Essential Amino Acids and Protein Synthesis: Insights into Maximizing the Muscle and Whole-Body Response to Feeding.

Ingesting protein-containing supplements and foods provides essential amino acids (EAA) necessary to increase muscle and whole-body protein synthesis (WBPS). Large variations exist in the EAA composition of supplements and foods, ranging from free-form amino acids to whole protein foods. We sought to investigate how changes in peripheral EAA after ingesting various protein and free amino acid formats altered muscle and whole-body protein synthesis. Data were compiled from four previous studies that used primed, constant infusions of L-(ring-2H5)-phenylalanine and L-(3,3-2H2)-tyrosine to determine fractional synthetic rate of muscle protein (FSR), WBPS, and circulating EAA concentrations. Stepwise regression indicated that max EAA concentration (EAACmax; R2 = 0.524, p < 0.001), EAACmax (R2 = 0.341, p < 0.001), and change in EAA concentration (ΔEAA; R = 0.345, p < 0.001) were the strongest predictors for postprandial FSR, Δ (change from post absorptive to postprandial) FSR, and ΔWBPS, respectively. Within our dataset, the stepwise regression equation indicated that a 100% increase in peripheral EAA concentrations increases FSR by ~34%. Further, we observed significant (p < 0.05) positive (R = 0.420-0.724) correlations between the plasma EAA area under the curve above baseline, EAACmax, ΔEAA, and rate to EAACmax to postprandial FSR, ΔFSR, and ΔWBPS. Taken together our results indicate that across a large variety of EAA/protein-containing formats and food, large increases in peripheral EAA concentrations are required to drive a robust increase in muscle and whole-body protein synthesis.

Intracellular Enzyme-Triggered Assembly of Amino Acid-Modified Gold Nanoparticles for Accurate Cancer Therapy with Multimode.

Multiple amino acid (glutamine and lysine)-modified gold nanoparticles a with pH-switchable zwitterionic surface were fabricated through coordination bonds using ferrous iron (Fe2+) as bridge ions, which are able to spontaneously and selectively assemble in tumor cells for accurate tumor therapy combining enzyme-triggered photothermal therapy and H2O2-dependent catalytic medicine. These gold nanoparticles showed electric neutrality at pH 7.4 (hematological system) to prevent endocytosis of normal cells, which could be positively charged at pH 6.8 (tumor microenvironment) to promote the endocytosis of tumor cells to these nanoparticles, performing great tumor selectivity. After cell uptake, the specific enzyme (transglutaminase) in tumor cells would catalyze the polymerization of glutamine and lysine to cause the intracellular assembly of these gold nanoparticles, resulting in an excellent photothermal property for accurate tumor therapy. Moreover, the Fe2+ ion could decompose excess hydrogen peroxide (H2O2) in tumor cells via the Fenton reaction, resulting in a large amount of hydroxyl radicals (·OH). These radicals would also cause tumor cell damage. This synergetic therapy associating with high tumor selectivity generated an 8-fold in vitro cytotoxicity against tumor cells compared with normal cells under 48 h incubation with 10 min NIR irradiation. Moreover, in vivo data from tumor-bearing nude mice models showed that tumors can be completely inhibited and gradually eliminated after multimode treatment combining catalytic medicine and photothermal therapy for 3 weeks. This system takes advantage of three tumor microenvironment conditions (low pH, enzyme, and H2O2) to trigger the therapeutic actions, which is a promising platform for cancer therapy that achieved prolonged circulation time in the blood system, selective cellular uptake, and accurate tumor therapy in multiple models.

Metabolic Availability of the Limiting Amino Acids Lysine and Tryptophan in Cooked White African Cornmeal Assessed in Healthy Young Men Using the Indicator Amino Acid Oxidation Technique.

Background: Maize is a staple food in many regions of the world, particularly in Africa and Latin America. However, maize protein is limiting in the indispensable amino acids lysine and tryptophan, making its protein of poor quality. Objective: The main objective of this study was to determine the protein quality of white African cornmeal by determining the metabolic availability (MA) of lysine and tryptophan. Methods: To determine the MA of lysine, 4 amounts of l-lysine (10, 13, 16, and 18 mg · kg-1 · d-1 totaling 28.6%, 37.1%, 45.7%, and 51.4% of the mean lysine requirement of 35 mg · kg-1 · d-1, respectively) were studied in 6 healthy young men in a repeated-measures design. To determine the MA of tryptophan, 4 amounts of l-tryptophan (0.5, 1, 1.5, and 2 mg · kg-1 · d-1 totaling 12.5%, 25.0%, 37.5%, and 50.0% of the mean tryptophan requirement of 4 mg · kg-1 · d-1, respectively) were studied in 7 healthy young men in a repeated-measures design. The MAs of lysine and tryptophan were estimated by comparing the indicator amino acid oxidation (IAAO) response with varying intakes of lysine and tryptophan in cooked white cornmeal compared with the IAAO response to l-lysine and l-tryptophan intakes in the reference protein (crystalline amino acid mixture patterned after egg protein) with the use of the slope ratio method. Results: The MAs of lysine and tryptophan from African cooked white cornmeal were 71% and 80%, respectively. Conclusion: Our study provides a robust estimate of the availability of lysine and tryptophan in African white maize to healthy young men. This estimate provides a basis for postproduction fortification or supplementation of maize-based diets. This trial was registered at www.clinicaltrials.gov as NCT02402179.

