Transmembrane transport and intracellular kinetics of amino acids in human skeletal muscle

Biolo, Gianni; Fleming, Rachel; Maggi, Sergio P.; Wolfe, Robert R.

doi:10.1152/ajpendo.1995.268.1.e75

Cited by 213 publications

(332 citation statements)

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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%

“…1) (28,29) and skin (30) makes it less suitable for more analytical biochemical studies with stable isotopes. Furthermore, although tissue glutamine levels in murine muscle appear to be ϳ10-fold lower than in human muscle (10,37,38), it will be necessary to infuse labeled glutamine for a prolonged period via a permanent jugular vein catheter to reach steady-state levels (38). Irrespective of these considerations, the use of a muscle-specific GS-deficient mouse has made it possible to quantify the contribution of muscle to glutamine production and ammonia detoxification in a fairly physiological condition.…”

Section: Discussionmentioning

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%

Section: Discussionmentioning

confidence: 99%

mentioning

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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“…This calculation shows that 67 % of the tracer administered must have entered the muscle and must have been trapped in the free glutamine pool in muscle, protected from further metabolism. This value seems unrealistically high given that the glutamine kinetic data of Biolo et al (1995a) indicate that only 19 % of the arterially-supplied glutamine is taken up by the muscle. Furthermore, glutamine tracer also enters the intracellular pools of all other organs and is used for protein synthesis in all organs of the body.…”

Section: Three-compartment Modelmentioning

confidence: 98%

Tracers to investigate protein and amino acid metabolism in human subjects

Wagenmakers

1999

Proc. Nutr. Soc.

110

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Abbreviations: FSR, fractional synthesis rate; IRMS, isotope-ratio mass spectrometry; α-KIC, α-ketoisocaproic acid; MS, mass spectrometry; nb, net balance of tracer; NB, net balance of tracee; Ra, rate of appearance; Rd, rate of disappearance. The contribution of whole-body methods to the future gain of knowledge is expected to be limited due to the fact that most physiological disturbances have been investigated extensively, and due to the lack of information on the relative contribution of various tissues and proteins to whole-body changes.Tracer amino acid-incorporation methods are most suited to investigate these latter aspects of protein metabolism. These methods have shown that some tissues (liver and gut) have much higher turnover rates and deposit much more protein than others (muscle). Massive differences also exist between the fractional synthesis rates of individual proteins. The incorporation methods have been properly validated, although minor disagreements remain on the identity of the true precursor pool (the enrichment of which should be used in the calculations). Arterio-venous organ balance studies have shown that little protein is deposited in skeletal muscle following a proteincontaining meal, while much more protein is deposited in liver and gut. The amount deposited in the feeding period in each of these tissues is released again during overnight fasting. The addition of tracers to organ balance studies allows the simultaneous estimation of protein synthesis and protein breakdown, and provides information on whether changes in net protein balance are caused primarily by a change in protein synthesis or in protein breakdown. In the case of a small arterio-venous difference in a tissue with a high blood flow, estimates of protein synthesis and breakdown become very uncertain, limiting the value of using the tracer. An additional measurement of the intracellular free amino acid pool enrichment allows a correction for amino acid recycling and quantification of the inward and outward transmembrane transport. However, in order to obtain reliable estimates of the intramuscular amino acid enrichment and, therefore, of muscle protein synthesis and breakdown in this so-called three-pool model, the muscle should be freeze-dried and the resulting fibres should be freed from connective tissue and small blood clots under a dissection microscope. Even when optimal precautions are taken, the calculations in these tracer balance methods use multiple variables and, therefore, are bound to lead to more variability in estimates of protein synthesis than the tracer amino acid incorporation methods. In the future, most studies should focus on the measurement of protein synthesis and breakdown in specific proteins in order to understand the mechanisms behind tissue adaptation in response to various stimuli (feeding, fasting, exercise, trauma, sepsis, disuse and disease). The tracer laboratories, therefore, should improve the methodology to allow the measurement of low tracer amino acid enrichments in small amount...

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“…25 As a consequence, the structure and function of essential organs such as skeletal muscle, skin, immune system, and cellular membrane transport functions are compromised. 26,27 Chang et al 28 delineated in their study that a 10% loss of LBM leads to an impairment of immune function, 20% loss of LBM decreased wound healing with 30% mortality, 30% loss of LBM to pneumonia and pressure sores with 50% mortality, and if 40% of LBM is lost, death will occur in 100%. Despite the identification and delineation of parts of the postburn response, no prospective large clinical study has ever fully characterized the major components during the acute phase postburn.…”

Section: Discussionmentioning

confidence: 99%

The Pathophysiologic Response to Severe Burn Injury.

Jeschke

Finnerty

Norbury

et al. 2006

Critical Care Medicine

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Objective-To improve clinical outcome and to determine new treatment options, we studied the pathophysiologic response postburn in a large prospective, single center, clinical trial.Summary Background Data-A severe burn injury leads to marked hypermetabolism and catabolism, which are associated with morbidity and mortality. The underlying pathophysiology and the correlations between humoral changes and organ function have not been well delineated.Methods-Two hundred forty-two severely burned pediatric patients [>30% total body surface area (TBSA)], who received no anabolic drugs, were enrolled in this study. Demographics, clinical data, serum hormones, serum cytokine expression profile, organ function, hypermetabolism, muscle protein synthesis, incidence of wound infection sepsis, and body composition were obtained throughout acute hospital course.Results-Average age was 8 ± 0.2 years, and average burn size was 56 ± 1% TBSA with 43 ± 1% third-degree TBSA. All patients were markedly hypermetabolic throughout acute hospital stay and had significant muscle protein loss as demonstrated by a negative muscle protein net balance (−0.05% ± 0.007 nmol/100 mL leg/min) and loss of lean body mass (LBM) (−4.1% ± 1.9%); P < 0.05. Patients lost 3% ± 1% of their bone mineral content (BMC) and 2 ± 1% of their bone mineral density (BMD). Serum proteome analysis demonstrated profound alterations immediately postburn, which remained abnormal throughout acute hospital stay; P < 0.05. Cardiac function was compromised immediately after burn and remained abnormal up to discharge; P < 0.05. Insulin resistance appeared during the first week postburn and persisted until discharge. Patients were hyperinflammatory with marked changes in IL-8, MCP-1, and IL-6, which were associated with 2.5 ± 0.2 infections and 17% sepsis.Conclusions-In this large prospective clinical trial, we delineated the complexity of the postburn pathophysiologic response and conclude that the postburn response is profound, occurring in a timely manner, with derangements that are greater and more protracted than previously thought.

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Transmembrane transport and intracellular kinetics of amino acids in human skeletal muscle

Cited by 213 publications

References 20 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

Tracers to investigate protein and amino acid metabolism in human subjects

The Pathophysiologic Response to Severe Burn Injury.

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