Plasma reciprocal pool specific activity predicts that of intracellular free leucine for protein synthesis

1989 ◽  
Vol 257 (3) ◽  
pp. E385-E399 ◽  
Author(s):  
F. F. Horber ◽  
C. M. Horber-Feyder ◽  
S. Krayer ◽  
W. F. Schwenk ◽  
M. W. Haymond
Keyword(s):  
Protein Synthesis ◽  
Striated Muscle ◽  
Specific Activity ◽  
Intracellular Space ◽  
Tissue Compartments ◽  
Venous Plasma

We previously proposed that, during the infusion of either labeled leucine or its alpha-ketoacid, alpha-ketoisocaproate (KIC), the plasma specific activity (SA) of the transaminated product of the infused tracer ("reciprocal pool SA") may better reflect the intracellular leucine SA than the plasma SA of either infused tracer ("primary pool SA"). To test this hypothesis, 14 dogs were simultaneously infused intravenously with [3H]leucine and [14C]KIC, and blood and tissue compartments were sampled. The ratios of [3H]-leucine to [14C]leucine [( 3H]/[14C]leucine) in mixed tissue proteins and in the intracellular space of striated muscle were the same as the ratio of the isotope infusion rates and similar, although slightly lower (P less than 0.01), than [3H]KIC/[14C]leucine SA (ratio of reciprocal pool SA) in plasma. Plasma [3H]KIC/[14C]leucine SA were essentially identical to the [3H]/[14C] of leucine in 1) mixed liver proteins, 2) intrahepatic free leucine, and 3) fibrin. The [3H]/[14C]leucine in mixed renal proteins and in the intracellular space of kidney and erythrocytes were similar to those of the venous plasma [3H]/[14C]leucine SA. The plasma [3H]KIC and [14C]leucine SA (the reciprocal pool SA) were similar to the SA of [3H]- and [14C]leucine in the intracellular space of all organs investigated with the exception of kidney. Therefore, in postabsorptive dogs, the plasma SA of the transaminated product of the infused labeled KIC or leucine is an excellent predictor of the intracellular leucine SA in all tissues investigated with the exception of kidney.

Stimulation of transport of alpha-aminoisobutyric acid into the testes of immature mice in vivo by follicle-stimulating hormone

1982 ◽  
Vol 100 (1) ◽  
pp. 137-142
Author(s):  
Nila Oza ◽  
Sarah J. Meanock ◽  
A. G. Davies
Keyword(s):  
Protein Synthesis ◽  
Amino Acid ◽  
Amino Acid Transport ◽  
Specific Activity ◽  
Follicle Stimulating Hormone ◽  
Acid Transport ◽  
Aminoisobutyric Acid ◽  
Old Mice ◽  
Stimulation Of

Abstract. Groups of immature mice were injected sc with radiocarbon-labelled alpha-aminoisobutyric acid (AIB) after being given a single sc injection of hFSH or of 0.9% saline. As an index of the transport of AIB, the specific activity of isotope was measured in homogenates of testis and of liver. FSH treatment caused statistically significant increases in the specific activity of isotope in the testes and in the ratio of testicular to liver specific activity. The effect was greatest in 9-day-old mice injected with FSH 16 h before removal of the testes. Uptake of labelled AIB was not stimulated after administration of hCG or testosterone. Doses of cycloheximide sufficient to reduce the rate of protein synthesis by over 99% did not impair testicular uptake of labelled AIB or the influence of FSH on AIB uptake. These results suggest that FSH stimulates amino acid transport into cells of the immature testis and that this action is independent of the stimulatory effect of FSH on testicular protein synthesis.

