Methods for studying protein synthesis and degradation in liver and skeletal muscle

Hasselgren, Per-Olof; Pedersen, Peter Volsted; Sax, Harry C.; Warner, Brad W.; Fischer, Josef E.

doi:10.1016/0022-4804(88)90136-9

Cited by 24 publications

(8 citation statements)

References 126 publications

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“…The most important feature of the catabolic state observed in patients following burn injury is the net release of amino acid nitrogen from skeletal muscle, resulting in a negative nitrogen balance and erosion of the lean body mass. Therefore, several experimental methods for measuring the rate skeletal muscle proteolysis have been developed in both patients and animal models (Hasselgren et al, 1988). In the non-isotopic approach, the rate of release of amino acids that are not metabolized by the muscle, such as phenylalanine, tyrosine or 3-methyl-histidine, as well as a total nitrogen balance are measured (Ardawi, 1988, Brooks et al, 1986Fukagawa et al, 1985;Solomon et al, 2002).…”

Section: Proteolysis and The Nitrogen Balancementioning

confidence: 99%

“…Although such studies are arguably the most physiologically relevant, they provide limited mechanistic information because one cannot decouple the effects of injury that are intrinsic to skeletal muscle vs. those related to exogenous factors (e.g., substrate and hormone levels), and it is not possible to precisely control the composition of the perfusing blood. The perfused hindquarter model, in contrast, allows the metabolic isolation of the skeletal muscle from the contributions of other organs and to control exogenous factors entering the tissue while preserving the tissue architecture, including the vascular control of metabolite transport (Hasselgren et al, 1988). Furthermore, the rates of many metabolic processes in isolated perfused organ preparations can be quantitatively determined by periodic sampling of the perfusate and the use of stoichiometric metabolic models (Lee et al, 1999;Yarmush and Banta, 2003).…”

Section: Introductionmentioning

confidence: 99%

“…Furthermore, the rates of many metabolic processes in isolated perfused organ preparations can be quantitatively determined by periodic sampling of the perfusate and the use of stoichiometric metabolic models (Lee et al, 1999;Yarmush and Banta, 2003). The in vitro incubated muscle approach provides the most flexible platform for metabolic investigations, as it isolates the skeletal muscle metabolism from systemic influences (Ardawi, 1988;Clark et al, 1984;Hasselgren et al, 1988;Odessey and Parr, 1982), and is also suitable for high-throughput analyses. However, this is also the least physiologically relevant system, since the tissue architecture is fundamentally altered, and the route of metabolite transport is changed from vascular delivery to external diffusion (Hasselgren et al, 1988).…”

Section: Introductionmentioning

confidence: 99%

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Quantitative effects of thermal injury and insulin on the metabolism of the skeletal muscle using the perfused rat hindquarter preparation

Banta¹,

Yokoyama²,

Berthiaume³

et al. 2004

Biotech & Bioengineering

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Injury from a severe burn or trauma can propel the body into a hypermetabolic state that can lead to the significant erosion of lean muscle mass. Investigations describing this process have been somewhat limited due to the lack of adequate experimental models. Here we report the use of a perfused rat hindquarter preparation to study the consequences of a moderate burn injury (approximately 20% total body surface area), with or without the addition of exogenous insulin (12.5 mU/mL), on the fluxes of major metabolites across the isolated skeletal muscle. The metabolic flux data was further analyzed using metabolic flux analysis (MFA), which allows for the estimation of the impact of these conditions on the intracellular muscle metabolism. Results indicate that this model is able to capture the increased rate of proteolysis, glutamine formation, and the negative nitrogen balance associated with the burn-induced hypermetabolic state. The inclusion of exogenous insulin resulted in significant changes in several fluxes, including an increase in the metabolism of glucose and the flux through the pentose phosphate pathway, as well as a reduction in the metabolism of glutamine, alanine, and leucine. However, insulin administration did not affect the nitrogen balance or the rate of proteolysis in the muscle, as has been suggested using other techniques. The use of the perfused hindquarter model coupled with MFA is a physiologically relevant and experimentally flexible platform for the exploration of skeletal muscle metabolism under catabolic conditions, and it will be useful in quantifying the specific metabolic consequences of other therapeutic advances.

