Protein Turnover In Liver Research Articles

Dietary L-arginine levels affect the liver protein turnover and alter the expression of genes related to protein synthesis and proteolysis of laying hens

Amino acids are considered to be anabolic factors that affect protein turnover. The aim of this study was to test the effects of dietary L-arginine (Arg) levels on protein metabolism in the liver of laying hens and the expression of genes related to protein synthesis and proteolysis. Xinyang black commercial laying hens (n = 864, 31 wk of age) were randomly allotted to 6 treatments with 4 replicates of 36 birds. The dietary treatments were corn-corn gluten meal based diets containing 0.64, 0.86, 1.03, 1.27, 1.42, and 1.66% L-Arg, respectively. Serum concentrations of total protein and albumin were maximized in the 1.27% L-Arg group, and serum concentration of urea acid was the lowest in the 1.27% L-Arg group. The 1.27% L-Arg group had the highest fractional protein synthesis rate and fractional protein gain rate in the liver. Consistent with the data on protein turnover, mRNA abundances of target of rapamycin (TOR) and ribosomal protein S6 kinase 1 increased in the liver of layers fed 1.27% L-Arg, while mRNA abundances of cathepsin B and 20S proteasome decreased at the same dietary L-Arg level. In conclusion, the dietary level of L-Arg increased the liver fractional protein synthesis rate and fractional protein gain rate of laying hens, and the action of an appropriate level of dietary L-Arg involves upregulating the gene expression of the TOR signaling pathway accompanied by suppressing the mRNA expression of cathepsin B and 20S proteasome in the liver.

A systems approach to determine the effect of changes in gene expression in adipose tissue on productive and reproductive efficiency in dairy cattle

In order to assess the patterns of metabolism that affect efficiency, an existing mechanistic metabolic model of dairy cows (Molly) was expanded to include transcriptional control of metabolism in adipose tissue and reproductive processes. Our objective was to test the effects of changes in gene transcription in adipose tissue on patterns of metabolism, reproductive processes and efficiency. The model describes the substrate sensitivity and maximum velocity for lipogenesis, esterification and lipolysis. Data were collected from first and second parity cows from various studies, and included nutrient intake, milk component output, changes in adipose tissue lipid, visceral and body protein and lipid, and metabolism rates in adipose tissue and gene transcription. The lipogenic pathways were primarily controlled by transcriptional changes, whereas lipolysis varied primarily by post-translational control. We used the difference in transcriptional and post-translational metabolic control to alter Vmax and Ks for lipogenesis and lipolysis; in addition, we modelled a range of energy costs for ion transport and protein turnover in liver, muscle and adipose tissues. Changing rates of lipogenesis and lipolysis (~four-fold) did not affect milk production or feed intake, but altered maintenance costs, fat accretion, energetic efficiency and reproduction. Increases in lipogenesis decreased postpartum interval to first ovulation; increases in rates of lipolysis increased the interval. Changes in metabolic rate from changing either adipose tissue metabolism or rate of ion transport and protein turnover affected rates of oestrogen and progesterone degradation, which affected follicular growth and corpus luteum function. This model may be used to help interpret genomic and transcriptomic data to pinpoint the most effective ways to select and manage for changes in productive and reproductive efficiency.

L-Arginine supplementation in pigs decreases liver protein turnover and increases hindquarter protein turnover both during and after endotoxemia

