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BACKGROUND Effects of high protein (HP) diets and prolonged energy restriction (ER) on integrated muscle protein kinetics have not been determined. OBJECTIVE The objective of this study was to measure protein kinetics in response to prolonged ER and HP on muscle protein synthesis (MPS; absolute rates of synthesis) and muscle protein breakdown (MPB; half-lives) for proteins across the muscle proteome. METHODS Female 6-wk-old obese Zucker rats (Leprfa+/fa+, n = 48) were randomly assigned to one of four diets for 10 wk: ad libitum-standard protein (AL-SP; 15% kcal from protein), AL-HP (35% kcal from protein), ER-SP, and ER-HP (both fed 60% feed consumed by AL-SP). During week 10, heavy/deuterated water (2H2O) was administered by intraperitoneal injection, and isotopic steady-state was maintained via 2H2O in drinking water. Rats were euthanized after 1 wk, and mixed-MPS as well as fractional replacement rate (FRR), relative concentrations, and half-lives of individual muscle proteins were quantified in the gastrocnemius. Data were analyzed using 2-factor (energy × protein) ANOVAs and 2-tailed t-tests or binomial tests as appropriate. RESULTS Absolute MPS was lower in ER than AL for mixed-MPS (-29.6%; P < 0.001) and MPS of most proteins measured [23/26 myofibrillar, 48/60 cytoplasmic, and 46/60 mitochondrial (P < 0.05)], corresponding with lower gastrocnemius mass in ER compared with AL (-29.4%; P < 0.001). Although mixed-muscle protein half-life was not different between groups, prolonged half-lives were observed for most individual proteins in HP compared with SP in ER and AL (P < 0.001), corresponding with greater gastrocnemius mass in HP than SP (+5.3%; P = 0.043). CONCLUSIONS ER decreased absolute bulk MPS and most individual MPS rates compared with AL, and HP prolonged half-lives of most proteins across the proteome. These data suggest that HP, independent of energy intake, may reduce MPB, and reductions in MPS may contribute to lower gastrocnemius mass during ER by reducing protein deposition in obese female Zucker rats.

BACKGROUND The Dietary Guidelines for Americans (DGAs) published an "ounce equivalents" recommendation to help consumers meet protein requirements with a variety of protein food sources. However, the metabolic equivalency of these varied protein food sources has not been established. OBJECTIVE We have investigated the hypothesis that the anabolic responses to consumption of ounce equivalents of protein food sources would be directly related to the essential amino acid (EAA) content of the protein food source. METHODS Following 3 d of dietary control, a total of 56 healthy young adults underwent an 8.5-h metabolic study using stable isotope tracer methodology. The changes from baseline following consumption of 1 of 7 different protein food sources were compared with the baseline value for that individual (n = 8 per group). RESULTS Consumption of ounce equivalents of animal-based protein food sources (beef sirloin, pork loin, eggs) resulted in a greater gain in whole-body net protein balance above baseline than the ounce equivalents of plant-based protein food sources (tofu, kidney beans, peanut butter, mixed nuts; P

Broiler chickens reared under heat stress (HS) conditions have decreased growth performance and show metabolic and immunologic alterations. This study aimed to evaluate the effect of supplementation with a standardized blend of plant-derived isoquinoline alkaloids (IQ) on the growth performance, protein catabolism, intestinal barrier function, and inflammatory status of HS-treated chickens. Three hundred sixty 0-day-old Ross 308 male broiler chickens were randomly distributed into 2 treatment groups: control diet (no additives) or diet supplemented with 100 ppm IQ. At day 14, the chicks in each diet group were further divided into 2 groups, each of which was reared under thermoneutral (TN) (22.4°C) or constant HS (33.0°C) conditions until day 42. Each group consisted of 6 replicates with 15 birds per replicate, and chickens were provided ad libitum access to water and feed. During days 15–21, the body weight gain (BWG) and feed intake (FI) were significantly lower in the HS treatment group than in the TN group, and feed conversion ratio was higher (P

Proteases are critical virulence determinants of Porphyromonas gingivalis, an emerging Alzheimer's disease, cancer, and arthritis pathogen and established agent of periodontitis. Transposon sequencing has been employed to define the core essential genome of this bacterium and genes conditionally essential in multiple environments - abscess formation; epithelial colonization; and cigarette smoke toxin exposure; as well as to elucidate genes required for iron acquisition and a functional type 9 secretion system. Validated and predicted protein catabolism genes identified include a combination of established virulence factors and a larger set of seemingly more mundane proteolytic genes. The functions and relevance of genes that share essentiality in multiple disease-relevant conditions are examined. These common stress-related genes may represent particularly attractive therapeutic targets for the control of P. gingivalis infections.

Activation of the mechanistic target of rapamycin (mTOR) pathway is known to promote protein synthesis by enhancing mRNA translation. However, there have been few literatures on the effect of mTOR on protein metabolism in non-mammals. The main source of ammonia in fish comes from protein catabolism. The key step of protein catabolism involves the deamination and/or transamination of amino acids. This study is aimed to explore the mechanism underlying mTOR pathway influencing protein retention from the perspective of protein catabolism. Chinese perch were fasted for 24 h and divided into 4 groups randomly before intracerebroventricular (ICV) injection: (1) control group for leucine; (2) leucine group; (3) control group for leucine and rapamycin; (4) leucine and rapamycin group. Food intake was equivalent between each control and treatment groups at each time point (0.5, 4, 12 and 24 h post-injection). Ammonia-N excretion rate, blood glucose, S6 phosphorylation level, and expression of relative genes of protein catabolism (GDH, AMPD, AST, ALT) were determined. The results indicated that the pS6 level was increased, and that the ammonia-N excretion rate, blood glucose, and mRNA level of protein catabolism genes (GDH and AMPD) were significantly decreased after injection with leucine, while those changes were reversed after injection with leucine and rapamycin. Our study not only reveals the mechanism by which mTOR mediates protein synthesis by inhibiting protein catabolism in Chinese perch, but also provides reference for improving the utilization of feed protein.

A meta-analysis was performed from 66 references retained from a total of 147 ones. The aim was to study the relationship between the THI and some physiologic, metabolic and oxidative responses in sheep. The effects of the breed, physiological state and heat relieving strategies were investigated as well. PCA, GLM, one nested ANOVA and global correlations were used to assess the fixed effects. Respiration rate, rectal temperature and heart pulse vary significantly in response to THI increase. However, the magnitude of this variance is defined by the breed and the physiological state of the animal. Moreover, Mechanical heat relieving strategies allowed alleviating the drastic effects of hyperthermia on sheep physiologic responses, whereas some nutritional heat abatement strategies increased metabolic heat load and respiration rate. Antioxidant and mineral supplementations were able to protect animals from heat stress. Environmental-induced hyperthermia reduced feed intake, body weight, circulating glucose and cholesterol levels, and increased lipid and protein catabolism. Significant increase of cortisol hormone was recorded. Indeed, ambient- induced hyperthermia resulted in oxidative stress marked by a decrease of Se contents, enhanced SOD activity, high oxidative stress index, increased protein oxidation, reactive oxygen species production and MDA levels. We report here for the first time that respiration is a second source of free radicals generation in sheep under environmental-induced heat stress. Heat stress disturbs rectal temperature, respiration and heart rates, exacerbates the post absorptive metabolism as well as the antioxidant status of sheep. Therefore, animals increase their maintenance requirements at the expense of production needs, which become not priorities at all.

