Effects of prostaglandin E(1) on the efficacy of xenogeneic extracorporeal pig liver perfusion in a canine model of acute liver failure.
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Xenogeneic extracorporeal liver perfusion (ECLP) has the potential to become an important tool in the management of patients with severe liver failure. We previously showed that xenogeneic pig liver perfusion may be prolonged for up to 9 hours by the administration of prostaglandin E(1) (PGE(1)). In this study, we used a canine model of acute liver failure to evaluate the effects of PGE(1) on the efficacy of ECLP as a liver-assist device. Liver failure was surgically induced in 12 beagle dogs, with a control group (group 1, n = 4) not connected to the ECLP circuit. Direct cross-circulation between the dogs and the ECLP circuit using a pig liver was performed without (group 2, n = 4) or with (group 3, n = 4) continuous administration of PGE(1) through the portal vein of the pig liver. The duration of cross-circulation in group 3 (9.4 +/- 1.2 hours) was significantly longer than in group 2 (4.3 +/- 1.0 hours). In addition, elevation of blood ammonia, total bile acid, and hyaluronic acid levels was less marked in group 3 compared with the other 2 groups. The ratio of branched-chain amino acids to aromatic amino acids was also improved in group 3. The mean survival time in group 3 (26.6 +/- 0.4 hours) was significantly longer than in group 1 (15.5 +/- 1.3 hours) or group 2 (17.1 +/- 2.9 hours). Continuous administration of PGE(1) to xenogeneic ECLP resulted in a significant improvement in both liver function and survival time of dogs with surgically induced liver failure. (+info)
Deficiency of dietary EAA preferentially inhibits mRNA translation of ribosomal proteins in liver of meal-fed rats.
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The goal of these studies was to investigate the mechanisms by which amino acid supply regulates global rates of protein synthesis as well as the translation of ribosomal protein (rp) mRNAs in liver. In the experiments conducted, male weanling rats were trained over a 2-wk period to consume their daily food intake within 3 h. On day 14, rats were fed the control diet or an isocaloric, isonitrogenous diet lacking glycine, tryptophan, leucine, or the branched-chain amino acids (BCAA) for 1 h. Feeding Trp-, Leu-, or BCAA-deficient diets resulted in significant reductions in serum insulin, hepatic protein synthesis, eukaryotic initiation factor 2B (eIF2B) activity, and phosphorylation of eIF4E-binding protein 1 (4E-BP1) and ribosomal protein S6 kinase (S6K1). Phosphorylation of eIF2alpha was inversely related to eIF2B activity under all conditions. Alterations in the hepatic synthesis of rp were assessed by changes in the distribution of rp (S4, S8, L26) mRNAs across sucrose density gradients and compared with non-rp (beta-actin, albumin) mRNAs. In all dietary treatments, non-rp mRNAs were mostly polysome associated. Conversely, the proportion of rp mRNAs residing in polysomes was two- to fivefold less in rats fed diets lacking tryptophan, leucine, or BCAA compared with rats fed the control diet. Total hepatic abundance of all mRNAs examined did not differ among treatment groups. For all parameters examined, there were no differences between rats fed the glycine-deficient diet and rats fed the control diet. The data suggest that essential amino acid (EAA) deficiency inhibits global rates of liver protein synthesis via a block in translation initiation. Additionally, the translation of rp mRNAs is preferentially repressed in association with decreased S6K1 phosphorylation. (+info)
Oral supplementation of branched-chain amino acid improves nutritional status in elderly patients on chronic haemodialysis.
