Role of glucose in the regulation of glutamine metabolism in health and in type 1 insulin-dependent diabetes

Régis G. Hankard1,2, Morey W. Haymond1,3, and Dominique Darmaun1,4

1 Nemours Children's Clinic, Jacksonville, Florida 32207; 2 Centre d'Investigation Clinique, Hôpital Robert-Debré, Paris, France; 3 US Department of Agriculture Children's Nutrition Research Center, Baylor College of Medicine, Houston, Texas 77030; and 4 Unité 539, Institut National de La Santé et de la Recherche Médicale, Centre de Recherche en Nutrition Humaine, Hôpital Hotel-Dieu, 44093 Nantes, France


    ABSTRACT
TOP
ABSTRACT
INTRODUCTION
SUBJECTS AND METHODS
RESULTS
DISCUSSION
REFERENCES

To determine the effect of glucose availability on glutamine metabolism, glutamine kinetics were assessed under conditions of hyperglycemia resulting from 1) intravenous infusion of 7.5% dextrose in healthy adults and 2) insulin deficiency in young adults with insulin-dependent diabetes mellitus (IDDM). Eight healthy adults and five young adults with IDDM were studied in the postabsorptive state by use of a primed continuous infusion of D-[U-14C]glucose, L-[5,5,5-2H3]leucine, and L-[3,4-13C]glutamine. Whether resulting from insulin deficiency or dextrose infusion, the rise in plasma glucose was associated with increased glucose turnover (23.5 ± 0.7 vs. 12.9 ± 0.3 µmol · kg-1 · min-1, P < 0.01 and 20.9 ± 2.5 vs. 12.8 ± 0.4 µmol · kg-1 · min-1, P = 0.03, in health and IDDM, respectively). In both cases, high blood glucose failed to alter glutamine appearance rate (Ra) into plasma [298 ± 9 vs. 312 ± 14 µmol · kg-1 · h-1, not significant (NS) and 309 ± 23 vs 296 ± 26 µmol · kg-1 · h-1, NS, in health and IDDM, respectively] and the estimated fraction of glutamine Ra arising from de novo synthesis (210 ± 7 vs. 217 ± 10 µmol · kg-1 · h-1, NS and 210 ± 16 vs. 207 ± 21 µmol · kg-1 · h-1, NS, in health and IDDM, respectively). When compared with the euglycemic day, the apparent contribution of glucose to glutamine carbon skeleton increased when high plasma glucose resulted from intravenous dextrose infusion in healthy volunteers (10 ± 0.8 vs. 4.8 ± 0.3%, P < 0.01) but failed to do so when hyperglycemia resulted from insulin deficiency in IDDM. We conclude that 1) the contribution of glucose to the estimated rate of glutamine de novo synthesis does not increase when elevation of plasma glucose results from insulin deficiency, and 2) the transfer of carbon from glucose to glutamine may depend on insulin availability.

leucine; isotope labeling; energy metabolism; substrate cycling; nutrition


    INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
SUBJECTS AND METHODS
RESULTS
DISCUSSION
REFERENCES

GLUTAMINE, THE MOST ABUNDANT free amino acid in the body (1), is at the crossroads of protein and carbohydrate metabolisms. Glutamine indeed serves as a major carbon donor for gluconeogenesis in humans (12, 17); conversely, glucose serves as precursor for glutamine's carbon skeleton (18). Thus, as is the case for alanine, a glucose-glutamine cycle might operate in humans (18). Yet it remains to be determined 1) whether glucose availability affects estimates of glutamine synthesis, and if so, 2) whether this effect is mediated via increased insulin secretion.

For this purpose, we used tracer dilution methodology to investigate the effect of elevations of plasma glucose on glutamine kinetics in 1) healthy volunteers and 2) patients with insulin-dependent diabetes mellitus (IDDM). In healthy subjects, high plasma glucose was achieved through intravenous infusion of 7.5% dextrose at a rate known to cause mild hyperglycemia within the physiological range. In patients with IDDM maintained on a continuous intravenous insulin drip, moderate hyperglycemia was obtained via a reduction in insulin infusion rate.