Background Elderly patients undergoing cardiac surgery often show poor nutritional status, muscle wasting, and sarcopenia, which are reported to affect postoperative functional recovery and incidence of complications. Amino acids are essential in maintaining nutritional status, synthesizing muscle protein, and promoting beneficial energy balance of the heart muscle. β-Hydroxy β-methylbutyric acid (HMB) is a leucine metabolite known to increase muscle protein synthesis and inhibit protein catabolism; it has been used to more effectively support patients with muscle wasting due to wearing diseases. However, the efficacy of amino acid administration comprising HMB in patients undergoing open heart surgery remains unclear. This study aims to examine whether preoperative short-term aggressive amino acid administration helps support postoperative recovery of physical function and prevent complications. Methods This is a single-center prospective randomized controlled trial (UMIN000030490). Patients aged ≥65 years who will be hospitalized for medical examination before cardiac surgery will be recruited. The participants will be randomly assigned to the experimental or control group. The experimental group will be administered with an amino acid supplement with HMB 1200 mg, l -glutamine 7000 mg, and l -arginine 7000 mg once or twice per day depending on the degree of renal dysfunction, for 14–28 days preoperatively. The control group will not receive any nutritional intervention. The main outcome will be a change in the 6-min walking test distance pre- and postoperatively as a sign of functional recovery. Secondary outcomes such as the incidence of complications; physical, nutritional, and psychological states; mortality; and length of hospital stay will also be evaluated. Conclusion This clinical study will determine the effects of preoperative short-term oral amino acid supplementation with HMB, l -glutamine, and l -arginine on postoperative physical function in elderly patients undergoing cardiac surgery.

Summary Background & aims Muscle wasting in cancer negatively affects physical function and quality of life. This study investigates amino acid metabolism and the association with muscle mass and function in patients with cancer. Methods In 16 patients with advanced cancer undergoing chemotherapy and 16 healthy controls, we administered an intravenous pulse and prime of stable amino acid tracers. We took blood samples to measure the Rate of appearance (Ra), whole body production (WBP), clearance (Cl), and post absorptive whole body net protein breakdown (WBnetPB). Plasma amino acid concentrations and enrichments were analysed by LC-MS/MS. We assessed muscle mass, handgrip/leg/respiratory muscle strength and reported physical activity, quality of life, and physical function. Results Muscle strength was lower in cancer patients than in healthy controls. Total and limb muscle mass, reported physical activity and WBnetPB were comparable. WBP and Cl of tau-methylhistidine, leucine, glutamine and taurine were higher in cancer patients as well as glycine Cl. Amino acid metabolism was correlated with low muscle mass, strength, physical function and quality of life. Conclusions Myofibrillar protein breakdown and production of amino acids involved in muscle contractility are up regulated in patients with cancer undergoing chemotherapy and related to muscle weakness and reduced physical outcomes.

The increase of dietary lipid utilization leads a supply of more energy to reduce protein breakdown and promote protein deposition in fish. Therefore, a search for feed additives to stimulate lipid metabolism interests fish nutritionists. Diacylglycerol oil (DGO), as a safe food additive in human, can increase lipid metabolism and prevent obesity in mammals, but it has not been studied in fish feed. The present study evaluated the role of DGO on lipid utilization and protein deposition in Nile tilapia (Oreochromis niloticus) by feeding fish with the diets containing three levels of dietary DGO (0%, 0.5% or 2.5%) for 8 weeks. The 0.5% dietary DGO increased weight gain and significantly decreased the hepatosomatic and mesenteric fat mass. The total lipid and triacylglycerol (TG) contents of the whole fish, liver and muscle tissues, as well as the serum free fatty acid content were significantly lower in the fish fed with both doses of the DGO diet compare to the control (0%). The fish fed DGO diets also exhibited lower lipid droplets in liver tissues than in the control. Moreover, dietary DGO increased the protein content in the whole fish and muscle tissues. The fish fed with the 0.5% DG diet showed an increasing trend on the gene expression of lipid catabolism and protein metabolism in the liver and muscle tissues. However, the 2.5% dietary DGO tended to downregulate metabolism gene expression and upregulate inflammation and stress genes expressions in liver. This study indicates that proper dose of dietary DGO can improve lipid utilization and help to promote protein deposition, and could be a potential additive in aquaculture feeds.

Acute intestinal failure is a reversible clinical condition characterised by reduced intestinal function, which requires the intravenous supplementation of nutrients, water and/or electrolytes for weeks or months to maintain an optimal state of health. Acute intestinal failure occurs mainly in postsurgical patients who have undergone abdominal operations or are critically ill and have organ dysfunction. These patients' medical and nutritional management can be a complex and prolonged process due to the associated complications such as electrolyte imbalances, malnutrition, sepsis and metabolic disorders. Medical treatment is focused on preventing and controlling sepsis, restoring fluids and electrolytes and rehabilitating the intestine. Nutritional therapy seeks to attenuate protein catabolism, prevent nutritional deficiencies and metabolic disorders and optimise the nutritional state. This review seeks to provide updated and useful information on the management of acute intestinal failure.

Bioturbators such as sediment-dwelling marine bivalves are ecosystem engineers that enhance sediment-water exchange and benthic-pelagic coupling. In shallow coastal areas, bivalves are exposed to frequent disturbance and salinity stress that might negatively affect their activity and physiological performance; however, the mechanisms underlying these effects are not fully understood. We investigated the effects of osmotic stress (low and fluctuating salinity) and repeated burrowing on aerobic and contractile capacity of the foot muscle (assessed by the activity of succinate dehydrogenase and myosin ATPase) as well as the levels of organic osmolytes (free amino acids) and biochemical markers of protein synthesis and proteolysis in key osmoregulatory and energy storing tissues (gills and hepatopancreas, respectively) in a common bioturbator, the soft shell clam Mya arenaria. Osmotic stress and exhaustive exercise altered the foot muscle capacity of soft shell clams and had a strong impact on protein and amino acid homeostasis in tissues not directly involved in locomotion. Acclimation to constant low salinity (5 practical salinity units) depleted the whole-body free amino acid pool and affected protein synthesis but not protein breakdown in the gill. In contrast, fluctuating (5-15) salinity increased protein breakdown rate, suppressed protein synthesis, caused oxidative damage to proteins in the gill and selectively depleted whole-body glycine pool. Clams acclimated to normal salinity (15) increased the aerobic capacity of the foot muscle upon repeated burrowing, whereas acclimation to low and fluctuating salinity reduced this adaptive muscle plasticity. Under the normal and low salinity conditions, exhaustive exercise induced protein conservation pathways (indicated by suppression of protein synthesis and catabolism), but this effect was disrupted by fluctuating salinity. These findings indicate that exhaustive exercise and osmotic stress interactively affect whole-body protein homeostasis and functional capacity of the foot muscle in soft shell clams which might contribute to reduced burrowing activity of bivalve bioturbators in osmotically challenging environments such as estuaries and shallow coastal zones.

Sandhoff disease (SD) results from mutations in the HEXB gene, subsequent deficiency of N-acetyl-β-hexosaminidase (Hex) and accumulation of GM2 gangliosides. SD leads to progressive neurodegeneration and early death. However, there is a lack of established SD biomarkers, while the pathogenesis etiology remains to be elucidated. To identify potential biomarkers and unveil the pathogenic mechanisms, metabolomics analysis with reverse phase liquid chromatography (RPLC) was conducted. A total of 177, 112 and 119 metabolites were found to be significantly dysregulated in mouse liver, mouse brain and human hippocampus samples, respectively (p .05, ID score 0.5). Principal component analysis (PCA) analysis of the metabolites showed clear separation of metabolomics profiles between normal and diseased individuals. Among these metabolites, dipeptides, amino acids and derivatives were elevated, indicating a robust protein catabolism. Through pathway enrichment analysis, we also found alterations in metabolites associated with neurotransmission, lipid metabolism, oxidative stress and inflammation. In addition, N-acetylgalactosamine 4-sulphate, key component of glycosaminoglycans (GAG) was significantly elevated, which was also confirmed by biochemical assays. Collectively, these results indicated major shifts of energy utilization and profound metabolic impairments, contributing to the pathogenesis mechanisms of SD. Global metabolomics profiling may provide an innovative tool for better understanding the disease mechanisms, and identifying potential diagnostic biomarkers for SD.

