, LD 75) of the pyrethroid insecticide deltamethrin were investigated on glucose catabolism pathways in the isolated fat body of starved 25-day-old male adult Locusta migratoria. Two methods of poisoning with deltamethrin were used. In the first case, the insecticide was injected into the locust in vivo and then the fat body was dissected either 30 min or 1hr after injection. The orientation of glucose with respect to its catabolic pathways was estimated in isolated fat body by means of radio-respirometric method using [1- 14C]glucose and [6- 14C]glucose. Deltamethrin reduced the relative participation of the pentose pathway, and this effect was maximal 1hr after insecticide treatment. Glucose degradation in the glycolysis-Krebs cycle was increased only 30 min after treatment. Deltamethrin significantly modified the orientation of glucose between the catabolic pathways. In the second case, deltamethrin was added directly into the fat body incubation medium after its sampling in in vitro conditions. Labeled glucose was added either simultaneously or 30 min after the insecticide dose. In both cases, deltamethrin diverted glucose from the pentose cycle; this effect was larger with simultaneous injections. The relative participation of the glycolysis-Krebs cycle was slightly increased, and this effect was observed only when the injections were separated by 30 min. A direct dose-dependent response was observed only on the pentose cycle. An indirect action of deltamethrin on glucose catabolism pathways in fat body might be the most probable explanation, but we cannot eliminate the possibility of a direct effect on these catabolic pathways.

An Important Role for Pentose Cycle in the Synthesis of Citrulline and Proline from Glutamine in Porcine Enterocytes

This study was designed to determine a role of pentose cycle in the provision of NADPH for the synthesis of citrulline and proline from glutamine in porcine enterocytes. Enterocytes from 4-day-old pigs were incubated at 37°C for 0 to 30 min in Krebs–Henseleit bicarbonate buffer (pH 7.4) containing 2 mMglutamine and 5 mMglucose in the presence of 0, 0.1, or 0.25 mMdehydroepiandrosterone (DHEA), a potent inhibitor of glucose-6-phosphate dehydrogenase which is the key regulatory enzyme of pentose cycle. The activity of this cycle was estimated with the use of [1-14C]glucose and [6-14C]glucose. In some experiments, the medium included 2 mMornithine and 2 mMNH4Cl (no glutamine). About 14% of glucose taken up by enterocytes was metabolized via pentose cycle. The flux from glucose into this cycle was decreased by 70 and 86%, respectively, in the presence of 0.1 and 0.25 mMDHEA compared with its absence. DHEA inhibited the synthesis of ornithine, citrulline, arginine, and proline from glutamine in a concentration-dependent manner, but had no effect on the formation of citrulline and arginine from ornithine. However, DHEA decreased the synthesis of proline from ornithine by 79%. DHEA had no effect on cellular ATP concentrations. These results provide the first line of evidence suggesting that glucose metabolism via pentose cycle plays an important role in providing NADPH for the conversion of glutamine into pyrroline-5-carboxylate in porcine enterocytes, which may explain the glucose-dependent synthesis of citrulline and proline from glutamine in these cells.