    SUBJECTS AND METHODS
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ABSTRACT
INTRODUCTION
SUBJECTS AND METHODS
RESULTS
DISCUSSION
REFERENCES

Materials

D-[U-14C]glucose was purchased from New England Nuclear (Boston, MA). L-[5,5,5-2H3]leucine (98% 2H3) and L-[3,4-13C2] glutamine (98% 13C2) were obtained from Cambridge Isotopes Laboratories (Andover, MA), as were the internal standards utilized in analytical procedures (d7-ketoisocaproic acid (d7-KIC), [3H]glucose, [2H3]glutamine). Solutions of tracers were prepared in sterile 0.45 g/dl NaCl, passed through 0.22-µm filters, and tested for sterility (plate culture) and absence of pyrogen (limulus lysate assay). Infusates were prepared <24 h before infusion and kept at 4°C until used.

Subjects

The present study was reviewed and approved by the Nemours Children's Clinic Research Committee and the Institutional Review and Radiation Safety Committees of the Baptist Medical Center in Jacksonville, FL.

Eight healthy adults [weight, 64 ± 3 kg, age, 28 ± 3 yr, body mass index (BMI), 22 ± 1 kg/m2, means ± SE] and five patients with IDDM (weight, 60 ± 4 kg, age, 19 ± 1 yr, BMI, 22 ± 1 kg/m2) gave their written consent after the aims, study design, and potential risks of the study were explained.

Protocol Design

Glucose, glutamine, and leucine kinetics were studied using two 4-h tracer infusion studies performed on two consecutive days in healthy volunteers (protocol 1) and in IDDM patients (protocol 2). In both protocols, volunteers were studied under conditions of mild hyperglycemia on one day, whereas normal postabsorptive plasma glucose levels were maintained on the other day.

Protocol 1. Volunteers received in random order an intravenous infusion of 7.5% dextrose (19.6 ± 0.6 µmol · kg-1 · min-1) on one day or 0.9% saline providing the same fluid load (3.2 ml · kg-1 · h-1) on the other day. On each study day, subjects ate dinner at 2000 and remained fasting until 1300 the next day. The tracer infusion procedure took place from 0900 to 1300.

Protocol 2. The night before the first study day, subjects ate dinner at 2000 with their normal subcutaneous injection of regular insulin and then remained fasted until 1200 the next day. Their blood glucose was kept between 200 and 250 mg/dl (10-14 mmol/l) with a continuous infusion of a solution of regular Humulin (0.125 UI/ml), adjusted on the basis of determinations of blood glucose measured every 30 min the first 5 h, every hour until 0800, and then every 30 min during tracer infusion between 0800 and 1200. On the second night, algorithms for the choice of insulin infusion rates were altered to maintain plasma glucose between 90 and 120 mg/dl (5.0-6.7 mmol/l) overnight and throughout the next morning until 1200. Tracer infusion protocol and plasma glucose monitoring were similar on both study days.

Tracer Infusion Procedure.

The first study day, two short catheters were placed, one in a forearm vein for isotope infusion and the other in a contralateral hand vein for arterialized venous blood sampling. Before the start of tracer infusion, two blood and air samples were obtained for determination of baseline isotopic enrichment and specific activity (SA) in plasma and expired air CO2. Expired air was collected in Douglas bags by having the subjects breathe through a one-way valve for 2-min time periods. For 13CO2 enrichment determination, triplicate 15-ml aliquots were transferred from the Douglas rubber bag into Vacutainer evacuated glass tubes with a 50-ml syringe. For 14CO2 SA determination, expired air was bubbled through hyamine and ethanolamine CO2 trapping solutions (24).

At 0900 (protocol 1) and 0800 (protocol 2), a primed continuous intravenous infusion of D-[U-14C]glucose (45 µCi/h), L-[5,5,5-d3]leucine (3.2 ± 0.3 µmol · kg-1 · h-1), and L-[3,4-13C2]glutamine (5.6 ± 0.3 µmol · kg-1 · h-1) was started. The priming dose was equal to a 1-h infusion rate. Arterialized venous blood samples were obtained at 20-min intervals the last 2 h of the infusion (i.e., 7 samples) for glucose and glutamine 14C SA and for [2H3]leucine and [13C2]glutamine enrichments in plasma. Additional blood samples were drawn at baseline and at the end of the tracer infusion for glucose and insulin concentrations in plasma. Over the same period of time, expired air was collected at 30-min intervals for 13CO2 enrichment and CO2 SA, and CO2 production rate (VCO2) was measured twice over a 30-min period using an indirect calorimeter (SensorMedics 2900, Yorba Linda, CA).