Accumulating evidence suggests that L-arginine, under conditions of septicemia, not only enhances immune function but also improves protein metabolism. Because the effect of L-arginine administration on the protein metabolism of different organs is unknown, the aim of the study was to elucidate the effects of exogenous supplementation of L-arginine during endotoxemia on the in vivo protein metabolism of individual organs and at the whole-body level. Female pigs were cannulated with catheters in the aorta and the splenic, caval, portal, hepatic, and renal veins, enabling measurements across the hindquarter, portal-drained viscera, liver, and kidneys. Endotoxemia was induced by a 24-h continuous intravenous infusion of endotoxin (3 microg x kg body wt(-1) x h(-1)). At 8 h, an intravenous infusion of L-arginine was started (n = 8). Control pigs (n = 6) received L-alanine. At 24 h, blood was sampled. After cessation of the endotoxin infusion, L-arginine and L-alanine infusions were continued as a supplement in the enterally infused diet. At 48 h, blood samples were obtained during the postendotoxemic and nutritionally supported conditions. Stable isotopes were used to assess protein metabolism and phenylalanine hydroxylation. Both during and after the endotoxin challenge, L-arginine administration enhanced protein synthesis and degradation across the hindquarter and simultaneously reduced protein synthesis and degradation in the liver at equal rates. Protein turnover across the kidneys and portal-drained viscera remained unaffected. After endotoxemia, L-arginine infusion decreased whole-body protein turnover without affecting the net protein balance. L-Arginine administration affects protein turnover of the muscle area and the liver oppositely.

Selective changes in the protein-turnover rates and nature of growth induced in trout liver by long-term starvation followed by re-feeding.

We report upon the effects of a cycle of long-term starvation followed by re-feeding on the liver-protein turnover rates and nature of protein growth in the rainbow trout (Oncorhynchus mykiss). We determined the protein-turnover rate and its relationship with the nucleic-acid concentrations in the livers of juvenile trout starved for 70 days and then re-fed for 9 days. During starvation the total hepatic-protein and RNA contents decreased significantly and the absolute protein-synthesis rate (A(S)) also fell, whilst the fractional protein-synthesis rate (K(S)) remained unchanged and the fractional protein-degradation rate (K(D)) increased significantly. Total DNA content, an indicator of hyperplasia, and the protein:DNA ratio, an indicator of hypertrophy, both fell considerably. After re-feeding for 9 days the protein-accumulation rates (K(G), A(G)) rose sharply, as did K(S), A(S), K(D)), protein-synthesis efficiency (K(RNA)) and the protein-synthesis rate/DNA unit (K(DNA)). The total hepatic protein and RNA contents increased but still remained below the control values. The protein:DNA and RNA:DNA ratios increased significantly compared to starved fish. These changes demonstrate the high response capacity of the protein-turnover rates in trout liver upon re-feeding after long-term starvation. Upon re-feeding hypertrophic growth increased considerably whilst hyperplasia remained at starvation levels.

Tumor growth influences skeletal muscle protein turnover in the pregnant rat.

The implantation of a fast growing tumor (the Yoshida AH-130 ascites hepatoma) to mid-pregnant rats resulted in no changes in fetus weight, in spite of an important body weight decrease observed in the mother. Tumor-bearing pregnant rats showed an accelerated muscle protein degradation that resulted in decreases in both gastrocnemius and soleus muscle weight and protein content. Although very slight changes were observed in liver protein turnover after tumor implantation, muscle protein degradation and ubiquitin gene expression were increased (in relation with the non-tumor-bearing pregnant rats) in the first postimplantation period (0-4 d), whereas it remained lower in the second studied period (4-7 d), compensating for the initial differences when the whole period (0-7 d) was considered. Similar results were observed when muscle protein synthesis was studied. On the whole, tumor growth resulted in a slightly decreased protein accumulation rate. The results presented suggest that the implantation of this tumor in the pregnant rat has little or no consequences in fetal growth but results in an important muscle waste in the mother.

Dose-dependent binding of trichloroethylene to hepatic DNA and protein at low doses in mice

