scholarly journals Measurement of protein synthesis in rat lungs perfused in situ

1980 ◽  
Vol 188 (1) ◽  
pp. 269-278 ◽  
Author(s):  
Clyde A. Watkins ◽  
D. Eugene Rannels
Keyword(s):  
Protein Synthesis ◽  
Amino Acid ◽  
Free Amino ◽  
Plasma Concentrations ◽  
Specific Radioactivity ◽  
Sole Source ◽  
Bicarbonate Buffer ◽  
Normal Plasma ◽  
Rat Lungs

Compartmentalization of amino acid was investigated to define conditions required for accurate measurements of rates of protein synthesis in rat lungs perfused in situ. Lungs were perfused with Krebs–Henseleit bicarbonate buffer containing 4.5% (w/v) bovine serum albumin, 5.6mm-glucose, normal plasma concentrations of 19 amino acids, and 8.6–690μm-[U-14C]phenylalanine. The perfusate was equilibrated with the same humidified gas mixture used to ventilate the lungs [O2/CO2 (19:1) or O2/N2/CO2 (4:15:1)]. [U-14C]Phenylalanine was shown to be a suitable precursor for studies of protein synthesis in perfused lungs: it entered the tissue rapidly (t½, 81s) and was not converted to other compounds. As perfusate phenylalanine was decreased below 5 times the normal plasma concentration, the specific radioactivity of the pool of phenylalanine serving as precursor for protein synthesis, and thus [14C]phenylalanine incorporation into protein, declined. In contrast, incorporation of [14C]histidine into lung protein was unaffected. At low perfusate phenylalanine concentrations, rates of protein synthesis that were based on the specific radioactivity of phenylalanyl-tRNA were between rates calculated from the specific radioactivity of phenylalanine in the extracellular or intracellular pools. Rates based on the specific radioactivities of these three pools of phenylalanine were the same when extracellular phenylalanine was increased. These observations suggested that: (1) phenylalanine was compartmentalized in lung tissue; (2) neither the extracellular nor the total intracellular pool of phenylalanine served as the sole source of precursor for protein; (3) at low extracellular phenylalanine concentrations, rates of protein synthesis were in error if calculated from the specific radioactivity of the free amino acid; (4) at high extracellular phenylalanine concentrations, the effects of compartmentalization were negligible and protein synthesis could be calculated accurately from the specific radioactivity of the free or tRNA-bound phenylalanine pool.

scholarly journals Reversible inhibition of protein synthesis in lung by halothane

1983 ◽  
Vol 210 (2) ◽  
pp. 379-387 ◽  
Author(s):  
D E Rannels ◽  
R Christopherson ◽  
C A Watkins
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Plasma Concentrations ◽  
Specific Radioactivity ◽  
Bicarbonate Buffer ◽  
Inhibition Of Protein Synthesis ◽  
Glucose Plasma ◽  
Dose Dependent ◽  
Synthesis And Degradation

Alterations in the synthesis and degradation of proteins were investigated in intact lungs exposed to the volatile anaesthetic halothane. In rat lungs perfused in situ with Krebs-Henseleit bicarbonate buffer containing 4.5% (w/v) bovine serum albumin, 5.6 mM-glucose, plasma concentrations of 19 amino acids and 690 microM-[U-14C]-phenylalanine and equilibrated with O2/N2/CO2 (4:15:1), protein synthesis, calculated based on the specific radioactivity of aminoacyl-tRNA, was inhibited by halothane. The anaesthetic did not affect degradation of lung proteins. The inhibition of protein synthesis was rapid in onset, dose-dependent, and quickly reversible. It did not appear to be associated with overall energy depletion, with non-specific changes in cellular permeability, or with decreased availability of amino acids as substrates for protein synthesis.

scholarly journals An analysis of errors in estimation of the rate of protein synthesis by constant infusion of a labelled amino acid

1978 ◽  
Vol 176 (2) ◽  
pp. 402-405 ◽  
Author(s):  
P J Garlick
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Detailed Analysis ◽  
Free Amino Acid ◽  
Free Amino ◽  
Specific Radioactivity ◽  
Constant Infusion ◽  
Simplifying Assumptions

Rates of protein synthesis in tissues can be calculated from the specific radioactivity of free and protein-bound amino acids at the end of a constant infusion of a labelled amino acid (Garlick, Millward & James (1973) Biochem. J. 136, 935–945]. The simplifying assumptions used in these calculations have been criticized [Madsen, Everett, Sparrow & Fowkes (1977) FEBS Lett. 79, 313–316]. A more detailed analysis using a programmable desk-top calculator is described, which shows that the errors introduced by the simplifying assumptions are small, particularly when the specific radioactivity of the free amino acid rises rapidly to a constant value.

