Amino Acid Oxidation Research Articles

Leucine requirement determined in healthy young adult males using the indicator amino acid oxidation method

BackgroundPrevious studies proposed varying leucine requirements for adults ranging from 25 to 40 mg⋅kg−1⋅d−1, but often these studies did not test intakes exceeding 40 mg⋅kg−1⋅d−1. Data using the indicator amino acid oxidation (IAAO) method suggest a higher requirement of 55 mg⋅kg−1⋅d−1 on the basis of the total branched-chain amino acids requirement, but not leucine independently. ObjectivesThe IAAO method was used to determine the leucine requirement in healthy young adult males. MethodsTen healthy adult males (26.9 ± 1.87 y, mean ± SEM) were studied at 7 leucine intakes; each studied over a 3-d period. Following 2-d of preadaptation to adequate protein intake (1.0 g⋅kg−1⋅d−1), subjects received experimental diets containing the randomly assigned test leucine intake (10, 20, 30, 40, 50, 65, and 75 mg⋅kg−1⋅d−1) for 8 h. The rate of the release of 13CO2 from the oxidation of L-[1-13C]phenylalanine (F13CO2) was measured on the third day, and the leucine requirement was determined using mixed-effect change-point regression and the F13CO2 data in R. The 95% confidence interval was calculated using parametric bootstrap. The effect of leucine intake on the concentration of plasma amino acids, insulin, and glucose were assessed using repeated measures analysis of variance and linear mixed effects. ResultsThe mean leucine requirement was 33.6 mg⋅kg−1⋅d−1 with a lower and upper 95% confidence of 26.16, 41.04 mg⋅kg−1⋅d−1. Higher leucine intakes were associated with increased plasma leucine, and decreased valine, isoleucine, and serine concentrations. ConclusionsThe leucine requirement of young adult males is ∼34 mg⋅kg−1⋅d−1, which aligns with previously published tracer balance experiments.This trial was registered at http://clinicaltrials.gov (https://clinicaltrials.gov/study/NCT05394155?term=leucine%20young%20adult&rank=1) as NCT05394155.

Impact of high temperature and drought stress on the microbial community in wolf spiders

High temperatures and drought present significant abiotic challenges that can limit the survival of many arthropods, including wolf spiders, which are ectothermic and play a crucial role in controlling pest populations. However, the impact of these stress factors on the microbiota of spiders remains poorly understood. In this study, we utilized 16 S rRNA gene sequencing to explore the diversity and composition of bacterial communities within Pardosa pseudoannulata under conditions of high temperature and drought stress. We found that Firmicutes, Bacteroidetes, and Proteobacteria were the predominant bacterial phyla present. Analyses of alpha diversity indicated an increase in bacterial diversity under combined stress conditions, as reflected by various diversity indices such as Ace, Chao1, Shannon, and Simpson. Furthermore, co-occurrence network analysis highlighted intricate interactions among the microbial taxa (e.g., Enterobacter, Chitinophaga, and Eubacterium), revealing the adaptive complexity of the spider's microbiome to environmental stress. Functional prediction analysis suggested that combined stress conditions might enhance key metabolic pathways, particularly those related to oxidative phosphorylation and amino acid metabolism. Using Random Forest analysis, we determined that changes in three heat shock proteins were largely attributed to variations in bacterial communities, with Firmicutes being notably influential. Collectively, this in-depth analysis offers novel insights into the responses of microbial communities within spider microbiomes to combined abiotic stresses, providing valuable information for understanding extreme climate impacts and informing ecological management strategies.

Glycolysis Changes in Alloreactive Memory B Cells in Highly Sensitized Kidney Transplant Recipients Undergonig Desensitization Therapy.

Despite the growing use of desensitization strategies, hyperimmune patients remain at high risk of antibody-mediated rejection suggesting that, even when donor-specific antibodies (DSA) are effectively depleted, anti-donor specific B cells persist. We included 10 highly sensitized recipients that underwent desensitization with plasmapheresis and B cell depletion prior to kidney transplantation. We quantified changes in DSA (luminex), total B-cell subsets (flow cytometry), anti-donor HLA B cells (fluorospot), and single-cell metabolism in serially collected samples before desensitization, at the time of transplant, and at 6 and 12months thereafter. Desensitization was associated with a decrease in DSA and total memory B cell and naive B cell percentage, while plasma cells and memory anti-donor HLA circulating B cells persisted up to 12months after transplant. At 12-month post-transplantation, memory B cells increased their glycolytic capacity, while proliferative KI67+ plasma cells modified their metabolism by increasing fatty acid and amino acid oxidation capacity and decreasing their glucose dependence. Despite effective DSA depletion, anti-donor B cells persist in kidney transplant recipients. Due to the reliance of these cells on glycolysis, glycolysis-targeting therapies might represent a valuable treatment strategy.