Sow productivity improvements continue to increase metabolic demands during lactation. During the peripartum period, energy requirements increase by 60%, and amino acid needs increase by 150%. As litter size has increased, research on peripartum sows has focused on increasing birth weight, shortening farrowing duration to reduce stillbirths and improving colostrum composition and yield. Dietary fibre can provide short-chain fatty acids to serve as an energy source for the uterus prior to farrowing; however, fat and glucose appear to be the main energy sources used by the uterus during farrowing. Colostrum immunoglobulin G concentration can be improved by increasing energy and amino acid availability prior to farrowing; however, the influence of nutrient intake on colostrum yield is unequivocal. As sows transition to the lactation period, nutrient requirements increase with milk production demands to support large, fast-growing litters. The adoption of automated feed delivery systems has increased feed supply and intake of lactating sows; however, sows still cannot consume enough feed to meet energy and amino acid requirements during lactation. Thus, sows typically catabolise body fat and protein to meet the needs for milk production. The addition of energy sources to lactation diets increases energy intake and energy output in milk, leading to a reduction in BW loss and an improvement in litter growth rate. The supply of dietary amino acids and CP close to the requirements improves milk protein output and reduces muscle protein mobilisation. The amino acid requirements of lactating sows are variable as a consequence of the dynamic body tissue mobilisation during lactation; however, lysine (Lys) is consistently the first-limiting amino acid. A regression equation using published data on Lys requirement of lactating sows predicted a requirement of 27 g/day of digestible Lys intake for each 1 kg of litter growth, and 13 g/day of Lys mobilisation from body protein reserves. Increases in dietary amino acids reduce protein catabolism, which historically leads to improvements in subsequent reproductive performance. Although the connection between lactation catabolism and subsequent reproduction remains a dogma, recent literature with high-producing sows is not as clear on this response. Many practical aspects of meeting the nutrient requirements of lactating sows have not changed. Sows with large litters should approach farrowing without excess fat reserves (e.g.

Increased plasma α-1 acid glycoprotein (AGP) is correlated with reduced growth rates in neonatal swine. The specific physiological mechanisms contributing to this relationship are unknown. This study was performed to determine if AGP can modify muscle metabolism by examining glucose oxidation and protein synthesis in the C2C12 muscle cell line. Cells were used for experiments 4 days post-fusion as myotubes. Myotubes were exposed to AGP for 24 h, with the last 4 h used to monitor 14C-glucose oxidation or to measure protein synthesis by incorporation of 3H-tyrosine. Treatment of C2C12 myotubes with mouse AGP (100 µg/ml) reduced glucose oxidation (P0.05, n=6 trials), whereas incubation with both AGP and insulin reduced 3H-tyrosine release by 15% (P

Gastric digestion is crucial for protein breakdown. Magnetic resonance techniques have a great deal of potential but remain underexplored with regard to their application in the study of food digestion via MRI-markers, such as transverse (R2) and longitudinal (R1) relaxation rates. R2 has been used to monitor gastric digestion of whey protein gels, but only in a static in vitro model. It is essential to investigate whether relaxation rates can be valid measures of digestion under dynamic circumstances. We developed a novel MRI-compatible semi-dynamic gastric simulator (MR-GAS) that includes controlled gastric secretion, emptying and mixing at body temperature. PH and protein hydrolysis were measured during protein gel digestion in the MR-GAS. R2 and R1 of the supernatant were measured by time-domain nuclear magnetic resonance (TD-NMR). The stomach chamber of the MR-GAS was also scanned with MRI to measure R2 and R1. For TD-NMR, 99% of the variance in R2 and 96% of variance in R1 could be explained as a function of protein concentration and [H+]. For MRI, the explained variances were 99% for R2 and 60% for R1. From these analysis, the obtained equations enabled the prediction of protein concentration and pH by R2 and R1. The normalised root mean squared deviation of the predictions for protein concentration were 0.15 (NMR) and 0.18 (MRI), and for pH were 0.12 (NMR) and 0.29 (MRI). In conclusion, the MR-GAS model may be used in a clinical MRI to monitor gastric digestion under in vitro dynamic circumstances, by measuring R2 and R1. These results underscore the potential of MRI to monitor nutrients hydrolysis and pH changes in future in vivo studies.

The aim of the present study was to evaluate changes in the composition of non-volatile metabolites in low-temperature sausage, stored for periods of 0, 2, 4, 6, 8, 10, or 12 d at 20 °C. Evaluations were conducted using untargeted liquid chromatography-mass spectrometry (LC–MS/MS)-based metabolomics. Compared with samples from day 0, significant differences (variable importance in projection (VIP) > 2, p < 0.05) were observed in 245 non-volatile metabolites, divided into 9 classes, in samples from the remaining storage periods. Moreover, an increased abundance of amino acids, peptides, and acylcarnitines potentially indicated increased protein catabolism and incomplete β-oxidation, followed by quality deterioration. These findings provide further understanding of the metabolic changes that occur in low-temperature meat products during storage, which could contribute to the identification of chemical biomarkers to assess meat quality.

Pancreatic cancer cells with limited access to free amino acids can grow by scavenging extracellular protein. In a murine model of pancreatic cancer, we performed a genome-wide CRISPR screen for genes required for scavenging-dependent growth. The screen identified key mediators of macropinocytosis, peripheral lysosome positioning, endosome-lysosome fusion, lysosomal protein catabolism, and translational control. The top hit was GCN2, a kinase that suppresses translation initiation upon amino acid depletion. Using isotope tracers, we show that GCN2 is not required for protein scavenging. Instead, GCN2 prevents ribosome stalling but without slowing protein synthesis; cells still use all of the limiting amino acids as they emerge from lysosomes. GCN2 also adapts gene expression to the nutrient-poor environment, reorienting protein synthesis away from ribosomes and toward lysosomal hydrolases, such as cathepsin L. GCN2, cathepsin L, and the other genes identified in the screen are potential therapeutic targets in pancreatic cancer.

Summary Background & aims It has been demonstrated that the relative content and profile of essential amino acids (EAA) play a determining role for stimulation of muscle protein synthesis (MPS) following intake of pure EAA or protein alone. Methods To test if this also holds in the context of mixed meals at both whole body and muscle levels, twelve older subjects (57–74 yrs) received primed continuous infusion of L-[ring-2H5]phenylalanine and L-[ring-2H2]tyrosine over a 9-h experimental period to determine whole body protein kinetics and MPS in the fasted state and following consumption of egg-based (EGG) or cereal-based (CEREAL) isocaloric and isonitrogenous breakfast. A standardized lunch, primarily consisting of beef protein was also consumed by each group. Whole body protein kinetics [protein synthesis (PS), breakdown (PB), and net balance (NB)] were expressed as changes from basal fasted period. Results We found that EGG breakfast resulted in a greater NB through a greater suppression of PB compared with the CEREAL breakfast. The greater NB during the post-breakfast period with the EGG was normalized following the standard lunch despite the sustained elevations in plasma EAA concentrations. However, the EGG breakfast stimulated both PS and PB compared with the CEREAL breakfast during the post-lunch period. MPS was not different between meals despite larges differences in the plasma EAA responses. Conclusions We conclude that in the context of mixed meals, quality of protein affects NB through changes in protein breakdown and affects protein turnover following subsequent meal intake.

Background: Acute inflammation, and subsequent release of bacterial products (e.g. LPS), inflammatory cytokines, and stress hormones, is catabolic, and the loss of lean body mass predicts morbidity and mortality. Lipid intermediates may reduce protein loss, but the roles of free fatty acids (FFAs) and ketone bodies during acute inflammation are unclear.Objective: We aimed to test whether infusions of 3-hydroxybutyrate (3OHB), FFAs, and saline reduce protein catabolism during exposure to LPS and Acipimox (to restrict and control endogenous lipolysis).Design: A total of 10 healthy male subjects were randomly tested 3 times, with: 1) LPS, Acipimox (Olbetam) and saline, 2) LPS, Acipimox, and nonesterified fatty acids (Intralipid), and 3) LPS, Acipimox, and 3OHB, during a 5-h basal period and a 2-h hyperinsulinemic, euglycemic clamp. Labeled phenylalanine, tyrosine, and urea tracers were used to estimate protein kinetics, and muscle biopsies were taken for Western blot analysis of protein metabolic signaling.Results: 3OHB infusion increased 3OHB concentrations (P < 0.0005) to 3.5 mM and decreased whole-body phenylalanine-to-tyrosine degradation. Basal and insulin-stimulated net forearm phenylalanine release decreased by >70% (P < 0.005), with both appearance and phenylalanine disappearance being profoundly decreased. Phosphorylation of eukaryotic initiation factor 2α at Ser51 was increased in skeletal muscle, and S6 kinase phosphorylation at Ser235/236 tended (P = 0.074) to be decreased with 3OHB infusion (suggesting inhibition of protein synthesis), whereas no detectable effects were seen on markers of protein breakdown. Lipid infusion did not affect phenylalanine kinetics, and insulin sensitivity was unaffected by interventions.Conclusion: During acute inflammation, 3OHB has potent anticatabolic actions in muscle and at the whole-body level; in muscle, reduction of protein breakdown overrides inhibition of synthesis. This trial was registered at clinicaltrials.gov as NCT01752348.

