Regulation of protein synthesis and degradation during in vitro cardiac work

1980 ◽  
Vol 238 (5) ◽  
pp. E431-E442 ◽  
Author(s):  
H. E. Morgan ◽  
B. H. Chua ◽  
E. O. Fuller ◽  
D. Siehl
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Protein Degradation ◽  
Nitrogen Balance ◽  
Cardiac Work ◽  
Amino Acid Availability ◽  
Intracellular Concentrations ◽  
Substrate Mixtures ◽  
Beta Hydroxybutyrate

Cardiac work increased protein synthesis in hearts supplied glucose (mixture 1), glucose-insulin-glucagon-lactate-beta-hydroxybutyrate (mixture 2) or palmitate-beta-hydroxybutyrate-glucose (mixture 3). In hearts provided mixture 1, acceleration of synthesis involved increased rates of peptide chain initiation. In these hearts intracellular concentrations of 5 amino acids decreased and 13 others were unchanged, indicating that faster protein synthesis did not depend on increased amino acid availability. In hearts supplied mixtures 2, 3, or 4 (lactate-glucose-insulin), intracellular concentrations of branched-chain amino acids were decreased by work, whereas intracellular levels of some acidic and neutral amino acids increased. Protein degradation was decreased by work in hearts supplied mixtures 1 and 2, but not mixtures 3 and 4. In hearts provided mixture 1, nitrogen balance was negative, but less so in working preparations. Nitrogen balance was zero or positive in working hearts provided mixtures 2 and 4. These studies indicated that in hearts supplied some, but not all, of the substrate mixtures, cardiac work maintained efficiently of protein synthesis and inhibited protein degradation. An improved method for perfusion of working hearts with albumin-containing buffer is described.

scholarly journals Effects of insulin in vitro on protein turnover in rat epitrochlearis muscle

1983 ◽  
Vol 210 (2) ◽  
pp. 323-330 ◽  
Author(s):  
W S Stirewalt ◽  
R B Low
Keyword(s):  
Protein Synthesis ◽  
Protein Degradation ◽  
Nitrogen Balance ◽  
Protein Metabolism ◽  
Serum Insulin ◽  
Specific Radioactivity ◽  
Physiological Concentration ◽  
Rat Serum ◽  
The Third

Rates of protein synthesis and degradation were measured in the isolated rat epitrochlearis muscle by radiotracer techniques, by using the specific radioactivity of tRNA-bound amino acid as precursor for protein synthesis. The tissue maintained linear rates of protein synthesis for 3 h of incubation in the presence of amino acids and glucose and in the absence of insulin. Under these conditions, however, the muscles were in negative nitrogen balance, with rates of protein degradation exceeding rates of protein synthesis. Under steady-state conditions of labelling, the specific radioactivities of tRNA-bound leucine, phenylalanine and valine were significantly less than their respective values in the incubation medium, at concentrations in the medium varying from 1 to 10 times those in normal rat serum. Insulin caused a dose- and time-dependent increase in tRNA-based protein synthesis rates, more than doubling rates at 5 and 50 ng of insulin/ml. At the lower, physiological, concentration of insulin, the stimulation of protein synthesis was not observed until the third hour of incubation with the hormone, whereas the rate of protein synthesis at the higher concentration was elevated during the second hour. There were no delays in the stimulation by insulin of glucose conversion into glycogen. The delayed stimulatory effects of insulin on the rate of protein synthesis brought the tissue to a nitrogen balance near zero. The presence of the hormone also prevented the increase in the rate of protein degradation seen in the third hour of incubation in the absence of the hormone. These studies demonstrate the viability of the incubated rat epitrochlearis muscle with respect to protein metabolism and sensitivity to the protein anabolic effects of physiological concentrations of insulin, and indicate that the preparation is a suitable experimental model for the study of the control of protein metabolism in fast-twitch skeletal muscle.

A role for leucine in regulation of protein turnover in working rat hearts

1980 ◽  
Vol 239 (6) ◽  
pp. E510-E514 ◽  
Author(s):  
B. H. Chua ◽  
D. L. Siehl ◽  
H. E. Morgan
Keyword(s):  
Protein Synthesis ◽  
Protein Degradation ◽  
Nitrogen Balance ◽  
Plasma Levels ◽  
Protein Turnover ◽  
Ribosomal Subunits ◽  
Rat Hearts ◽  
Plasma Leucine ◽  
Beta Hydroxybutyrate