Metabolic flux in macrophages in obesity and type-2 diabetes.

Recent literature extensively investigates the crucial role of energy metabolism in determining the inflammatory response and polarization status of macrophages. This rapidly expanding area of research highlights the importance of understanding the link between energy metabolism and macrophage function. The metabolic pathways in macrophages are intricate and interdependent, and they can affect the polarization of macrophages. Previous studies suggested that glucose flux through cytosolic glycolysis is necessary to trigger pro-inflammatory phenotypes of macrophages, and fatty acid oxidation is crucial to support anti-inflammatory responses. However, recent studies demonstrated that this understanding is oversimplified and that the metabolic control of macrophage polarization is highly complex and not fully understood yet. How the metabolic flux through different metabolic pathways (glycolysis, glucose oxidation, fatty acid oxidation, ketone oxidation, and amino acid oxidation) is altered by obesity- and type 2 diabetes (T2D)-associated insulin resistance is also not fully defined. This mini-review focuses on the impact of insulin resistance in obesity and T2D on the metabolic flux through the main metabolic pathways in macrophages, which might be linked to changes in their inflammatory responses. We closely evaluated the experimental studies and methodologies used in the published research and highlighted priority research areas for future investigations.

Species-specific metabolome changes during salinity downshift in sub-Arctic populations of Mytilus edulis and M. trossulus

The blue mussels Mytilus edulis and Mytilus trossulus are ecologically and economically important species distributed widely across the Northern Hemisphere. Understanding their behavioral and physiological disparities is crucial for assessing their ecological success and aquacultural value. The recent finding of non-native M. trossulus in the White Sea raises concerns regarding its potential competition with native M. edulis and its prospective spread in light of climate change and surface water freshening. We investigated the responses of M. edulis and M. trossulus to salinity variations by examining shell closure thresholds and tissue levels of 35 metabolic intermediates in mussels acclimated to different salinities (25, 16, and 10). The salinity threshold for valve closure was similar in both studied species, but M. trossulus consistently opened at lower salinities (by 0.2–0.7 practical salinity units) compared to M. edulis. Salinity-induced changes in metabolite levels were similar between the two species. Taurine emerged as the dominant osmolyte, comprising over 50% of the total free amino acid pool, with aspartate and glycine contributing 15–30%. Concentrations of taurine, glycine, and total free amino acids declined with decreasing salinity. Taurine to glycine ratios were higher in M. edulis and increased in both species with declining salinity. Acclimation salinity significantly influenced urea cycle intermediates and methionine sulfoxide content, a cellular biomarker of amino acid oxidation. Species-specific differences were observed in purine metabolism, with higher levels of GMP and AMP found in M. edulis. Likewise, aromatic amino acids and histidine levels were higher in M. edulis compared to M. trossulus. However, no evidence suggests superior adaptation of M. trossulus metabolism to hypoosmotic stress compared to M. edulis. Further research is necessary to elucidate the functional implications of subtle metabolic differences between these Mytilus congeners and their ecological consequences in changing marine environments.

Molecular genetic analysis of candidate genes for glutaric aciduria type II in a cohort of patients from Queensland, Australia

Glutaric aciduria type II (GAII) is a heterogeneous genetic disorder affecting mitochondrial fatty acid, amino acid and choline oxidation. Clinical manifestations vary across the lifespan and onset may occur at any time from the early neonatal period to advanced adulthood. Historically, some patients, in particular those with late onset disease, have experienced significant benefit from riboflavin supplementation. GAII has been considered an autosomal recessive condition caused by pathogenic variants in the gene encoding electron-transfer flavoprotein ubiquinone-oxidoreductase (ETFDH) or in the genes encoding electron-transfer flavoprotein subunits A and B (ETFA and ETFB respectively). Variants in genes involved in riboflavin metabolism have also been reported. However, in some patients, molecular analysis has failed to reveal diagnostic molecular results. In this study, we report the outcome of molecular analysis in 28 Australian patients across the lifespan, 10 paediatric and 18 adult, who had a diagnosis of glutaric aciduria type II based on both clinical and biochemical parameters. Whole genome sequencing was performed on 26 of the patients and two neonatal onset patients had targeted sequencing of candidate genes. The two patients who had targeted sequencing had biallelic pathogenic variants (in ETFA and ETFDH). None of the 26 patients whose whole genome was sequenced had biallelic variants in any of the primary candidate genes. Interestingly, nine of these patients (34.6%) had a monoallelic pathogenic or likely pathogenic variant in a single primary candidate gene and one patient (3.9%) had a monoallelic pathogenic or likely pathogenic variant in two separate genes within the same pathway. The frequencies of the damaging variants within ETFDH and FAD transporter gene SLC25A32 were significantly higher than expected when compared to the corresponding allele frequencies in the general population. The remaining 16 patients (61.5%) had no pathogenic or likely pathogenic variants in the candidate genes. Ten (56%) of the 18 adult patients were taking the selective serotonin reuptake inhibitor antidepressant sertraline, which has been shown to produce a GAII phenotype, and another two adults (11%) were taking a serotonin-norepinephrine reuptake inhibitor antidepressant, venlafaxine or duloxetine, which have a mechanism of action overlapping that of sertraline. Riboflavin deficiency can also mimic both the clinical and biochemical phenotype of GAII. Several patients on these antidepressants showed an initial response to riboflavin but then that response waned. These results suggest that the GAII phenotype can result from a complex interaction between monoallelic variants and the cellular environment. Whole genome or targeted gene panel analysis may not provide a clear molecular diagnosis.