Trichloroethylene (TCE) is a widely used industrial chemical and a low level contaminant of surface and ground water in industrialized areas. It is weakly mutagenic in several test systems and carcinogenic in rodents. However, the mechanism for its carcinogenicity is not known. We investigated the binding of [1,2- 14C]TCE ([ 14C]TCE) to liver DNA and proteins in male B6C3F1 mice at doses more relevant to humans than used previously. The time course for the binding was studied in animals dosed with 4.1 μg [ 14C]TCE/kg body weight (b.w.) and sacrificed between 0.5 and 120 h after i.p. injection. A dose response study was carried out in mice given [ 14C]TCE at doses between 2 μg/kg and 200 mg/kg b.w. and sacrificed 2 h post-treatment. [ 14C]TCE associated with the DNA and protein extracts was measured using accelerator mass spectrometry. The highest level of protein binding (2.4 ng/g protein) was observed 1 h after the treatment followed by a rapid decline, indicating pronounced instability of the adducts and/or rapid turnover of liver proteins. DNA binding was biphasic with the first peak (75 pg/g DNA) at 4 h. However, the highest binding (120 pg/g DNA) was found between 24 and 72 h after the treatment. Dose response curves were linear for both protein and DNA binding. The binding of TCE metabolites to DNA was ca. 100-fold lower than to proteins when calculated per unit weight of macromolecules and when measured 2 h post-exposure. This study shows that TCE metabolites bind to DNA and proteins in a dose-dependent manner in liver, one of the target organs for its tumorigenicity. Thus, protein and DNA adduct formation should be considered as a factor in the tumorigenesis of TCE.

Dietary lysine deficiency greatly affects muscle and liver protein turnover in growing chickens.

We analysed the respective influences of age and lysine deficiency on skeletal muscle and liver protein turnover. Growing male broilers were fed ad libitum on isoenergetic diets containing 200 g crude protein/kg which varied in their lysine content (7.7 or 10.1 g/kg). Fractional rates of protein synthesis (FSR) were measured in vivo in the liver and the pectoralis major muscle of 2-, 3- and 4-week-old chickens (flooding dose of L-[4-3H]phenylalanine). Fractional rates of proteolysis (FBR) were estimated for the same tissues as the difference between synthesis and growth. Over the 2-week period liver FSR and FBR were unchanged, whereas muscle FSR decreased with age. This developmental decline was related to the lower capacity for protein synthesis (Cs) without any modifications of the translational efficiency. Whatever the age, lysine deficiency resulted in significant decreases in body weight, tissue protein content and tissue protein deposition, apparently because of reduced amounts of proteins synthesized. We recorded a difference in the response of the two tissues to lysine deficiency, the pectoralis major being more sensitive than the liver. When comparing birds of the same age, liver FSR and FBR were not modified by the diet, whereas muscle FSR, Cs and FBR were higher in chicks fed on a lysine-deficient diet than in the controls. Conversely, when chicks of similar weights were compared, the main effect of the dietary deficiency was an increase in muscle FBR. The results suggest that lysine deficiency not only delayed chick development so that protein turnover was affected, but also induced greater changes in metabolism. Thus, the principal mechanism whereby muscle mass decreased appeared to be a change in FBR.

Age-Related Changes of Protein Turnover in Specific Tissues of the Chick

The effect of age on muscle and liver protein turnover was analyzed in growing broilers. Protein turnover was compared in the Pectoralis major (PM), the Anterior Latissimus dorsi (ALD), and the Sartorius (SART) muscles, as well as in the liver in broilers at three ages (2, 3, and 4 wk). Protein fractional synthesis rates (FSR, in percentage per day) were measured by a flooding dose of L-[4-3 H] Phe. Protein fractional breakdown rates (FBR, in percentage per day) were estimated as the difference between FSR and the gain rates of tissue protein. When expressed in absolute rates (grams per day), tissue protein deposition increased during chick growth (approximately 1.7-fold, whatever the tissue). This phenomenon was accompanied by increased protein synthesis (ASR) and proteolysis (ABR) in the PM and in the ALD muscles as well as in the liver. However, ASR and ABR did not seem to be significantly modified in SART. The FSR in skeletal muscles significantly decreased with age. This developmental decline was associated with decreases in RNA content and RNA:protein ratio (Cs, capacity for protein synthesis). In contrast to muscle, we found no significant variation in liver FSR between 2 and 4 wk of age. In addition, liver Cs and translational efficiencies were not decreased. Finally, in muscle as in liver, FBR did not show any clear age-related pattern.