Effect of leucine on protein turnover was examined in perfused hearts provided with 1 (0.2 mM) or 5 times (1 mM) plasma levels of leucine and normal plasma levels of other amino acids. When hearts were perfused as Langendorff or working preparations with buffer that contained 15 mM glucose, protein degradation was 2–3 times faster than protein synthesis. As a result, the heart was in marked negative nitrogen balance. Addition of 1 mM leucine significantly improved the nitrogen balance (24–33%) by stimulating protein synthesis in Langendorff preparations (25%) and inhibiting protein degradation in both preparations (14–29%). The stimulatory effect of leucine on protein synthesis was associated with a reduction in levels of ribosomal subunits. In hearts supplied physiological levels of glucose, lactate, beta-hydroxybutyrate, insulin, and glucagon, protein synthesis was more nearly equal to protein degradation. Provision of 1 mM leucine stimulated protein synthesis only in Langendorff preparations (32%) but did not have a significant effect on protein degradation in either preparation. Although leucine did not have a significant effect on either protein synthesis or degradation in working hearts, nitrogen balance became positive with addition of 1 mM leucine. These results suggest that leucine may exert an effect on myocardial nitrogen balance in vivo under conditions that elevate plasma leucine concentrations.

Protein Degradation in Human Skeletal Muscle Tissue: The Effect of Insulin, Leucine, Amino Acids and Ions

1981 ◽  
Vol 60 (3) ◽  
pp. 319-326 ◽  
Author(s):  
K. Lundholm ◽  
S. Edström ◽  
L. Ekman ◽  
I. Karlberg ◽  
P. Walker ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Amino Acids ◽  
Protein Synthesis ◽  
Protein Degradation ◽  
Muscle Tissue ◽  
Human Skeletal Muscle ◽  
Degradation Rate ◽  
Rectus Abdominus ◽  
Protein Degradation Rate

1. The protein degradation rate of human skeletal muscle was evaluated in vitro in isolated fibre bundles from the rectus abdominus muscle by measuring the tyrosine released from muscle tissue proteins. Protein metabolism in this semi-intact preparation was compared with that of the intact extensor digitorum longus muscles from rats under the same experimental conditions. 2. Protein balance was negative in both preparations, but protein synthesis and degradation were two to three times higher in the rat muscles. Tyrosine was released at a constant rate for at least 3 h of incubation independent of whether protein synthesis was inhibited or not. Disintegration of the muscle fibres more than doubled the tyrosine release rate. Human red gastrocnemius muscle showed 37% higher degradation rate compared with the predominantly white rectus abdominus muscle. The half-life of human skeletal muscle protein in vitro was estimated to be 20 days when calculated from the rate of tyrosine release. 3. The addition of leucine to the incubation medium decreased the rate of protein degradation, which was further decreased by the addition of other amino acids. Insulin did not influence the protein degradation rate during 2 h of incubation. This did not reflect a lack of sensitivity to insulin of the preparation, since protein synthesis responded to insulin. Calcium (5 mmol/l) stimulated and zinc (0.1 mmol/l) inhibited the protein degradation. 4. This experimental system may be suitable as an additional tool for evaluating protein degradation in human skeletal muscles.

Turnover of total proteins and ornithine aminotransferase during liver regeneration in rats

1980 ◽  
Vol 238 (1) ◽  
pp. E46-E52
Author(s):  
S. L. Augustine ◽  
R. W. Swick
Keyword(s):  
Amino Acids ◽  
Liver Regeneration ◽  
Protein Degradation ◽  
Partial Hepatectomy ◽  
Free Amino ◽  
Liver Protein ◽  
Ornithine Aminotransferase ◽  
Fractional Rate

The recovery of approximately 40% of the total liver protein during the first day after partial hepatectomy was shown to be due to the near cessation of protein breakdown rather than to an increase in protein synthesis. The decrease in degradation of total protein was less if rats were adrenalectomized or protein-depleted prior to partial hepatectomy. The effect of these treatments originally suggested that changes in free amino acid levels in liver might be related to the rate of protein degradation. However, no correlation was found between levels of total free amino acids and rates of breakdown. Measurements of individual amino acids during liver regeneration suggested that levels of free methionine and phenylalanine, amino acids that have been found to lower rates of protein degradation in vitro, are not correlated with rates of breakdown in vivo. The difference between the fractional rate of ornithine aminotransferase degradation (0.68/day and 0.28/day in sham-hepatectomized and partially hepatectomized rats, respectively) was sufficient to account for the higher level of this protein 3 days after surgery in the latter group.

scholarly journals Effect of long-term refeeding on protein metabolism in patients with cirrhosis of the liver

1997 ◽  
Vol 77 (2) ◽  
pp. 197-212 ◽  
Author(s):  
Jens Kondrup ◽  
Klaus Nielsen ◽  
Anders Juul
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Protein Degradation ◽  
Protein Metabolism ◽  
Cirrhosis Of The Liver ◽  
N Balance ◽  
Whole Body ◽  
Protein Requirement ◽  
Protein Utilization ◽  
Igf I