Assessment of whole body protein turnover (WBPT) can provide fundamental information about protein kinetics which underpins the conservation of lean tissue. Reliability and methodology studies on the measurement of WBPT are scarce. This study aimed to assess the effects of urine collection duration (9 versus 12 h) and the reproducibility of WBPT with the end product method calculated from ammonia as the end product.WBPT was assessed in 21 healthy participants (11M, 10F) on 2 test days. WBPT was assessed using the end product method with a single dose ofThe CV for protein flux averaged 10% and 12% for 9 and 12-h urine collections respectively. Protein flux, synthesis and balance were significantly higher and protein breakdown significantly lower with 9-h urine collections compared to 12-h collections (P 0.01) and there was a trend towards increasingly greater overestimation of 9-h calculated WBPT kinetics with greater overall rates of WBPT. Correlations between the 9 and 12-h values were strong (r 0.962, P 0.001 for all variables).The reproducibility of WBPT with ammonia as the end product was similar to previously reported reproducibility of the gold standard precursor technique. The use of a 12-h urine collection is more effective to achieve full turnover of the ammonia free amino acid (AA) pool.

Surgical stress provokes protein catabolism and hyperglycaemia that is enhanced in patients with type 2 diabetes (T2DM), and increases perioperative morbidity. This study hypothesized that perioperative administration of high dose intravenous (IV) amino acids (AA) will augment protein balance in T2DM patients receiving tight plasma glucose control via continuous IV insulin compared to standard plasma glucose control via subcutaneous (SC) insulin sliding scale.Eighteen patients with well-controlled T2DM (HbA1C% 7.1) undergoing colorectal surgery were assigned randomly to receive standard glucose control (6-10 mmol/l, SC insulin, n = 9) or tight glucose control (4-6 mmol/l, IV insulin, n = 9). Both groups received general anaesthesia and epidural analgesia. AA (1 ml/kg h Aminoven™ 10%, ∼2.4 g/kg d) were infused via a peripheral vein for two 3-h periods: at the beginning of surgery and in the post-operative care unit. Whole-body protein and glucose kinetics were assessed by stable isotope tracers, L-[1-Whole-body protein balance was positive after surgery in all patients. Since protein synthesis, breakdown and leucine oxidation were comparable in both groups, whole body protein balance was not different (p = 0.605). Tight glucose control suppressed endogenous glucose production (EGP, p 0.001) and increased glucose clearance (p 0.001) compared to standard glucose control during both study periods. No episode of hypoglycaemia occurred in either group.High-dose perioperative AA administration under optimal anti-catabolic care with epidural analgesia was effective in achieving a positive protein balance in T2DM patients undergoing surgery that was independent of glycaemic control strategy. Continuous IV insulin maintained normoglycaemia by inhibiting EGP and increasing glucose clearance. Improved glucose control, without a pronounced increase in protein balance with the intravenous insulin regimen, suggests perioperative protein metabolism may be less sensitive to insulin than is glucose.

Introduction: Early studies have shown that patients with chronic kidney disease (CKD) are able to maintain nitrogen balance despite significantly lower protein intake, but how and to what extent muscle protein metabolism adapts to a low-protein diet (LPD) or to a supplemented very LPD (sVLPD) is still unexplored. Methods: We studied muscle protein turnover by the forearm perfusion method associated with the kinetics of 2H-phenylalanine in patients with CKD: (i) in a parallel study in subjects randomized to usual diet (1.1 g protein/kg, n = 5) or LPD (0.55 g protein/kg, n = 6) (Protocol 1); (ii) in a crossover, self-controlled study in subjects on a 0.55 g/kg LPD followed by a sVLPD (0.45 g/kg + amino/ketoacids 0.1 g/kg, n = 6) (Protocol 2). Results: As compared with a 1.1 g/kg containing diet, a 0.55 g/kg LPD induced the following: (i) a 17% to 40% decrease in muscle protein degradation and net protein balance, respectively, (ii) no change in muscle protein synthesis, (iii) a slight (by approximately 7%, P < 0.06) decrease in whole-body protein degradation, and (iv) an increase in the efficiency of muscle protein turnover. As compared with an LPD, an sVLPD induced the following: (i) no change in muscle protein degradation, and (ii) an approximately 50% decrease in the negative net protein balance, and an increase in the efficiency of muscle protein turnover. Conclusion: The results of these studies indicate that in patients with CKD the adaptation of muscle protein metabolism to restrained protein intake can be obtained via combined responses of protein degradation and the efficiency of recycling of amino acids deriving from protein breakdown. Keywords: amino acids, chronic kidney disease, ketoacids, low-protein diet, nutrition

The acquisition of rapid desiccation tolerance for the resurrection plant Boea hygrometrica requires a cycle of drought acclimation. Here, a gas chromatography-mass spectrometry based global untargeted metabolite profiling analysis was conducted on leaves of B. hygrometrica during dehydration and rehydration. Metabolic profiles between non-acclimated and drought acclimated plants were clearly distinguished in principal-component and hierarchical cluster analysis, indicating that drought acclimation response required the process in metabolic reprogramming. Accumulation of numerous metabolites involved in carbon and energy metabolism as well as amino acid and fatty acid biosynthesis was observed to be significantly changed upon drought acclimation and dehydration treatments. Using partial least-squares discriminant analysis, 25 known metabolites and 51 compounds with unknown structures were identified as putative biomarkers contributed to the discrimination between drought acclimated and non-acclimated plants. Remarkably, as revealed by transcriptome data analysis, some up-accumulated metabolites in acclimated plants, such as maltose, glutaric acid, L-tryptophan and α-tocopherol, which are in good correlation with the increased desiccation tolerance, and were precisely controlled by the transcript changes in multigene family members of related enzymes. Furthermore, genes positively correlated with the up-accumulated biomarkers in transcription abundances were enriched in multiple biological processes, such as “ubiquitin-dependent protein catabolism” and “abscisic acid-activated signaling pathway”. Taken together, our observations indicate that the accumulation of important metabolites correlates with the transcriptional activity of biosynthetic related enzymes and putative regulators involved in ubiquitination and ABA signal transduction during a short period of drought acclimation might contribute to the acquisition of rapid desiccation tolerance in B. hygrometrica.

Amino acids are essential components of plant metabolism, not only as constituents of proteins, but also as precursors of important secondary metabolites and as carriers of organic nitrogen between the organs of the plant. Transport across intracellular membranes and translocation of amino acids within the plant is mediated by membrane amino acid transporters. The past few years have seen the identification of a new family of amino acid transporters in Arabidopsis, the characterization of intracellular amino acid transporters, and the discovery of new roles for already known proteins. While amino acid metabolism needs to be tightly coordinated with amino acid transport activity and carbohydrate metabolism, no gene involved in amino acid sensing in plants has been unequivocally identified to date. This review aims at summarizing the recent data accumulated on the identity and function of amino acid transporters in plants, and discussing the possible identity of amino acid sensors based on data from other organisms.