Aminoacyl-tRNA and tissue free amino acid pools are equilibrated after a flooding dose of phenylalanine

1999 ◽  
Vol 277 (1) ◽  
pp. E103-E109 ◽  
Author(s):  
Teresa A. Davis ◽  
Marta L. Fiorotto ◽  
Hanh V. Nguyen ◽  
Douglas G. Burrin
Keyword(s):  
Skeletal Muscle ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Free Amino Acid ◽  
Free Amino ◽  
Specific Radioactivity ◽  
Isotopic Labeling ◽  
Precursor Pool ◽  
Free Amino Acid Pools

The flooding dose method, which is used to measure tissue protein synthesis, assumes equilibration of the isotopic labeling between the aminoacyl-tRNA pool and the tissue and blood free amino acid pools. However, this has not been verified for a phenylalanine tracer in an in vivo study. We determined the specific radioactivity of [3H]phenylalanine in the aminoacyl-tRNA and the tissue and blood free amino acid pools of skeletal muscle and liver 30 min after administration of a flooding dose of phenylalanine along with [3H]phenylalanine. Studies were performed in neonatal pigs in the fasted and refed states and during hyperinsulinemic-euglycemic-amino acid clamps. The results showed that, 30 min after the administration of a flooding dose of phenylalanine, there was equilibration of the specific radioactivity of phenylalanine among the blood, tissue, and tRNA precursor pools. Equilibration of the specific radioactivity of the three precursor pools for protein synthesis occurred in both skeletal muscle and liver. Neither feeding nor insulin status affected the aminoacyl-tRNA specific radioactivity relative to the tissue free amino acid specific radioactivity. The results support the assumption that the tissue free amino acid pool specific radioactivity is a valid measure of the precursor pool specific radioactivity and thus can be used to calculate protein synthesis rates in skeletal muscle and liver when a flooding dose of phenylalanine is administered.

scholarly journals The specific radioactivity of the tissue free amino acid pool as a basis for measuring the rate of protein synthesis in the rat in vivo

1974 ◽  
Vol 142 (2) ◽  
pp. 413-419 ◽  
Author(s):  
Edward B. Fern ◽  
Peter J. Garlick
Keyword(s):  
Protein Synthesis ◽  
Amino Acid ◽  
Free Amino Acid ◽  
Free Amino ◽  
Gastrocnemius Muscle ◽  
Specific Radioactivity ◽  
Amino Acid Pool ◽  
Free Amino Acid Pool ◽  
Acid Pool

1. Rats were infused in vivo with [U-14C]glycine for periods of 2–6h, during which time the specific radioactivity of the free glycine in plasma and tissue approached a constant value. 2. Free serine also became labelled. The ratio of specific radioactivity of serine to that of glycine in the protein of liver, kidney, brain, jejunum, heart, diaphragm and gastrocnemius muscle was closer to the ratio in the free amino acid pool of the tissue than that of the plasma. 3. The kinetics of incorporation of [14C]glycine and [14C]serine into the protein of gastrocnemius muscle further suggested that the plasma free amino acids were not the immediate precursors of protein. 4. Infusion of rats with [U-14C]serine resulted in labelling of free glycine. The ratio of specific radioactivity of glycine to serine in the protein of liver, kidney, brain, jejunum and heart again suggested incorporation from a pool similar to the free amino acid pool of the tissue. 5. Rates of tissue protein synthesis calculated from the incorporation into protein of both radioactive glycine and serine, either infused or derived, were very similar when the precursor specific radioactivity was taken to be that in the total free amino acids of the tissue. Except for gastrocnemius muscle and diaphragm during the infusion of radioactive serine, the rates of tissue protein synthesis calculated from the specific radioactivity of the free glycine and serine in plasma differed markedly.

scholarly journals Effects of progesterone on protein metabolism in chicken oviduct tissue pretreated with oestrogen

1970 ◽  
Vol 120 (2) ◽  
pp. 337-344 ◽  
Author(s):  
K. R. Muller ◽  
R. F. Cox ◽  
N. H. Carey
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Free Amino ◽  
Specific Radioactivity ◽  
Dna And Rna ◽  
Progesterone Treatment ◽  
Chicken Oviduct ◽  
Total Dna