Measurement of the Rate of Protein Synthesis in Muscle of Postabsorptive Young Men by Injection of a ‘Flooding Dose’ of [1-13C]leucine

1989 ◽  
Vol 77 (3) ◽  
pp. 329-336 ◽  
Author(s):  
Peter J. Garlick ◽  
Jan Wernerman ◽  
Margaret A. McNurlan ◽  
Pia Essen ◽  
Gerald E. Lobley ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Amino Acid ◽  
Muscle Protein ◽  
Muscle Protein Synthesis ◽  
Isotopic Enrichment ◽  
Plasma Leucine ◽  
Convenient Technique ◽  
Tissue Compartments ◽  
Muscle Biopsies

1. The ‘flooding dose’ technique for measuring the rate of protein synthesis in tissues in vivo involves the injection of a large amount of unlabelled amino acid together with the tracer to minimize differences in isotopic enrichment of the free amino acid in plasma and tissue compartments. This approach has been investigated in human muscle by taking biopsies from postabsorptive male volunteers given [1-13C]leucine. 2. Intravenous injection of 4 g of unlabelled leucine resulted in a rapid rise in free leucine concentration of seven- to eleven-fold in plasma and five-fold in muscle. Values were still elevated by two-fold after 2 h. 3. Five minutes after injection of [1-13C]leucine (0.05 g/kg) the isotopic enrichment of plasma leucine was 82% that of the injected material, falling to 44% at 120 min. The enrichment of free leucine in sequential muscle biopsies was close to that in plasma and almost identical to that for plasma α-ketoisocaproate. 4. The rate of protein synthesis was determined from the increase in leucine enrichment in protein of muscle biopsies taken before and 90 min after injection of [1-13C]leucine (0.05 g/kg; 19 or 39 atom% excess) and the average plasma α-ketoisocaproate enrichment over this period (taken to represent muscle free leucine). The mean rate of muscle protein synthesis in 10 subjects was 1.95 (sem 0.12)%/day. Rates of protein synthesis calculated from plasma leucine as precursor enrichment were only 5% lower than those calculated from plasma α-ketoisocaproate. 5. It is concluded that a ‘flooding dose’ of 13C-labelled amino acid is a useful and convenient technique for determining the rate of protein synthesis in tissues of human volunteers and patients.

Insulin stimulation of protein synthesis in cultured skeletal and cardiac muscle cells

1982 ◽  
Vol 243 (1) ◽  
pp. C81-C86 ◽  
Author(s):  
J. Airhart ◽  
J. A. Arnold ◽  
W. S. Stirewalt ◽  
R. B. Low
Keyword(s):  
Protein Synthesis ◽  
Specific Activity ◽  
Muscle Cells ◽  
Cell Types ◽  
Cardiac Cells ◽  
Cardiac Muscle Cells ◽  
Significant Stimulation ◽  
Chick Heart ◽  
Embryonic Chick Heart ◽  
Stimulation Of

The effects of acute exposure to insulin on protein synthesis were examined in primary, differentiated cultures of embryonic chick heart and skeletal muscle cells. Synthetic rates were calculated using the specific activity of tRNA-bound leucine as precursor, a specific activity that was significantly less than that of extracellular leucine but greater than that of free, intracellular leucine at 0.2 mM external leucine. Insulin did not alter these relationships. Doses of insulin in the physiological range produced significant stimulation of protein synthesis in both cell types. Maximal responses, involving approximately 30% increases in both absolute and fractional rates, were observed at higher insulin concentrations. Significant stimulation by insulin was seen in cardiac cells after only 1 h of insulin treatment, and the effects of the hormone were observed both in the presence and absence of serum in the culture medium.

Regulation of cardiac protein balance by hydrocortisone: interaction with insulin.

1978 ◽  
Vol 234 (3) ◽  
pp. E306
Author(s):  
E E Griffin ◽  
K Wildenthal
Keyword(s):  
Protein Synthesis ◽  
Protein Degradation ◽  
Cathepsin D ◽  
Specific Activity ◽  
Branched Chain Amino Acids ◽  
Protein Balance ◽  
Net Uptake ◽  
Specific Activities ◽  
Branched Chain ◽  
Net Release

In fetal mouse hearts in organ culture the rate of protein synthesis was substantially reduced and the rate of protein degradation slightly increased by hydrocortisone in the absence of insulin, but in the presence of insulin the steroid caused a small increase in protein synthesis and a significant reduction in protein degradation. Hydrocortisone promoted the net uptake (or reduced the net release) of branched-chain amino acids independent of insulin and independent of simultaneous changes in protein balance. The specific activities of the lysosomal enzymes cathepsin D and glucosaminidase were reduced by hydrocortisone in all media, whereas the specific activity of creatine kinase increased when the medium contained insulin but decreased in the absence of insulin. It is concluded that hydrocortisone regulates cardiac protein balance via alterations both in synthesis and in degradation. Some of the hormone's myocardial effects are influenced by insulin so that hydrocortisone is anabolic in its presence but catabolic in its absence.

