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1.
Talanta ; 223(Pt 2): 121872, 2021 Feb 01.
Artículo en Inglés | MEDLINE | ID: mdl-33298292

RESUMEN

Metabolic phenotyping using mass spectrometry (MS) is being applied to ever increasing sample numbers in clinical and epidemiology studies. High-throughput and robust methods are being developed for the accurate measurement of metabolites associated with disease. Traditionally, quantitative assays have utilized triple quadrupole (QQQ) MS based methods; however, the use of such focused methods removes the ability to perform discovery-based metabolic phenotyping. An integrated workflow for the hybrid simultaneous quantification of 34 biogenic amines in combination with full scan high-resolution accurate mass (HRAM) exploratory metabolic phenotyping is presented. Primary and secondary amines are derivatized with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate prior to revered-phase liquid chromatographic separation and mass spectrometric detection. Using the HRAM-MS data, retrospective phenotypic data mining could be performed, demonstrating the versatility of HRAM-MS instrumentation in a clinical and molecular epidemiological environment. Quantitative performance was assessed using two MS detector platforms: Waters TQ-XS (QQQ; n = 3) and Bruker Impact II QToF (HRAMS-MS; n = 2) and three human biofluids (plasma, serum and urine). Finally, each platform was assessed using a certified external reference sample (NIST SRM 1950 plasma). Intra- and inter-day accuracy and precision were comparable between the QQQ and QToF instruments (<15%), with excellent linearity (R2 > 0.99) over the quantification range of 1-400 µmol L-1. Quantitative values were comparable across all instruments for human plasma, serum and urine samples, and calculated concentrations were verified against certified reference values for NIST SRM 1950 plasma as an external reference. As a real-life biological exemplar, the method was applied to plasma samples obtained from SARS-CoV-2 positive patients versus healthy controls. Both the QQQ and QToF approaches were equivalent in being able to correctly classify SARS-CoV-2 positivity. Critically, the use of HRAM full scan data was also assessed for retrospective exploratory mining of data to extract additional biogenic amines of biomarker interest beyond the 34 quantified targets.


Asunto(s)
Aminoácidos/metabolismo , Aminas Biogénicas/metabolismo , Aminoácidos/sangre , Aminas Biogénicas/sangre , /orina , Cromatografía Líquida de Alta Presión , Humanos , Espectrometría de Masas , Metabolómica , Fenotipo , Control de Calidad , Estándares de Referencia , Reproducibilidad de los Resultados , Estudios Retrospectivos
2.
Int J Mol Sci ; 21(22)2020 Nov 16.
Artículo en Inglés | MEDLINE | ID: mdl-33207699

RESUMEN

The novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spread to nearly every continent, registering over 1,250,000 deaths worldwide. The effects of SARS-CoV-2 on host targets remains largely limited, hampering our understanding of Coronavirus Disease 2019 (COVID-19) pathogenesis and the development of therapeutic strategies. The present study used a comprehensive untargeted metabolomic and lipidomic approach to capture the host response to SARS-CoV-2 infection. We found that several circulating lipids acted as potential biomarkers, such as phosphatidylcholine 14:0_22:6 (area under the curve (AUC) = 0.96), phosphatidylcholine 16:1_22:6 (AUC = 0.97), and phosphatidylethanolamine 18:1_20:4 (AUC = 0.94). Furthermore, triglycerides and free fatty acids, especially arachidonic acid (AUC = 0.99) and oleic acid (AUC = 0.98), were well correlated to the severity of the disease. An untargeted analysis of non-critical COVID-19 patients identified a strong alteration of lipids and a perturbation of phenylalanine, tyrosine and tryptophan biosynthesis, phenylalanine metabolism, aminoacyl-tRNA degradation, arachidonic acid metabolism, and the tricarboxylic acid (TCA) cycle. The severity of the disease was characterized by the activation of gluconeogenesis and the metabolism of porphyrins, which play a crucial role in the progress of the infection. In addition, our study provided further evidence for considering phospholipase A2 (PLA2) activity as a potential key factor in the pathogenesis of COVID-19 and a possible therapeutic target. To date, the present study provides the largest untargeted metabolomics and lipidomics analysis of plasma from COVID-19 patients and control groups, identifying new mechanisms associated with the host response to COVID-19, potential plasma biomarkers, and therapeutic targets.