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Section: Proteolysis and The Nitrogen Balancementioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Quantitative effects of thermal injury and insulin on the metabolism of the skeletal muscle using the perfused rat hindquarter preparation

Banta¹,

Yokoyama²,

Berthiaume³

et al. 2004

Biotech & Bioengineering

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“…The rate of protein synthesis was determined by the modified methods of Hasselgren et al (22) and Tyson and Green (23). In brief, hepatocyte suspensions were incubated with 5 mmol/L Lleucine (cold) and 2.5 pCi of I4C leucine (DuPont, NEN Products, Boston, MA) for 30 min.…”

Section: Thamentioning

confidence: 99%

Tetrahydroaminoacridine–Induced Ribosomal Changes and Inhibition of Protein Synthesis in Rat Hepatocyte Suspensions

et al. 1994

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Tacrine (tetrahydroaminoacridine) is currently the only drug approved for the treatment of Alzheimer's disease. Unfortunately, tetrahydroaminoacridine therapy is often limited by this drug's propensity to induce reversible hepatotoxicity. Using suspensions of freshly isolated rat hepatocytes, we investigated the mechanism of tetrahydroaminoacridine cytotoxicity by examining the effect of tetrahydroaminoacridine on hepatocyte viability, protein synthesis, protein, DNA and RNA levels and ultrastructure. Our experimental findings support the explanation that tetrahydroaminoacridine-induced hepatotoxicity results from tetrahydroaminoacridine's adverse effect on protein synthesis and ribosomal structure and function. We found that viable, tetrahydroaminoacridine-treated hepatocytes (1.0 to 2.0 mmol/L or 118 to 235 pg/lOs cells) demonstrated a dose-dependent and dramatic aggregation of ribosomes on endoplasmic reticulum as well as the aggregation of other nucleic acids found in the nucleus (chromatin) and in mitochondria. These electron microscopy data suggest that tetrahydroaminoacridine treatment results in severe ribosomal dysfunction. This was confirmed by the observed rapid loss of cellular RNA content (but not DNA or protein) and the rapid and complete inhibition of protein synthesis in tetrahydroaminoacridine-treated cells (lowest concentration tested was 0.5 mmol/L or 58 pg/106 cells). Thus tetrahydroaminoacridine treatment appears to aggregate hepatocellular nucleic acids, and in doing so adversely affects ribosomal function and protein synthesis. We propose that these adverse effects of exposure to tetrahydroaminoacridine are responsible for tetrahydroaminoacridine-induced hepatotoxicity.

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“…The molecular mechanisms that may participate in the regulation of protein turnover of skeletal muscle have been reviewed recently [1]. The processes involved in synthesis and degradation and the evaluation of the results obtained have been discussed in detail [2][3][4]. In human subjects surgical trauma is accompanied by loss of nitrogen from the body.…”

mentioning

confidence: 99%

Degradation of Proteins in Human Skeletal Muscle. Characterization of the Biopsy System and Its Application to Patients Undergoing Abdominal Surgery.

Decken

Hammarqvist

Vinnars

et al. 1993

J. Clin. Biochem. Nutr.

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Methods for studying protein synthesis and degradation in liver and skeletal muscle

Cited by 24 publications

References 126 publications

Quantitative effects of thermal injury and insulin on the metabolism of the skeletal muscle using the perfused rat hindquarter preparation

Quantitative effects of thermal injury and insulin on the metabolism of the skeletal muscle using the perfused rat hindquarter preparation

Tetrahydroaminoacridine–Induced Ribosomal Changes and Inhibition of Protein Synthesis in Rat Hepatocyte Suspensions

Degradation of Proteins in Human Skeletal Muscle. Characterization of the Biopsy System and Its Application to Patients Undergoing Abdominal Surgery.

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