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BACKGROUND: Anorexia may be associated with decreased plasma levels of branched-chain amino acids (BCAA). In malnourished elderly haemodialysis (HD) patients, oral BCAA supplementation may improve anorexia, resulting in improved nutritional status. METHODS: Among 44 elderly (age >70 years) patients on chronic HD, 28 patients with low plasma albumin concentration (<3.5 g/dl) were classified as the malnourished group; they also suffered from anorexia. The other 16 patients did not complain of anorexia and were classified as the well-nourished group. We performed a 12-month, placebo-controlled, double-blind study on the malnourished group. Fourteen patients each received daily oral BCAA supplementation (12 g/day) or a placebo in random order in a crossover trial for 6 months. Body fat percentage, lean body mass, plasma albumin concentration, dietary protein and caloric intakes, and plasma amino acid profiles were monitored. RESULTS: Lower plasma levels of BCAA and lower protein and caloric intakes were found in the malnourished group as compared to the well-nourished group. In BCAA-treated malnourished patients, anorexia and poor oral protein and caloric intakes improved within a month concomitant with the improvement in plasma BCAA levels over the values in well-nourished patients. After 6 months of BCAA supplementation, anthropometric indices showed a statistically significant increase and mean plasma albumin concentration increased from 3.31 g/dl to 3.93 g/dl. After exchanging BCAA for a placebo, spontaneous oral food intake decreased, but the favourable nutritional status persisted for the next 6 months. In 14 patients initially treated with a placebo, no significant changes in nutritional parameters were observed during the first 6 months. However, positive results were obtained by BCAA supplementation during the subsequent 6 months, and mean plasma albumin concentration increased from 3.27 g/dl to 3.81 g/dl. CONCLUSIONS: Normalization of low plasma levels of BCAA by oral supplementation can reduce anorexia and significantly improve overall nutritional status in elderly malnourished HD patients. (+info)
Five ruminally cannulated Holstein steers (176 kg) were used in a 5 x 5 Latin square to evaluate the effects of branched-chain AA supplementation on N retention and plasma AA concentrations of steers. Steers were limit-fed (3.0 kg/d of DM) twice daily diets low in ruminally undegradable protein (72% soybean hulls, 19% alfalfa, 5% molasses, and 4% vitamins and minerals). Acetate (400 g/d) was continuously infused into the rumen. Treatments were continuous abomasal infusions of 1) 115 g/d of a mixture of 10 essential AA designed to exceed the steers' requirements (10AA), 2) 10AA with Leu removed, 3) 10AA with Ile removed, 4) 10AA with Val removed, and 5) 10AA with all three branched-chain AA removed. Experimental periods were 7 d, with 3 d for adaptation to treatments and 4 d for total fecal and urinary collections for N balance. Blood samples were collected 5 h after feeding on d 7. Retained N decreased in response to removal of Leu (P < 0.06), Val (P < 0.05), or all three branched-chain AA (P < 0.05). Plasma Leu concentrations decreased (P < 0.05) in response to removal of Leu and all three branched-chain AA. Plasma Ile concentrations decreased (P < 0.05) in response to removal of Ile and all three branched-chain AA but increased (P < 0.05) in response to removal of Leu. Plasma Val concentrations decreased (P < 0.05) in response to removal of Val and all three branched-chain AA but increased (P < 0.05) in response to removal of Leu. Responses in N balance and plasma AA concentrations of growing cattle limit-fed soybean hull-based diets demonstrate limitations in the basal supply of Leu and Val but not Ile provided that supplies of all other essential AA are met. (+info)
Dietary L-carnitine suppresses mitochondrial branched-chain keto acid dehydrogenase activity and enhances protein accretion and carcass characteristics of swine.
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A trial was conducted to biochemically explain the decreased lipid deposition and increased protein accretion observed in pigs fed carnitine. Our hypothesis was that an increase in the ratio of acetyl CoA:CoA-SH produced by stimulation of fatty acid oxidation by supplemental L-carnitine may decrease branched-chain alpha-keto acid dehydrogenase activity and increase pyruvate carboxylase activity. Such changes could reduce oxidative loss of branched-chain amino acids and provide more carbons for amino acid biosynthesis. Yorkshire gilts (n = 36; 12 per treatment) were fed a control diet or diets containing either 50 or 125 ppm of added L-carnitine during growth from 56 to 120 kg. After slaughter, the semitendinosus muscle and liver were collected for isolation of mitochondria and hepatocytes. Increasing dietary L-carnitine did not influence growth performance (P > 0.10) but linearly decreased (P < 0.05) 10th rib backfat thickness and increased (linear, P < 0.05) percentages of lean and muscle. The rates of [1-(14)G]palmitate oxidation in isolated hepatocytes and isolated mitochondria, and incorporation of [35S]methionine into the acid insoluble fraction of isolated hepatocytes were increased (linear, P < 0.01) in pigs fed L-carnitine. Flux through branched-chain alpha-keto acid dehydrogenase linearly decreased (P < 0.01) in isolated liver and muscle mitochondria with increasing dietary carnitine. Flux through pyruvate carboxylase was increased (linear, P < 0.01) in isolated mitochondria from liver of pigs fed carnitine, and assays with particle-free extracts indicated that the amount of mitochondrial pyruvate carboxylase was tripled by feeding carnitine (linear, P < 0.01). The association of increased protein accretion and reduced backfat thickness with greater rates of palmitate oxidation, more rapid flux through pyruvate carboxylase, and reduced flux through branched-chain alpha-keto acid dehydrogenase suggests pigs fed carnitine are more able to use fat for energy, divert carbon toward synthesis of amino acids, and spare branched-chain amino acids for protein synthesis. (+info)
Nutrition in CAPD: serum bicarbonate and the ubiquitin-proteasome system in muscle.