Patients with cirrhosis of the liver require an increased amount of protein to achieve N balance. However, the utilization of protein with increased protein intake, i.e. the slope from regression analysis of N balance v. intake, is highly efficient (Nielsen et al. 1995). In the present study, protein requirement and protein utilization were investigated further by measuring protein synthesis and degradation. In two separate studies, five or six patients with cirrhosis of the liver were refed on a balanced diet for an average of 2 or 4 weeks. Protein and energy intakes were doubled in both studies. Initial and final whole-body protein metabolism was measured in the fed state by primed continous [15N]glycine infusion. Refeeding caused a statistically significant increase of about 30% in protein synthesis in both studies while protein degradation was only slightly affected. The increase in protein synthesis was associated with significant increases in plasma concentrations of total amino acids (25%), leucine (58%), isoleucine (82%), valine (72%), proline (48%) and triiodothyronine (27%) while insulin, growth hormone, insulin-like growth factor (IGF)-I and IGF-binding protein-3 were not changed significantly. The results indicate that the efficient protein utilization is due to increased protein synthesis, rather than decreased protein degradation, and suggest that increases in plasma amino acids may be responsible for the increased protein synthesis. A comparison of the patients who had a normal protein requirement with the patients who had an increased protein requirement suggests that the increased protein requirement is due to a primary increase in protein degradation. It is speculated that this is due to low levels of IGF-I secondary to impaired liver function, since initial plasma concentration of IGF-I was about 25% of control values and remained low during refeeding.

scholarly journals Identification in skeletal muscle of a distinct extracellular pool of amino acids, and its role in protein synthesis

1971 ◽  
Vol 121 (5) ◽  
pp. 817-827 ◽  
Author(s):  
R. C. Hider ◽  
E. B. Fern ◽  
D. R. London
Keyword(s):  
Skeletal Muscle ◽  
Amino Acids ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Incubation Medium ◽  
Extensor Digitorum Longus ◽  
Extensor Digitorum ◽  
Longus Muscle ◽  
Kinetics Of

1. The kinetics of radioactive labelling of extra- and intra-cellular amino acid pools and protein of the extensor digitorum longus muscle were studied after incubations with radioactive amino acids in vitro. 2. The results indicated that an extracellular pool could be defined, the contents of which were different from those of the incubation medium. 3. It was concluded that amino acids from the extracellular pool, as defined in this study, were incorporated directly into protein.

Effect of Cycloheximide on In Vitro and In Vivo Protein Synthesis by Certain Tissues of Rats and Pigeons with Special Reference to Muscle

1973 ◽  
Vol 51 (12) ◽  
pp. 933-941 ◽  
Author(s):  
Njanoor Narayanan ◽  
Jacob Eapen
Keyword(s):  
Amino Acids ◽  
Body Weight ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Amino Acid Incorporation ◽  
Contrast Administration ◽  
Acid Incorporation ◽  
Tissue Homogenates

The effect of cycloheximide in vitro and in vivo on the incorporation of labelled amino acids into protein by muscles, liver, kidneys, and brain of rats and pigeons was studied. In vitro incorporation of amino acids into protein by muscle microsomes, myofibrils, and myofibrillar ribosomes was not affected by cycloheximide. In contrast, administration of the antibiotic into intact animals at a concentration of 1 mg/kg body weight resulted in considerable inhibition of amino acid incorporation into protein by muscles, liver, kidneys, and brain. This inhibition was observed in all the subcellular fractions of these tissues during a period of 10–40 min after the administration of the precursor. Tissue homogenates derived from in vivo cycloheximide-treated animals did not show significant alteration in in vitro amino acid incorporation with the exception of brain, which showed a small but significant enhancement.

Effects of glucose, pyruvate, lactate, and amino acids on muscle protein synthesis

1982 ◽  
Vol 242 (3) ◽  
pp. E184-E192 ◽  
Author(s):  
M. P. Hedden ◽  
M. G. Buse
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Muscle Protein ◽  
Muscle Protein Synthesis ◽  
Creatine Phosphate ◽  
Serum Amino Acids ◽  
14Co2 Production ◽  
Beta Hydroxybutyrate ◽  
Free Media ◽  
Effect Of Glucose