Asunto(s)
Infecciones por Coronavirus/metabolismo , Metaboloma , Neumonía Viral/metabolismo , Anciano , Anciano de 80 o más Años , Aminoácidos/sangre , Ácido Araquidónico/sangre , Biomarcadores/sangre , Ciclo del Ácido Cítrico , Infecciones por Coronavirus/sangre , Infecciones por Coronavirus/patología , Femenino , Gluconeogénesis , Humanos , Masculino , Persona de Mediana Edad , Ácido Oléico/sangre , Pandemias , Fosfatidilcolinas/sangre , Fosfatidiletanolaminas/sangre , Fosfolipasas A2/sangre , Neumonía Viral/sangre , Neumonía Viral/patología , Triglicéridos/sangre
3.
Ann Biol Clin (Paris) ; 78(5): 555-564, 2020 Oct 01.
Artículo en Inglés | MEDLINE | ID: mdl-33026350

RESUMEN

Biochemical diagnosis of hereditary metabolic diseases requires the detection and simultaneous identification of a large number of compounds, hence the interest in metabolic profiles. Amino acid chromatography allows the identification and quantification of more than forty compounds. As part of the accreditation process for medical biology examinations according to standard NF EN ISO 15189, the group from SFEIM recommends an approach to accredit amino acid chromatography. Validation parameters and recommendations are discussed in this specific framework.


Asunto(s)
Aminoácidos/análisis , Cromatografía/normas , Pruebas Diagnósticas de Rutina/normas , Errores Innatos del Metabolismo/diagnóstico , Acreditación/normas , Adulto , Aminoácidos/sangre , Aminoácidos/líquido cefalorraquídeo , Aminoácidos/orina , Amniocentesis/normas , Líquido Amniótico/química , Análisis Químico de la Sangre/métodos , Análisis Químico de la Sangre/normas , Recolección de Muestras de Sangre/normas , Niño , Cromatografía/métodos , Cromatografía Liquida/normas , Pruebas Diagnósticas de Rutina/métodos , Femenino , Humanos , Recién Nacido , Errores Innatos del Metabolismo/sangre , Errores Innatos del Metabolismo/líquido cefalorraquídeo , Errores Innatos del Metabolismo/orina , Tamizaje Neonatal/métodos , Tamizaje Neonatal/normas , Fase Preanalítica , Embarazo , Diagnóstico Prenatal/métodos , Diagnóstico Prenatal/normas , Espectrometría de Masas en Tándem/normas , Urinálisis/métodos , Urinálisis/normas , Toma de Muestras de Orina/normas
4.
BMC Med Genet ; 21(1): 183, 2020 09 21.
Artículo en Inglés | MEDLINE | ID: mdl-32957924

RESUMEN

BACKGROUND: Disorders of the metabolism and absorption of vitamin B12 can lead to decrease in activity of methionine synthetase and methylmalonate coenzyme A mutase (MMUT), which results in increased levels of methylmalonic acid and homocysteine in blood and urine. Often, combined methylmalonic acidemia (MMA) and homocysteinemia is misdiagnosed due to a lack of specific symptoms. The clinical manifestations are diverse, but proteinuria as the initial presentation is rare. CASE PRESENTATION: Two cases of MMA with homocysteinemia in children are reported. Proteinuria were a primary presenting symptom, followed by anemia and neurologic symptoms (frequent convulsions and unstable walking, respectively). Screening of amino acids and acyl carnitine in serum showed that the propionyl carnitine:acetylcarnitine ratio increased. Profiling of urinary organic acids by gas chromatography-mass spectrometry revealed high levels of methylmalonic acid. Homocysteine content in blood was increased. Comprehensive genetic analyses of peripheral blood-derived DNA demonstrated heterozygous variants of methylmalonic aciduria type C and homocystinuria (MMACHC) and amnionless (AMN) genes in our two patients, respectively. After active treatment, the clinical manifestations in Case 1 were relieved and urinary protein ceased to be observed; Case 2 had persistent proteinuria and was lost to follow-up. CONCLUSIONS: Analyses of the organic acids in blood and urine suggested MMA combined with homocysteinemia. In such diseases, reports of renal damage are uncommon and proteinuria as the initial presentation is rare. Molecular analysis indicated two different genetic causes. Although the pathologic mechanisms were related to vitamin B12, the severity and prognosis of renal lesions were different. Therefore, gene detection provides new insights into inherited metabolic diseases.