1. The effect of simultaneous injections of oestrogen benzoate and progesterone (0.5mg/day each) on immature chicken oviduct tissue pretreated with oestrogen benzoate (0.5mg/day) was studied. 2. After 3 days of treatment with both hormones, the weight of the tissue doubles as compared with tissue treated only with oestradiol benzoate. 3. The progesterone-induced weight increase had no effect on total DNA content of the tissue, but greatly increased the protein/DNA and RNA/DNA ratios. 4. Amino acid incorporation in vivo after progesterone treatment was elevated as measured by using 2h pulses of amino acids; this effect could be accounted for by observed alterations in the concentrations of free amino acids in the tissue. 5. With longer pulses of amino acid the specific radioactivity of total protein remained high in tissue treated with progesterone, at times when specific radioactivity of protein in oestrogen-treated animals was diminishing. 6. From a knowledge of the specific radioactivity of labelled amino acids in the free amino acid pool and in newly synthesized protein, the rate of protein synthesis was estimated in differently treated tissues. 7. The results suggest that progesterone treatment does not cause an increase in protein synthesis. It is concluded that the observed accumulation of oviduct protein is achieved via an effect of progesterone on protein catabolism.

scholarly journals Protein turnover in pulmonary macrophages Utilization of amino acids derived from protein degradation

1981 ◽  
Vol 198 (1) ◽  
pp. 53-65 ◽  
Author(s):  
J A Hammer ◽  
D E Rannels
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Protein Degradation ◽  
Plasma Concentrations ◽  
Specific Radioactivity ◽  
Pulmonary Macrophages ◽  
Protein Synthesis And Degradation ◽  
Synthesis And Degradation ◽  
Extracellular Amino Acid

Conditions were defined under which rates of protein synthesis and degradation could be estimated in alveolar macrophages isolated from rabbits by pulmonary lavage and incubated in the presence of plasma concentrations of amino acids and 5.6 mM-glucose. Phenylalanine was validated as suitable precursor for use in these studies: it was not metabolized appreciably, except in the pathways of protein synthesis and degradation; it entered the cells rapidly; it maintained a stable intracellular concentration; and it was incorporated into protein at measurable rates. When extracellular phenylalanine was raised to a concentration sufficient to minimize dilution of the specific radioactivity of the precursor for protein synthesis with amino acid derived from protein degradation, the specific radioactivity of phenylalanyl-tRNA was only 60% of that of the extracellular amino acid. This relationship was unchanged in cells where proteolysis increased 2.5-fold after uptake and degradation of exogenous bovine serum albumin. In contrast, albumin prevented the decrease in phenylalanine incorporation observed in macrophages deprived of an exogenous source of amino acids. These observations suggested that macrophages preferentially re-utilized amino acids derived from the degradation of endogenous, but not from exogenous (albumin), protein. However, when the extracellular supply of amino acids was restricted, substrates derived from albumin catabolism could support the protein-synthetic pathway.

Evaluation of plasma free amino acid and carnitine levels in patients with cesarean scar pregnancy

2020 ◽  
Vol 23 ◽  
Author(s):  
Omer Tammo ◽  
Hacer Uyanikoglu ◽  
İsmail Koyuncu
Keyword(s):  
Amino Acid ◽  
Pregnant Women ◽  
Free Amino Acid ◽  
Free Amino ◽  
Plasma Concentrations ◽  
Myometrial Invasion ◽  
Controlled Study ◽  
Control Group ◽  
Cesarean Scar Pregnancy ◽  
Cesarean Scar

Aim and Objective: This study aimed to explore the plasma free amino acid (FAA) and carnitine levels in pregnant women with cesarean scar pregnancy (CSP), and to compare them with those of healthy pregnant women. Materials and Methods: This prospective and randomized controlled study was conducted in patients admitted to Harran University Medical Faculty Hospital Obstetrics Clinic between January 2018 and January 2019. A total of 60 patients were included in the study, and the patients were divided into two groups: CSP group (n = 30) and healthy pregnant group as the control group (n = 30). The blood samples were taken from the participants between 7 - 12 weeks of gestation. Twentyseven carnitines and their esters and 14 FAAs were analysed by liquid chromatography – mass spectrometry (LC-MS/MS). Results: The mean plasma concentrations of some carnitines, including C2, C5, C5-OH, C5-DC, C6, C8-1, C12, C14, C14- 1, C14-2, C16, C16-1, C18, and C18-1 were significantly higher in CSP group than in the control group. However, other carnitines, including C0, C3, C4, C4-DC, C5-1, C6-DC, C8, C8-DC, C10, C10-1, C18-1-OH, and C18-2 were similar in both groups. The plasma levels of some FAAs, including Methyl Glutaryl, Leu, Met, Phe, Arg, Orn, and Glu values were significantly higher in CSP group than in the control group. However, there was no statistically significance in other FAA levels, including Val, Asa, Tyr, Asp, Ala, Cit, and Gly between the two groups. Additionally, Pearson’s correlation analysis showed that there were significantly positive correlations between many FAA and carnitine values. Conclusion: Since several plasma carnitine and FAA levels were higher in CSP group than in the control group, we think that scar pregnancy increases metabolic need for myometrial invasion. Also, we think that these results may be useful in clinical practice for CSP diagnosis.