Summary Objectives The use of proper diet in athletes is one of the most important elements of sports training. Moreover, the essentials in planning diet for athletes among other things should be taken into account: energy balance, the nutritional value of meals and supplements of diet. The main aim of this study was to estimate the level of whole body proteins catabolism, nitrogen and energy balance in young gymnasts. Methods In the present study participated young gymnastics (13.0 ± 0.5 year), which have been trained artistic gymnastics since 6.8 ± 0.45 year. The investigation contained two times measuring urea in 24 h urine collection, measuring energy expenditure indirect method by 24 h monitoring heart rate, and supplementation by 7 days products contain protein and carbohydrate. Results Concentration of urea nitrogen in urine at the first test before supplementation was 8.0 ± 0.75 g/d. The energy expenditure in young boys before supplementation was negative (−915 ± 130 kcal). The concentration of urea nitrogen was lower in 4 out of 5 examined boys as compared to the first test after supplementation. The difference between the level of nitrogen balance output and the level after supplementation was statistically significant (P ≤ 0.05). Conclusion The supplementation with proteins and carbohydrates products immediately after training may lead to decrease concentration of urea nitrogen and whole body protein catabolism. Monitoring energy expenditure, food nutrients, and calories in diet during and after training should be an integral part of planning for long-term training program.

Highlights • Poor nutritional status and decreased lean body mass. • Decline in pulmonary function. • Increased mortality from lung disease. • Microvascular complications. • Macrovascular complications (not currently a significant complication but this may change with modulators)., Cystic fibrosis related diabetes (CFRD) occurs in at least 40–50% of adults with CF. With other forms of diabetes, microvascular and macrovascular disease are the major causes of morbidity and mortality. Macrovascular disease is rare in CF. While microvascular disease does occur in this population, there are CF-specific diabetes complications that have a more important impact on prognosis. The additional diagnosis of diabetes in CF is associated with decreased lung function, poor nutritional status, and an overall increase in mortality from lung disease. These negative findings start even before the clinical diagnosis of CFRD, during the period when patients experience abnormal glucose tolerance related to insulin insufficiency. The main mechanisms by which CFRD negatively affects prognosis are thought to be a combination of 1) protein catabolism, decreased lean body mass and undernutrition resulting from insulin insufficiency, and 2) an increased pro-inflammatory and pro-infectious state related to intermittent hyperglycemia. With the introduction of CFTR modulators, the care of CF patients has been revolutionized and many aspects of CF health such as BMI and lung function are improving. The impact of these drugs on the adverse prognosis related to the diagnosis of diabetes in CF, as well as the potential to delay or prevent onset of CFRD remain to be determined.

Background The aging process is often associated with the presence of sarcopenia. Although changes in the plasma concentration of several amino acids have been observed in older adults, it remains unclear whether these changes are related to disturbances in whole body production and/or interconversions. Methods We studied 10 healthy young (~ 22.7 y) and 17 older adults (~ 64.8 y) by administering a mixture of stable amino acid tracers in a pulse and in a primed constant infusion. We calculated whole body production (WBP) and metabolite to metabolite interconversions. In addition, we measured body composition, muscle function, and provided questionnaires to assess daily dietary intake, physical activity, mood (anxiety, depression) and markers of cognitive function. Plasma enrichments and metabolite concentrations were measured by GC- and LC-MS/MS and statistics were performed by student t-test. Results Older adults had a 11% higher body mass index (p = 0.04) and 27% reduced peak leg extension force (p = 0.02) than the younger group, but comparable values for muscle mass, mood and cognitive function. Although small differences in several plasma amino acid concentrations were observed, we found older adults had about 40% higher values of WBP for glutamine (221 ± 27 vs. 305 ± 21 μmol/kg ffm/h, p = 0.03) and tau-methylhistidine (0.15 ± 0.01 vs. 0.21 ± 0.02 μmol/kg ffm/h, p = 0.04), 26% lower WBP value for arginine (59 ± 4 vs. 44 ± 4 μmol/kg ffm/h, p = 0.02) and a reduction in WBP (50%; 1.23 ± 0.15 vs. 0.69 ± 0.06 μmol/kg ffm/h, p = 0.001) and concentration (25%; 3.5 ± 0.3 μmol/l vs. 2.6 ± 0.2 μmol/l, p = 0.01) for β-Hydroxy β-Methylbutyrate. No differences were observed in protein catabolism. Clearance of arginine was decreased (27%, p = 0.03) and clearance of glutamine (58%, p = 0.01), leucine (67%, p = 0.001) and KIC (76%, p = 0.004) were increased in older adults. Conclusions Specific differences exist between young and older adults in amino acid metabolism.

CBL (Casitas B cell lymphoma), an important ubiquitin protein ligase, is involved in protein folding, protein maturation, and proteasome-dependent protein catabolism in different cells. However, its role in cardiac hypertrophy is still unclear. In this study, we found that expression of CBL is increased in an Ang II-induced mouse cardiac hypertrophy animal model and in Ang II-treated H9C2 cells. Interference with CBL expression attenuates the degree of myocardial hypertrophy as well as the expression of hypertrophy-related genes in H9C2 cells. Further research found that CBL aggravates myocardial hypertrophy by activating HIF-1α, which is an aggravating factor for hypertrophy. The effect of CBL on promoting myocardial hypertrophy was reversed by interference with HIF-1α. Mechanistically, we found that CBL directly interacted with and degraded VHL by increasing its ubiquitination level, which is a widely accepted regulatory factor of HIF-1α. Finally, our results showed that CBL was partially dependent on degradation of VHL and that activation of HIF-1α promoted myocardial hypertrophy. Collectively, these findings suggest that strategies based on activation of the CBL/HIF-1α axis might be promising for the treatment of hypertrophic cardiomyopathy.

The emergence of resistance to albendazole has encouraged the search for effective alternatives for cysticercosis and other parasitosis treatment. RCB15 is a benzimidazole derivative that may be used against such diseases. The aim of this study was to determine the in vitro effect of RCB15 on the alternative energetic pathways of Taenia crassiceps cysticerci. The cysticerci were in vitro exposed to albendazole sulphoxide (ABZSO) or RCB15 at different concentrations during 24h. The cysticerci extract and the culture medium were analyzed through spectrophotometry and high performance liquid chromatography as to detect glucose, urea, creatinine and organic acids of the energetic metabolism. The drugs did not influence the protein catabolism. Fatty acids oxidation was enhanced through significantly higher acetate concentrations in the groups treated with RCB15 and ABZSO. Beta-hydroxybutyrate concentrations were decreased which indicates the use of fatty acids towards acetyl-CoA synthesis. There was a decrease in glucose uptake and pyruvate concentrations. The absence of lactate indicates the use of pyruvate in gluconeogenesis. Therefore it is possible to conclude that RCB15 enhanced the alternative energetic pathways of cysticerci in vitro exposed to different concentration, with emphasis on the fatty acids catabolism.

The main objective of the study was to evaluate the bioaccessibility of rapeseed soluble proteins (RSPs) by simulated gastrointestinal digestion and isolated intestine methods. In the gastric phase, the degree of hydrolysis (DH) of RSPs increased from 2.6% to 8.2%, while the surface hydrophobicity (H0) decreased from 62.8% to 59.1%. Further digestion in the intestinal phase brought DH and H0 to 18.2% and 50.5%, respectively. Scavenging DPPH and hydroxyl radical abilities and reducing power of RSPs increased slightly after digestion by pepsin, however, a distinct increment was achieved after subsequent digestion by trypsin. Molecular weight distributions indicated that RSPs above 3000 Da decreased significantly from 87.0% to 24.1% after digestion, revealing these fractions were more easily digested by gastrointestinal proteases than those below 3000 Da. The absorption of RSPs by intestine reached the maximum (28.6%) at the concentration of 5 mg/mL after incubation for 60 min. For the fraction below 307 Da, the absorption efficiency increased with incubation from 20 to 60 min.

