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Autoimmune hepatitis (AIH) is a chronic inflammatory condition of the liver characterized by parenchymal destruction, hypergammaglobulinemia, specific autoantibody production, and hepatic fibrosis and necrosis. Murine models of AIH have been described; however, little is known about the immunologic mechanisms of tissue destruction. In this study, a new murine model of type 2 AIH was developed using recombinant human cytochrome P450 (CYP) 2D6 emulsified with complete Freund's adjuvant (CFA). BALB/c mice were immunized with 2 μg/mL i.p. of CYP2D6 in CFA. The control group received CFA or phosphate-buffered saline alone. Alanine aminotransferase activity, autoantibody production, IgG concentrations, histologic damage, and specific T-cell response were evaluated. Persistent AIH, characterized by cellular infiltration, hepatic fibrosis, elevated alanine aminotransferase, and the production of anti-liver kidney microsomal antibody type 1 developed in CFA/CYP2D6-immunized mice. This group presented high levels of IgG and its subclasses IgG1, IgG2a, and IgG2b against liver self-proteins. Interestingly, IL-2+ and interferon γ-positive Cyp2d6-specific T cells were present in greater concentrations in mice immunized with CFA/CYP2D6 compared with controls. Immunization with CFA, in combination with a natural human autoantigen like CYP2D6, was demonstrated to break tolerance, resulting in a chronic form of autoimmune-related liver damage. This murine model of type 2 AIH is expected to be instrumental in understanding the immunologic mechanisms of the pathogenesis of this autoimmune liver disease.

Sucralose, one kind of "sugar-free" artificial sweeteners, is widely used as food and drinks additives. It is generally considered that sucralose is safe because majority of ingested sucralose is not metabolized and absorbed by human body. However, increasing evidence shows the negative effects of artificial sweeteners in perturbations of gut microbiota which plays an important role in a variety of processes related to host health such as immune system development. Specifically, sucralose uptake can alter the homeostasis of mouse gut microbiota, resulting in the significant changes of gut bacterial genera diversity, metabolic patterns, and fecal metabolite profiles as well as inducing host liver inflammation. Therefore, there is a need to study liver proteome changes which may be potentially affected by sucralose-induced dysbiosis. In this study, isobaric labeling-based quantitative proteomics was performed to reveal the liver functional proteome changes in male C57BL/6J mice with sucralose administration in drinking water for six-month period. The labeled tryptic peptides were off-line fractionated before LC-MS/MS analysis to improve proteome coverage detected. SIGNIFICANCE: We demonstrated the first quantitative proteomics for mice liver proteome to evaluate the effect of sucralose consumption. In total, >5700 protein groups were identified from 18 mouse liver tissues (9 from control group; 9 from sucralose-treated group), and 4327 protein groups were quantified in all samples without any missing values. Among them, 113 protein groups were identified with statistical significance (q value

This work aimed at exploring the potential of partial substitution of the back-fat with fat mimetic, a novel polysaccharide-protein complex of xanthan gum and succinylated chicken liver protein, to improve the physicochemical characteristics and fatty acid composition of emulsified sausages. The quality characteristics of the sausages demonstrated that low-fat emulsified sausages with the addition of fat mimetic or its sunflower oil pre-emulsion presented similar microstructure and texture to high-fat sausages, while pre-emulsion increased the gel strength and whiteness of products. The rheological behavior characterization indicated that fat replacers caused a significant increase in storage modulus and loss modulus, explaining well the decreased cooking loss and the improved emulsification stability, water holding capacity (WHC) and heating stability. Although the back-fat replacement led to a great decrease in fatty acids, the nutritional values of fatty acids were optimized due to the incorporation of sunflower oil, as revealed by a 10% increase of polyunsaturated fatty acids and the decreased TBARS values within the storage. This study suggests the technological feasibility to simultaneously improve the quality and nutritional features of emulsified sausages with partial back-fat replacement by the proposed polysaccharide-protein complex, as well as the way to achieve chicken liver valorization.

Background: The phosphorylation of eukaryotic initiation factor 2 (p-eIF2) during dietary amino acid insufficiency reduces protein synthesis and alters gene expression via the integrated stress response (ISR).Objective: We explored whether a Met-restricted (MR) diet activates the ISR to reduce body fat and regulate protein balance.Methods: Male and female mice aged 3-6 mo with either whole-body deletion of general control nonderepressible 2 (Gcn2) or liver-specific deletion of protein kinase R-like endoplasmic reticulum kinase (Perk) alongside wild-type or floxed control mice were fed an obesogenic diet sufficient in Met (0.86%) or an MR (0.12% Met) diet for ≤5 wk. Ala enrichment with deuterium was measured to calculate protein synthesis rates. The guanine nucleotide exchange factor activity of eIF2B was measured alongside p-eIF2 and hepatic mRNA expression levels at 2 d and 5 wk. Metabolic phenotyping was conducted at 4 wk, and body composition was measured throughout. Results were evaluated with the use of ANOVA (P < 0.05).Results: Feeding an MR diet for 2 d did not increase hepatic p-eIF2 or reduce eIF2B activity in wild-type or Gcn2-/- mice, yet many genes transcriptionally regulated by the ISR were altered in both strains in the same direction and amplitude. Feeding an MR diet for 5 wk increased p-eIF2 and reduced eIF2B activity in wild-type but not Gcn2-/- mice, yet ISR-regulated genes altered in both strains similarly. Furthermore, the MR diet reduced mixed and cytosolic but not mitochondrial protein synthesis in both the liver and skeletal muscle regardless of Gcn2 status. Despite the similarities between strains, the MR diet did not increase energy expenditure or reduce body fat in Gcn2-/- mice. Finally, feeding the MR diet to mice with Perk deleted in the liver increased hepatic p-eIF2 and altered body composition similar to floxed controls.Conclusions: Hepatic activation of the ISR resulting from an MR diet does not require p-eIF2. Gcn2 status influences body fat loss but not protein balance when Met is restricted.