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BACKGROUND: Metabolic acidosis in chronic renal failure (CRF) induces loss of lean body mass while elimination of acidosis during a one year trial improved anthropometric indices in continuous ambulatory peritoneal dialysis (CAPD) patients. In rats with CRF, the mechanisms causing loss of lean body mass have been linked to acidosis-induced destruction of the essential, branched-chain amino acids (BCAA) and activation of the ubiquitin-proteasome system that degrades muscle protein; the latter response includes increased transcription of the ubiquitin gene. METHOD: Our aim was to determine if increasing the serum bicarbonate (HCO3) concentration of CAPD patients would improve their nutritional status, increase plasma BCAA levels, and reduce ubiquitin mRNA in their muscle as an index of suppressed activity of the ubiquitin-proteasome system. Eight, stable, long-term CAPD patients underwent vastus lateralis muscle biopsy before being randomized to continue 35 mmol/L lactate dialysate or convert to a 40 mmol/L lactate dialysate. After four weeks, measurements were repeated. RESULTS: Serum HCO3 increased in all patients and final values did not differ statistically between the two groups so results for all patients were combined. Weight and body mass index increased significantly as did plasma BCAA. Muscle levels of ubiquitin mRNA decreased significantly; serum tumor necrosis factor-alpha (TNF-alpha) also decreased. CONCLUSION: Our results indicate that even a small correction of serum HCO3 improves nutritional status, and provide evidence for down-regulation of BCAA degradation and muscle proteolysis via the ubiquitin-proteasome system. Whether acidosis and inflammatory cytokines (such as, TNF-alpha) interact to impair nutrition is unknown. (+info)
Rhizobium leguminosarum has a second general amino acid permease with unusually broad substrate specificity and high similarity to branched-chain amino acid transporters (Bra/LIV) of the ABC family.
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Amino acid uptake by Rhizobium leguminosarum is dominated by two ABC transporters, the general amino acid permease (Aap) and the branched-chain amino acid permease (Bra(Rl)). Characterization of the solute specificity of Bra(Rl) shows it to be the second general amino acid permease of R. leguminosarum. Although Bra(Rl) has high sequence identity to members of the family of hydrophobic amino acid transporters (HAAT), it transports a broad range of solutes, including acidic and basic polar amino acids (L-glutamate, L-arginine, and L-histidine), in addition to neutral amino acids (L-alanine and L-leucine). While amino and carboxyl groups are required for transport, solutes do not have to be alpha-amino acids. Consistent with this, Bra(Rl) is the first ABC transporter to be shown to transport gamma-aminobutyric acid (GABA). All previously identified bacterial GABA transporters are secondary carriers of the amino acid-polyamine-organocation (APC) superfamily. Also, transport by Bra(Rl) does not appear to be stereospecific as D amino acids cause significant inhibition of uptake of L-glutamate and L-leucine. Unlike all other solutes tested, L-alanine uptake is not dependent on solute binding protein BraC(Rl). Therefore, a second, unidentified solute binding protein may interact with the BraDEFG(Rl) membrane complex during L-alanine uptake. Overall, the data indicate that Bra(Rl) is a general amino acid permease of the HAAT family. Furthermore, Bra(Rl) has the broadest solute specificity of any characterized bacterial amino acid transporter. (+info)
Cloning and inactivation of a branched-chain-amino-acid aminotransferase gene from Staphylococcus carnosus and characterization of the enzyme.
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Staphylococcus carnosus and Staphylococcus xylosus are widely used as aroma producers in the manufacture of dried fermented sausages. Catabolism of branched-chain amino acids (BCAAs) by these strains contributes to aroma formation by production of methyl-branched aldehydes and carboxy acids. The first step in the catabolism is most likely a transamination reaction catalyzed by BCAA aminotransferases (IlvE proteins). In this study, we cloned the ilvE gene from S. carnosus by using degenerate oligonucleotides and PCR. We found that the deduced amino acid sequence was 80% identical to that of the corresponding enzyme in Staphylococcus aureus and that the ilvE gene was constitutively expressed as a monocistronic transcript. To study the influence of ilvE on BCAA catabolism, we constructed an ilvE deletion mutant by gene replacement. The IlvE protein from S. carnosus was shown mainly to catalyze the transamination of isoleucine, valine, leucine, and, to some extent, methionine using pyridoxal 5'-phosphate as a coenzyme. The ilvE mutant degraded less than 5% of the BCAAs, while the wild-type strain degraded 75 to 95%. Furthermore, the mutant strain produced approximately 100-fold less of the methyl-branched carboxy acids, 2-methylpropanoic acid, 2-methylbutanoic acid, and 3-methylbutanoic acid, which derived from the BCAA catabolism, clearly emphasizing the role of IlvE in aroma formation. In contrast to previous reports, we found that IlvE was the only enzyme that catalyzed the deamination of BCAAs in S. carnosus. The ilvE mutant strain showed remarkably lower growth rate and biomass yield compared to those of the wild-type strain when grown in rich medium. Normal growth rate and biomass yield were restored by addition of the three BCAA-derived alpha-keto acids, showing that degradation products of BCAAs were essential for optimal cell growth. (+info)