In vivo amino acid metabolism of gut and liver during short and prolonged starvation.

During starvation, splanchnic organs are proportionally more affected by protein loss than other organs. Amino acid membrane transport is one of the regulating mechanisms of protein turnover, but until now in vivo data were lacking. To study in vivo phenylalanine and tyrosine membrane transport and protein turnover in splanchnic organs, a primed continuous infusion of L-[2,6-3H]phenylalanine was given to control rats (postabsorptive) and after short (40 h) and prolonged (112 h) starvation. Data were analyzed using a three-compartment model previously used in muscle membrane transport studies. Inward and outward amino acid plasma-tissue membrane transport rates in both the liver and gut were upregulated after prolonged starvation. Metabolic shunting of phenylalanine and tyrosine increased in the gut but decreased to zero in the liver after prolonged starvation. In conjunction with this, gut and liver protein turnover increased after prolonged starvation. In the liver the net uptake of gluconeogenic precursors also increased, indicative for increased gluconeogenesis. The observed changes in amino acid metabolism in both splanchnic organs after prolonged starvation may reflect an adaptation of the gut and liver to nutritional deprivation and could be of benefit during refeeding.

ZONATION OF TOTAL AND PLASMA PROTEIN TURNOVER IN RAT LIVER: EFFECTS OF CHRONIC TREATMENT

To study the acinar distribution of protein turnover, rats were given a tracer dose of 14C-valine intraperitoneally. At 30 min or 20 hr later, livers were perfused in situ and periportal (pp) or perivenous (pv) cell lysates were collected after digitonin pulse infusion. In control rats the post-label radioactivity of cell lysates showed no zonation. The radioactivity of the pp and pv cell lysates was significantly higher in livers of chronically ethanol-fed rats than in the controls, and the increase was more prominent in the pp region. Furthermore, after chronic ethanol feeding, an enrichment of plasma protein in the pv region was observed. The results support the view that in the control rats liver protein turnover is not zonated. Chronic ethanol feeding may, however, induce such zonation, as evidenced by both a slower rate of protein turnover in the pp cells and preferential pv accumulation of plasma proteins.

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

Different methods used for measuring protein turnover in liver and skeletal muscle are described, with special emphasis on technical and practical aspects and the advantages and limitations of different techniques. In the first part of the review, the concept of precursor specific radioactivity and its importance for accurate determination of protein synthesis rate is discussed. In the second part, different in vivo techniques for protein turnover measurements are reviewed, including continuous administration of tracer amino acid, flooding dose technique, indirect measurement of protein synthesis, and estimation of protein degradation in vivo. In the third part of the report, in vitro techniques are described, including measurement of protein turnover in incubated liver slices, perfused liver, isolated hepatocytes, incubated isolated muscles or muscle biopsies, and perfused rat hemicorpus. In vivo techniques are preferred when accurate absolute values of protein turnover rates are desired. In vitro techniques offer the advantage of standardized conditions, maintaining strict control of substrate and hormone concentrations, and eliminating complicating interactions with other tissues. For several in vitro techniques, a good correlation has been demonstrated between relative changes in protein turnover in vitro and in vivo in different conditions.

Effect of protein starvation on protein turnover in liver, oviduct and whole body of laying hens

1. 1. Whole body protein turnover rates in White Leghorn laying hens were reduced by protein starvation for 7 days, followed by complete restoration by protein repletion for 7 days. 2. 2. Protein starvation considerably reduced fractional and absolute rates of protein synthesis both in the liver and, to greater extent, in the oviduct. 3. 3. It was suggested that a considerable portion of the reduced whole body protein synthesis could be accounted for by the reduced protein synthesis in these organs when laying hens were subjected to protein starvation.