World ageing is a growing world concern to which dietary solutions may offer effective measures to tackle malnutrition and support a high quality of life. This study applied a dynamic in vitro model mimicking adult or elderly gastro-duodenal digestion, coupled with proteomic analyses to interrogate the differences in the breakdown of an enteral nutrition formula as well as purified α-lactalbumin (α-lac), β-lactoglobulin (β-lg) and lactoferrin (LF). First, the importance of gastric lipase and various age-related differences in gastro-intestinal conditions (e.g. gastric pH profiles) is shown. Droplet sizing and fluorescent microscopy substantiate emulsion coalescence is favored under adult conditions while flocculation is dominant under elderly conditions. SDS-PAGE analyses of digesta also substantiate differences between adults and seniors in terms of protein breakdown. LC-MS proteomic analyses reveal that digesta samples differ in the incidence of amino acids therein. Comparing proteomic data to databases of known bioactive peptides reveals that β-lg and α-lac may generate more bioactive peptides under adult conditions contrary to LF, which was found to be a better source for bioactive peptides under elderly gut conditions. In addition, proteomic data was mined using PeptideRanker, a bioinformatic tool, in an attempt to identify novel peptide candidates. These efforts generated lists of potentially novel bioactive peptides with a bioactivity probability exceeding 0.6 and in some cases even having a bioactivity probability exceeding 0.9. This study brings forward new information regarding lipids and proteins digestive fate of enteral formula and whey proteins in the elderly gut that stimulate the need for further investigations.

The purpose of this study was to investigate whether low-intensity pulsed ultrasound (LIPUS) promotes exercise-induced muscle hypertrophy. Twenty-four adult Sprague-Dawley (SD) rats were randomly assigned to three groups (n = 8 per group): normal control group (NC), treadmill exercise group (TE) and treadmill exercise + LIPUS group (TE + LIPUS). The TE + LIPUS group received a LIPUS treatment (1 MHz, 30 mW/cm2) at the gastrocnemius for 20 min/d after treadmill exercise. The TE group was sham-treated. Eight weeks of treadmill training successfully established the exercise-induced muscle hypertrophy model. Muscle strength, muscle mass and muscle fiber cross-sectional area were significantly increased in the TE + LIPUS group compared with the TE group. Moreover, LIPUS treatment significantly upregulated the expression of Akt, mTOR, p-Akt and p-mTOR and significantly downregulated the expression of MSTN, ActRIIB, FoxO1 and its phosphorylation. The results indicated that LIPUS promotes exercise-induced muscle hypertrophy by facilitating protein synthesis and inhibiting the protein catabolism pathway.

This research investigated how ploidy level (diploid versus triploid) affects the heat shock protein (HSP) response in erythrocytes under different thermal stress regimes, both in vivo and in vitro, in Atlantic salmon (Salmo salar) and brook charr (Salvelinus fontinalis) in order to address the question of why triploids typically have reduced thermal tolerance. A preliminary study confirmed that identical volumes of diploid and triploid erythrocytes (which equates to a smaller number of larger cells for triploids compared to diploids) did not differ in total protein synthesis rates. After chronic (100 d) acclimation of fish to 5, 15 and 25 °C, triploid erythrocytes had lower HSP70, HSP90, heat shock factor 1 (HSF1) and ubiquitin (free and total) levels than diploids in both species. Furthermore, Atlantic salmon erythrocytes showed significantly higher protein breakdown (based on conjugated ubiquitin levels) in triploids than diploids after acute heat stress in vitro, but no significant difference was detected between ploidies after acute cold stress. These results indicate that: 1) triploid erythrocytes synthesize more total protein per cell than diploids as a result of increased cell size; 2) triploids have sufficient total HSP levels for survival under low stress conditions; and 3) the lower basal titres of HSPs in triploids may be a handicap when combating acute stress. Taken together, this suggests that triploids are limited in their ability to withstand thermal stress because of a reduced ability to maintain proteostasis under stressful conditions.

Over the past four decades the epidemiology of multiple organ failure (MOF) has evolved as a result of advances in care. A series of paradigms have been described to explain the pathophysiology of the new emerging predominant phenotypes. With the most recent improvements in the delivery of critical care, patients are less-frequently expiring early in their clinical course and in-hospital MOF-related mortality is on the decline. Unfortunately, this has resulted in a dramatic increase in the number of chronic critically ill patients (CCI) who linger in the intensive care unit (ICU), have high resource utilization, are discharged to non-home locations, experience sepsis recidivism requiring readmission, have persistent cognitive and functional impairments, and poor long-term survival. Within this population, we have proposed that a substantial subset of these patients suffer from a new phenotype termed Persistent Inflammation, Immunosuppression, and Catabolism Syndrome (PICS) which underlies these poor outcomes. While the mechanism(s) of PICS are under investigation, there is evidence that myelodysplasia with expansion of myeloid derived suppressor cells, innate and adaptive immune suppression and protein catabolism with malnutrition are major contributors. Optimal care of these patients will require a novel multimodality intervention using pharmacotherapy, physiotherapy and nutritional support.

Preterm infants, especially very low birth weight (VLBW

Sex and age differences in hepatic expression of drug-metabolizing enzyme genes could cause variations in drug metabolism, but has not been fully elucidated, especially in Asian population. In this study, the global expression of human hepatic genes was analyzed by microarrays in 40 Japanese subjects (27 males and 13 females). Thirty-five sex-biased genes were identified (P

Protein modifications occurring during sterilisation of infant formulas can affect protein digestibility and release of bioactive peptides. The effect of glycation and cross-linking on protein breakdown and release of β-casomorphins was evaluated during in vitro gastro-intestinal digestion (GID) of six sterilised model systems of infant formula. Protein degradation during in vitro GID was evaluated by SDS-PAGE and by measuring the nitrogen content of ultrafiltration (3kDa) permeates before and after in vitro GID of model IFs. Glycation strongly hindered protein breakdown, whereas cross-linking resulting from β-elimination reactions had a negligible effect. Only β-casomorphin 7 (β-CM7) was detected (0.187-0.858mgL(-1)) at the end of the intestinal digestion in all untreated IF model systems. The level of β-CM7 in the sterilised model systems prepared without addition of sugars ranged from 0.256 to 0.655mgL(-1). The release of this peptide during GID was hindered by protein glycation.

An unexpected observation among the COVID-19 pandemic is that smokers constituted only 1.4-18.5% of hospitalized adults, calling for an urgent investigation to determine the role of smoking in SARS-CoV-2 infection. Here, we show that cigarette smoke extract (CSE) and carcinogen benzo(a)pyrene (BaP) increase ACE2 mRNA but trigger ACE2 protein catabolism. BaP induces an aryl hydrocarbon receptor (AhR)-dependent upregulation of the ubiquitin E3 ligase Skp2 for ACE2 ubiquitination. ACE2 in lung tissues of non-smokers is higher than in smokers, consistent with the findings that tobacco carcinogens downregulate ACE2 in mice. Tobacco carcinogens inhibit SARS-CoV-2 Spike protein pseudovirions infection of the cells. These data indicate that recommendation of cessation of tobacco smoking remains valid because it is the carcinogens that are responsible for ACE2 degradation.

Fat is the major fuel for migratory flight of birds, but protein is also catabolized. Flight range could be reduced if protein is used too quickly from muscles and organs, and it is important to understand factors that influence protein catabolism. Previous correlative studies suggested high protein diets may increase protein use in flight, although a wind tunnel study with yellow-rumped warblers (Setophaga coronata) did not support this relationship. We tested the hypothesis that diet composition affects nutrient oxidation in resting, fasted yellow-rumped warblers. For method development, we gavaged or subcutaneously injected warblers with 13C labelled glucose or leucine, and measured δ13C of breath CO2 in real time using infrared laser spectrometry. Regardless of route of administration, leucine had greater instantaneous and cumulative oxidation than glucose. Compared to subcutaneous injection, gavaged birds reached maximum oxidation rate faster for leucine and glucose, respectively, had a higher maximum oxidation rate, and reached final cumulative oxidation approximately faster for leucine or glucose, respectively, indicating immediate oxidation of the substrates by the digestive system. Warblers (N = 10 each) were fed isocaloric 60% carbohydrate or 60% protein diets for minimum 2 weeks, and subcutaneously injected with 13C labelled glucose or leucine. Diet composition had little effect on oxidation kinetics except that warblers fed high-carbohydrate reached final cumulative oxidation of leucine more quickly than those fed high-protein. The findings do not support the hypothesis that high protein diets increase the oxidation of protein during negative energy states in migratory birds, and provide methodology that could be applied to test it in flight.

Recent research has shown that salt levels of minimal 3.5 g/kg diet result in best performance in piglets between 7 and 11 kg. In the US, pig diets tend to have higher salt levels and higher crude protein (CP) levels than in Europe. Higher CP levels may lead to higher protein breakdown, while higher salt levels may lead to increased water consumption, which might facilitate nitrogen clearance through the kidneys. Therefore, we hypothesized that high salt levels are more important when the pigs receive a high CP diet. For this experiment, 96 pens of 5 piglets were divided over 8 treatments in a 2 × 4 factorial design with 2 protein levels (189 vs 223 g/kg) and 4 salt levels (1.9, 2.5, 3.1, and 3.8 g/kg in the low protein diet and 1.9, 2.6, 3.2, and 3.9 g/kg in the high protein diet). Different salt levels were obtained by exchanging SiO2 for NaCl. The low protein diet was formulated to contain 11.8 g SID lysine and 9.85 MJ net energy/kg diet, while the high protein diet had 12.5 g SID lysine and 10.40 MJ net energy/kg diet. Salt level did not affect performance nor did it interact with crude protein level. Fecal consistency worsened as salt levels increased during the first week after weaning. The latter might be a result of either increased water consumption or increased osmotic pressure in combination with the piglets’ limited absorption capacity shortly after weaning, especially when receiving diets without zinc oxide or antibiotics. The higher nutrient density in the high protein diet resulted in lower feed intake with similar growth, leading to improved feed efficiency per kg diet, but not per MJ of net energy consumed. Based on the present experiment, we could not confirm our hypothesis that increased protein levels lead to higher sodium requirements in weaned piglets.

Few studies have investigated the effects of long-term low-concentration nitrite exposure and detoxification on growth and physiology of fish. Considering this, the present study focuses on the effect of nitrite exposure and detoxification on Chinese perch. Chinese perch were exposed to water with 0 mg/L (C group) and 0.3 mg/L nitrite (N group) for 30 and 60 days, respectively (C30, C60, N30, N60). No significant differences in the average daily growth (ADG), food intake (FI), feed conversion ratio (FCR), and histology were observed between C30 group and N30 group, while N30 group exhibited significantly lower protein retention and crude protein content and significantly higher levels of blood urea nitrogen (BUN) and higher activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) than C30 group. The ADG, FI, plasma glucose, total triglyceride, total cholesterol, the activities of SOD, CAT and GSH-Px were significantly decreased in N60 group, whereas the levels of methemoglobin, BUN, and immune responses-related indicators (globulin, aspartate aminotransferase, and alanine aminotransferase) were significantly increased. The damages to the gill and liver of fish and the increased expressions of pro-inflammatory cytokines genes (IL-Iβ, p65-NF-κB, TNFα and PAI) were observed in N60 group. In addition, fish exposed to nitrite for 30 and 60 days were detoxified in nitrite-free water for 30 days (DN30 and DN60). The detoxification significantly increased the protein retention and protein content. However, detoxification did not improve the appetite or fully restore histopathological injury in DN60 group. Those results indicated that 30-day exposure to 0.3 mg/L of nitrite promoted protein catabolism for providing energy to activate the defense, and that the detoxification could completely reverse the effects. The 60-day exposure to nitrite could disrupt defense system, cause histopathological damage, and affect energy metabolism. Moreover, nitrite exposure significantly decreased appetite. Above factors might be responsible for growth inhibition. The detoxification post 60-day nitrite exposure only restored partial physiological functions. Our findings provide a reference for the concentration control of nitrite in feed farming of Chinese perch.

It is well known that scorpions are highly adapted to thermal temperatures. However, little is known of the metabolic and respiration adaptations caused by temperature fluctuations in these animals. Therefore we used the African burrowing scorpion Opistophthalmus latimanus to measure the effect of temperature on its metabolism and respiration. Radioactive d-glucose was injected into the ventral sinus of the circulatory system and metabolites of d-glucose were determined after six hour incubation at four temperatures (7, 17, 25 and 37°C). The oxygen consumption rate (ṀO2) and carbon dioxide production rate (ṀCO2) were measured simultaneously at 17, 25 and 37°C. The metabolomics investigation included LC-MS, GC-MS and NMR analytical platforms. The average radioactivity recovered after the carbon-14 d-glucose injection, glycogen precipitation and column fractionation at the four temperatures was between 92.4% and 95.0%. Strong acids, CO2 and neutral compounds all increased with temperature, while glycogen and neutral sugars decreased as the temperature increased. Weak acids initially increased with temperature, then decreased again as the temperature was increased to 37°C. Respiration also gradually increased as the temperature was increased. Metabolomics identified 23 metabolites that were significantly influenced by temperature. Pathway analysis of these metabolites indicated numerous metabolic pathways that were affected by temperature, clearly demonstrating that the scorpion uses proteins, lipids and carbohydrates at higher temperatures to generate energy. However, protein catabolism seems to be the main source of energy at higher temperatures in these animals, although this needs to be confirmed in a more targeted metabolomics study.

BACKGROUND The neonatal methionine requirement must consider not only the high demand for rapid tissue protein expansion but also the demands as the precursor for a suite of critical transmethylation reactions. However, methionine metabolism is inherently complex because upon transferring its methyl group during transmethylation, methionine can be reformed by the dietary methyl donors choline (via betaine) and folate. OBJECTIVE We sought to determine whether dietary methyl donors contribute to methionine availability for protein synthesis in neonatal piglets. METHODS Yucatan miniature piglets aged 4-8 d were fed a diet that provided 38 μg folate/(kg·d), 60 mg choline/(kg·d), and 238 mg betaine/(kg·d) [methyl-sufficient (MS); n = 8] or a diet devoid of these methyl precursors [methyl-deficient (MD); n = 8]. After 5 d, dietary methionine was reduced from 0.30 to 0.20 g/(kg·d) in both groups. On day 6, piglets received a constant [1-13C]phenylalanine infusion to measure whole-body protein kinetics, and on day 8 they received a constant [3H-methyl]methionine infusion to measure tissue-specific protein synthesis in skeletal muscle, the liver, and the jejunum. RESULTS Whole-body phenylalanine flux, protein synthesis, and protein breakdown were 13%, 12%, and 22% lower, respectively, in the MD group than in the MS group (P < 0.05). Reduced whole-body protein synthesis in the MD piglets was attributed to 50% lower protein synthesis in skeletal muscle and the jejunum than in the MS piglets (P < 0.05). Furthermore, methionine availability in skeletal muscle was halved in piglets fed the MD diet (P < 0.05), and the specific radioactivity of methionine was doubled in the jejunum of MD piglets (P < 0.05), suggesting lower intestinal remethylation. Liver protein synthesis did not significantly differ between the groups, but secreted proteins were not measured. CONCLUSIONS Dietary methyl donors can affect whole-body and tissue-specific protein synthesis in neonatal piglets and should be considered when determining the methionine requirement.

Background: Chronic obstructive pulmonary disease (COPD) is a condition characterized by systemic low-grade inflammation that could increase the production of nitric oxide (NO), of which arginine is the sole precursor. Arginine is derived from the breakdown of protein and through the conversion of citrulline to arginine (de novo arginine production). Objective: Our objective was to study whole-body arginine and citrulline and related metabolism in stable COPD patients. Design: With the use of stable isotope methodology, we studied whole-body arginine and citrulline rates of appearance, de novo arginine (citrulline-to-arginine flux) and NO (arginine-to-citrulline flux) production, protein synthesis and breakdown rates, and plasma amino acid concentrations in a heterogeneous group of patients with moderate-to-severe COPD [n = 23, mean ± SE age: 65 ± 2 y, forced expiratory volume in 1 s (FEV1): 40% ± 2% of predicted], and a group of healthy older adults (n = 19, mean ± SE age: 64 ± 2 y, FEV1: 95% ± 4% of predicted). Results: Although plasma arginine and citrulline concentrations were comparable between COPD patients and controls, whole-body arginine (P = 0.015) and citrulline (P = 0.026) rates of appearance were higher in COPD patients and related to a 57% greater de novo arginine production (P < 0.0001). Despite a higher whole-body arginine clearance in COPD patients (P < 0.0001), we found no difference in NO production. Conclusion: In stable patients with moderate-to-severe COPD, endogenous arginine production is upregulated to support a higher arginine utilization that is unrelated to whole-body NO production. This trial was registered at clinicaltrials.gov as {"type":"clinical-trial","attrs":{"text":"NCT01173354","term_id":"NCT01173354"}}NCT01173354 and {"type":"clinical-trial","attrs":{"text":"NCT01172314","term_id":"NCT01172314"}}NCT01172314.

Replacement of canonical amino acids with unnatural amino acids (UAAs) can provide proteins with novel physicochemical properties and biological functions. In this study, as an alternative option to conventional cell-based methods, we used a cell-free protein synthesis system as a flexible platform for facile and efficient production of UAA-containing proteins. We designed a cell-free protein synthesis system derived from the extract of Escherichia coli cells to maximize the selective incorporation of UAAs into the protein structure. First, for the purpose of avoiding competitive incorporation of canonical amino acids and UAAs, the cell extract was extensively washed using a diafiltration process to remove residual amino acids, thereby making the protein synthesis reaction completely dependent upon the exogenous addition of amino acids. In addition, the relatively low affinity of UAAs for cognate aminoacyl-tRNA synthetase was kinetically overcome by increasing the concentration of UAAs to nonphysiological levels. As a result of these modifications of the cell-free synthesis systems, we were able to produce UAA-containing proteins at comparable yields to those of proteins made of canonical amino acids.

Chronic metabolic acidosis is not uncommon in patients with chronic kidney disease (CKD). Clinical practice guidelines suggest that clinicians administer alkali to maintain serum bicarbonate level at a minimum of 22 mEq/L to prevent the effects of acidosis on bone demineralization and protein catabolism. Small interventional studies support the notion that correcting acidosis slows CKD progression as well. Furthermore, alkaline therapy in persons with CKD and normal bicarbonate levels may also preserve kidney function. Observational studies suggest that targeting a serum bicarbonate level near 28 mEq/L may improve clinical outcomes above and beyond targeting a value ≥ 22 mEq/L, yet values > 26 mEq/L have been reported to be associated with incident heart failure and mortality in the CRIC (Chronic Renal Insufficiency Cohort) Study. Furthermore, correcting acidosis may provoke vascular calcification. This teaching case discusses several uncertainties regarding the management of acidosis in CKD, such as when to initiate alkali treatment, potential side effects of alkali, and the optimum serum bicarbonate level based on current evidence in CKD. Suggestions regarding the maximum sodium bicarbonate dose to administer to patients with CKD to achieve the target serum bicarbonate concentration are offered.

Rationale & Objectives Previously we reported a cohort of patients with coronavirus disease 2019 (COVID-19)–associated acute kidney injury (AKI) with striking biochemical evidence of tissue breakdown in the absence of apparent rhabdomyolysis. We sought to quantify the extent of tissue catabolism in similar patients. Study Design During acute peritoneal dialysis (PD) in patients with COVID-19–associated AKI, we measured urea Kt/V adequacy and calculated the daily urea nitrogen generation rate while quantifying daily protein intake. Setting & Population We did calculations in 8 patients with COVID-9–associated AKI undergoing acute PD at Mount Sinai Hospital in New York City. As a comparator, we obtained urea kinetic parameters from our database of ambulatory patients receiving maintenance PD. Exposure or Predictors 8 patients with COVID-19–associated AKI undergoing acute PD. Outcomes Urea nitrogen generation rate in relation to daily protein intake. Analytical Approach Urea nitrogen generation rate from urea kinetics was related to measured daily dietary protein intake in these patients and we compared it with this relationship in ambulatory maintenance PD patients for whom both parameters were calculated from urea kinetics. Results Urea nitrogen generation rate in patients with AKI was 10.2±5g/d, which is more than 2-fold higher than for stable outpatients receiving maintenance PD (4.7±3g/d) despite similar dietary protein intake (74.8±11 vs 67.2±29g/d, respectively). This strongly suggests endogenous protein breakdown, probably from muscle. Urea nitrogen generation rate in these patients with AKI corresponds to 315g/d of ongoing muscle breakdown and cumulative 2.5kg of muscle breakdown during the early course of AKI. Limitations Small number of participants and assumptions in comparing urea nitrogen generation rate with protein intake. Conclusions In highly catabolic patients, an endogenous source of urea generation such as muscle protein breakdown seems to be the most likely explainable cause for our findings. This is the first study that we are aware of to quantify the degree of endogenous protein breakdown induced by COVID-19–related cytokine storm.

Nannochloropsis oceanica is a microalga, highly concentrated in protein and EPA (eicosapentaenoic acid). It has a recalcitrant cell wall that decreases nutrient digestibility. The objective of this study is to evaluate the effect of Nannochloropsis oceanica feed inclusion on the muscle proteome of fattening rabbits. Twenty rabbits were housed in individual cages for 5 weeks and were fed a control diet (n = 10) and one with 4.45% inclusion of microalga, replacing whole soybean meal (n = 10). After slaughter, samples of gastrocnemius muscle were taken for label-free proteomic analysis. A total of 1497 proteins were identified, 46 with differential abundance. Control rabbits had high abundance of proteins related to protein metabolism, suggesting higher muscle protein turnover. They also had higher abundance of structural proteins, suggesting a less tender meat by comparison with algae-fed rabbits. These had high abundance of proteins related to amino acid catabolism (Phe, Tyr) and synthesis (Gln). In addition, they had high abundance of proteins related to protein breakdown, overall suggesting that more tender meat may result from algae feeding. Biological significance Microalgae are interesting feedstuffs particularly from a sustainability point of view. Their use in farm animal diets could simultaneously enhance the nutritional value and quality of meat. However, the biochemical pathways behind such improvement remain uncharacterized, namely in the rabbit species. By using a proteomics approach, this study will enlighten such biochemical pathways and relate them with productive performance parameters. In addition, results reflect some anti-nutritional properties of the recalcitrant cell wall over amino acid digestibility and consequently, its availability for protein synthesis.

High energy diets (HED) have been widely used in farmed fish, but HED also caused some metabolic diseases. The rapamycin-induced inhibition of mechanistic target of rapamycin (mTOR) signalling has been verified to alleviate metabolic syndromes in mammals, however, the functions of rapamycin in regulating metabolism in the fish fed with HED has not been well investigated. In the present 8-week feeding trial, Nile tilapia were fed with a normal diet with medium fat and medium carbohydrate (MFMC), a high fat and high carbohydrate (HFHC) diet, or the HFHC diet supplemented with rapamycin (Rap). Compared to MFMC diet, the HFHC diet significantly increased phosphorylation of mTOR protein, lipid deposition in tissues, glycogen degradation, oxidative stress and inflammation, but decreased lipid catabolism capability. The supplementation of Rap in the HFHC diet inhibited phosphorylation of mTOR protein and protein synthesis, but elevated protein catabolism. Rap addition also increased catabolism of lipid and carbohydrate, reduced oxidative stress and inflammation, and promoted autophagic activities, meaning Rap addition could alleviate the most of the adverse effects caused by the HFHC diet. All these indicate that the elevated mTOR activity was highly correlated to the adverse effects of HFHC diet on physiological functions in fish, and the inhibition of mTOR activity could largely help to maintain normal physiological status in the HED-fed fish. Therefore, mTOR could be a promising regulatory target to alleviate the metabolic dysfunctions caused by HED in fish, however, the possible negative effects of TOR pathway regulation on protein retention and growth should also be carefully evaluated in practices.