Analytical Procedures

Glucose assay. Plasma glucose concentration was determined by means of automated glucose oxidase reaction (Beckman II glucose analyzer).

For determination of plasma [2H2]glucose enrichment, 50 µl of plasma were deproteinized using 300 µl ice-cold acetone, and the deproteinized sample was derivatized with acetic anhydride to form the glucose pentacetate derivative. Glucose pentacetate was analyzed for mass-to-charge (m/z) fragments 98 and 100 using gas chromatography-mass spectometry (GC-MS) (24).

For determination of plasma glucose 14C SA (Glc SA, dpm/µmol), a known aliquot of a [3H]glucose internal standard was first added to plasma. The plasma was deproteinized with barium hydroxide and zinc sulfate, and the sample was submitted to cation and anion exchange chromatography. The neutral eluate was counted for 3H and 14C radioactivity with a liquid scintillation counter (LS9800 series, Beckman Instruments, Palo Alto, CA) with correction for quenching and 14C contribution into the 3H spectrum. Plasma glucose SA was calculated as the ratio of glucose dpm to plasma glucose concentration, with a correction for glucose recovery, by use of the [3H]glucose as an internal standard.

Plasma alpha -KIC assay. alpha -KIC was isolated from plasma by ion exchange chromatography and derivatized to its t-butyldimethylsilyl-oxime derivative. Plasma alpha -KIC enrichment was determined by GC-MS monitoring ions with m/z 316 and 319 (16). Plasma alpha -KIC concentration was determined by the reverse dilution method with d7-KIC as an internal standard (16).

Glutamine assay. Plasma [13C]glutamine enrichment: glutamine was first extracted from plasma by ion exchange chromatography and then derivatized to N-acetyl-n-propyl (NAP) -glutamate, as described previously (4). Injections were made into a 0.25 × 25 mm HP1 capillary column by use of an isothermal program and the split mode. Ions at m/z ratios of 186 and 188, corresponding to natural glutamine and [13C2]glutamine, respectively, were selectively monitored with GC-MS (HP5971, Hewlett-Packard, Palo Alto, CA). Plasma glutamine concentration was determined by the reverse dilution method with [2H3]glutamine as internal standard and GC-MS (4).

Plasma glutamine 14C SA: glutamine was extracted from plasma by ion exchange chromatography, and the glutamine-containing eluate was specifically converted to glutamate using glutaminase (EC 3.5.1.2, Sigma, St. Louis, MO) to remove labeled compounds other than glutamine. The glutamate was extracted by ion exchange chromatography and then counted for disintegrations per minute (dpm) with a scintillation counter with quenching correction.

Breath CO2. CO2 SA (dpm/mmol) was determined by counting the dpm in hyamine and ethanolamine CO2 trapping solutions (24). Breath 13CO2 was measured by gas chromatography-isotope ratio mass spectrometry (GC-IRMS; VG-Isochrom-III, VG Isogas, Ipswich, UK).

Insulin assay. Plasma insulin concentration was determined by radioimmunoassay (Endocrine Science, Calabasas Hills, CA).

Calculations

Stable isotopes. Rates of appearance (Ra) of glutamine (Ra,Gln), leucine (Ra,Leu), and glucose (Ra,Glc) in plasma were calculated, in micromoles per kilogram per minute, as
R<SUB>a</SUB><IT>=</IT>i<IT>×</IT>[(E<SUB>i</SUB><IT>/</IT>E<SUB>p</SUB>)<IT>−1</IT>]<IT>,</IT>
where i is the tracer infusion rate, Ei is the tracer enrichment in the intravenous infusate (mol % excess), and Ep is the tracer enrichment in the relevant plasma substrate at steady state (mol % excess) (3, 4, 5, 15).

The apparent rate of glutamine release from whole body protein breakdown (BGln; µmol · kg-1 · h-1) was roughly estimated from Ra,leu, an index of whole body protein degradation, as
B<SUB>Gln</SUB><IT>=</IT>R<SUB>a,Leu</SUB><IT> · 0.07 · 131/</IT>(<IT>146 · 0.08</IT>)
where 0.07 and 0.08 are grams of glutamine and leucine contents per gram of protein, respectively (10), and 146 and 131 are, respectively, glutamine and leucine molecular weights.

The fraction of glutamine Ra arising from de novo synthesis (DGln; µmol · kg-1 · h-1) was estimated as
D<SUB>Gln</SUB><IT>=</IT>R<SUB>a,Gln</SUB><IT>−</IT>B<SUB>Gln</SUB>
Endogenous glucose Ra in plasma (Endo Ra,Glc) was calculated as
Endo R<SUB>a,Glc</SUB><IT>=</IT>R<SUB>a,Glc</SUB><IT>−</IT>Exo R<SUB>a,Glc</SUB>
where Exo Ra,Glc is the intravenous unlabeled glucose infusion rate (µmol · kg-1 · min-1).

Radioactive isotopes. Glucose oxidation rate (OxGlc, µmol · kg-1 · min-1) was estimated from the 14CO2 excretion rate in expired air (F14CO2, µmol · kg-1 · min-1) by two methods (hyamine and ethanolamine) and averaged.

F14CO2 was calculated from ethanolamine CO2 trapping solution as
F<SUP><IT>14</IT></SUP>CO<SUB><IT>2</IT></SUB><IT>=</IT>CO<SUB><IT>2</IT></SUB>A(E)<IT> · 250/</IT>(W<IT> · 2</IT>)
where CO2 A (E) is CO2 activity in a 1-ml ethanolamine aliquot (dpm/ml), 250 corrects 1 ml to 250 ml ethanolamine total volume of trapping solution, 2 is the 2-min collecting period, and W is the weight (kg).

F14CO2 was calculated from hyamine CO2 trapping solution as
F<SUP><IT>14</IT></SUP>CO<SUB><IT>2</IT></SUB><IT>=</IT>CO<SUB><IT>2</IT></SUB>SA(H)<IT> · </IT>VCO<SUB><IT>2</IT></SUB><IT>/</IT>(W<IT> · 22.4</IT>)<IT>,</IT>
where CO2 SA (H) is CO2 SA in 1 mmol hyamine (dpm/mmol).

OxGlc was then calculated from mean F14CO2 as
Ox<SUB>Glc</SUB><IT>=</IT>F<SUP><IT>14</IT></SUP>CO<SUB><IT>2</IT></SUB><IT>/</IT>(Glc SA<IT> · </IT>R<IT> · 10<SUP>−2</SUP></IT>)
where R is labeled carbon fractional recovery in expired CO2. R was set at 70% in basal conditions and at 82% during glucose intravenous infusion in protocol 1 (13) and at 70% on both days in protocol 2 (3).

The fraction of glutamine arising from glucose was estimated from the glutamine-to-glucose SA ratio.

Indirect calorimetry. VCO2 and VO2 were calculated during the two 20- to 30-min sets from CO2 and O2 concentrations in inspired and expired air and dilution airflow using Haldane's transformation (23).

Statistical Analysis

Data are expressed as means ± SE. Comparisons within groups were performed using paired t-tests. Significance was established at P < 0.05.


    RESULTS
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ABSTRACT
INTRODUCTION
SUBJECTS AND METHODS
RESULTS
DISCUSSION
REFERENCES

Glucose Metabolism

In healthy volunteers, plasma glucose concentration was higher (~34%) during dextrose than during saline infusion (6.3 ± 0.2 vs. 4.7 ± 0.6 mmol/l, P < 0.01). During saline infusion, plasma insulin concentration decreased from 11 ± 1 to 7 ± 1 mU/l (P < 0.01) over the course of the infusion, whereas insulin increased from 12 ± 2 to 18 ± 3 mU/l (P < 0.01) during dextrose infusion.

In IDDM patients, plasma glucose concentration was significantly higher on the first study day (13.5 ± 3.8 vs. 7.2 ± 1.3 mmol/l, P < 0.01) resulting from the lower insulin infusion rate (0.6 ± 0.7 vs. 1.6 ± 0.6 UI/h, P < 0.01; Fig. 1). On both study days, plasma glucose concentration remained stable throughout the tracer infusion period (Fig. 1).


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Fig. 1.   Plasma glucose concentration and insulin infusion rate in the insulin-dependent diabetes mellitus (IDDM) experiment. Time course (clock time, hh:mm) of plasma glucose concentration (A) and insulin infusion rate (B) the day plasma glucose was set high (H) and normal (N). Results are means ± SE.

Glucose SA and enrichments were at isotopic steady state on both study days (Table 1). In healthy volunteers, intravenous dextrose infusion resulted in increased glucose turnover (23.5 ± 0.7 vs. 12.9 ± 0.3 µmol · kg-1 · h-1, P < 0.01) and oxidation rate (6.8 ± 0.2 vs. 4.8 ± 0.3 µmol · kg-1 · min-1, P < 0.01; Fig. 2), and decreased endogenous glucose production rate (3.9 ± 0.7 vs. 12.9 ± 0.3 µmol · kg-1 · h-1, P < 0.01). Similarly, in IDDM patients, glucose turnover rate was higher when blood glucose level was maintained high, compared with the euglycemic day (20.9 ± 2.5 vs. 12.8 ± 0.4 µmol · kg-1 · min-1, P = 0.03). However, elevation of blood glucose resulting from insulin deficiency was not associated with an increase in glucose oxidation rate (3.7 ± 0.4 vs. 3.9 ± 0.5 µmol · kg-1 · min-1, NS; Fig. 2).

                              
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Table 1.   Plasma substrates specific activity and enrichments in the 2 experiments



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Fig. 2.   Glucose metabolism under conditions of high and normal plasma glucose concentrations in healthy (A) and IDDM (B) volunteers. Results are means ± SE; solid bars, hyperglycemia (6.3 mmol/l in health and 13.5 mmmol/l in IDDM); open bars, normal plasma glucose conditions (4.7 mmol/l in health and 7.2 mmol/l in IDDM). Glucose Ra, glucose turnover; Glucose Ox, glucose oxidation rate; P, significance of observed differences using paired t-test; NS, not significant.

The minimum fraction of plasma glutamine carbon arising from glucose (estimated from the ratio of glutamine SA to glucose SA) doubled between the saline and the dextrose infusion days in healthy subjects: 10.4 ± 0.8 vs. 4.8 ± 0.3% (P < 0.01).

In contrast, the apparent glucose-to-glutamine carbon transfer did not increase when high blood glucose resulted from a lower insulin infusion rate (4.4 ± 0.5 vs. 4.6 ± 0.7%, NS) in IDDM subjects.

Leucine Metabolism

On both days, plasma alpha -KIC enrichments were at isotopic steady state over the last 2 h of isotope infusion (Table 1). In healthy volunteers, leucine turnover rate, an index of protein degradation, and plasma alpha -KIC concentration were lower during intravenous dextrose compared with the saline infusion (112 ± 5 vs. 120 ± 7 µmol · kg-1 · h-1, P = 0.03, and 18 ± 1 vs. 24 ± 2 µmol/l, P = 0.006, respectively). In contrast, in IDDM patients, high blood glucose resulting from a reduction in the insulin infusion rate was associated with a higher leucine turnover rate (126 ± 11 vs. 112 vs. 7 µmol · kg-1 · h-1, P = 0.03) and plasma concentrations of alpha -KIC, the intracellular metabolite of leucine (48 ± 2 vs. 43 ± 2 µmol/l, P = 0.03).

Glutamine Metabolism

Regardless of the group studied (healthy volunteers or IDDM patients), elevation of plasma glucose failed to alter plasma glutamine concentration, glutamine Ra into plasma, and the estimated fraction of glutamine Ra arising from de novo synthesis (Table 2). A decrease in glutamine arising from whole body protein breakdown resulting from lower rate of protein degradation was observed in healthy volunteers receiving 7.5% dextrose (88 ± 4 vs. 94 ± 6 µmol · kg-1 · h-1, P = 0.03) and in normoglycemic IDDM patients (88 ± 6 vs. 99 ± 9 µmol · kg-1 · h-1, P = 0.03).

                              
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Table 2.   Effect of "high blood glucose" on glutamine metabolism in H and D subjects


    DISCUSSION
TOP
ABSTRACT
INTRODUCTION
SUBJECTS AND METHODS
RESULTS
DISCUSSION
REFERENCES

We determined the effect of elevation of plasma glucose on glutamine metabolism in two different "models": hyperglycemia resulting from intravenous dextrose infusion in healthy adults on one hand and hyperglycemia resulting from insulinopenia in type 1 diabetic subjects on the other hand. We observed that 1) high plasma glucose level, whether resulting from either insulin deficiency or exogenous glucose infusion, failed to increase the estimated rate of glutamine de novo synthesis, and 2) the apparent contribution of glucose to glutamine carbon skeleton increased when exogenous glucose was infused but failed to do so when high plasma glucose resulted from insulinopenia.

In the present study, as in previous ones (8), leucine turnover rate, an index of whole body proteolysis, increased by 12% when high blood glucose resulted from insulin deficiency. Conversely, the enhanced insulin secretion induced by glucose most likely accounts for the decrease in protein degradation observed during dextrose infusion in healthy subjects. These data highlight the anticatabolic effect of insulin on protein metabolism (2).

The methodology used in the present study yields estimates of the fraction of overall glutamine appearance into plasma that arises from de novo synthesis. These estimates are, however, based on the assumptions that 1) in the postabosptive state, leucine appearance rate reflects whole body protein breakdown, and 2) the release of amino acids from proteolysis is proportional to their relative abundance as bound residues in body protein. The glutamine content of muscle protein was recently reevaluated and found to be lower than the value used in the present study (14). This reevaluation emphasizes the fact that estimates of DGln are imperfect and do not measure the true rates of glutamine synthesis. Although lowering the glutamine content of body protein in calculations would result in higher estimates of DGln, it would not alter the conclusions reached in the current study.

Whether hyperglycemia resulted from insulin deficiency or 7.5% dextrose intravenous infusion, it failed to stimulate the estimated fraction of glutamine Ra arising from de novo synthesis. These results support those obtained in a previous study using [15N]glutamine as a tracer (8) in IDDM. Lack of carbon and nitrogen availability cannot account for the lack of stimulation of DGln observed in the current study. Indeed, abundant carbon was available because of hyperglycemia. Nitrogen availability may have been a limiting factor in healthy subjects, in whom the release of precursor amino acids from proteolysis was depressed by insulin. However, in IDDM subjects, nitrogen was not limiting, because precursor amino acid availability was enhanced as a result of accelerated protein breakdown on the hyperglycemic day.

Taken together with findings from earlier studies, the results of the current study are consistent with the view that glutamine appearance into plasma and the estimated release of glutamine from de novo synthesis are more responsive to increased demand than they are to increased precursor availability. Glutamine is indeed released mainly by skeletal muscle and is utilized in tissues with a high cell turnover rate such as gut and bone marrow. Estimated rates of glutamine synthesis clearly increased during hypercortisolemia (6), a condition that enhances glutamine utilization in the gut (22), as well as in children suffering from sickle cell disease (21), a condition associated with increased cell replication in the bone marrow. In contrast, glutamine Ra and estimated glutamine synthesis were found to be depressed in patients with short bowel syndrome (7, 11), a condition associated with a reduced intestinal cell mass.

The current findings also suggest that glutamine synthesis may not be insulin dependent in vivo. They are consistent with in vitro studies showing that glutamine synthetase is not dependent on the presence of insulin (9). Further studies using hyperinsulinemic euglycemic clamp and/or somatostatin infusion would be warranted to further delineate the role of substrate (i.e., glucose) availability on glutamine synthesis in vivo. Interestingly, the synthetic rate of alanine, the other major amino acid precursor for gluconeogenesis, failed to increase in IDDM patients under conditions of insulinopenia (20), whereas hyperglycemia was able to stimulate alanine synthesis in the presence of insulin (19). Taken together, these data suggest that, although both glutamine and alanine are nonessential gluconeogenic amino acids, alanine metabolism is dependent on insulin, whereas glutamine synthesis is not.

Even though glucose infusion failed to alter the overall rate of glutamine de novo synthesis, glucose infusion enhanced the apparent contribution of glucose carbon to glutamine synthesis in healthy subjects. Carbons arising from glucose oxidation can appear into glutamine because they enter the tricarboxylic acid cycle and end up in alpha -ketoglutarate, which is a precursor of glutamate and glutamine. A rise in glucose oxidation may therefore enhance the incorporation of glucose-derived carbons into the glutamine molecule. This should, however, be interpreted with caution, because glutamine labeling mostly reflects 14C loading of metabolic intermediates rather than a true glucose-to-glutamine conversion. This increased carbon availability may nevertheless help spare other precursors for glutamine synthesis. In fact, we observed an apparent increase in the contribution of glucose-to-glutamine's carbon skeleton when high blood glucose resulted from 7.5% dextrose intravenous infusion. Interestingly, the rise in the apparent carbon transfer from glucose to glutamine was not observed in IDDM when high blood glucose resulted from insulin deficiency. In the latter situation, high blood glucose failed to elicit an increase in glucose oxidation. Insulin might therefore play a key, albeit indirect, role in regulating the carbon transfer from glucose to glutamine through its effect on glucose utilization and/or glucose oxidation to CO2.


    ACKNOWLEDGEMENTS

We acknowledge the superb technical assistance of Brenda K. Sager, W. Reed Parsons, Astride Altomare, and Ed Jones. We are grateful to Pennye Arehart for coordinating the study, and to Susan Welch, Annie Rini, Bernice Rutledge, and the nursing staff at Wolfson Children's Hospital for their excellent care of the patients.


    FOOTNOTES

This work was supported in part by grants from the Juvenile Diabetes Foundation International (Grant 193117 to D. Darmaun), the Nemours Foundation, Jacksonville, FL, and Grant DK-51477 from the National Institutes of Health. Dr. Hankard was supported by a fellowship grant from the Nemours Foundation.

Address for reprint requests and other correspondence: Dominique Darmaun, Centre de Recherche en Nutrition Humaine, Hôpital Hotel-Dieu, Place Alexis Ricordeau, 3ème étage aile nord, 44093 Nantes cedex 1, France (E-mail: ddarmaun{at}nantes.inserm.fr).

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

Received 2 November 1999; accepted in final form 11 April 2000.


    REFERENCES
TOP
ABSTRACT
INTRODUCTION
SUBJECTS AND METHODS
RESULTS
DISCUSSION
REFERENCES

1.   Bergstrom, J, Furst P, Noree LO, and Vinnars E. Intracellular free amino acid concentration in human muscle tissue. J Appl Physiol 36: 693-697, 1974[Free Full Text].

2.   Castellino, P, Luzi L, Simonson DC, Haymond M, and DeFronzo RA. Effect of insulin and plasma amino acid concentrations on leucine metabolism in man. Role of substrate availability on estimates of whole body protein synthesis. J Clin Invest 80: 1784-1793, 1987[ISI][Medline].

3.   Darmaun, D, Cirillo D, Koziet J, Chauvet D, Young VR, and Robert JJ. Whole body glucose kinetics in type I diabetes studied with [6,6-2H] and [U-13C]-glucose and the artificial beta -cell. Metabolism 37: 491-498, 1988[ISI][Medline].

4.   Darmaun, D, Manary MJ, and Matthews DE. A method for measuring both glutamine and glutamate levels and stable isotopic enrichments. Anal Biochem 147: 92-102, 1985[ISI][Medline].

5.   Darmaun, D, Matthews DE, and Bier DM. Glutamine and glutamate kinetics in humans. Am J Physiol Endocrinol Metab 251: E117-E126, 1986[Abstract/Free Full Text].

6.   Darmaun, D, Matthews DE, and Bier DM. Physiological hypercortisolemia increases proteolysis, glutamine, and alanine production. Am J Physiol Endocrinol Metab 255: E366-E373, 1988[Abstract/Free Full Text].

7.   Darmaun, D, Messing B, Just B, Rongier M, and Desjeux JF. Glutamine metabolism after small intestinal resection in humans. Metabolism 40: 42-44, 1991[ISI][Medline].

8.   Darmaun, D, Rongier M, Koziet J, and Robert JJ. Glutamine nitrogen kinetics in insulin-dependent diabetic humans. Am J Physiol Endocrinol Metab 261: E713-E718, 1991[Abstract/Free Full Text].

9.   Durschlag, RP, and Smith RJ. Regulation of glutamine production by skeletal muscle cells in culture. Am J Physiol Cell Physiol 248: C442-C448, 1985[Abstract].

10.   Hankard, RG, Darmaun D, Sager BK, D'Amore D, Parsons WR, and Haymond MW. Response of glutamine metabolism to exogenous glutamine in humans. Am J Physiol Endocrinol Metab 269: E663-E670, 1995[Abstract/Free Full Text].

11.   Hankard, R, Goulet O, Ricour C, Rongier M, Colomb V, and Darmaun D. Glutamine metabolism in children with short-bowel syndrome: a stable isotope study. Pediatr Res 36: 202-206, 1994[Abstract].

12.   Hankard, RG, Haymond MW, and Darmaun D. Role of glutamine as a glucose precursor in fasting humans. Diabetes 46: 1535-1541, 1997[Abstract].

13.   Hoerr, RA, Yu YM, Wagner DA, Burke JF, and Young VR. Recovery of 13C in breath from NaH13CO3 infused by gut and vein: effect of feeding. Am J Physiol Endocrinol Metab 257: E426-E438, 1989[Abstract/Free Full Text].

14.   Kuhn, KS, Schuhmann K, Stehle P, Darmaun D, and Furst P. Determination of glutamine in muscle protein facilitates accurate assessment of proteolysis and de novo synthesis-derived endogenous glutamine production. Am J Clin Nutr 70: 484-489, 1999[Abstract/Free Full Text].

15.   Matthews, DE, Motil KJ, Rohrbaugh DK, Burke JF, Young VR, and Bier DM. Measurement of leucine metabolism in man from a primed continuous infusion of L-[1-13C]leucine. Am J Physiol Endocrinol Metab 238: E473-E479, 1980[Abstract/Free Full Text].

16.   Matthews, DE, Schwarz HP, Yang RD, Motil KJ, Young VR, and Bier DM. Relationship of plasma leucine and alpha -ketoisocaproate during a L-[1-13C]leucine infusion in man: a method for measuring human intracellular leucine tracer enrichment. Metabolism 31: 1105-1112, 1982[ISI][Medline].

17.   Nurjhan, N, Bucci A, Perriello G, Stumvoll M, Dailey G, Bier DM, Toft I, Jenssen TG, and Gerich JE. Glutamine: a major gluconeogenic precursor and vehicle for interorgan carbon transport in man. J Clin Invest 95: 272-277, 1995[ISI][Medline].

18.   Perriello, G, Jorde R, Nurjhan N, Stumvoll M, Dailey G, Jenssen T, Bier DM, and Gerich JE. Estimation of glucose-alanine-lactate-glutamine cycles in postabsorptive humans: role of skeletal muscle. Am J Physiol Endocrinol Metab 269: E443-E450, 1995[Abstract/Free Full Text].

19.   Robert, JJ, Bier DM, Zhao XH, Matthews DE, and Young VR. Glucose and insulin effects on the novo amino acid synthesis in young men: studies with stable isotope labeled alanine, glycine, leucine, and lysine. Metabolism 31: 1210-1218, 1982[ISI][Medline].

20.   Robert, JJ, Beaufrere B, Koziet J, Desjeux JF, Bier DM, Young VR, and Lestradet H. Whole body de novo amino acid synthesis in type I (insulin-dependent) diabetes studied with stable isotope-labeled leucine, alanine, and glycine. Diabetes 34: 67-73, 1985[Abstract].

21.   Salman, EK, Haymond MW, Bayne E, Sager BK, Wiisanen A, Pitel P, and Darmaun D. Protein and energy metabolism in prepubertal children with sickle cell anemia. Pediatr Res 40: 34-40, 1996[Abstract].

22.   Sarantos, P, Abouhamze Z, Copeland EM, and Souba WW. Glucocorticoids regulate glutaminase gene expression in human intestinal epithelial cells. J Surg Res 57: 227-231, 1994[ISI][Medline].

23.   Simonson, DC, and DeFronzo RA. Indirect calorimetry: methodological and interpretative problems. Am J Physiol Endocrinol Metab 258: E399-E412, 1990[Abstract/Free Full Text].

24.   Wolfe, RR. Radioactive and Stable Isotope Tracers in Biomedicine: Principles and Practice of Kinetic Analysis. New York: Wiley-Liss, 1992.


Am J Physiol Endocrinol Metab 279(3):E608-E613
0193-1849/00 $5.00 Copyright © 2000 the American Physiological Society




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