Transport of oestrone sulphate by the mammary gland in the goat

1984 ◽  
Vol 101 (2) ◽  
pp. 221-230 ◽  
Author(s):  
R. B. Heap ◽  
M. Hamon ◽  
I. R. Fleet
Keyword(s):  
Mammary Gland ◽  
Specific Activity ◽  
Kinetic Studies ◽  
Total Radioactivity ◽  
Oestrone Sulphate ◽  
Lactating Goats ◽  
Endogenous Compound ◽  
Tracer Kinetic ◽  
Arterial Plasma ◽  
Venous Plasma

ABSTRACT During pregnancy in goats the concentration of endogenous oestrone sulphate in milk increased more than twofold, and that in arterial and mammary venous plasma 10- and 20-fold respectively. The concentration in milk was higher than that in arterial plasma, particularly in lactating goats during mid-gestation. This was partly related to mammary production of oestrone sulphate (or of a closely related steroid which cross-reacted in the radioimmunoassay) since in tracer infusion studies the specific activity of oestrone sulphate in milk was significantly lower than that in arterial or mammary venous plasma. It was also related to the existence of a mechanism within the gland which concentrates oestrone sulphate in milk since when infused close-arterially into the mammary gland of a non-pregnant goat with undetectable levels of the endogenous compound in the circulation, a concentration ratio of 7·4:1·0 was reached for oestrone sulphate in milk: arterial plasma. Tracer kinetic studies showed that mammary extraction of [3H]oestrone sulphate was variable (up to 41·3 ± 30·6%, mean ± s.e.m.). During intravenous or close-arterial infusion, radioactivity in arterial and mammary venous plasma at steady state was mainly in the form of [3H]oestrone sulphate (range, 64± 10·6 to 80·2 ± 5·9% of total radioactivity in plasma). The remainder was in the form of compounds chromatographically similar to oestradiol-17β-3-monosulphate, oestradiol-17α-3-monosulphate and unconjugated oestrogens. The distribution of radioactivity between these different steriods was similar in arterial mammary venous plasma indicating a low level of selective mammary metabolism or extraction. The amount of labelled oestrone sulphate transferred into milk was low, and it was significantly less in pregnant (range, 0·11 ±0·07 to 0·27 ± 0·16% of total infusate) than in non-pregnant animals (3·23±0·50%). Studies of the rate of transfer of [3H]oestrone sulphate from blood to milk indicated the presence of a transcellular route with peak activity in milk occurring about 110 min after the start of the infusion. J. Endocr. (1984) 101, 221–230

scholarly journals Adaptation of the rat to a low-protein diet: the effect of a reduced protein intake on the pattern of incorporation of L-[14C]lysine

1966 ◽  
Vol 20 (3) ◽  
pp. 461-484 ◽  
Author(s):  
J. C Waterlow ◽  
Joan M. L. Stephen
Keyword(s):  
Protein Synthesis ◽  
Protein Intake ◽  
Specific Activity ◽  
Muscle Protein ◽  
Normal Diet ◽  
Internal Organs ◽  
Low Protein ◽  
Total Body ◽  
Protein Free Diet ◽  
Total Muscle

1. Rats were chronically depleted of protein by being kept on a 6% casein diet for 5–8 weeks. Control rats were fed on a normal diet. Both groups were injected intraperitoneally with L-[U-14C]lysine. Some rats from each group were then put on a protein-free diet to produce acute depletion. The animals were killed 3 days after the injection. 2. The organs and tissues were analysed for total nitrogen and radioactivity. Free lysine, total amino N and specific activity of free lysine were measured in muscle, liver and serum. The total muscle mass of the animals was determined. Samples of muscle and skin were fractionated and the sp. ac. of the fractions was measured. 3. The main loss of N in acute depletion was found in the viscera and carcass residue; the percentage of total body N contributed by muscle was increased in protein-depleted rats. 4. The depleted rats retained relatively more radioactivity in the internal organs and less in the carcass than normal rats. 5. The ratio of the sp. ac. in protein-bound lysine to the sp. ac. of free lysine showed that protein synthesis was reduced in the muscle of the protein-depleted rats, although there was no decrease in the amount or sp. ac. of free lysine even in severe depletion. 6. Sarcoplasmic and fibrillar proteins of muscle were equally affected by protein depletion, but there was some indication of a preferential decrease in protein synthesis in one of the skin fractions. 7. The results for muscle protein are compared with those given in the literature for liver proteins. It is suggested that the rat adapts to a low-protein intake by an alteration in the pattern of protein synthesis.