Effect of diet and infusion of volatile fatty acids into the rumen on the concentration of plasma free amino acids in sheep

1982 ◽  
Vol 48 (3) ◽  
pp. 519-526 ◽  
Author(s):  
J. R. Mercer ◽  
E. L. Miller
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Free Amino ◽  
Volatile Fatty Acids ◽  
Plasma Concentrations ◽  
Essential Amino Acids ◽  
Protein Nitrogen ◽  
Post Infusion ◽  
Limiting Amino Acid ◽  
The Mean

1. The effect of supplementing barley diets with urea (U), extracted decorticated groundnut meal (GNM) or Peruvian fish meal (PFM) on plasma free amino acid concentrations in sheep have been examined and the first limiting amino acid has been indicated by measuring the changes in the concentration of the plasma essential amino acids (PEAA) during a rumen infusion of a volatile fatty acid (VFA) mixture.2. Three wethers fitted with rumen and re-entrant duodenal cannulas were given isonitrogenous, isoenergetic diets containing (g/kg dry matter (DM)) U 20, GNM 106 or PFM 78, the crude protein (nitrogen × 6.25) contents being 139, 145 and 148 respectively. The sheep were fed hourly, the mean daily dm intake being 0.634 kg.3. Plasma concentrations of valine, threonine, lysine, isoleucine and leucine were linearly related to their concentrations in duodenal digesta.4. A VFA mixture was infused into the rumen for 6 h to supply (mmol/min) acetate 1.47, propionate 0.22 and n-butyrate 0.27. Blood samples were taken 6 h before, during and 12 h after the end of the infusion.5. The concentration of all PEAA decreased relative to the pre-infusion and post-infusion controls but there were no significant differences between diets.6. The mean decreases in concentration averaged over all three diets showed that the decrease in concentration of methionine (41.5%) was far greater than for any other essential amino acid suggesting that under these conditions methionine was the first limiting amino acid.

scholarly journals The regulation of protein synthesis in the liver of rats. Mechanisms of dietary amino acid control in the immature animal

1968 ◽  
Vol 107 (5) ◽  
pp. 615-623 ◽  
Author(s):  
R. W. Wannemacher ◽  
W. K. Cooper ◽  
M. B. Yatvin
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Specific Radioactivity ◽  
Protein Diet ◽  
Low Protein Diet ◽  
Deficient Diet ◽  
Low Protein ◽  
Rna Content ◽  
Cytoplasmic Rna

Weanling (23-day-old) rats were fed either on an amino acid-deficient diet (6% of casein, which in effect represents an ‘amino acid-deficient’ diet) or on a diet containing an adequate amount of protein (18% of casein) for 28 days. The hepatic cells from the animals fed on the low-protein diet were characterized by low amino acid content, almost complete inhibition of cell proliferation and a marked decrease in cell volume, protein content and concentration of cytoplasmic RNA compared with cells from control rats. The lower concentration of cytoplasmic RNA was correlated with a decreased ribosomal-RNA content, of which a larger proportion was in the form of free ribosomes. The protein-synthetic competence and messenger-RNA content of isolated ribosomes from liver cells of protein-deprived animals were 40–50% of those noted in controls. At 1hr. after an injection of radioactive uridine, the specific radioactivity of liver total RNA was greater in the group fed on the low-protein diet, but the amount of label that was associated with cytoplasmic RNA or ribosomes was significantly less than that noted in control animals. From these data it was concluded that dietary amino acids regulate hepatic protein synthesis (1) by affecting the ability of polyribosomes to synthesize protein and (2) by influencing the concentration of cytoplasmic ribosomes. It is also tentatively hypothesized that the former process may be directly related to the concentration of cellular free amino acids, whereas the latter could be correlated with the ability of newly synthesized ribosomal sub-units to leave the nucleus.

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