Characterization of zinc amino acid complexes for zinc delivery in vitro using Caco-2 cells and enterocytes from hiPSC.

Zn is essential for growth and development. The bioavailability of Zn is affected by several factors such as other food components. It is therefore of interest, to understand uptake mechanisms of Zn delivering compounds to identify ways to bypass the inhibitory effects of these factors. Here, we studied the effect of Zn amino acid conjugates (ZnAAs) on the bioavailabilty of Zn. We used Caco-2 cells and enterocytes differentiated from human induced pluripotent stem cells from a control and Acrodermatitis enteropathica (AE) patient, and performed fluorescence based assays, protein biochemistry and atomic absorption spectrometry to characterize cellular uptake and absorption of ZnAAs. The results show that ZnAAs are taken up by AA transporters, leading to an intracellular enrichment of Zn mostly uninhibited by Zn uptake antagonists. Enterocytes from AE patients were unable to gain significant Zn through exposure to ZnCl2 but did not show differences with respect to ZnAAs. We conclude that ZnAAs may possess an advantage over classical Zn supplements such as Zn salts, as they may be able to increase bioavailability of Zn, and may be more efficient in patients with AE.

Whole-body and splanchnic amino acid metabolism in sheep during an acute endotoxin challenge.

Supplemented protein or specific amino acids (AA) are proposed to help animals combat infection and inflammation. The current study investigates whole-body and splanchnic tissue metabolism in response to a lipopolysaccharide (LPS) challenge with or without a supplement of six AA (cysteine, glutamine, methionine, proline, serine and threonine). Eight sheep were surgically prepared with vascular catheters across the gut and liver. On two occasions, four sheep were infused through the jugular vein for 20 h with either saline or LPS from Escherichia coli (2 ng/kg body weight per min) in a random order, plus saline infused into the mesenteric vein; the other four sheep were treated with saline or LPS plus saline or six AA infused via the jugular vein into the mesenteric vein. Whole-body AA irreversible loss rate (ILR) and tissue protein metabolism were monitored by infusion of [ring-2H2]phenylalanine. LPS increased (P<0·001) ILR (+17 %), total plasma protein synthesis (+14 %) and lymphocyte protein synthesis (+386 %) but decreased albumin synthesis (-53 %, P=0·001), with no effect of AA infusion. Absorption of dietary AA was not reduced by LPS, except for glutamine. LPS increased the hepatic removal of leucine, lysine, glutamine and proline. Absolute hepatic extraction of supplemented AA increased, but, except for glutamine, this was less than the amount infused. This increased net appearance across the splanchnic bed restored arterial concentrations of five AA to, or above, values for the saline-infused period. Infusion of key AA does not appear to alter the acute period of endotoxaemic response, but it may have benefits for the chronic or recovery phases.

Determining the functional role of waterborne amino acid uptake in hagfish nutrition: a constitutive pathway when fasting or a supplementary pathway when feeding?

Hagfish are unique among aquatic "vertebrates" in their ability to absorb amino acids directly from the water via skin and gill epithelia, but it is unknown whether this phenomenon extends beyond a few studied substrates; what effect fed state has on absorption; and what functional role this may play in hagfish nutrition. Using in vivo and in vitro transport assays, uptake and tissue distribution of the waterborne amino acids L-alanine, L-lysine, and L-phenylalanine were examined as a function of fed state. All three amino acids were shown to be taken up from the water (lysine and phenylalanine for the first time). Following immersion in radiolabelled solutions for 24 h, phenylalanine was the amino acid that accumulated at the highest levels in almost all tissues, with the highest accumulation noted in red blood cells and bile, followed by gill and liver. In general, tissues of fed hagfish displayed a significantly reduced phenylalanine accumulation compared to tissues of hagfish fasted for 3 weeks. An in vitro assay showed that phenylalanine was transported across the skin at the highest rate, with the uptake of lysine occurring at the lowest rate. Feeding status had no significant effect on in vitro transport. These data indicate that dissolved organic nutrients are a significant source of nutrition to hagfish, and may be relatively more important during periods of fasting than during periods of feeding when immersed in decaying carcasses.