Asunto(s)
Errores Innatos del Metabolismo de los Aminoácidos/complicaciones , Hiperhomocisteinemia/complicaciones , Proteinuria/diagnóstico , Adolescente , Errores Innatos del Metabolismo de los Aminoácidos/genética , Aminoácidos/sangre , Secuencia de Bases , Carnitina/análogos & derivados , Carnitina/sangre , Preescolar , ADN/sangre , ADN/genética , Cromatografía de Gases y Espectrometría de Masas , Homocisteína/sangre , Humanos , Hiperhomocisteinemia/genética , Masculino , Ácido Metilmalónico/orina , Proteinuria/etiología
5.
PLoS One ; 15(9): e0238522, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32946478

RESUMEN

The effects of feeding frequency on postprandial response of circulating appetite-regulating hormones, insulin, glucose and amino acids, and on physical activity, energy expenditure, and respiratory quotient were studied in healthy adult cats. Two experiments were designed as a 2 x 3 replicated incomplete Latin square design. Eight cats, with an average body weight (BW) of 4.34 kg ± 0.04 and body condition score (BCS) of 5.4 ± 1.4 (9 point scale), were fed isocaloric amounts of a commercial adult maintenance canned cat food either once (0800 h) or four times daily (0800 h, 1130 h, 1500 h, 1830 h). Study 1 consisted of three 21-d periods. On day 14, two fasted and 11 postprandial blood samples were collected over 24 hours to measure plasma concentrations of ghrelin, GLP-1, GIP, leptin, PYY, insulin and amino acids, and whole blood glucose. Physical activity was monitored from day 15 to 21 of each period. In Study 2 indirect calorimetry was performed on the last day of each period. Body weight was measured weekly and feed intake recorded daily in both experiments. No effect of feeding regimen on BW was detected. Cats eating four times daily had lesser plasma concentrations of GIP and GLP-1 (P<0.05) and tended to have lesser plasma PYY concentrations (P<0.1). Plasma leptin and whole blood glucose concentrations did not differ between regimens (P>0.1). Cats fed once daily had a greater postprandial plasma amino acid response, and greater plasma ghrelin and insulin concentrations (P<0.05). Physical activity was greater in cats fed four times (P<0.05), though energy expenditure was similar between treatments at fasting and in postprandial phases. Finally, cats eating one meal had a lower fasting respiratory quotient (P<0.05). Overall, these data indicate that feeding once a day may be a beneficial feeding management strategy for indoor cats to promote satiation and lean body mass.


Asunto(s)
Aminoácidos/metabolismo , Regulación del Apetito , Gatos/fisiología , Conducta Alimentaria , Hormonas/metabolismo , Aminoácidos/sangre , Alimentación Animal/análisis , Animales , Apetito , Glucemia/análisis , Glucemia/metabolismo , Gatos/sangre , Metabolismo Energético , Femenino , Ghrelina/sangre , Ghrelina/metabolismo , Hormonas/sangre , Insulina/sangre , Insulina/metabolismo , Masculino , Fotoperiodo , Condicionamiento Físico Animal , Respiración
6.
Nat Commun ; 11(1): 4643, 2020 09 16.
Artículo en Inglés | MEDLINE | ID: mdl-32938935