Adaptive Effects of Endocrine Hormones on Metabolism of Macronutrients during Fasting and Starvation: A Scoping Review.

Food deprivation can occur for different reasons. Fasting (<24 h duration) occurs to meet religious or well-being goals. Starvation (>1-day duration) occurs when there is intentional (hunger strike or treatment of a medical condition) or unintentional (anorexia nervosa, drought, epidemic famine, war, or natural disaster) food deprivation. A scoping review was undertaken using the PubMed database to explore 1805 abstracts and review 88 eligible full-text articles to explore the adaptive relationships that emerge between cortisol, insulin, glucagon, and thyroid hormones on the metabolic pathways of macronutrients in humans during fasting and starvation. The collected data indicate that fasting and starvation prime the human body to increase cortisol levels and decrease the insulin/glucagon ratio and triiodothyronine (T3) levels. During fasting, increased levels of cortisol and a decreased insulin/glucagon ratio enhance glycogenolysis and reduce the peripheral uptake of glucose and glycogenesis, whereas decreased T3 levels potentially reduce glycogenolysis. During starvation, increased levels of cortisol and a decreased insulin/glucagon ratio enhance lipolysis, proteolysis, fatty acid and amino acid oxidation, ketogenesis, and ureagenesis, and decreased T3 levels reduce thermogenesis. We present a potential crosstalk between T3 and the above hormones, including between T3 and leptin, to extend their adaptive roles in the metabolism of endogenous macronutrients during food deprivation.

Valine and Inflammation Drive Epilepsy in a Mouse Model of ECHS1 Deficiency.

ECHS1 Deficiency (ECHS1D) is a rare and devastating pediatric disease that currently has no defined treatments. This disorder results from missense loss-of-function mutations in the ECHS1 gene that result in severe developmental delays, encephalopathy, hypotonia, and early death. ECHS1 enzymatic activity is necessary for the beta-oxidation of fatty acids and the oxidation of branched-chain amino acids within the inner mitochondrial matrix. The pathogenesis of disease remains unknown, however it is hypothesized that disease is driven by an accumulation of toxic metabolites from impaired valine oxidation. To expand our knowledge on disease mechanisms, a novel mouse model of ECHS1D was generated that possesses a disease-associated knock-in (KI) allele and a knock-out (KO) allele. To investigate the behavioral phenotype, a battery of testing was performed at multiple time points, which included assessments of learning, motor function, endurance, sensory responses, and anxiety. Neurological abnormalities were assessed using wireless telemetry EEG recordings, pentylenetetrazol (PTZ) seizure induction, and immunohistochemistry. Metabolic perturbations were measured within the liver, serum, and brain using mass spectrometry and magnetic resonance spectroscopy. To test disease mechanisms, mice were subjected to disease pathway stressors and then survival, body weight gain, and epilepsy were assessed. Mice containing KI/KI or KI/KO alleles were viable with normal development and survival, and the presence of KI and KO alleles resulted in a significant reduction in ECHS1 protein. ECHS1D mice displayed reduced exercise capacity and pain sensation. EEG analysis revealed increased slow wave power that was associated with perturbations in sleep. ECHS1D mice had significantly increased epileptiform EEG discharges, and were sensitive to seizure induction, which resulted in death of 60% of ECHS1D mice. Under basal conditions, brain structure was grossly normal, although histological analysis revealed increased microglial activation in aged ECHS1D mice. Increased dietary valine only affected ECHS1D mice, which significantly exacerbated seizure susceptibility and resulted in death. Lastly, acute inflammatory challenge drove regression and early lethality in ECHS1D mice. In conclusion, we developed a novel model of ECHS1D that may be used to further knowledge on disease mechanisms and to develop therapeutics. Our data suggests altered metabolic signaling and inflammation may contribute to epilepsy in ECHS1D, and these alterations may be attributed to impaired valine metabolism.

An insight into trichomes-deficiency and trichomes-rich black teas by comparative metabolomics: The impact of oxidized trichomes on metabolic profiles and infusion color

Tea trichomes were regarded as an essential evaluation index for reflecting tea flavor quality in terms of aroma and influence on infusion color. This study reveals the impact of golden oxidized trichomes on the color, volatile and non-volatile metabolites of black teas through comparative metabolomics combined quantitative analysis on hongbiluo (trichomes-deficiency black teas), hongjinluo (trichomes-rich black teas), and trichomes (from hongjinluo). Forty-six volatile components were detected using headspace solid-phase microextraction gas chromatography-mass spectrometry, while the results suggested that the contribution of trichomes to black teas is limited. A total of 60 marker non-volatile compounds were identified, including catechins, catechin oxidation products, flavonoid glycosides, organic acids, hydrolysable tannins and amino acids. Notably, p-coumaroyl-kaempferol glucosides, and catechin dimers demonstrated high levels in independent trichomes and showed a positive correlation with the brightness and yellow hue of black tea infusions, specifically kaempferol 3-O-di-(p-coumaroyl)-hexoside. Furthermore, results from fractional extraction analysis of separated trichomes provided that N-ethyl-2-pyrrolidinone-substituted epicatechin gallates, acylated kaempferol glycosides, and chromogenic catechins dimers, such as theaflavins, were primary color contributors in oxidized trichomes. Especially, we found that epicatechin gallate (ECG) and its derivates, 3′-O-methyl-ECG and N-ethyl-2-pyrrolidinone-substituted ECG, highly accumulated in trichomes, which may be associated with the varieties of hongbiluo and hongjinluo black teas. Eventually, addition tests were applied to verify the color contribution of trichome mixtures. Our findings employed comprehensive information revealing that golden oxidized trichomes contributed significantly to the brightness and yellow hue of black tea infusion, but their contribution to the aroma and metabolic profile is limited. These findings may contribute to the effective modulation of the infusion color during black tea production by regulating the proportion of tea trichomes or screening trichomes-rich or deficiency varieties.

Dietary protein load affects the energy and nitrogen balance requiring liver glutamate dehydrogenase to maintain physical activity

Provision of amino acids to the liver is instrumental for gluconeogenesis while it requires safe disposal of the amino group. The mitochondrial enzyme glutamate dehydrogenase (GDH) is central for hepatic ammonia detoxification by deaminating excessive amino acids towards ureagenesis and preventing hyperammonemia. The present study investigated the early adaptive responses to changes in dietary protein intake in control mice and liver-specific GDH knockout mice (Hep-Glud1-/-). Mice were fed chow diets with a wide coverage of protein contents; i.e. suboptimal 10%, standard 20%, over optimal 30%, and high 45% protein diets; switched every 4 days. Metabolic adaptations of the mice were assessed in calorimetric chambers before tissue collection and analyses. Hep-Glud1-/- mice exhibited impaired alanine induced gluconeogenesis and constitutive hyperammonemia. The expression and activity of GDH in liver lysates were not significantly changed by the different diets. However, applying an in situ redox-sensitive assay on cryopreserved tissue sections revealed higher hepatic GDH activity in mice fed the high-protein diets. On the same section series, immunohistochemistry provided corresponding mapping of the GDH expression. Cosinor analysis from calorimetric chambers showed that the circadian rhythm of food intake and energy expenditure was altered in Hep-Glud1-/- mice. In control mice, energy expenditure shifted from carbohydrate to amino acid oxidation when diet was switched to high protein content. This shift was impaired in Hep-Glud1-/- mice and consequently the spontaneous physical activity was markedly reduced in GDH knockout mice. These data highlight the central role of liver GDH in the energy balance adaptation to dietary proteins.

#49 Uremic toxins and inflammation: Metabolic pathways affected in stage 5 kidney disease

Abstract Background and Aims Chronic kidney disease (CKD) is a global health threat, affecting around 12% of the population. The uremic phenotype associated with CKD is considered a predictor for the high cardiovascular and overall mortality in these patients and inflammation is regarded as an essential contributor to it. We seek to discover new inflammation-related metabolites in the CKD population through the use of semitargeted metabolomics, in order to gain insight into the pathways involved and identify potential biomarkers. Method The study employed an observational prospective design to investigate metabolite impact on inflammation in 50 non-dialysis-dependent stage 5 CKD patients. Blood samples were collected before the start of the first haemodialysis session. A semitargeted metabolomic approach was followed, utilising liquid chromatography and high-resolution mass spectrometry with an in-house library of polar compounds. Univariate (Mann-Whitney test) and multivariate (logistic regression with LASSO regularization) analyses identified metabolomic variables associated with inflammation. Results Univariate analysis revealed that adenosine-5′-phosphosulfate (APS) (p-value (p) 0.0211), dimethyglycine (p 0.0015), pyruvate (p 0.0066), lactate (p 0.0156) and 2-ketobutyric acid (p 0.0458) were significantly increased with inflammation, with differences over 1,5-fold changes (inflammation = yes vs inflammation = no). Cholic acid (p 0.01655), homogentisic acid (p 0.0007), 2-phenylpropionic acid (p 0.0351) and 3-indoxylsulfate (p 0.0087) show similar patterns but in the opposite direction, as observed in the volcano plot in Fig. 1. Multivariate analysis revealed significant associations between metabolites and inflammation in CKD stage 5 patients. Specifically, odds ratios for N-Butyrylglycine (OR: 1.87), dimethylglycine (OR: 1.81), 2-Oxoisopentanoic acid (OR: 1.70), and pyruvate (OR: 1.71) were related to a higher probability of experiencing inflammation. Conversely, homogentisic acid (OR: 072), 2-Phenylpropionic acid (OR: 0.77), and 2-Methylglutaric acid (OR: 0.78) exhibited an OR&amp;lt;1, suggestive of decreased probability. This is graphically represented in Fig. 2. Conclusion In our semi-targeted metabolomics study on stage 5 CKD patients, we pinpointed various metabolites linked to inflammation. Notably, dimethylglycine, a uremic toxin, may contribute to vascular damage, while pyruvate's abnormal metabolism is a known feature in heart failure and chronic kidney disease. N-butyrilglycine and 2-oxoisopentanoic acid were also associated with inflammation, potentially tied to mitochondrial fatty acid beta oxidation and incomplete breakdown of branched-chain amino acids. APS, impacting bone development and intestinal permeability, showed a positive correlation with inflammation. Conversely, homogentisic acid and phenylpropionic acid were negatively correlated. While homogentisic acid's anti-inflammatory role isn't clear, phenylpropionic acid exhibits protective effects against hepatotoxicity by reducing immune response. 2-methylglutaric acid and cholic acid were inversely correlated with inflammation, suggesting a protective effect related to fatty acid metabolism and bile acid regulation. These identified metabolites enhance our understanding of CKD-associated inflammation and present strong candidates for future biomarker validation with potential for clinical translation.

Trichodelphinine A alleviates pulmonary fibrosis by inhibiting collagen synthesis via NOX4-ARG1/TGF-β signaling pathway

BackgroundPulmonary fibrosis, a progressive and fatal lung disease with no effective treatment medication, is characterized by lung remodeling and fibroblastic foci caused by an oxidative imbalance with an overloading deposition of collagen. Trichodelphinine A, a hetisine-type C20-diterpenoid alkaloid, was found anti-fibrotic activity in vitro, but its effect and mechanism on pulmonary fibrosis still unknown. PurposeOur study aimed to investigate and validate the anti-fibrotic properties of trichodelphinine A in pulmonary fibrosis animals induced by bleomycin (BLM), and its mechanism whether via NOX4-ARG1/TGF-β signaling pathway. MethodsThe anti-fibrotic effects of trichodelphinine A were evaluated using BLM-induced rats through indicators of lung histopathology and collagen synthesis. Dynamic metabolomics evaluated the metabolic disorder and therapeutic effect of trichodelphinine A. The interaction between trichodelphinine A and NOX4 receptor was confirmed using CETSA and molecular dynamics experiments. Molecular biology experiments were conducted in NOX4 gene knockout mice to investigate the intervention effect of trichodelphinine A. ResultsTrichodelphinine A could suppress histopathologic changes, collagen deposition and proinflammatory cytokine release pulmonary fibrosis in bleomycin induced rats. Dynamic metabolomics studies revealed that trichodelphinine A could correct endogenous metabolic disorders of arachidonic acid, arginine and proline during fibrosis development, which revealed that the regulation of oxidative stress and amino acid metabolism targeting NOX4 and ARG1 may be the main pharmacological mechanisms of trichodelphinine A on pulmonary fibrosis. We further determined that trichodelphinine A inhibited over oxidative stress and collagen deposition by suppressing Nrf2-keap1 and ARG1-OAT signaling pathways, respectively. Molecular dynamics studies showed that trichodelphinine A was directly binds with NOX4, in which PHE354 and THR355 residues of NOX4 are critical binding sites for trichodelphinine A. Mechanistic validation in cells or mice with NOX4 knockout or silencing suggested that the anti-fibrotic effects of trichodelphinine A depended on inhibition of NOX4 to suppress ARG1/OAT activation and TGF-β/Smads signaling pathway. ConclusionCollectively, our findings indicate a powerful anti-fibrotic function of trichodelphinine A in pulmonary fibrosis via targeting NOX4. NOX4 mediates the activation of ARG1/OAT to regulate arginase-proline metabolism, and promotes TGF-β/Smads signaling pathway, thereby affecting the collagen synthesis in pulmonary fibrosis, which is a novel finding and indicates that inhibition of NOX4 is a novel therapeutic strategy for pulmonary fibrosis.

#67 Is Sodium bicarbonate a better alternative in kidney transplantation for better early renal function? A systematic review and meta-analysis

Abstract Background and Aims Kidney transplantation is a choice treatment for end-stage renal disease. It is the most cost-effective treatment and significantly improves the quality of life. However, metabolic acidosis increased following the transplantation. It can lead to several undesirable effects, such as disturbing the protein balance, resulting in a negative nitrogen equilibrium, increased protein degradation and essential amino acid oxidation, reduced physical functioning, and consequently decreased quality of life. The intraoperative crystalloid used may have significant effects on early function and acid-base balance. Also, sodium bicarbonate may enhance early graft function. This study aims to compare the effect of sodium bicarbonate in enhancing early graft function post-renal transplantation. Method We conducted a comprehensive search on multiple databases, including PubMed, MEDLINE, Embase, Scopus, WOS, and the Central Cochrane Registry. We included randomized controlled trials (RCTs) that compared sodium bicarbonate to normal saline during intraoperative kidney transplantation. Results Our search retrieved three RCTs, with 246 patients eligible for our criteria. We used either fixed- or random-effects models in the quantitative synthesis of the eligible studies accounting for clinical heterogeneity. Pooled data revealed that the sodium bicarbonate group was associated with a significantly improve in PH level (MD = 0.1; 95% CI: 0.06, 0.13; P &amp;lt; 0.01), Base Excess level (MD = 4.60 mEq/L; 95% CI: 3.43, 5.77; P = 0.003), and urine volume (MD = 2.59 L; 95% CI: 0.88, 4.29; P &amp;lt; 0.01). However, there was no significant difference between two groups in decreasing creatinine level after first day (MD = −0.46 mg/dl; 95% CI: −1.68, 0.76; P = 0.46), but the creatinine level significantly decreases after 2nd day (MD = −1.11 mg/dl; 95% CI: −1.96, −0.26; P = 0.01), and 3rd day (MD = −1 mg/dl; 95% CI: −1.17, −0.72; P &amp;lt; 0.01). Conclusion Our study suggests a potential advantage of sodium bicarbonate over the normal saline solution as an intraoperative fluid in improving metabolic acidosis during kidney transplantation.

Metabolic availability of amino acids in humans.

Knowledge of amino acid bioavailability and the effect of combining complementary protein sources are required to determine how to best meet an individual's protein and indispensable amino acid needs. Traditionally, protein quality of foods has been assessed using digestibility data. Digestibility may overestimate bioavailability of some amino acids particularly those more susceptible to heat and processing. The indicator amino acid oxidation (IAAO) method has been validated and applied to determine amino acid bioavailability termed metabolic availability of the first limiting amino acid of a proteinaceous food. The metabolic availability of the limiting amino acid in the test protein is determined as a ratio of the indicator amino acid oxidation response to graded intakes of the test protein compared to the indicator response to a reference protein (crystalline amino acid patterned after egg protein). The IAAO method has also been applied to assess the effect of protein complementation directly in humans on the overall protein quality of the diet. The results demonstrate that protein complementation augments the limiting amino acid supply and increases protein synthesis.

Dietary Lysine Requirements of Older Adults Stratified by Age and Sex

BackgroundCurrent recommendation for lysine in older adults, 30 mg/kg/d, is based on young adult data. Evidence suggests that amino acid requirements may differ between young and old adults with both sex and age having an effect in the elderly. ObjectivesThis study aimed to define the lysine requirements in healthy older adults using the indicator amino acid oxidation (IAAO) method with L-[1-13C] phenylalanine as the indicator and to compare the derived estimates based on age: 60–69 y and >70 y. MethodsFourteen healthy males and 16 healthy females [>60 y, body mass index (BMI) = 26.3 kg/m2] were randomly assigned to receive 3–7 lysine intakes from 10 to 80 mg/kg/d. Subjects were adapted to a standard liquid diet providing 1.0 g/kg/d protein and adequate energy, for 2 d, with indicator oxidation measurements performed on day 3. The rate of release of 13CO2 from the oxidation of L-[1-13C] phenylalanine was measured in breath. A 2-phase linear mixed-effect model, and parametric bootstrap were used to determine mean lysine requirements and the 95% confidence intervals (CIs). The overlap of the 95% CI between the 2 age groups were used to compare the requirement estimates. The null hypothesis was accepted if the interval contained zero. ResultsThe mean and upper 95% CI of the lysine requirement for females were 32.9 and 40.9 and 46.2 and 53.7 mg/kg/d for those aged 60–69 y and >70 y, respectively. The mean and upper 95% CI of the lysine requirement for the 2 groups of males were not different so was combined to yield a mean and 95% CI of 32.2 and 38.2 mg/kg/d. ConclusionsTo our knowledge, this is the first study to report on the lysine requirement in adults aged >60 y. These results provide a basis from which the adequacy of diets to meet lysine needs of older adults can be assessed.The trial was registered at clinicaltrials.gov as NCT02008955 (https://clinicaltrials.gov/study/NCT02008955).

Dietary CP and digestion kinetics influence BW loss, litter weight gain, and reproduction by affecting postprandial amino acid metabolism in lactating sows

To avoid a high body protein mobilization in modern lean sows during lactation, an adequate dietary amino acid (AA) supply and an efficient AA utilization are crucial. This study evaluated the effects of dietary CP and in vitro protein digestion kinetics on changes in sow body condition, litter weight gain, milk composition, blood metabolites, protein utilization efficiency and subsequent reproductive performance. We hypothesized that a slower digestion of dietary protein would improve AA availability and utilization. In total, 110 multiparous sows were fed one of four lactation diets in a 2 × 2 factorial design, with two CP concentrations: 140 g/kg vs 180 g/kg, and two protein digestion kinetics, expressed as a percentage of slow protein (in vitro degradation between 30 and 240 min): 8 vs 16% of total protein. Feeding sows the high CP diets reduced sow weight loss (Δ = 7.6 kg, P < 0.01), estimated body fat loss (Δ = 2.6 kg, P = 0.02), and estimated body protein loss (Δ = 1.0 kg, P = 0.08), but only at a high percentage of slow protein. A higher percentage of slow protein increased litter weight gain throughout lactation (Δ = 2.6 kg, P = 0.04) regardless of CP concentrations, whereas a higher CP only increased litter weight gain during week 3 of lactation (Δ = 1.2 kg, P = 0.01). On Day 15 postfarrowing, serial blood samples were taken from a subsample of sows fed with the high CP diets. In these sows, a high percentage of slow protein resulted in higher plasma AA concentrations at 150 and 180 min after feeding (Δ = 0.89, P = 0.02, Δ = 0.78, P = 0.03, mmol/L, respectively) and lower increases in urea at 90 and 120 min after feeding (Δ = 0.67, P = 0.04, Δ = 0.70, P = 0.03, mmol/L, respectively). The higher dietary CP concentration increased total nitrogen loss to the environment (Δ = 604 g, P < 0.01) with a reduction of protein efficiency (Δ = 14.8%, P < 0.01). In the next farrowing, a higher percentage of slow protein increased subsequent liveborn litter size (Δ = 0.7, P < 0.05). In conclusion, feeding sows with a high dietary CP concentration alleviated maternal weight loss during lactation when the dietary protein digestion rate was slower, but lowered protein efficiency. A slower protein digestion improved litter weight gain, possibly by reducing AA oxidation and improving plasma AA availability, thus, improving protein efficiency.

Intestinal carbapenem-resistant Klebsiella pneumoniae undergoes complex transcriptional reprogramming following immune activation

ABSTRACT Carbapenem-resistant Klebsiella pneumoniae (CR-Kp) is a significant threat to public health worldwide. The primary reservoir for CR-Kp is the intestinal tract. There, the bacterium is usually present at low density but can bloom following antibiotic treatment, mostly in hospital settings. The impact of disturbances in the intestinal environment on the fitness, survival, expansion, and drug susceptibility of this pathogen is not well-understood, yet it may be relevant to devise strategies to tackle CR-Kp colonization and infection. Here, we adopted an in vivo model to examine the transcriptional adaptation of a CR-Kp clinical isolate to immune activation in the intestine. We report that as early as 6 hours following host treatment with anti-CD3 antibody, CR-Kp underwent rapid transcriptional changes including downregulation of genes involved in sugar utilization and amino acid biosynthesis and upregulation of genes involved in amino acid uptake and catabolism, antibiotic resistance, and stress response. In agreement with these findings, treatment increased the concentration of oxidative species and amino acids in the mouse intestine. Genes encoding for proteins containing the domain of unknown function (DUF) 1471 were strongly upregulated, however their deletion did not impair CR-Kp fitness in vivo upon immune activation. Transcription factor enrichment analysis identified the global regulator cAMP-Receptor Protein, CRP, as a potential orchestrator of the observed transcriptional signature. In keeping with the recognized role of CRP in regulating utilization of alternative carbon sources, crp deletion in CR-Kp resulted in strongly impaired gut colonization, although this effect was not amplified by immune activation. Thus, following intestinal colonization, which occurs in a CRP-dependent manner, CR-Kp can rapidly respond to immune cues by implementing a well-defined and complex transcriptional program whose direct relevance toward bacterial fitness warrants further investigation. Additional analyses utilizing this model may identify key factors to tackle CR-Kp colonization of the intestine.

Role of reactive carbonyls and superoxide radicals in protein damage by cigarette smoke extracts: Comparison of Heat-not-Burn e-cigarettes to conventional cigarettes

Oxidative protein damage involving carbonylation of respiratory tract proteins typically accompanies exposure to tobacco smoke. Such damage can arise via multiple mechanisms, including direct amino acid oxidation by reactive oxygen species or protein adduction by electrophilic aldehydes. This study investigated the relative importance of these pathways during exposure of a model protein to fresh cigarette emission extracts. Briefly, protein carbonyl adducts were estimated in bovine serum albumin following incubation in buffered solutions with whole cigarette emissions extracts prepared from either a single 1R6F research cigarette or a single “Heat-not-Burn” e-cigarette. Although both extracts caused concentration-dependent protein carbonylation, conventional cigarette extracts produced higher adduct yields than e-cigarette extracts. Superoxide radical generation by conventional and e-cigarette emissions was assessed by monitoring nitro blue tetrazolium reduction and was considerably lower in extracts made from “Heat-Not-Burn” e-cigarettes. The superoxide dismutase/catalase mimic EUK-134 strongly suppressed radical production by whole smoke extracts from conventional cigarettes, however, it did not diminish protein carbonyl adduction when incubating smoke extracts with the model protein. In contrast, edaravone, a neuroprotective drug with strong carbonyl-trapping properties, strongly suppressed protein damage without inhibiting superoxide formation. Although these findings require extension to appropriate cell-based and in vivo systems, they suggest reactive aldehydes in tobacco smoke make greater contributions to oxidative protein damage than smoke phase radicals.

Post-prandial tracer studies of protein and amino acid utilisation: what can they tell us about human amino acid and protein requirements?

Nitrogen balance (NB), the principal methodology used to derive recommendations for human protein and amino acid requirements, has been widely criticised, and calls for increased protein and amino acid requirement recommendations have been made, often on the basis of post-prandial amino acid tracer kinetic studies of muscle protein synthesis, or of amino acid oxidation. This narrative review considers our knowledge of the homeostatic regulation of the FFM throughout the diurnal cycle of feeding and fasting and what can and has been learnt from post-prandial amino acid tracer studies, about amino acid and protein requirements. Within the FFM, muscle mass in well fed weight-stable adults with healthy lifestyles appears fixed at a phenotypic level within a wide range of habitual protein intakes. However homoeostatic regulation occurs in response to variation in habitual protein intake, with adaptive changes in amino acid oxidation which influence the magnitude of diurnal losses and gains of body protein. Post-prandial indicator amino acid oxidation (IAAO) studies have been introduced as an alternative to NB and to the logistically complex 24 h [13C-1] amino acid balance studies, for assessment of protein and amino acid requirements. However, a detailed examination of IAAO studies shows both a lack of concern for homeostatic regulation of amino acid oxidation and major flaws in their design and analytical interpretation, which seriously constrain their ability to provide reliable values. New ideas and a much more critical approach to existing work is needed if real progress is to be made in the area.

Protein requirements may be lower on a training compared to rest day but are not influenced by moderate training volumes in endurance trained males.

The impact of training volume on protein requirements in endurance trained males was investigated with indicator amino acid oxidation (IAAO) methodology on a recovery day (REST) or after a 10 or 20 km run while consuming a single suboptimal protein intake (0.93g/kg/day). Phenylalanine excretion (F13CO2; inverse proxy for whole body protein synthesis) was greatest and phenylalanine net balance was lowest on REST compared to post-exercise recovery with no difference between training volumes. Single point F13CO2 was indistinguishable from past IAAO studies using multiple protein intakes. Our results suggest that protein requirements may be greatest on recovery days but are not influenced by moderate training volumes in endurance athletes.