Effect of the Level of Dietary Lysine on Turnover Rate of Liver Protein in the Chick

The effect of .36, .86, and 1.24% dietary lysine on the rate of turnover of liver proteins in chicks in the first 15 days of life was studied. Half-lives of 18.4 ± .2, 5.7 ± 1.0, and 6.3 ± .1 days corresponding to 26.6 ± .2, 8.2 ± 1.4, and 9.1 ± .1 days turnover time and 3.8 ± .02, 12.3 ± 1.9, and 11.0 ± .2% net daily degradation for liver proteins of chicks fed .36, .86, and 1.24% dietary lysine, respectively, were obtained. Turnover rate of the liver proteins increased in a manner similar to the gain in body weight with the levels of dietary lysine.

EFFECTS OF α-KETOISOCAPROATE AND OF LEUCINE ON NITROGEN METABOLISM IN POSTOPERATIVE PATIENTS

21 patients undergoing major abdominal surgery were randomly assigned to one of three groups. On the day of surgery and for the succeeding 4 days each group received a daily infusion of one of the following: 10 g glucose plus 70 mmol NaHCO 3, 70 mmol leucine plus 70 mmol NaHCO 3, or 70 mmol of sodium α-ketoisocaproate (KIC). No other calories were given. Leucine infusions had no significant effect on nitrogen (N) balance, 3-methylhistidine excretion, or plasma concentrations of pre-albumin or retinol-binding protein, but they increased blood acetoacetate concentration (p=0·004). N balance was less negative (p = 0·002) and 3-methylhistidine excretion lower (p = 0·002) in the group receiving KIC than in those receiving glucose. Blood ketone bodies, plasma pre-albumin, and plasma retinol-binding protein concentrations at the end of the study were significantly higher in the KIC group than in the others. These N-sparing effects of KIC may be related to the heightened ketosis that followed its administration, to suppression of protein degradation, or to an effect on liver protein turnover. KIC alone in small doses diminishes N wastage in postoperative patients but under the same conditions leucine does not.

Turnover of liver proteins in Phenoclor DP6-treated rats

The effect of a diet containing Phenoclor DP6 on the average turnover of protein in subcellular fractions of the liver was studied in young rats. In experiment A, rats were given an intraperitoneal injection of [ 14 C]- guanidinolabeled arginine, were then fed a diet containing 100 ppm of Phenoclor DP6 for 24 h, and then resumed a control diet. In experiment B rats were fed a DP6 diet for 4 days prior to receiving the isotope injection, and then resumed the DP6 diet for the entire experiment. Afterward, these rats were killed 1, 3, 5, and 8 days after pulse-labeling. Mitochondrial, microsomal, microsomal membrane, and ribosomal fractions were prepared from their livers, and these fractions were analyzed for acid-precipitable protein and radioactivity. Our results indicate no significant difference in the average rates of 14 C degradation between rats receiving a single DP6 dose and the control group. In rats ingesting a DP6 diet every day, the half-lives of total liver proteins were significantly shorter than in the control group. This difference is perhaps due to the rate of breakdown of the microsomal proteins. We found that in the microsomal fraction only the microsomal membrane proteins were affected by the DP6 treatment. Our results suggest that under our experimental conditions microsomal membrane protein metabolism (synthesis and breakdown) is enhanced by DP6 ingestion.

The State of Lysosomes and Protein Turnover in Rat Liver Effect of Excess Vitamin A

Administration of excess vitamin A to rats induces labilization of liver lysosomal membranes, as shown by the release of lysosomal cathepsins upon tissue homogenization. The effect of lysosomal labilization on liver protein turnover was investigated. The apparent turnover rate of liver proteins in the hypervitaminotic animals, as measured by a double isotope-labeling technique (Glass and Doyle (1972) J, Biol. Chem, 247, 5234-5242), was found to be the same as that in control animals. Neutral and alkaline fructose-1, 6-biphosphatase [EC 3.3.3.11] activities in the liver were also found to be unchanged in hypervitaminosis A. These data indicate that the rate of intracellular protein degradation is not determined by the level of "free cathepsins. Protein synthesis in the livers of the hypervitaminotic animals was partially imparied, as shown by the shift of polysomal profiles toward lighter aggregates.

Measurement of Protein Turnover in Rat Liver with [14C]Carbonate: PROTEIN TURNOVER DURING LIVER REGENERATION

Abstract Several isotopic methods for the measurement of liver protein turnover were compared in a study of plasma albumin and the mixed hepatic proteins during liver regeneration. In albumin after the administration of [14C]carbonate, the rate of decay of the radioactivity of the intact protein was similar to that of arginine isolated therefrom, whereas after the administration of [guanidino-14C]arginine, the decay rate was 30% slower than that obtained with [14C]carbonate. When [guanidino-14C]arginine is given, the extrahepatic tissues, such as muscle and the plasma globulins, are labeled more heavily than the liver proteins. It is likely that the reutilization of this extrahepatic arginine by the liver accounts for the slower disappearance of radioactivity from albumin. Similar results were obtained with the mixed, indigenous hepatic proteins. In animals that had been partially hepatectomized, the decay in radioactivity in arginine after [14C]carbonate administration showed that the rate of protein breakdown was reduced by 30 % during the first 5 days of liver regeneration. There was no decay in radioactivity in similar animals given [guanidino-14C]arginine, suggesting a massive transfer of labeled extrahepatic protein arginine into the liver during regeneration.

Effects of Glucagon on General Protein Degradation and Synthesis in Perfused Rat Liver

Abstract The direct influence of glucagon on rates of valine incorporation into and release from protein was assessed in the perfused rat liver. Glucagon alone significantly increased net valine release, an effect which was ascribed largely to the stimulation of proteolysis. The magnitude of this increase in absolute terms remained virtually unchanged in the presence of an amino acid mixture that suppressed proteolysis. The increase was also obtained in the presence of sufficient cycloheximide to inhibit valine incorporation by 93 %. In addition, free intracellular valine was elevated relative to external values over an 8-fold range of perfusate valine concentration. Rates of valine incorporation into liver protein were reduced by glucagon alone to a small but significant degree. Of interest, however, was the finding that the inhibition was potentiated by additions of amino acids. In the presence of a complete amino acid mixture, simulating the composition of plasma amino acids at 10 times their normal concentrations, glucagon strongly depressed valine incorporation, and the rate was comparable to the value obtained previously when the amino acid supply was severely limited. These findings suggest that the turnover of liver proteins is regulated by glucagon at sites of both protein degradation and synthesis.

Synthesis and turnover of liver proteins during extrahepatic cholestasis in the rat.

The protein-bound radioactivity in liver and kidney, heart and brain was measured 30 and 90 min after an intraperitoneal injection of 3H-leuine. The incorporated radioactivity was slightly higher in liver and in brain proteins in cholestatic animals after 30 min, while no appreciable difference was observed after 90 min. A higher labeling was seen in serum proteins at 30 min. The trichloroacetic acid soluble radioactivity in liver, serum and other organs was also measured. The effects of cholestasis on the rate of degradation of liver protein were measured by following the decay in radioactivity after single pulse labeling and by the application of the double isotope technique. The results indicated a slightly lowered half-life of cholestatic proteins. The results are discussed in connection with previous findings of highly increased amino acid incorporation in vitro.

Incorporation of cystine into the proteins of regenerating wound tissue

A procedure to distinguish between cystine bound into protein by peptide bonds and that associated with the protein as prosthetic groups by disulfide bonds only has been devised. This distinction is achieved by reaction of the cysteine and cystine in the protein with sodium sulfite in alkaline medium and simultaneous separation of the resulting thiosulfate derivatives by dialysis. Of the total cystine in regenerating wound tissue proteins, 6.6% is bound by disulfide bonds only. This fraction of the half-cystine residues turns over very much more rapidly than do the cystine residues in the protein bound by peptide bonds. The formation and turnover of liver proteins, as indicated by cystine-S 35 incorporation, is much more rapid in wounded than in normal rats. Wounding also appears to have the effect of increasing the proportion of half-cystine residues linked to the liver proteins by disulfide bonds only.