RESUMEN

Time-restricted feeding (TRF) improves metabolism independent of dietary macronutrient composition or energy restriction. To elucidate mechanisms underpinning the effects of short-term TRF, we investigated skeletal muscle and serum metabolic and transcriptomic profiles from 11 men with overweight/obesity after TRF (8 h day-1) and extended feeding (EXF, 15 h day-1) in a randomised cross-over design (trial registration: ACTRN12617000165381). Here we show that muscle core clock gene expression was similar after both interventions. TRF increases the amplitude of oscillating muscle transcripts, but not muscle or serum metabolites. In muscle, TRF induces rhythmicity of several amino acid transporter genes and metabolites. In serum, lipids are the largest class of periodic metabolites, while the majority of phase-shifted metabolites are amino acid related. In conclusion, short-term TRF in overweight men affects the rhythmicity of serum and muscle metabolites and regulates the rhythmicity of genes controlling amino acid transport, without perturbing core clock gene expression.


Asunto(s)
Aminoácidos/sangre , Ritmo Circadiano/genética , Ayuno , Lípidos/sangre , Músculo Esquelético/metabolismo , Adulto , Relojes Circadianos/genética , Estudios Cruzados , Expresión Génica , Humanos , Metabolismo de los Lípidos , Masculino , Músculo Esquelético/fisiología , Sobrepeso/dietoterapia , Sobrepeso/metabolismo
7.
J Proteome Res ; 19(11): 4442-4454, 2020 11 06.
Artículo en Inglés | MEDLINE | ID: mdl-32806897

RESUMEN

The metabolic effects of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection on human blood plasma were characterized using multiplatform metabolic phenotyping with nuclear magnetic resonance (NMR) spectroscopy and liquid chromatography-mass spectrometry (LC-MS). Quantitative measurements of lipoprotein subfractions, α-1-acid glycoprotein, glucose, and biogenic amines were made on samples from symptomatic coronavirus disease 19 (COVID-19) patients who had tested positive for the SARS-CoV-2 virus (n = 17) and from age- and gender-matched controls (n = 25). Data were analyzed using an orthogonal-projections to latent structures (OPLS) method and used to construct an exceptionally strong (AUROC = 1) hybrid NMR-MS model that enabled detailed metabolic discrimination between the groups and their biochemical relationships. Key discriminant metabolites included markers of inflammation including elevated α-1-acid glycoprotein and an increased kynurenine/tryptophan ratio. There was also an abnormal lipoprotein, glucose, and amino acid signature consistent with diabetes and coronary artery disease (low total and HDL Apolipoprotein A1, low HDL triglycerides, high LDL and VLDL triglycerides), plus multiple highly significant amino acid markers of liver dysfunction (including the elevated glutamine/glutamate and Fischer's ratios) that present themselves as part of a distinct SARS-CoV-2 infection pattern. A multivariate training-test set model was validated using independent samples from additional SARS-CoV-2 positive patients and controls. The predictive model showed a sensitivity of 100% for SARS-CoV-2 positivity. The breadth of the disturbed pathways indicates a systemic signature of SARS-CoV-2 positivity that includes elements of liver dysfunction, dyslipidemia, diabetes, and coronary heart disease risk that are consistent with recent reports that COVID-19 is a systemic disease affecting multiple organs and systems. Metabolights study reference: MTBLS2014.


Asunto(s)
Aminoácidos/sangre , Infecciones por Coronavirus , Lipoproteínas/sangre , Modelos Biológicos , Insuficiencia Multiorgánica , Pandemias , Neumonía Viral , Anciano , Betacoronavirus , Biomarcadores , Glucemia/análisis , Infecciones por Coronavirus/sangre , Infecciones por Coronavirus/complicaciones , Infecciones por Coronavirus/epidemiología , Infecciones por Coronavirus/metabolismo , Femenino , Humanos , Espectroscopía de Resonancia Magnética , Masculino , Espectrometría de Masas , Metaboloma , Persona de Mediana Edad , Insuficiencia Multiorgánica/sangre , Insuficiencia Multiorgánica/etiología , Insuficiencia Multiorgánica/metabolismo , Neumonía Viral/sangre , Neumonía Viral/complicaciones , Neumonía Viral/epidemiología , Neumonía Viral/metabolismo
8.
Arch Biochem Biophys ; 691: 108508, 2020 09 30.
Artículo en Inglés | MEDLINE | ID: mdl-32712289

RESUMEN

BACKGROUND: The clinical and diagnostic significance of systemic amino acids in sepsis and septic shock is unclear. Hence, the purpose of our study was to assess amino acids relationship with sepsis-related clinical data and to analyze whether they might have prognostic and discriminative value in sepsis and septic shock. MATERIALS AND METHODS: Prospective and observational study with 5-day follow-up. Circulating amino acids were measured in 20 patients with sepsis or septic shock diagnosis and 30 healthy volunteers by means of targeted metabolomics (LC-MS/MS). RESULTS: Non-survivors were distinguished by significant elevated concentration of hPro (1st and 2nd day) and by mHis (5th day). Septic shock was associated with significant increased concentration of hPro (1st and 5th day) and Gly-Pro, His, Sarc and Phe (2nd day), Gly-Pro (3rd day) and Gly-Pro and mHis (5th day). In non-survivors was observed the rising trend in concentration of His (P = 0.04; 2nd day) and declining trend in concentration of Asn (P = 0.004; 5th day) and Pro (P = 0.03; 3rd day). In septic shock was observed mainly the declining trend in concentration of Arg (P = 0.03; 5th day), APA (P = 0.04; 2nd day), Lys (P = 0.02; 5th day), Sarc (P = 0.04; 5th day), Ser (P = 0.02; 5th day), Val (P = 0.04; 5th day), Trp (P = 0.03; 5th day) and Gly-Pro (P = 0.03; 2nd day; P = 0.02; 3rd day). CONCLUSION: Sepsis and septic shock are associated with altered concentration of serum amino acids indicative particularly of the intensified breakdown of muscle and connective tissue proteins leading to the accumulation of their characteristic degradation products. Some amino acids hold potential as predictors of sepsis progression and outcome but, in the light of discrepancies between studies, should be assessed in more numerous cohort study.


Asunto(s)
Aminoácidos/sangre , Choque Séptico/diagnóstico , Anciano , Anciano de 80 o más Años , Aminoácidos/farmacocinética , Biomarcadores/sangre , Femenino , Humanos , Masculino , Metaboloma/fisiología , Metabolómica , Persona de Mediana Edad , Pronóstico , Estudios Prospectivos , Choque Séptico/sangre , Choque Séptico/metabolismo
9.
Int Heart J ; 61(4): 739-747, 2020 Jul 30.
Artículo en Inglés | MEDLINE | ID: mdl-32684600

RESUMEN

The metabolism of branched-chain amino acids (BCAAs) is reported to change in heart failure (HF) and correlate with cardiac function. However, the effect of BCAAs on HF remains controversial. We investigate the prognostic value of the plasma BCAA level in nonischemic dilated cardiomyopathy (NIDCM).This study enrolled 39 NIDCM patients, who underwent plasma amino acid (AA) analysis. The ratio of BCAAs to total AAs was calculated. All patients were divided into two groups at the median of BCAA/total AA ratio; high BCAA/total AA group (≥ 0.15, n = 20) and low BCAA/total AA group (< 0.15, n = 19). A cardiac event was defined as a composite of cardiac death, hospitalization for worsening HF, and lethal arrhythmia.The mean age was 51.1 ± 12.3 years and left ventricular ejection fraction (LVEF) was 32.7 ± 10.1%. In the low BCAA/total AA group, the body mass index and the total cholesterol level were lower than in the high BCAA/total AA group. The BCAA/total AA ratio was positively correlated with LVEF (r = 0.35, P = 0.031) and negatively correlated with brain natriuretic peptide (r = -0.37, P = 0.020). The low BCAA/total AA group had a lower cardiac event-free rate (5-year: 100% versus 73%; P = 0.019). In univariate analysis, angiotensin converting enzyme inhibitor or angiotensin II receptor blocker (hazard ratio: 0.045, P = 0.0014), hemoglobin (hazard ratio: 0.49 per 1 g/dL, P = 0.0022), and BCAA/total AA ratio < 0.15 (hazard ratio: not available, P = 0.0066) were major predictors for cardiac events.The BCAA/total AA ratio might be a useful predictor for future cardiac events in patients with NIDCM.


Asunto(s)
Aminoácidos/sangre , Cardiomiopatía Dilatada/sangre , Adulto , Anciano , Cardiomiopatía Dilatada/diagnóstico por imagen , Ecocardiografía , Femenino , Humanos , Masculino , Persona de Mediana Edad
10.
BMC Psychiatry ; 20(1): 333, 2020 06 24.
Artículo en Inglés | MEDLINE | ID: mdl-32580709

RESUMEN

BACKGROUND: The alterations of biological markers are thought to be effective tools to understand the pathophysiology and management of major depressive disorder (MDD). A lot of researches has implied many markers for depression, but any of them fully discovered the association between the markers and depression. The present study investigated the serum levels of amino acids and non-enzymatic antioxidants in major depression, and also explained their association with depression. METHODS: This study examined 247 MDD patients and 248 healthy controls (HCs) matched by age and sex. The Hamilton Depression Rating Scale (Ham-D) was used to all the participants to measure the severity of depression. Quantification of serum amino acids, vitamin A and E were carried out using the HPLC system whereas vitamin C levels were measured by UV-spectrophotometer. All the statistical analysis was performed by SPSS statistical software (version 23.0). The independent sample t-test, the Mann-Whitney U test, and the Fisher's exact test were applied to detect the group differences where a Bonferroni correction applied to the p value. RESULTS: It was observed that serum levels of four amino acids (methionine, phenylalanine, tryptophan, and tyrosine) along with three non-enzymatic antioxidants (vitamin A, E, and C) were significantly dropped in MDD patients compared to HCs (Cohen's d (d): - 0.45, - 0.50, - 0.68, - 0.21, - 0.27, - 0.65, and - 0.24, respectively). Furthermore, Ham-D scores of cases were negatively correlated with serum levels of methionine (r = - 0.155, p = 0.015) and tyrosine (r = - 0.172, p = 0.007). CONCLUSION: The present study suggests that lowered serum methionine, phenylalanine, tryptophan, tyrosine, and non-enzymatic antioxidants are associated with depression. The reduction of these parameters in MDD patients may be the consequence, and not the cause, of major depression.


Asunto(s)
Aminoácidos/sangre , Antioxidantes/análisis , Trastorno Depresivo Mayor/sangre , Adolescente , Adulto , Cromatografía Líquida de Alta Presión , Femenino , Humanos , Masculino , Persona de Mediana Edad , Adulto Joven
11.
PLoS One ; 15(6): e0234404, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32555688

RESUMEN

Myocardial energy deprivation plays a causal role in the development of heart failure. A cardiac protection blend (CPB) of nutrients including medium chain triglycerides, fish oil and other key nutrients was developed to slow the progression of canine myxomatous mitral valve disease (MMVD). A six-month dietary intervention demonstrated efficacy of CPB in slowing MMVD progression. Untargeted metabolomic analysis of serum from these dogs identified 102 differential metabolites (adjusted P < 0.05). The ratios of omega-6 to omega-3 fatty acid (FA) changed from 2.41 and 1.46 in control and CPB groups at baseline to 4.30 and 0.46 at 6 months respectively. A 2.7-fold increase of α-aminobutyrate, a myocardial modulator of glutathione homeostasis, was found in CPB dogs compared to 1.3-fold increase in control dogs. Arginine and citrulline, precursors of nitric oxide biosynthesis, were both increased 2-fold; caprate, a medium chain FA, was increased 3-fold; and deoxycarnitine, precursor of carnitine biosynthesis, was increased 2.5-fold in CPB dogs. Margarate and methylpalmitate decreased in response to CPB, a potential benefit in MMVD dogs as positive correlations were found between changes in both these FAs and left atrial diameter (r = 0.69, r = 0.87 respectively, adjusted P < 0.05). Sphingomyelins with very long chain saturated FAs associated with decreased risk of heart failure in humans were increased in MMVD dogs fed the CPB diet. Our data supports the hypothesis that CPB improves FA utilization and energetics, reduces oxidative stress and inflammation in MMVD dogs. More studies are needed to understand the roles of specific metabolites in MMVD.


Asunto(s)
Enfermedades de los Perros/sangre , Enfermedades de los Perros/dietoterapia , Metaboloma , Insuficiencia de la Válvula Mitral/veterinaria , Aminoácidos/sangre , Alimentación Animal/análisis , Animales , Dietoterapia/veterinaria , Enfermedades de los Perros/patología , Perros , Ácidos Grasos/sangre , Femenino , Atrios Cardíacos/patología , Masculino , Metabolómica , Insuficiencia de la Válvula Mitral/sangre , Insuficiencia de la Válvula Mitral/dietoterapia , Fosfolípidos/sangre
12.
Biomed Khim ; 66(2): 124-129, 2020 Feb.
Artículo en Ruso | MEDLINE | ID: mdl-32420892

RESUMEN

The study of the role of neurotransmitter systems in the pathogenesis of epilepsy is one of the priorities of epileptology. New data on the functions of free neurotransmitter-like amino acid in the central nervous system are of the greatest importance and determine the prospects for the development of novel effective anticonvulsants. It is widely believed in clinical medicine that epilepsy has distinct gender characteristics. The aim of this study was to investigate the gender peculiarities in the content of neurotransmitter amino acids in the brain of Krushinsky-Molodkina (KM) rats, which were used as model organisms for the study of genetically induced audiogenic epilepsy. The content of Asp, Glu, GABA, Gly, and Tau of the medulla oblongata, hippocampus and cerebral cortex were determined using high-performance liquid chromatography (HPLC) in intact KM rats, KM rats exposed to a series of epileptiform seizures, and Wistar rats (control group). Both the Wistar and KM rats had gender distinctions in the distribution of free amino acids among the investigated brain parts. The audiogenic epilepsy was characterized by smoothing gender differences as well as differences between the concentrations of free amino acids in the cortex and medulla oblongata, specific for Wistar rats. The changes observed in male rats after the set of seizures included the increase in GABA concentration and a decrease in the Gly level in all investigated brain parts, as well as the decrease of the Tau content in the cortex and hippocampus. At the same time, the Glu content in cortex increased, while the Asp level decreased. After 6 days of audiogenic stimulations the female KM rats demonstrated the increase in the Glu level in all investigated brain parts, the increase in Gly and Asp levels in hippocampus, and no changes in the GABA content. Thus, after the set of epileptiform seizures the KM rats achieved a new steady state of the studied amino acids pool, which differed in males and females. In this case, gender differences significantly changed after the seizures.


Asunto(s)
Aminoácidos/sangre , Epilepsia Refleja/sangre , Neurotransmisores/sangre , Factores Sexuales , Estimulación Acústica , Animales , Femenino , Masculino , Ratas , Ratas Wistar
13.
Am J Clin Nutr ; 112(2): 303-317, 2020 08 01.
Artículo en Inglés | MEDLINE | ID: mdl-32359142

RESUMEN

BACKGROUND: Protein ingestion increases skeletal muscle protein synthesis rates during recovery from endurance exercise. OBJECTIVES: We aimed to determine the effect of graded doses of dietary protein co-ingested with carbohydrate on whole-body protein metabolism, and skeletal muscle myofibrillar (MyoPS) and mitochondrial (MitoPS) protein synthesis rates during recovery from endurance exercise. METHODS: In a randomized, double-blind, parallel-group design, 48 healthy, young, endurance-trained men (mean ± SEM age: 27 ± 1 y) received a primed continuous infusion of l-[ring-2H5]-phenylalanine, l-[ring-3,5-2H2]-tyrosine, and l-[1-13C]-leucine and ingested 45 g carbohydrate with either 0 (0 g PRO), 15 (15 g PRO), 30 (30 g PRO), or 45 (45 g PRO) g intrinsically l-[1-13C]-phenylalanine and l-[1-13C]-leucine labeled milk protein after endurance exercise. Blood and muscle biopsy samples were collected over 360 min of postexercise recovery to assess whole-body protein metabolism and both MyoPS and MitoPS rates. RESULTS: Protein intake resulted in ∼70%-74% of the ingested protein-derived phenylalanine appearing in the circulation. Whole-body net protein balance increased dose-dependently after ingestion of 0, 15, 30, or 45 g protein (mean ± SEM: -0.31± 0.16, 5.08 ± 0.21, 10.04 ± 0.30, and 13.49 ± 0.55 µmol phenylalanine · kg-1 · h-1, respectively; P < 0.001). 30 g PRO stimulated a ∼46% increase in MyoPS rates (%/h) compared with 0 g PRO and was sufficient to maximize MyoPS rates after endurance exercise. MitoPS rates were not increased after protein ingestion; however, incorporation of dietary protein-derived l-[1-13C]-phenylalanine into de novo mitochondrial protein increased dose-dependently after ingestion of 15, 30, and 45 g protein at 360 min postexercise (0.018 ± 0.002, 0.034 ± 0.002, and 0.046 ± 0.003 mole percentage excess, respectively; P < 0.001). CONCLUSIONS: Protein ingested after endurance exercise is efficiently digested and absorbed into the circulation. Whole-body net protein balance and dietary protein-derived amino acid incorporation into mitochondrial protein respond to increasing protein intake in a dose-dependent manner. Ingestion of 30 g protein is sufficient to maximize MyoPS rates during recovery from a single bout of endurance exercise.This trial was registered at trialregister.nl as NTR5111.


Asunto(s)
Proteínas en la Dieta/metabolismo , Proteínas Musculares/biosíntesis , Músculo Esquelético/metabolismo , Adulto , Aminoácidos/sangre , Aminoácidos/metabolismo , Proteínas en la Dieta/análisis , Método Doble Ciego , Entrenamiento Aeróbico , Ejercicio Físico , Humanos , Masculino
14.
J Chromatogr A ; 1622: 461135, 2020 Jul 05.
Artículo en Inglés | MEDLINE | ID: mdl-32360058

RESUMEN

Here we describe a new HPLC-MS/MS method using a mixed mode stationary phase and a binary gradient of elution for the rapid separation and quantification of AAs in human plasma without derivatization or ion pairing reagent addition. The sample preparation procedure consists in a single dilution step after protein precipitation with sulfosalicylic acid. The proposed method allows for the unambiguous identification and analysis of 52 AAs and related compounds including the separation of isomers and isobars in an 18 min chromatographic run including the conditioning and the equilibration times. AAs were detected by selective reaction monitoring. Internal calibration was used for the quantification of 37 AAs, including 25 using the corresponding isotopically labeled internal standards. External calibration (no internal standard) was used for five additional analytes. Qualitative detection was achieved for the remaining compounds. Validation studies evaluated accuracy, linearity, within- and between-run precision, lower limits of detection and quantification for 37 amino acids present in commonly used quality control samples. For within-run precision CVs averaged 3.8 % (n = 30) for all compounds. For between-run precision, CVs averaged 8.6 % for all compounds (n = 20). Correlation with the common standard ion-exchange chromatography with post-column derivatization method was also performed for 32 plasma samples. While the proposed method is at least 50 times more sensitive, the data showed good correlation with slopes equal or higher than 0.9 and correlation coefficients mostly higher than 0.90. The method was successfully applied for analysis of plasma samples for detection of inherited disorders of amino acid metabolism.


Asunto(s)
Aminoácidos/sangre , Cromatografía Liquida/métodos , Espectrometría de Masas en Tándem/métodos , Aminoácidos/química , Calibración , