Determination of glutamine in muscle protein facilitates accurate assessment of proteolysis and de novo synthesis–derived endogenous glutamine production

Kuhn, Katharina S.; Schuhmann, Karin; Stehle, Peter; Darmaun, Dominique; Fürst, Peter

doi:10.1093/ajcn/70.4.484

Cited by 72 publications

(48 citation statements)

References 27 publications

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“…Apparently, muscle glutamine levels do not directly reflect the flux through GS in mice. In this respect, it is of interest to note that muscle glutamine levels in mice (ϳ1 mmol/kg) are 3-7-fold lower than those in rats (25,36) and even 9 -12-fold lower than those in humans (1,10,37,38). Because plasma glutamine concentrations are similar in these species, the differences must arise from differences in glutamine transport across the sarcolemma and/or metabolism.…”

Section: Discussionmentioning

confidence: 99%

“…Almost 90% of the daily glutamine production originates from endogenous sources, because 30 -35% of all nitrogen derived from protein catabolism is transported in the form of glutamine (1,2). This glutamine can serve, after transport via the vasculature, as an oxidative fuel for enterocytes and immune cells, a precursor for purine and pyrimidine synthesis, a modulator of protein turnover, or an intermediate for gluconeogenesis and acid-base balance.…”

mentioning

confidence: 99%

“…To be able to study the tissue-specific functions of GS, we have generated a mouse strain in which the protein-coding region of the GS allele is flanked by loxP sequences (GS fl ). Skeletal muscle accounts for ϳ70% of the endogenous production of glutamine in man (10,11), ϳ15% of which arises from proteolysis and the remainder from de novo synthesis (1,10). Because of this quantitatively large contribution, muscular glutamine synthesis is thought to become crucial when circulating glutamine levels fall under catabolic conditions (12,13), including fasting (11,14), and when circulating ammonia levels rise because of liver failure (15,16).…”

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confidence: 99%

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Glutamine Synthetase in Muscle Is Required for Glutamine Production during Fasting and Extrahepatic Ammonia Detoxification

Hakvoort

Köhler³

et al. 2010

Journal of Biological Chemistry

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The main endogenous source of glutamine is de novo synthesis in striated muscle via the enzyme glutamine synthetase (GS). The mice in which GS is selectively but completely eliminated from striated muscle with the Cre-loxP strategy (GS-KO/M mice) are, nevertheless, healthy and fertile. Compared with controls, the circulating concentration and net production of glutamine across the hindquarter were not different in fed GS-KO/M mice. Only a ϳ3-fold higher escape of ammonia revealed the absence of GS in muscle. However, after 20 h of fasting, GS-KO/M mice were not able to mount the ϳ4-fold increase in glutamine production across the hindquarter that was observed in control mice. Instead, muscle ammonia production was ϳ5-fold higher than in control mice. The fasting-induced metabolic changes were transient and had returned to fed levels at 36 h of fasting. Glucose consumption and lactate and ketone-body production were similar in GS-KO/M and control mice. Challenging GS-KO/M and control mice with intravenous ammonia in stepwise increments revealed that normal muscle can detoxify ϳ2.5 mol ammonia/g muscle⅐h in a muscle GS-dependent manner, with simultaneous accumulation of urea, whereas GS-KO/M mice responded with accumulation of glutamine and other amino acids but not urea. These findings demonstrate that GS in muscle is dispensable in fed mice but plays a key role in mounting the adaptive response to fasting by transiently facilitating the production of glutamine. Furthermore, muscle GS contributes to ammonia detoxification and urea synthesis. These functions are apparently not vital as long as other organs function normally.Glutamine is among the most abundant free amino acids in mammals. Almost 90% of the daily glutamine production originates from endogenous sources, because 30 -35% of all nitrogen derived from protein catabolism is transported in the form of glutamine (1, 2). This glutamine can serve, after transport via the vasculature, as an oxidative fuel for enterocytes and immune cells, a precursor for purine and pyrimidine synthesis, a modulator of protein turnover, or an intermediate for gluconeogenesis and acid-base balance. The only enzyme capable of glutamine synthesis is glutamine synthetase (L-glutamate: ammonia ligase (ADP); EC 6.3.1.2). Because of the prominent role of glutamine in the interorgan transport of carbon and nitrogen, the plasma glutamine pool is turning over very rapidly (3). Glutamine tracer kinetic studies in humans have shown that ϳ50% of plasma glutamine is oxidized and that of the remainder, 10 -20% is used for gluconeogenesis, and most of the rest is used for protein synthesis and incorporation into macromolecules (2).Thus far, the irreversible GS 5 inhibitor methionine sulfoximine (MSO) was the tool of choice to interfere with cellular glutamine synthesis. Administration of MSO for 4 -6 days results in a 40 -50% decrease in plasma glutamine levels, a 55-65% decrease in intracellular muscle glutamine, and a 50% increase in muscle ammonia levels (4 -6). An inherent drawback of the...

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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confidence: 99%

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Glutamine Synthetase in Muscle Is Required for Glutamine Production during Fasting and Extrahepatic Ammonia Detoxification

Hakvoort

Köhler³

et al. 2010

Journal of Biological Chemistry

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“…Recently, by using a novel analytical tool, human skeletal muscle protein was, however, shown to contain only 4.32 g glutamine per 100 g of bound amino acid residues. B GLN was therefore estimated as 0.423 ϫ Ra LEU (28). This approach assumes that 1) the release of an amino acid from proteolysis is proportional to its abundance in body protein; 2) 100 g of body protein contain 9,180 mg leucine, i.e., 70.1 mmol leucine (as 1 mmol leucine ϭ 131 mg, and 9,180/131 ϭ 70.1); and ϳ4,320 mg glutamine (ϭ29.6 mmol glutamine since 1 mmol glutamine ϭ 146 mg, and 4,320/146 ϭ 29.6), or 0.423 mol glutamine per mol leucine (29.6/70.1 ϭ 0.423).…”

Section: Methodsmentioning

confidence: 99%

“…The fraction of glutamine Ra that cannot be accounted for by release of glutamine from protein breakdown was attributed to D GLN. DGLN was therefore estimated by DGLN ϭ Ra GLN Ϫ BGLN (28).…”

Section: Methodsmentioning

confidence: 99%

Corticosteroids increase glutamine utilization in human splanchnic bed

Thibault

Welch²,

Mauras³

et al. 2008

American Journal of Physiology-Gastrointestinal and Liver Physi

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Thibault R, Welch S, Mauras N, Sager B, Altomare A, Haymond M, Darmaun D. Corticosteroids increase glutamine utilization in human splanchnic bed. Am J Physiol Gastrointest Liver Physiol 294: G548-G553, 2008. First published December 27, 2007 doi:10.1152/ajpgi.00461.2007.-Glutamine is the most abundant amino acid in the body and is extensively taken up in gut and liver in healthy humans. To determine whether glucocorticosteroids alter splanchnic glutamine metabolism, the effect of prednisone was assessed in healthy volunteers using isotope tracer methods. Two groups of healthy adults received 5-h intravenous infusions of L-[1-14 C]leucine and L-[ 2 H5]glutamine, along with q. 20 min oral sips of tracer doses of L-[1-13 C]glutamine in the fasting state, either 1) at baseline (control group; n ϭ 6) or 2) after a 6-day course of 0.8 mg ⅐ kg Ϫ1 ⅐ day Ϫ1 prednisone (prednisone group; n ϭ 8). Leucine and glutamine appearance rates (Ra) were determined from plasma [1-14 C]ketoisocaproate and [ 2 H5]glutamine, respectively, and leucine and glutamine oxidation from breath 14 CO2 and 13 CO2, respectively. Splanchnic glutamine extraction was estimated by the fraction of orally administered [ 13 C]glutamine that failed to appear into systemic blood. Prednisone treatment 1) did not affect leucine Ra or leucine oxidation; 2) increased plasma glutamine Ra, mostly owing to enhanced glutamine de novo synthesis (medians Ϯ interquartiles, 412 Ϯ 61 vs. 280 Ϯ 190 mol ⅐ kg Ϫ1 ⅐ h Ϫ1 , P ϭ 0.003); and 3) increased the fraction of orally administered glutamine undergoing extraction in the splanchnic territory (means Ϯ SE 64 Ϯ 6 vs. 42 Ϯ 12%, P Ͻ 0.05), without any change in the fraction of glutamine oxidized (means Ϯ SE, 75 Ϯ 4 vs. 77 Ϯ 4%, not significant). We conclude that high-dose glucocorticosteroids increase in splanchnic bed the glutamine requirements. The role of such changes in patients receiving chronic corticoid treatment for inflammatory diseases or suffering from severe illness remains to be determined. nutrition; protein metabolism; stable isotopes; stress; gut GLUTAMINE PLAYS AN IMPORTANT role in the maintenance of intestinal function (39,43,46). Glutamine indeed is both a preferred fuel (9, 47) for enterocytes and a precursor for the intestinal synthesis of purines, pyrimidines (9), and glutathione (37), and it may regulate gut protein synthesis (21,30,32). In addition, glutamine is a major donor of carbon for gluconeogenesis (19). This explains why glutamine is extensively extracted in the splanchnic tissues in several animal species (39,47).In humans, the concomitant infusion of two tracers of glutamine (one infused intravenously, the other one enterally) can be used to assess the extraction of glutamine by the splanchnic bed. By using this technique, ϳ50 to 75% of enterally administered glutamine undergoes first-pass extraction within the splanchnic bed, presumably by the gut and liver, in healthy humans (4, 17, 18, 34). Additionally, the major fate of splanchnic glutamine after enteral administration was oxidation, ...

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Analysis of the role of GADD153 in the control of apoptosis in NS0 myeloma cells

Lengwehasatit

Dickson²

2002

Biotech & Bioengineering

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Apoptosis can limit the maximum production of recombinant protein expression from cultured mammalian cells. This article focuses on the links between nutrient deprivation, ER perturbation, the regulation of (growth arrest and DNA damage inducible gene 153) GADD153 expression and apoptosis. During batch culture, decreases in glucose and glutamine correlated with an increase in apoptotic cells. This event was paralleled by a simultaneous increase in GADD153 expression. The expression of GADD153 in batch culture was suppressed by the addition of nutrients and with fed-batch culture the onset of apoptosis was delayed but not completely prevented. In defined stress conditions, glucose deprivation had the greatest effect on cell death when compared to glutamine deprivation or the addition of tunicamycin (an inhibitor of glycosylation), added to generate endoplasmic reticulum stress. However, the contribution of apoptosis to overall cell death (as judged by morphology) was smaller in conditions of glucose deprivation than in glutamine deprivation or tunicamycin treatment. Transient activation of GADD153 expression was found to occur in response to all stresses and occurred prior to detection of the onset of cell death. These results imply that GADD153 expression is either a trigger for apoptosis or offers a valid indicator of the likelihood of cell death arising from stresses of relevance to the bioreactor environment.

show abstract

Determination of glutamine in muscle protein facilitates accurate assessment of proteolysis and de novo synthesis–derived endogenous glutamine production

Cited by 72 publications

References 27 publications

Glutamine Synthetase in Muscle Is Required for Glutamine Production during Fasting and Extrahepatic Ammonia Detoxification

Glutamine Synthetase in Muscle Is Required for Glutamine Production during Fasting and Extrahepatic Ammonia Detoxification

Corticosteroids increase glutamine utilization in human splanchnic bed

Analysis of the role of GADD153 in the control of apoptosis in NS0 myeloma cells

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