Protein synthesis was measured in rat diaphragms incubated with serum amino acids + 0.35 mM L-[2,6-3H]tyrosine and different energy-yielding substrates. Muscles incubated with 5.5 mM glucose (with or without actinomycin D) synthesized more protein than those incubated with 11 mM pyruvate or 11 mM lactate. Tissue ATP decreased during incubation with lactate, but pyruvate maintained ATP, ADP, and creatine phosphate as well as glucose. Glucose 6-phosphate decreased in muscles incubated in glucose-free media. 14CO2 production from substrates was [1-14C]pyruvate greater than [1-14C]lactate greater than [3,4-14C]glucose. Intracellular lactate/pyruvate was measured to assess cytoplasmic free NADH/NAD+; the effect of different media on these ratios was lactate greater than glucose = lactate + pyruvate greater than pyruvate + glucose greater than pyruvate. Lactate + pyruvate (8.8 + 2.2 mM) supported protein synthesis better than pyruvate and as well as glucose. Adding glucose to pyruvate accelerated protein synthesis and increased NADH/NAD+. Iodoacetate (0.1 mM) inhibited glycolytic NAD reduction and abolished the stimulatory effect of glucose on protein synthesis in the presence of pyruvate. Supplementation of pyruvate media with 1 mM leucine or isoleucine stimulated protein synthesis, but beta-hydroxybutyrate, malate, alpha-ketoisocaproate, and all other amino acids were ineffective. The cytoplasmic redox potential may act as a translational modulator of protein synthesis in skeletal muscle.

scholarly journals The rate-limiting step of protein synthesis in vivo and in vitro and the distribution of growing peptides between the puromycinlabile and puromycin-non-labile sites on polyribosomes

1971 ◽  
Vol 122 (3) ◽  
pp. 267-276 ◽  
Author(s):  
D. C. N. Earl ◽  
Susan T. Hindley
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Donor Site ◽  
Peptide Bond ◽  
Rate Limiting Step ◽  
Limiting Step ◽  
Donor And Acceptor ◽  
Rate Limiting

1. At 3 min after an intravenous injection of radioactive amino acids into the rat, the bulk of radioactivity associated with liver polyribosomes can be interpreted as growing peptides. 2. In an attempt to identify the rate-limiting step of protein synthesis in vivo and in vitro, use was made of the action of puromycin at 0°C, in releasing growing peptides only from the donor site, to study the distribution of growing peptides between the donor and acceptor sites. 3. Evidence is presented that all growing peptides in a population of liver polyribosomes labelled in vivo are similarly distributed between the donor and acceptor sites, and that the proportion released by puromycin is not an artifact of methodology. 4. The proportion released by puromycin is about 50% for both liver and muscle polyribosomes labelled in vivo, suggesting that neither the availability nor binding of aminoacyl-tRNA nor peptide bond synthesis nor translocation can limit the rate of protein synthesis in vivo. Attempts to alter this by starvation, hypophysectomy, growth hormone, alloxan, insulin and partial hepatectomy were unsuccessful. 5. Growing peptides on liver polyribosomes labelled in a cell-free system in vitro or by incubating hemidiaphragms in vitro were largely in the donor site, suggesting that either the availability or binding of aminoacyl-tRNA, or peptide bond synthesis, must be rate limiting in vitro and that the rate-limiting step differs from that in vivo. 6. Neither in vivo nor in the hemidiaphragm system in vitro was a correlation found between the proportion of growing peptides in the donor site and changes in the rate of incorporation of radioactivity into protein. This could indicate that the intracellular concentration of amino acids or aminoacyl-tRNA limits the rate of protein synthesis and that the increased incorporation results from a rise to a higher but still suboptimum concentration.

scholarly journals Action of growth hormone in vitro on the net uptake and incorporation into protein of amino acids in muscle from intact rabbits given protein-deficient diets

1976 ◽  
Vol 35 (1) ◽  
pp. 1-10 ◽  
Author(s):  
M. R. Turner ◽  
P. J. Reeds ◽  
K. A. Munday
Keyword(s):  
Amino Acids ◽  
Growth Hormone ◽  
Protein Synthesis ◽  
Amino Acid ◽  
De Novo ◽  
Amino Acid Uptake ◽  
Acid Uptake ◽  
Amino Acid Incorporation ◽  
Acid Incorporation

1. Net amino acid uptake, and incorporation into protein have been measured in vitro in the presence and absence of porcine growth hormone (GH) in muscle from intact rabbits fed for 5 d on low-protein (LP), protein-free (PF) or control diets.2. In muscle from control and LP animals GH had no effect on the net amino acid uptake but stimulated amino acid incorporation into protein, although this response was less in LP animals than in control animals.3. In muscle from PF animals, GH stimulated both amino acid incorporation into protein and the net amino acid uptake, a type of response which also occurs in hypophysectomized animals. The magnitude of the effect of GH on the incorporation of amino acids into protein was reduced in muscle from PF animals.4. The effect of GH on the net amino acid uptake in PF animals was completely blocked by cycloheximide; the uptake effect of GH in these animals was dependent therefore on de novo protein synthesis.5. It is proposed that in the adult the role of growth hormone in protein metabolism is to sustain cellular protein synthesis when there is a decrease in the level of substrate amino acids, similar to that which occurs during a short-term fast or when the dietary protein intake is inadequate.

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