International audience; Objectives Severely low-protein diets (LP) induce behavioral and metabolic changes including a decrease in body weight, an increase in relative food intake (FI) and alterations in hepatic metabolism. During such protein restriction, changes in hepatic anabolic and catabolic protein pathways could transitory participate to compensate for amino acid (AA) deficiency. In the present study, liver expression of gene involved in proteosynthesis and proteolysis pathways, were related to FI, blood AA levels and body composition in rats fed LP diet. Methods Growing rats were fed for three weeks different diets containing 3-5-8-12-15 or 20% energy of milk protein. Body weight and FI were measured daily. At the end of the experiment, tissues and biological fluids were removed for gene expression measurement and blood AA UPLC analysis. Statistical analysis was done by 1- or 2-factor ANOVA, when data were repeated. Results Despite an increase in relative food intake under P3 and P5% diets, P3, P5 and P8% diets resulted in significant growth retardation compared to other groups. Lean mass was significantly decreased in rats under P3, P5 and P8% compared to P12, P15 and P20% diets, while there was no difference in fat mass between all groups. P3, P5 and P8% diets induced a decrease in essential amino acid concentrations in portal vein, whereas there was no significant difference between groups in veina cava. Severely protein restricted P3% and P5% diets induced an increase in hepatic gene expression involved in proteolysis as calpain 2 and ubiquitin, and an activation of ATF4-CHOP-TRB3 pathway. Conclusions These results suggested that under severe protein restriction, hepatic protein catabolism became a source of plasma amino acid that could partially compensate for the AA not provided by the diet. These observations confirm that liver plays a major role in the adaptation of the body to dietary protein restriction and highlight that severe dietary protein restriction induced liver protein catabolism by inducing an activation of ATF4-CHOP-TRB3 pathway in order to provide amino acids to body tissues. Funding Sources ABIES, AlimH-INRAE.

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.

We conducted two experiments to evaluate the flooding dose method for measuring intestinal and liver protein synthesis in sheep. Experiment 1 showed that large doses of phenylalanine did not cause marked metabolic disturbances. Experiment 2 examined the effectiveness of flooding with phenylalanine and the time dependency of the protein synthesis calculation. Rams were injected with 1.2 MBq L-[ring 2,6-3H]phenylalanine/kg body wt and slaughtered 20, 40 or 60 min later. Plasma specific radioactivity reached a plateau within 2.5 min and did not change significantly (P0.05) throughout the experiment. Tissue intracellular free pool specific radioactivity also remained constant from 20 to 60 min postinfusion. Flooding conditions were achieved in the intracellular free pool of intestinal tissues (specific radioactivity 70-96% of plasma specific radioactivity), although liver flooding was less successful (57-67%). Protein synthesis rates measured after 20 min were significantly (P0.05) higher in the liver, jejunum and ileum than those measured at 60 min. Protein synthesis rates also tended to decline with time in the duodenum and colon (P0.05). There were no significant differences (P0.05) between protein synthesis rates calculated using the intracellular specific radioactivity vs. plasma specific radioactivity in the duodenum, ileum or colon. Therefore, this method represents an improvement over continuous infusion methods for measurements of protein synthesis in visceral tissues.

Seasonal variations in water temperature are a natural stressor of temperate fish that affect growth performance and metabolism globally. Gilthead sea bream is one of the most economically interesting species in the Mediterranean; but its liver metabolism is affected by the cold season. However, the effects of cold on protein turnover mechanisms have hardly been studied. Here, we study the relationship between liver oxidative status and protein homeostasis pathways during a 50-day low temperature period at 14 °C, and subsequent recovery at two times: 7 days (early recovery) and 30 days (late recovery). Liver redox status was determined by measuring oxidised lipids and proteins, the glutathione redox cycle and major antioxidant enzymes activities. Protein turnover was analysed via liver protein expression of HSP70 and HSP90; proteasome 26S subunits and polyubiquitination, as markers of the ubiquitin–proteasome system (UPS); and cathepsin D, as a lysosomal protease. Low temperature exposure depressed antioxidant enzyme activities, affecting the glutathione redox cycle and reducing total glutathione levels. Both the UPS and lysosomal pathways were also depressed and consequently, oxidised protein accumulated in liver. Interestingly, both protein oxidation and polyubiquitination tagging depended on protein molecular weight. Despite all these alterations, temperature recovery reverted most consequences of the cold at different rates: with delayed recovery of total glutathione levels and oxidised protein degradation with respect to enzyme activities recovery. All these findings demonstrate that protein liver homeostasis is compromised after chronic cold exposure and could be the cause of liver affectations reported in aquaculture of temperate fish.

In vivo studies confirmed that chlorogenic acid (CGA) improved glucose tolerance and mineral pool distribution in obese Zucker (fa/fa) rats. We found a significant decrease (P.05) in postprandial blood glucose concentrations, which may have been due to an improved sensitivity to insulin. Impaired glucose tolerance and insulin resistance have been associated with differences in the hepatic mRNA expression of the spliced variants of the insulin receptor at exon 11. Spliced variants of the insulin receptor have not been studied in obese Zucker (fa/fa) rats, and no information exists about the effects of CGA in vivo as a possible insulin sensitizer. Thus, we studied the in vivo effect of CGA on plasma insulin concentrations during a glucose tolerance test, liver protein and DNA concentrations, the hepatic activity of glucose-6-phosphatase (G-6-PASE) and the mRNA expression of the two variants of the insulin receptor at exon 11. Zucker (fa/fa) rats were implanted with jugular vein catheters. Chlorogenic acid was administered (5 mg/kg body weight per day) for 3 weeks via intravenous infusion. In the CGA-treated group, areas under the curve (AUC) for blood glucose and plasma insulin improved (P.005), and the protein and DNA concentrations in the liver increased (P.05). No significant differences (P.05) were found between groups for the hepatic G-6-PASE activity. The insulin receptor exon 11(+) and the exon 11(-) variants were expressed in the liver of Zucker (fa/fa) rats without significant changes (P.05). Chlorogenic acid improved some cellular mechanisms that are stimulated by insulin.

International audience; Epidemiologic and experimental studies showed that folate deficiency is associated with increased risk of degenerative diseases by enhancing abnormal one-carbon metabolism. We studied the changes in the proteome of liver, the main tissue of folate storage and metabolism, in a rat model of dietary folate depletion. Four-month-old rats were fed for 4 wk an amino acid-defined diet without folate and compared with pair-fed rats given the same diet adequately supplemented with folic acid. Folate deprivation decreased plasma and hepatic folate concentrations dramatically, while increasing homocysteinemia significantly. Using 2-dimensional electrophoresis and matrix-assisted laser desorption/ionization time-of-flight MS, we identified 9 spots corresponding to differentially expressed proteins in the liver of folate-deficient rats compared with controls. Among those spots, 4 had a significantly increased volume, whereas the volume of the 5 other spots was decreased. Upregulated proteins included glutathione peroxidase (GPx) 1 and peroxiredoxin 6, 2 enzymes involved in the response to oxidative stress, and MAWD binding protein (MAWDBP), which has been associated with cancer. MAWDBP was simultaneously identified as a second spot with a lower isoelectric point (pI) that vanished almost completely after folate deficiency. Decreased abundance was also observed for cofilin 1, a protein linked to tumorigenesis, and for the GRP 75 precursor and preproalbumin, both of which are responsive to oxidative stress and/or inflammation. Moreover, an enzyme activity assay and/or Western blot analysis of GPx-1 and MAWDBP confirmed the proteomic findings. Our results show that folate deficiency modifies the abundance of several liver proteins consistently with adaptive tissue responses to oxidative and degenerative processes.

In eukaryotic cells, amino acid depletion reduces translation by a mechanism involving phosphorylation of eukaryotic initiation factor-2 (eIF2). Herein we describe that mice lacking the eIF2 kinase, general control nonderepressible 2 (GCN2) fail to alter the phosphorylation of this initiation factor in liver, and are moribund in response to dietary leucine restriction. Wild-type (GCN2(+/+)) and two strains of GCN2 null (GCN2(-/-)) mice were provided a nutritionally complete diet or a diet devoid of leucine or glycine for 1 h or 6 days. In wild-type mice, dietary leucine restriction resulted in loss of body weight and liver mass, yet mice remained healthy. In contrast, a significant proportion of GCN2(-/-) mice died within 6 days of the leucine-deficient diet. Protein synthesis in wild-type livers was decreased concomitant with increased phosphorylation of eIF2 and decreased phosphorylation of 4E-BP1 and S6K1, translation regulators controlled nutritionally by mammalian target of rapamycin. Whereas translation in the liver was decreased independent of GCN2 activity in mice fed a leucine-free diet for 1 h, protein synthesis in GCN2(-/-) mice at day 6 was enhanced to levels measured in mice fed the complete diet. Interestingly, in addition to a block in eIF2 phosphorylation, phosphorylation of 4E-BP1 and S6K1 was not decreased in GCN2(-/-) mice deprived of leucine for 6 days. This suggests that GCN2 activity can also contribute to nutritional regulation of the mammalian target of rapamycin pathway. As a result of the absence of these translation inhibitory signals, liver weights were preserved and instead, skeletal muscle mass was reduced in GCN2(-/-) mice fed a leucine-free diet. This study indicates that loss of GCN2 eIF2 kinase activity shifts the normal maintenance of protein mass away from skeletal muscle to provide substrate for continued hepatic translation.

Background: Protein tyrosine phosphatases (PTPases) appear to play an essential role in the regulation of steady-state phosphorylation of the insulin receptor and other proteins in the insulin-signaling pathway. Objectives: This study determined the role of PTPases in skeletal muscle and liver in an insulin-resistant model and the in vivo effect of troglitazone on PTPases. Methods: We used 26-week-old male Otsuka Long-Evans Tokushima Fatty (OLETF) rats as models of insulin resistance. PTPase activity was assessed with reduced carboxyamido-methylated and -maleylated lysozyme as substrates. The protein levels of 3 major candidate PTPases—leukocyte common antigen-related PTPase (LAR), PTPase 1B (PTP1B), and src homology 2-containing PTPase (SH-PTP2)—in skeletal muscle and liver of OLETF rats were analyzed using Western blot. The activities and amounts of PTPases in skeletal muscle and liver of OLETF rats treated with troglitazone for 7 days also were examined. Results: Particulate and cytosolic PTPase activities in skeletal muscle of OLETF rats were increased compared with those in control rats. Similar changes were observed in the liver. The protein levels of LAR, PTP1B, and SH-PTP2 in particulate fractions of skeletal muscle of OLETF rats were increased, but levels of PTP1B and SH-PTP2 in cytosolic fractions were unchanged. Similar findings were observed in the liver. Troglitazone normalized the increased PTPase activity and each protein level except cytosolic fraction of the liver. Conclusions: The findings suggest that troglitazone decreases elevated activities and levels of PTPases in the particulate fraction of the skeletal muscle and liver of OLETF rats, and that PTPase plays important roles through regulating the insulin-signaling system in the pathogenesis of insulin resistance in OLETF rats.

Rat intoxication with a single dose of the hepatotoxin carbon tetrachloride induces a significant modification of liver protein kinase C total activity which depends on the degree of the intrahepatocyte oxidatiye unbalance provoked by various concentrations of the haloalkane. Low carbon tetrachloride amounts stimulate total protein kinase C activity, while one order of magnitude higher amounts exert strong enzyme inhibition. The latter effect is due to an early inactivation followed with progress of time by a proteolytic degradation of the enzyme. A pathological recruitment of the calcium-dependent protein kinase C regulatory enzymes calpain and calpastatin appears responsible for protein kinase C loss. The prolonged excess of cytosolic calcium which characterizes the single high dose carbon tetrachloride poisoning also leads to inactivation of calpain II and calpastatin in a time-dependent manner.

Fresh water fish, Tilapia mossambica (Peters) were acclimated to sublethal acidic (pH 5.0) and alkaline (pH 9.0) media to study liver proteolysis. In low pH medium the hepatic tissue showed activated acidic protease activity with inhibited neutral and basic proteases, while a reverse pattern of changes was observed in alkaline medium. The proteolysis was observed in both the media, with an elevated soluble proteins in the basic medium. The asidic stress was associated with hyperalbuminic and hypoglobulinic conditions, and in alkaline medium towards hyperalbuminic and hyperglobulinic conditions. The acidic stress induced amino acid catabolism with inhibition of the same in alkaline stress. Such changes in the fish may contribute to survival under altered environmental pH conditions.

Polypeptide assembly rates during in vivo hepatic protein synthesis were studied as a function of age and restricted feeding in male rats. With ageing the time to assemble the average peptide in the liver of fully-fed rats significantly increased. In young rats maintained on a restricted feeding regime known to retard ageing, the time to assemble the average polypeptide was increased 2.5 times. With ageing the rate of peptide elongation increased so that at 2 years of age the underfed animals assembled peptides at a significantly faster rate than their age-matched controls. The rate of elongation of peptides during hepatic protein synthesis was shown to be directly dependent upon circulating T3 levels rather than the dietary status of the animal. On refeeding young diet restricted rats, polypeptide assembly kinetics did not immediately return to control values although the rate of protein synthesis was significantly increased. Total liver RNA content increased significantly in refed animals allowing for a greater rate of chain initiation to offset the slow rate of chain elongation. A period of 28 days of ad libitum feeding was required before assembly kinetics returned to control values and is probably indicative of a persistent impaired monodeiodination of T4 to T3.

At 6 wk of age, male Fischer F344 rats were fed a purified, casein-based diet either ad libitum or in the amount of 60% of the diet consumed by the rats fed ad libitum (restricted diet). Hepatocytes were isolated from the rats between 2.5 and 19 mo of age. The protein content of the hepatocytes isolated from the rats fed the restricted amount of diet was significantly lower than that of hepatocytes isolated from rats fed ad libitum. The DNA and RNA content of the hepatocytes were similar for the rats fed the two dietary regimens. The absolute rate of protein synthesis for hepatocytes isolated from rats fed ad libitum decreased 55 % between 2.5 and 19 mo of age. However, the rate of protein synthesis by hepatocytes from rats fed the restricted amount of diet decreased only slightly with increasing age. At 19 mo of age, the rate of protein synthesis by hepatocytes from the rats fed the restricted amount of diet was significantly higher than the rate of protein synthesis for hepatocytes from rats fed ad libitum. Therefore, dietary restriction retards the age-related decline in liver protein synthesis. Reproduced by permission. Maria C. Birchenall-Sparks, Michael S. Roberts, Jeffrey Staeker, James E. Hardwick, Arlan Richardson, Effect of Dietary Restriction on Liver Protein Synthesis in Rats. J. Nutr. 115 , 944-950 (1985).

Two groups of six mice each were injected intraperitoneally with L-arginine (guanido-14C); 36 hours later 23 of the liver was removed from the animals in one group. The removed tissue served as control for both groups. Three days after the operation the regenerating livers still contained 104% of the total protein radioactivity whereas the normal livers had lost 50% of it.

Partial hepatectomy and acute stress (intraperitoneal injection of Celite) cause increases in the rates of synthesis of serum albumin and fibrinogen, proteins that are formed mainly or entirely by the liver. The rises in protein synthesis after removal of part of the liver differ kinetically from those after acute stress. In addition, only the response to partial hepatectomy is independent of the adrenal glands. Both responses seem to be related to an increase in the ability of liver polyribosomes to form protein.

Rats pretreated with phenobarbitone and their controls were exposed to 0.15% atmospheric CS 2 for 2 h. Liver protein metabolism and microsomal drug metabolizing enzyme activities were analyzed 1, 4 and 46 h later. In the pretreated rats the [ 14 C]leucine uptake was at first inhibited, then liver RNA tended to increase. This increase was followed by a decline in the [ 14 C]leucine uptake, while RNA content diminished to the control level. In the control rats (not pretreated with phenobarbital) the effects of CS 2 on liver protein metabolism were less; only at 4 h after the exposure was liver RNA increased and [ 14 C]leucine uptake slightly stimulated. In the pretreated rats CS 2 had decreased microsomal P-450 by about 50% at 1 and 4 h, and the activity of 7-O-dealkylase of ethoxycoumarin had decreased even more. The measurable UDPglucuronosyltransferase of the liver microsomes of the pretreated rats had increased by 26% at 1 h and by 80% at 4 h after the CS 2 exposure. The in vivo activation of microsomal UDPglucuronosyltransferase may result from a stimulated lipid peroxidation of reticuloendothelial membranes by CS 2 metabolites in vivo, as suggested by the diene conjugation spectra. In control rats CS 2 depressed only the ethoxycoumarin deethylase activity. All the microsomal changes caused by CS 2 exposure were restored within 46 h. It is suggested that the different action of CS 2 on the liver protein metabolism of rats pretreated with phenobarbitone and controls results from the different stage of protein turnover in the two groups, i.e. in the barbituratetreated animals there is a phase of increased protein synthesis and accordingly the protein turnover is more sensitive to the action of CS 2 .

Mechanisms responsible for diabetes-induced alterations in liver protein synthesis were investigated in vivo and in perfused liver using Bio-Breeding Worcester (BB/W) control rats, spontaneously diabetic BB/W rats maintained on insulin therapy, and diabetic BB/W rats withdrawn from insulin therapy for 48 h. Withdrawal of insulin therapy in the diabetic rats resulted in marked alterations in a number of parameters related to liver protein synthesis compared to BB/W control or insulin-maintained diabetic rats. Alterations seen in vivo following withdrawal of insulin included changes in the relative concentrations of several plasma proteins, a 40% reduction in total liver RNA relative to DNA, a 5-fold reduction in albumin synthesis relative to the synthesis of total liver proteins, a 5-fold reduction in albumin mRNA relative to total RNA, reductions in the relative abundance of mRNAs for at least four plasma proteins other than albumin, and a relative increase in mRNA for at least one plasma protein. Alterations observed in perfused liver included reductions in total liver protein synthesis (60% of control), albumin production (24% of control), and total secretory protein production (44% of control). All parameters studied were essentially unchanged from BB/W control values when the diabetic rats were maintained on insulin therapy. The results indicate that insulin deficiency leads to marked reductions in liver protein synthesis, particularly the synthesis of albumin and other plasma proteins. The mechanisms responsible for these alterations include changes in the relative abundance of specific mRNAs and a decrease in total cellular RNA.

Effects of single oral doses of aflatoxin B 1 (2 mg/kg) to monkeys Macaco irus on total liver protein synthesis and accumulation of liver lipid were investigated. Incorporation of [ 14 C]leucine into total liver protein in vivo was inhibited 50 per cent by aflatoxin B 1 3 hr after oral administration. This degree of inhibition was maintained up to 13 hr. Concomitant with this change there was an extensive disaggregation of polysomes in liver preparations. A progressive accumulation of lipid in liver following single dose administration of aflatoxin B 1 was observed. This increase in liver lipid was primarily triglyceride. The role of aflatoxin induced inhibition of hepatic protein synthesis in vivo to increase liver lipid is discussed.

Since S-adenosylhomocysteine is a potent inhibitor of rat liver protein methylase III in vitro , this enzyme appears to be regulated by modulation of the intracellular molar ratio of S-adenosylmethionine to S-adenosylhomo-cysteine (SAM:SAH ratio). The intraperitoneal administration of D,L-homocysteine thiolactone to animals (500 or 1000 mg/kg), followed by sacrifice 20 minutes later, decreased the SAM:SAH ratio in vivo from 4.35 in the control rats to 1.33 and 0.56 and caused a 62 and 82% inhibition of protein methylase III activity, respectively. Similarily, rats maintained on a pyridoxine-deficient diet for 7 weeks displayed a 5-fold reduction in the hepatic SAM:SAH ratio, when compared either to control or pair-fed counterparts, and a significant 26% reduction in the activity of protein methylase III. Thus, it would appear that a pyridoxine deficiency affects protein methylase III activity via altered SAM:SAH ratios.

We assessed the effects of total enteral nutrition with long-chain triacylglycerides (LCT), medium-chain triacylglycerides (MCT), or two structured lipids, modified dairy fat (MDF) and modified MCT (Captex 810B, Capital City Products, Columbus, OH), on protein and energy metabolism in hypermetabolic burned rats (25% to 30% body surface area). Male Sprague-Dawley rats (200 +/- 10 g) were continuously gastrostomy-fed isovolemic diets that provided 50 kcal/d, 2 g amino acids/d and 40% nonprotein calories as lipid for three days. Changes in body weight, nitrogen balance, serum albumin, indirect calorimetry, whole body leucine kinetics, and rectus muscle and liver protein kinetics were determined. Whole body leucine kinetics and tissue fractional protein synthetic rates (FSR, percent per day) were estimated using a four-hour constant intravenous infusion of L-[1-14C]leucine on day 3. The group of rats enterally fed MDF lost less body weight than the other groups (P less than or equal to .05). MDF and Captex 810B produced a positive and significantly greater (P less than or equal to .05) daily and cumulative nitrogen balance than either LCT or MCT. Oxygen consumption (P less than or equal to .05) and total energy expenditure (P less than or equal to .05) were elevated approximately 22% with MDF as compared with LCT or MCT. Rectus muscle FSR and absolute rate of protein synthesis were increased 19% with MDF (P less than or equal to .05) as compared with LCT or MCT.(ABSTRACT TRUNCATED AT 250 WORDS)

Female rats were fed on leucine supplemented diet during pregnancy and lactation and the changes in body weight and liver protein status of the animals were studied. When female rats received leucine supplemented diet during pregnancy and lactation, their body weights were reduced, however, these animals showed liver cell hyperplasia. It was interesting to correlate this decrease in body weight with the increased lactational performance of the animal.

Partial hepatectomy in the rat is known to cause rises in the rates of incorporation by subcellular liver preparations of 3H-puromycin into nascent peptide and 14C-leucine into protein. The operation has also been shown to be followed by a rise in the rate of liver protein synthesis in vivo. The kinetics of the three rises after partial hepatectomy are the same, suggesting that the changes are interrelated. The effects of actinomycin D on the rises were studied. When the antibiotic (5 µg per hour) was given immediately after partial hepatectomy, it blocked the appearance of the rises in vitro and in vivo and, as a function of dose, affected the three changes in an indistinguishable manner. When actinomycin (10 µg per hour) was given to partially hepatectomized animals after the rises had occurred, the reactivities in vitro and in vivo were gradually reduced to the normal levels. The halfway points in the return to the normal levels were at 4 hours from the time of the first injection of the antibiotic.

Total, free and membrane-bound polysomes from the liver of rats were assayed for mRNA activity in vitro. Controls (n = 3) and rats 1 (day 2, n = 3) or 3 days (day 4, n = 3) after a 25% body surface burn were used. The sedimentation profiles of total and bound polysomes shifted to a more aggregated state on day 2, and the peaks of monosomes and their sub-units in the free polysomal fraction decreased markedly. Significant changes in mRNA activity were observed in 23 and 17 polypeptides out of 170 on days 2 and 4, respectively. Most of the polypeptides with an increased activity were synthesised by bound polysomes. The mRNA activity for albumin was decreased by 60% on day 4. The activity for carbamoyl-phosphate synthetase I was increased two-fold on day 2. Although treatments of normal animals with counter-regulatory hormones produced characteristic patterns of in vitro products in each case, they mimicked only partially the effect of burn injury. These results indicate that burn injury induces marked changes in the level of translatable mRNAs associated with both free and membrane-bound polysomes in the liver, and suggest that other factors in addition to the counter-regulatory hormones are involved during the course of these changes.

The action of the nitrogen mustard di-(2-chloroethyl) methylamine hydrochloride (HN2) on the in vitro incorporation of labelled amino acids into the proteins of rat liver slices has been compared with its action on other biosynthetic processes in the liver. Uptake of 14 C-glycine was found to be less sensitive to HN2 than was incorporation of 32 P into ribonucleic acid. Liver slices were treated with different concentrations of HN2 and nuclear and cytoplasmic fractions isolated separately after incubation. Uptake of 14 C-glycine by the proteins of these two fractions was similarly affected by HN2. The action of HN2 on 14 C-phenylalanine incorporation and on p -aminohippuric acid synthesis by liver slices were then compared. At all concentrations of the inhibitor, essentially the same response was obtained with both systems. It has been concluded that the inhibition of amino acid incorporation produced by HN2 acting on liver slices is due to general inhibition of synthetic reactions rather than to a specific action on the mechanism of protein synthesis.

The time constant for assembly and release of polypeptide chains in the process of protein synthesis in rat liver in vivo has been determined by a new technique which is independent of most sources of variation and error in amino acid incorporation experiments. In this method the ratio of incorporation of 14C-amino acids into completed soluble proteins to that in total protein (including ribosome-bound chains) is measured as a function of short incubation periods (15 to 105 s) after hepatic portal vein injection to obtain tc, the average polypeptide chain assembly time. For normal male Long-Evans rats, ages 2 to 2½ months, tc in liver in vivo is 1.16 ± 0.16 min. Induction of hyperthyroidism by three injections of triiodothyronine (40 µg/100 g body weight) over 72 hours led to a value of tc = 0.92 ± 0.08 min, representing a 27% increase in synthetic rate. Surgically thyroidectomized rats showed a markedly longer time for polypeptide chain assembly, tc = 1.92 ± 0.22 min, corresponding to a 39% depression in protein synthetic output compared with sham-operated and normal controls of the same age. Almost complete recovery of protein synthetic rate in thyroidectomized rats was achieved by replacement doses of triiodothyronine (20 µg/100 g body weight) given daily for 2 days prior to assay. The results are discussed in relation to the mechanism of control of the protein synthetic pathway, the anabolic action of the thyroid hormones, and other methodology which has been used in attempts to estimate protein synthetic capacity.

Objective: We evaluated the effects of a new combined hormonal contraceptive vaginal ring (CVR) delivering the nonandrogenic progestin Nestorone® (NES) and ethinyl estradiol (EE) on several key estrogen-sensitive hepatic proteins that may be markers for the risk of arterial or venous disease events and on blood pressure (BP). Because the pharmacologic androgenicity of the progestin in these formulations influences the hepatic impact of EE, we selected an oral contraceptive (OC) delivering the androgenic progestin levonorgestrel (LNG) and EE as the comparator. We also investigated the effect of delivery route, which is known to modify the hepatic effects of estradiol, but has not been widely studied with EE. Study Methods: Women, aged 18-34 years, with no contraindications to the use of combined OCs, were randomized to three cycles of treatment with a CVR delivering NES/EE (150/15 μg/day) or a combined OC providing LNG and EE (150/30 μg per tablet). Each cycle consisted of 21 days of active treatment, followed by 7 days without treatment. During the last weeks of the pretreatment and third treatment cycles, blood samples were obtained for determinations of plasma concentrations of angiotensinogen, an estrogen-sensitive hepatic protein, and serum concentrations of sex hormone-binding globulin (SHBG), total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), triglycerides (TG) and estrogen- and androgen-sensitive proteins. BP was also measured. Results: Of 47 women randomized, 45 completed the study (CVR: 23; OC: 22). Within-group comparisons over time by repeated-measure analysis of variance demonstrated statistically significant changes over time with both treatments for all hepatic proteins (p

Effects of protein and energy intake on tissue protein synthesis and degradation were studied in rats fed 12, 24 and 36% protein diets at 100, 70, 50 and 30% of ad libitum intake begining on d 7 of lactation. Rates of protein synthesis were measured in liver, mammary, abdominal viscera and remaining carcass of rats on d 10 of lactation. Tissue protein loss was estimated as the difference in tissue protein from d 7 to 14. Data were analyzed using stepwise multiple quadratic regression to define response surfaces of protein loss, synthesis and degradation to intake of protein and energy. Protein loss increased with restriction of energy and protein intake in all tissues. However, carcass protein synthesis and degradation did not vary systematically with either energy or protein intake. Energy intake restriction increased visceral protein turnover while protein intake restriction decreased rate of protein turnover. Liver protein loss increased with decreased dietary protein due to increased liver protein degradation. Decreased energy intake also increased liver protein loss without directly affecting rate of synthesis or degradation. Mammary protein synthesis decreased very slightly with energy restriction while decreased protein intake sharply inhibited protein synthesis.

Human liver samples obtained from 343 autopsies at the Instituto Medico Legal Nina Rodrigues in Salvador, capital of Bahia State, Brazil, were studied for protein determination in parallel with ADH activity levels. There is no effect of age, race, or sex on the level of protein concentration, or on ADH levels. However, cause of death was found to be an important factor on total liver protein concentration in a similar manner as previously reported for ADH. Livers from patients dying from sudden traumatic accident had higher levels of protein concentration than those dying from acute illness. Also the acute illness group had higher protein concentration than those patients dying of chronic illness. Similar types of association were also confirmed for ADH activity levels in the present data. The possible mechanisms responsible for lowering liver protein during diseases are discussed.

Weaning albino rats were fed diet containing different amounts of vitamin D. Their body and liver weights, liver protein, free α-amino nitrogen, RNA and DNA contents were determined. When compared with the animals receiving vitamin D free diet, those receiving vitamin D excess diet showed significantly higher body and liver weights, liver protein, RNA and DNA contents. The data suggest the role of vitamin D in hepatic cellular growth.

Mice and rats receiving puromycin dihydrochloride intraperitoneally (6.0 and 60 mg. per animal, respectively) in four equally divided doses over a 4-hr, period showed complete depletion of hepatic glycogen and inhibition of incorporation of glycine-C 14 into liver protein. Administration of a single dose of puromycin to mice was followed by rapid inhibition of synthesis of liver protein with concurrent loss of glycogen from the liver. Carbohydrate levels in liver, blood, and muscle were followed in mice up to 4 hr. after a single injection of puromycin. The decrease in liver glycogen was accompanied by an increase in blood sugar levels while muscle glycogen content was not appreciably affected during this period.

Effects of orotic acid on liver lipid accumulation and incorporation of methionine [methyl-14C] into liver phosphatidylcholine and protein, and into serum beta-lipoprotein were studied. Male and female rats of Wistar strain were fed a semisynthetic diet supplemented with 1 per cent orotic acid for 7 days. Feeding of orotic acid induced a marked fatty liver in female rats, but not in males. In female rats, radioactivity in liver phosphatidylcholine was significantly decreased by orotic acid, and that in liver protein was slightly decreased. In male rats, incorporation of methionine [methyl-14C] into liver phosphatidylcholine and protein was unchanged between the control and the rats fed orotic acid. Radioactivity in serum beta-lipoprotein was decreased to a greater extent in female rats than in males. These results suggest that sex difference in the development of fatty liver may be due to the difference in the effect of orotic acid on liver phosphatidylcholine biosynthesis.

Primary cultures of rat hepatocytes were exposed to various concentrations of L-asparaginase derived from Escherichia Coli. Protein synthesis was inhibited by about 33% and cellular glutamine was reduced proportionally to the enzyme concentration. However, protein synthesis was inhibited only by amounts of enzyme able to reduce glutamine to critical levels below 10 nmol/mg cell protein. These data suggest that the glutaminase activity which probably contaminates E. coli asparaginase may be responsible for reduced liver protein synthesis.

Young male albino rats were fed 1.5 mg T-2 toxin/kg body weight daily for 4 days and the effects on body weight, liver weight and protein, RNA, and DNA contents of liver and intestinal mucosa were studied. A significant decrease in body weight and an increase in liver weight were observed. Liver protein and DNA and intestinal mucosal protein contents were significantly decreased, whereas intestinal mucosal RNA content was increased. Decreased liver and intestinal mucosal protein synthesis in T-2 toxin-fed rats was inferred from the [14C]leucine incorporation studies.

Body weight, liver weight, and protein of mice were significantly decreased after four days of starvation. However, liver protein concentration (milligram protein/gram tissue) did not show any significant changes. Liver acid phosphatase and β-glucuronidase specific activities were increased by about 40 and 50%, respectively. On the other hand, acid phosphatase, β-glucuronidase and cathepsin A specific activities of peritoneal exudates showed a significant decrease. The decreased specific activities of the enzymes in peritoneal exudates may partly be responsible for the decreased resistance to infection generally observed in starved animals.

We investigated the effect of non-protein calorie source on the repletion by total parenteral nutrition (TPN) of protein-depleted rats. Rats were depleted by feeding them a protein free diet for 4 weeks. Control (non-depleted) and depleted rats were given one of three isocaloric, isonitrogenous TPN regi mens for 6 days. They differed in the proportion of non-protein calories given as glucose and fat: diet 1, 100% glucose; diet 2, 75% glucose, 25% fat; and diet 3, 100% fat. The liver, skeletal muscle and whole body protein synthesis and break down rates were measured by using (15N)glycineas a tracer. The fatty acid distri bution in the liver was measured by gas chromatography-mass spectrometry. Results: (a) Omission of fat leads to a decrease in the fraction of linoleate in the liver, (b) Giving all the non-protein calories as a fat emulsion causes fatty acid accumulation in the liver and a significant increase in the liver protein fractional synthesis rate in the control rats and killed about half the depleted rats, (c) For TPN regimens 1 and 2 both the muscle and whole body synthesis rates were in creased during repletion, (d) Of the three TPN regimens, the second one which contained both fat and glucose was superior to the other two. J. Nutr. Ill: 154-165, 1981.

Two groups of rat pups were gastrostomized at 24 hours of age and formula-fed each hour or once every 6 hours a daily intake equal to 33% of their body weight. They were killed at age 5 to 7 days when body weight had reached twice that at time of gastrostomy; organ weights, body composition and hepatic activity of glucose-6-phosphate-dehydrogenase (G-6-PD) and alpha-glycerophosphate dehydrogenase (alpha-GPD) were determined. The stomach, small intestine, liver and spleen of pups fed 4 times each day were significantly larger than those of pups fed hourly. Food efficiency (gain/kcal) of pups fed each hour was significantly greater than that of pups fed every 6 hours; however, because pups fed each hour gained more water and less fat, an estimate of the percentage of energy intake utilized for growth was significantly less for these animals than for those fed every 6 hours. Concentration of calcium was greater and concentration of sodium was less in the carcasses of pups fed each hour than in the carcasses of pups fed 4 times daily. Total liver activities of G-6-PD and alpha-GPD were significantly less in pups fed each hour but the activity of these enzymes expressed per milligram of liver protein did not differ significantly between the two feeding groups.