Novel insights on intake of meat and prevention of sarcopenia: all reasons for an adequate consumption / Nuevos conocimientos sobre el consumo de carne en la prevención de la sarcopenia: todos los motivos de un consumo adecuado

Introducion: sarcopenia is defined as a syndrome characterized by progressive and generalized loss of muscle mass and strength. The main cause of sarcopenia is the alteration of protein metabolism, in which the proteolytic processes are not accompanied by an appropriate protein synthesis and muscle cells lose progressively the sensitivity to the anabolic stimulus. The most rational approach to delay the progression of sarcopenia and counteract the anabolic resistance is proper nutrition. Meat contains biologically active compounds, such as creatine, carnitine, Conjugated Linoleic Acid (CLA) which have significant impacts upon human protein metabolism. Methods: we performed a narrative literature review to evaluate the till-now evidence regarding: 1. adequate intake of meat in elderly as a topic for prevention of sarcopenia; 2. the correct intake of biologically active compounds contain in meat, which have significant impacts upon human protein metabolism and so have beneficial effects on prevention of sarcopenia. This review included 62 eligible studies. Results: the results demonstrated that in elderly the optimum diet therapy for the sarcopenia prevention and treatment, which must aim at achieving specific metabolic goals, must recommend the consumption of 113 g of meat (220 kcal; 30 g protein) five time a week. Conclusion: in a varied and balanced diet, for preventing sarcopenia, it is recommended to assume meat 4-5 times a week (white meat 2 times per week, lean red meat less than 2 times per week, processed meat less than 1 time per week), as suggested in the diet pyramid for elderly (AU)

Introducción: la sarcopenia se define como un síndrome caracterizado por la pérdida progresiva y generalizada de la masa muscular y de la fuerza. La principal causa de la sarcopenia es la alteración del metabolismo de las proteínas, en la que los procesos proteolíticos no van acompañados de una síntesis de proteínas y células musculares adecuadas, con lo que se pierde progresivamente la sensibilidad al estímulo anabólico. El enfoque más racional para retrasar la progresión de la sarcopenia y contrarrestar la resistencia anabólica es una nutrición adecuada. La carne contiene compuestos biológicamente activos, tales como creatina, carnitina y ácido linoleico conjugado (CLA) que tienen impactos significativos sobre el metabolismo de la proteína humana. Métodos: se realizó una revisión de la literatura narrativa para evaluar la evidencia hasta ahora, en relación con: 1. ingesta adecuada de carne en ancianos como prevención de la sarcopenia; 2. la ingesta correcta de compuestos biológicamente activos que contiene la carne, que tienen impactos significativos sobre el metabolismo de la proteína humana y para así obtener efectos beneficiosos en la prevención de la sarcopenia. Esta revisión incluyó 62 estudios elegibles. Resultados: los resultados demostraron que en personas de edad avanzada la terapia óptima con dieta para la prevención y tratamiento de la sarcopenia, que debe apuntar al logro de los objetivos metabólicos específicos, debe recomendar el consumo de 113 g de carne (220 kcal; 30 g de proteínas) cinco veces a la semana. Conclusión: en una dieta variada y equilibrada, para prevenir la sarcopenia, se recomienda consumir la carne 4-5 veces a la semana (carne blanca 2 veces por semana, carne roja magra menos de 2 veces por semana, carne procesada menos de 1 vez por semana), como se sugiere en la pirámide de la dieta para personas mayores (AU)

Glutamina como coadyuvante en la recuperación de la fuerza muscular: revisión sistemática de la literatura / Glutamine as an aid in the recovery of muscle strength: systematic review of literature

Antecedentes: la pérdida de fuerza del músculo esquelético es frecuente tras una lesión traumática o en el postquirúrgico ortopédico. Además de los esquemas de ejercicio de fuerza y/o resistencia para su tratamiento, ha sido propuesto como auxiliar el uso de algunos aminoácidos como la glutamina (Gln), de manera aislada o combinada con otros nutrimentos. Sin embargo, la información sobre la eficacia de la suplementación oral con Gln durante los esquemas de ejercicio de fuerza y/o resistencia en adultos con déficit de fuerza es inconsistente. Objetivo: evaluar la solidez de la evidencia disponible del efecto de la suplementación oral con Gln sobre la fuerza muscular, junto con esquemas de ejercicio de fuerza y/o resistencia en adultos con déficit de fuerza muscular. Métodos: se realizó una búsqueda sistemática en diferentes bases de datos, de ensayos clínicos reportados desde el año 1980 a 2014, en idioma inglés y español, sobre suplementación oral con Gln aislada o combinada con otros nutrimentos, con grupo control, en adultos con déficit de fuerza, bajo esquemas de ejercicio de fuerza y/o resistencia, seguimiento menor a un año y fuerza muscular como desenlace primario. Resultados: de 661 artículos, se identificaron seis estudios relevantes. El estudio con más participantes que evaluó la Gln aislada no sugiere cambios entre los grupos, solo una mejoría en la percepción de la debilidad muscular. Los estudios que evaluaron la Gln con otros nutrimentos reportan resultados a favor de esta. No fue posible realizar un metanálisis. Conclusiones: actualmente no se dispone de suficientes datos de los efectos relacionados con la Gln sobre el déficit de fuerza muscular durante esquemas de ejercicio en adultos. Se requiere mayor investigación al respecto para responder con mayor solidez sobre este hecho (AU)

Background: after a traumatic injury or post surgical orthopedic, the loss of skeletal muscle strength is common. In addition to strength training schemes and/or resistance to treatment, it has been proposed as an additional treatment, the use of some amino acids such as glutamine (Gln) in isolation or combination with other nutrients. However, the information on the effectiveness of oral Gln supplementation during exercise strength schemes and / or endurance in adults with strength deficit is inconsistent. Objective: to evaluate the strength of the evidence at hand about the effect of oral supplementation on muscle strength Gln set to strength training schemes and / or resistance in adult muscle strength deficit. Methods: a systematic search was conducted in different databases, in clinical trials reported from the year 1980-2014, both in English and Spanish, about oral Gln supplementation alone or in combination with other nutrients, with a control group, in adults with strength deficits under exercise schemes of strength and / or endurance, tracking under a year and muscle power as the primary outcome. Results: of 661 articles, six relevant studies were identified. The study participants in Gln isolation evaluation did not suggest changes between the groups, only an improvement in the perception of muscle weakness. Studies evaluating Gln with other nutrients, have reported results in favor of it. No meta-analysis was posible. Conclusions: nowadays there are insufficient data on the effects related to the Gln on the deficit of muscular force during exercise schemes in adults. It is required more research in this topic to respond more accurately about this fact (AU)

Insulin resistance and protein metabolism in chronic hemodialysis patients.

OBJECTIVE: Loss of lean body mass (sarcopenia) is associated with increased morbidity and mortality in patients receiving chronic hemodialysis (CHD). Insulin resistance (IR), which is highly prevalent in patients receiving CHD, has been proposed to play a critical role in the development of sarcopenia. The aim of this study was to examine the effect of IR on amino acid metabolism in patients receiving CHD. DESIGN: This was a cross-sectional study. SUBJECTS: The study included 12 prevalent (i.e., patients that have been on dialysis for more than 90 days) African American patients receiving CHD. METHODS: IR was measured as glucose disposal rate (GDR) determined from hyperinsulinemic euglycemic clamp (HGEC) studies performed 3 consecutive times. Plasma amino acid (AA) concentrations were measured by real-time high-performance liquid chromatography (HPLC) throughout the clamp study. The primary outcome was percentage change in leucine concentrations during the clamp study. The main predictor was the GDR measured simultaneously during the HGEC studies. Mixed model analysis was used to account for repeated measures. RESULTS: All individual AA concentrations declined significantly in response to high-dose insulin administration (P < .001). There was a significant direct association between GDR by HECG studies and the percentage change in leucine concentration (P = .02). Although positive correlations were observed between GDR values and concentration changes from baseline for other AAs, these associations did not reach statistical significance. CONCLUSIONS: Our results suggest that the severity of IR of carbohydrate metabolism is associated with a lesser decline in plasma leucine concentrations, suggesting a similar resistance to protein anabolism. Insulin resistance represents a potential mechanism for sarcopenia commonly observed in patients receiving CHD.

La suplementación de la L-arginina no afecta la producción de óxido nítrico ni la recuperación muscular / No effect of L-arginine supplementation on nitric oxide production and muscle recovery

Introducción: La L-arginina (L-Arg) es un aminoácido semiesencial y precursor de la síntesis de óxido nítrico (NO). Recientemente, los suplementos nutricionales que contiene L-Arg son comercializados con la pretensión de promover vasodilatación, debido al aumento de la producción de NO en músculo. El resultado de la vasodilatación elevaría la perfusión sanguínea, promoviendo un mayor aporte de nutrientes y oxigeno, los cuales pueden mejorar el rendimiento y la recuperación muscular. Propósito: Identificar el efecto agudo de la suplementación con L-Arg sobre la ratio de recuperación del trabajo (WRR), potencia media, trabajo total (TW3S) y los indicadores de producción de NO, nitrito y nitrato plasmático (NOx), durante el ejercicio de contra-resistencia. Métodos: Diecisiete hombres sanos y entrenados, participaron en un estudio, doble ciego, controlado con placebo. Se tomaron muestras de sangre antes y 90 min después de la ingesta de 6 g de LArg o placebo. El protocolo de ejercicio (3 series de 10 repeticiones máximas de extensión de codo isocinético concéntrico en 60º.s-1 con 2 min de descanso entre las series) se inició 80 minutos después de la suplementación. Mediciones de NOx se realizaron por el método de Griess usando un espectrofotómetro de absorción a 540 nm. Resultados: No se encontraron diferencias significativas entre el grupo suplementado con L-Arg vs placebo en lo referente parámetros de WRR, potencia media y TW3S (2630,4 ± 758,0vs 2573,1 ± 669,9 Joules). Además, no se observó diferenciasignificativa en el NOx plasmático, en ningún momento, entre el grupo suplementado con L-Arg vs placebo, antes de la suplementación (9,8 ± 2,3vs 9,5 ± 1,4 micro mol/L) o inmediatamente después del ejercicio (11,9 ± 5,5 vs 10,2 ±2,3 micro mol/L). Conclusión: Nuestros resultados indican que la ingesta aguda de L-Arg no aumenta la producción de NO, ni mejora el rendimiento recuperación muscular. En base a estos resultados, es precipitado recomendar suplementos nutricionales que contienen L-Arg como ayuda ergogénica para optimizar la recuperación muscular después del ejercicio de contra-resistencia en individuos sanos y entrenados (AU)

Introduction: L-arginine (L-Arg) is a semi-essential aminoacid precursor to nitric oxide (NO) synthesis. Recently, nutritional supplements containing L-Arg have been marketed with the purpose of promote vasodilation, due to an increased production of NO in the exercising muscle. The resulting vasodilation would elevate blood perfusion, leading to a higher nutrient and oxygen delivery, which may enhance exercise performance and muscular recovery. Purpose: Identify the acute effect of L-Arg supplementation on work recovery ratio (WRR), average power, total work (TW3S) and indicators of NO production, plasma nitrite and nitrate(NOx), during a resistance exercise protocol. Methods: Seventeen healthy and resistance-trained males participated in a randomized, double-blind, placebo-controlled study. Blood samples were collected before and 90 min (immediately post-exercise) after ingestion of oral 16g of L-Arg or placebo. The exercise protocol (3 sets of 10 maximal voluntary contractions of isokinetic concentric elbow extension at 60 o.s-1with 2-min of rest between sets) was initiated 80 min after supplementation. NOx measurements were made by a traditional Griess reaction colorimetric method using a spectrophotometer monitoring absorbance at 540 nm. Results: No significant difference between L-Arg versus placebo supplemented groups was observed on WRR, average power and TW3S (2630.4 ± 758.0 versus 2573.1 ± 669.9 Joules). Furthermore, no significant difference was observed in plasma NOx at any time point between L-Arg versus placebo supplemented groups at baseline (9.8 ± 2.3 vs. 9.5 ± 1.4 micre mol/L) and immediately post-exercise (11.9 ± 5.5 vs. 10.2 ±2.3 micremol/L). Conclusion: Our data indicates that acute ingestion of L-Arg does not increase NO production nor enhances muscle performance and recovery. Based on this fact, it is still premature to recommend nutritional supplements containing L-Arg as an ergogenic aid to optimize muscle recovery after resistance exercise bouts in healthy and resistance-trained subjects (AU)

Acute supplementation of amino acids increases net protein accretion in IUGR fetal sheep.

Placental insufficiency decreases fetal amino acid uptake from the placenta, plasma insulin concentrations, and protein accretion, thus compromising normal fetal growth trajectory. We tested whether acute supplementation of amino acids or insulin into the fetus with intrauterine growth restriction (IUGR) would increase net fetal protein accretion rates. Late-gestation IUGR and control (CON) fetal sheep received acute, 3-h infusions of amino acids (with euinsulinemia), insulin (with euglycemia and euaminoacidemia), or saline. Fetal leucine metabolism was measured under steady-state conditions followed by a fetal muscle biopsy to quantify insulin signaling. In CON, increasing amino acid delivery rates to the fetus by 100% increased leucine oxidation rates by 100%. In IUGR, amino acid infusion completely suppressed fetal protein breakdown rates but increased leucine oxidation rate by only 25%, resulting in increased protein accretion rates by 150%. Acute insulin infusion, however, had very little effect on amino acid delivery rates, fetal leucine disposal rates, or fetal protein accretion rates in CON or IUGR fetuses despite robust signaling of the fetal skeletal muscle insulin-signaling cascade. These results indicate that, when amino acids are given directly into the fetal circulation independently of changes in insulin concentrations, IUGR fetal sheep have suppressed protein breakdown rates, thus increasing net fetal protein accretion.

A mass balance approach to identify and compare differential routing of 13C-labeled carbohydrates, lipids, and proteins in vivo.

All animals route assimilated nutrients to their tissues where they are used to support growth or are oxidized for energy. These nutrients are probably not allocated homogeneously among the various tissue and are more likely to be preferentially routed toward some tissues and away from others. Here we introduce an approach that allows researchers to identify and compare nutrient routing among different organs and tissues. We tested this approach by examining nutrient routing in birds. House sparrows Passer domesticus were fed a meal supplemented with one of seven (13)C-labeled metabolic tracers representing three major classes of macronutrients, namely, carbohydrates, amino acids, and fatty acids. While these birds became postabsorptive (2 h after feeding), we quantified the isotopic enrichment of the lean and lipid fractions of several organs and tissues. We then compared the actual (13)C enrichment of various tissue fractions with the predictions of our model to identify instances where nutrients were differentially routed and found that different classes of macronutrients are uniquely routed throughout the body. Recently ingested amino acids were preferentially routed to the lean fraction of the liver, whereas exogenous carbohydrates were routed to the brain and the lipid fraction of the liver. Fatty acids were definitively routed to the heart and the liver, although high levels of palmitic acid were also recovered in the adipose tissue. Tracers belonging to the same class of molecules were not always routed identically, illustrating how this technique is also suited to examine differences in nonoxidative fates of closely related molecules. Overall, this general approach allows researchers to test heretofore unexamined predictions about how animals allocate the nutrients they ingest.

Amino acid uptake profiling of wild type and recombinant Streptomyces lividans TK24 batch fermentations.

Streptomyces lividans is considered an interesting host for the secretory production of heterologous proteins. To obtain a good secretion yield of heterologous proteins, the availability of suitable nitrogen sources in the medium is required. Often, undefined mixtures of amino acids are used to improve protein yields. However, the understanding of amino acid utilization as well as their contribution to the heterologous protein synthesis is poor. In this paper, amino acid utilization by wild type and recombinant S. lividans TK24 growing on a minimal medium supplemented with casamino acids is profiled by intensive analysis of the exometabolome (metabolic footprint) as a function of time. Dynamics of biomass, substrates, by-products and heterologous protein are characterized, analyzed and compared. As an exemplary protein mouse Tumor Necrosis Factor Alpha (mTNF-α) is considered. Results unveil preferential glutamate and aspartate assimilation, together with glucose and ammonium, but the associated high biomass growth rate is unfavorable for protein production. Excretion of organic acids as well as alanine is observed. Pyruvate and alanine overflow point at an imbalance between carbon and nitrogen catabolism and biosynthetic fluxes. Lactate secretion is probably related to clump formation. Heterologous protein production induces a slowdown in growth, denser clump formation and a shift in metabolism, as reflected in the altered substrate requirements and overflow pattern. Besides glutamate and aspartate, most amino acids are catabolized, however, their exact contribution in heterologous protein production could not be seized from macroscopic quantities. The metabolic footprints presented in this paper provide a first insight into the impact and relevance of amino acids on biomass growth and protein production. Type and availability of substrates together with biomass growth rate and morphology affect the protein secretion efficiency and should be optimally controlled, e.g., by appropriate medium formulation and substrate dosing. Overflow metabolism as well as high biomass growth rates must be avoided because they reduce protein yields. Further investigation of the intracellular metabolic fluxes should be conducted to fully unravel and identify ways to relieve the metabolic burden of plasmid maintenance and heterologous protein production and to prevent overflow.

Prolonged maternal amino acid infusion in late-gestation pregnant sheep increases fetal amino acid oxidation.

Protein supplementation during human pregnancy does not improve fetal growth and may increase small-for-gestational-age birth rates and mortality. To define possible mechanisms, sheep with twin pregnancies were infused with amino acids (AA group, n = 7) or saline (C group, n = 4) for 4 days during late gestation. In the AA group, fetal plasma leucine, isoleucine, valine, and lysine concentrations were increased (P < 0.05), and threonine was decreased (P < 0.05). In the AA group, fetal arterial pH (7.365 +/- 0.007 day 0 vs. 7.336 +/- 0.012 day 4, P < 0.005), hemoglobin-oxygen saturation (46.2 +/- 2.6 vs. 37.8 +/- 3.6%, P < 0.005), and total oxygen content (3.17 +/- 0.17 vs. 2.49 +/- 0.20 mmol/l, P < 0.0001) were decreased on day 4 compared with day 0. Fetal leucine disposal did not change (9.22 +/- 0.73 vs. 8.09 +/- 0.63 micromol x min(-1) x kg(-1), AA vs. C), but the rate of leucine oxidation increased 43% in the AA group (2.63 +/- 0.16 vs. 1.84 +/- 0.24 micromol x min(-1) x kg(-1), P < 0.05). Fetal oxygen utilization tended to be increased in the AA group (327 +/- 23 vs. 250 +/- 29 micromol x min(-1) x kg(-1), P = 0.06). Rates of leucine incorporation into fetal protein (5.19 +/- 0.97 vs. 5.47 +/- 0.89 micromol x min(-1) x kg(-1), AA vs. C), release from protein breakdown (4.20 +/- 0.95 vs. 4.62 +/- 0.74 micromol x min(-1) x kg(-1)), and protein accretion (1.00 +/- 0.30 vs. 0.85 +/- 0.25 micromol x min(-1) x kg(-1)) did not change. Consistent with these data, there was no change in the fetal skeletal muscle ubiquitin ligases MaFBx1 or MuRF1 or in the protein synthesis regulators 4E-BP1, eEF2, eIF2alpha, and p70(S6K). Decreased concentrations of certain essential amino acids, increased amino acid oxidation, fetal acidosis, and fetal hypoxia are possible mechanisms to explain fetal toxicity during maternal amino acid supplementation.

Glutathione synthesis rates after amino acid administration directly after birth in preterm infants.

BACKGROUND: The availability of glutathione, the main intracellular antioxidant, is compromised in preterm neonates. A possible explanation is the low availability of substrate for synthesis, because many neonatologists are reluctant to administer amino acids in the direct postnatal period for fear of intolerance. OBJECTIVE: The objective of the study was to determine the effects of amino acid administration directly after birth on glutathione synthesis rates and markers of oxidative stress. DESIGN: Premature infants (<1500 g) received from birth onward either dextrose (control group; n = 10) or dextrose plus 2.4 g amino acids . kg (- 1) . d(-1) (intervention group; n = 10). On postnatal day 2, [1-(13)C]glycine was administered to determine glutathione fractional synthesis rates (FSR(GSH)) and absolute synthesis rates (ASR(GSH)) in erythrocytes. In plasma, advanced oxidized protein products and dityrosine, both markers of oxidative stress, were measured. The results are expressed as means +/- SDs. RESULTS: The FSR(GSH) was not different between groups: 44 +/- 6 and 48 +/- 9%/d in the control and intervention groups, respectively (P = 0.28). The concentration of erythrocyte glutathione was higher (P < 0.001) in the intervention group (2.28 +/- 0.35 mmol/L) than in the control group (1.73 +/- 0.37 mmol/L). ASR(GSH) values were 6.5 +/- 1.5 and 11.3 +/- 1.9 mg . kg(-1) . d(-1) in the control and intervention groups, respectively (P < 0.001). Advanced oxidized protein products and dityrosine concentrations were not significantly different between groups. CONCLUSIONS: Amino acid administration directly after birth increases ASR(GSH) in preterm infants. Our data are consistent, however, with higher glutathione concentrations rather than a higher FSR(GSH). Greater availability of glutathione, nevertheless, did not decrease markers of oxidative stress.

The influence of caecectomy on amino acid availability of three feedstuffs for ganders.

1. Experiments were conducted to assess the influence of caecectomy on amino acid availability (AAA) of three feedstuffs for goose. 2. Nine caecectomised and 9 intact Yangzhou ganders, 24 weeks old, were used in these experiments. Fish meal, soybean meal and cottonseed meal were used as the sole source of protein. The endogenous amino acid (AA) losses were evaluated by a nitrogen (N)-free diet method. The influence of caecectomy on apparent amino acid availability (AAAA) in fish meal, soybean meal and cottonseed meal was assessed in experiment 1 and true amino acid availability (TAAA) of three protein diets was determined in experiment 2. 3. Results showed that, in the soybean meal and cottonseed meal, the AAAA and TAAA of most AA determined by the intact ganders were higher than in the caecectomised ganders; in the fish meal, the AAAA and TAAA of most AA determined by the intact ganders were lower than in the caecectomised ganders. 4. Results of the present study suggest that the effect of caecectomy on AAA in geese was dependent on the feedstuff assayed, and it was better to use caecectomised poultry for AAA assessment.

A comparison of methods to determine amino acid availability of feedstuffs in cecectomized ganders.

Experiments were conducted to compare endogenous amino acid losses and the true amino acid availability (TAAA) of 3 feedstuffs by using methods involving a short-term fasting and an N-free diet with cecectomized ganders. Diets were formulated to contain soybean meal, fish meal, and cottonseed meal as the sole source of protein. A precision-fed assay was used in which each feed sample was precise-fed (60 g) to geese and excreta were collected for 48 h. A N-free diet and fasting methods were used to evaluate the endogenous amino acid losses. Endogenous losses of 3 amino acids were significantly different (P < 0.01) with the N-free diet and fasting methods. The TAAA of soybean meal, fish meal, and cottonseed meal determined by N-free diet method ranged from 84.49 to 97.09%, 89.18 to 98.16%, and 77.09 to 98.32%, respectively. The TAAA of these 3 diets determined by the fasting method ranged from 83.50 to 97.77%, 88.08 to 99.60%, and 76.09 to 98.09%, respectively. However, there were only a few small differences (P > 0.05) between methods in each amino acid. In conclusion, there was no difference in determination of the amino acid availability of these feedstuffs using cecectomized ganders between the N-free diet and fasting methods.

La glutamina, un aminoácido casi indispensable en el enfermo crítico / Glutamine, an almost essential amino acidin the critically ill patient

La glutamina es el aminoácido más abundante del organismo y está implicada en numerosos procesos del metabolismo intermediario, sobre todo en la síntesis de aminoácidos y purinas, en el ciclo de los ácidos tricarboxílicos y en la generación de urea. Aunque se ha considerado un aminoácido no esencial debido a que puede ser sintetizado por el organismo, existen situaciones clínicas graves que cursan con una depleción marcada, por lo que ha sido considerado como condicionalmente esencial. Los niveles bajos de glutamina se asocian con alteraciones de la función inmune, con cambios en la estructura y función de la mucosa intestinal y del tejido linfático asociado, con disminución de la capacidad oxidante y con modificaciones de la sensibilidad a la insulina en el enfermo grave. La administración de suplementos clínicos de glutamina, tanto por vía enteral como parenteral, han dado resultados contradictorios pero, en su mayor parte, apoyan la hipótesis de que los aportes de glutamina pueden modificar la morbimortalidad de los enfermos graves. Quedan cuestiones pendientes de resolver como la dosis adecuada y la vía de administración, y más importante, definir aquellos subgrupos de pacientes que pueden beneficiarse más de su empleo

Glutamine is the most abundant amino acid in the human body and plays an important role in a number of metabolic pathways. Specifically, it is involved in amino acid and nucleotide synthesis, in the tricarboxylic acid cycle and in ureagenesis. Glutamine has been classified as a non-essential amino acid because the body can synthesize it, but under severe clinical conditions, the pool of glutamine is depleted and could be considered as conditionally essential. Low levels of glutamine are associated with a decrease in the immune response, changes in the structure and function of the intestinal mucose and the gut associated lymphoid tissue, a decreased anti-oxydant capacity and changes of the insulin sensitivity in critically ill patients. Administration of supplemental glutamine by enteral or parenteral route has produced controversial results. Most of the studies published support the hypothesis that glutamine can change the morbidity-mortality of the critically ill patients. There are unresolved questions related to the dose of glutamine and the best way to administer it, and particularly the subgroups of patients who will really benefit from this treatment

Early segregation of layered projections from the lateral superior olivary nucleus to the central nucleus of the inferior colliculus in the neonatal cat.

The central nucleus of the inferior colliculus (IC) is a laminated structure that receives multiple converging afferent projections. These projections terminate in a layered arrangement and are aligned with dendritic arbors of the predominant disc-shaped neurons, forming fibrodendritic laminae. Within this structural framework, inputs terminate in a precise manner, establishing a mosaic of partially overlapping domains that likely define functional compartments. Although several of these patterned inputs have been described in the adult, relatively little is known about their organization prior to hearing onset. The present study used the lipophilic carbocyanine dyes DiI and DiD to examine the ipsilateral and contralateral projections from the lateral superior olivary (LSO) nucleus to the IC in a developmental series of paraformaldehyde-fixed kitten tissue. By birth, the crossed and uncrossed projections had reached the IC and were distributed across the frequency axis of the central nucleus. At this earliest postnatal stage, projections already exhibited a characteristic banded arrangement similar to that described in the adult. The heaviest terminal fields of the two inputs were always complementary in nature, with the ipsilateral input appearing slightly denser. This early arrangement of interdigitating ipsilateral and contralateral LSO axonal bands that occupy adjacent sublayers supports the idea that the initial establishment of this highly organized mosaic of inputs that defines distinct synaptic domains within the IC occurs largely in the absence of auditory experience. Potential developmental mechanisms that may shape these highly ordered inputs prior to hearing onset are discussed.

Effects of various dietary amino acid preparations for phenylketonuric patients on the metabolic profiles along with postprandial insulin and ghrelin responses.

AIM: We investigated the metabolic profiles along with insulin and ghrelin responses following ingestion of various amino acid (AA) substitutes commonly used in the treatment of phenylketonuria to study the effects of added macronutrients. METHODS: Twenty healthy and 6 phenylketonuric adults ingested AA mixtures with or without carbohydrates and fat (Anamix, Easiphen, or p-am 3; 0.35 g AA/kg body weight); milk powder shakes were used for control purposes. Serum AA, glucose, urea, insulin, and ghrelin were measured over 5 h. RESULTS: Peak AA concentrations were achieved at around 60 min postprandially for supplemented AA powders and control shakes, significantly later than for pure AA. Of interest, the mean Phe/Tyr ratio declined by 40-50% in phenylketonuric patients following intake of Easiphen, Anamix, or p-am 3. The insulin peaks, up to 500% as compared with baseline, occurred at 30 min and were approximately 100% higher after intake of AA plus macronutrients. Glucose and urea remained constant. Ghrelin showed a nadir at 60 min, followed by a rise leading to a 30% increase of initial concentrations for pure AA as compared with more constant levels for preparations with macronutrients. CONCLUSION: An oral AA bolus together with macronutrients retards hyperaminoacidemia, displays a higher insulin secretion, normoglycemia, and more stable ghrelin concentrations, whereas the pure AA tested here exerted weaker anabolic effects.