scholarly journals Properties and developmental regulation of an α-d-galactosidase from Dictyostelium discoideum

1976 ◽  
Vol 158 (2) ◽  
pp. 409-417 ◽  
Author(s):  
D C Kilpatrick ◽  
J L Stirling
Keyword(s):  
Protein Synthesis ◽  
Dictyostelium Discoideum ◽  
Early Development ◽  
Subcellular Distribution ◽  
De Novo ◽  
Specific Activity ◽  
Soluble Fraction ◽  
Developmental Regulation ◽  
Cellular Slime Mould ◽  
Cellular Slime

An alpha-D-galactosidase was detected in cells of the cellular slime mould, Dictyostelium discoideum, at all stages of development. Its specific activity was highest during early development (interphase), and this accumulation of enzyme appears to require protein synthesis de novo. Its subcellular distribution differs from that of other D. discoideum glycosidases, since most activity was recovered in the soluble fraction. No evidence was obtained for more than one isoenzymic form after subjection of extracts to electrophoresis and various chromatographic procedures. It is excreted from the cell during development, but no evidence was found for an extracellular function for the enzyme.

Effect of glutamine on leucine metabolism in humans

1996 ◽  
Vol 271 (4) ◽  
pp. E748-E754 ◽  
Author(s):  
R. G. Hankard ◽  
M. W. Haymond ◽  
D. Darmaun
Keyword(s):  
Protein Synthesis ◽  
Protein Degradation ◽  
Protein Turnover ◽  
Specific Activity ◽  
Excretion Rate ◽  
The Other ◽  
Anabolic Effect ◽  
Protein Breakdown ◽  
Glutamine Concentration ◽  
Putative Protein

The aim of this study was to determine whether the putative protein anabolic effect of glutamine 1) is mediated by increased protein synthesis or decreased protein breakdown and 2) is specific to glutamine. Seven healthy adults were administered 5-h intravenous infusions of L-[1-14C]leucine in the postabsorptive state while receiving in a randomized order an enteral infusion of saline on one day or L-glutamine (800 mumol.kg-1.h-1, equivalent to 0.11 g N/kg) on the other day. Seven additional subjects were studied using the same protocol except they received isonitrogenous infusion of glycine. The rates of leucine appearance (RaLeu), an index of protein degradation, leucine oxidation (OxLeu), and nonoxidative leucine disposal (NOLD), an index of protein synthesis, were measured using the 14C specific activity of plasma alpha-ketoisocaproate and the excretion rate of 14CO2 in breath. During glutamine infusion, plasma glutamine concentration doubled (673 +/- 66 vs. 1,184 +/- 37 microM, P < 0.05), whereas RaLeu did not change (122 +/- 9 vs. 122 +/- 7 mumol. kg-1.h-1), OxLeu decreased (19 +/- 2 vs. 11 +/- 1 mumol.kg-1.h-1, P < 0.01), and NOLD increased (103 +/- 8 vs. 111 +/- 6 mumol. kg-1.h-1, P < 0.01). During glycine infusion, plasma glycine increased 14-fold (268 +/- 62 vs. 3,806 +/- 546 microM, P < 0.01), but, in contrast to glutamine, RaLeu (124 +/- 6 vs. 110 +/- 4 mumol. kg-1.h-1, P = 0.02), OxLeu (17 +/- 1 vs. 14 +/- 1 mumol.kg-1.h-1, P = 0.03), and NOLD (106 +/- 5 vs. 96 +/- 3 mumol.kg-1.h-1, P < 0.05) all decreased. We conclude that glutamine enteral infusion may exert its protein anabolic effect by increasing protein synthesis, whereas an isonitrogenous amount of glycine merely decreases protein turnover with only a small anabolic effect resulting from a greater decrease in proteolysis than protein synthesis.

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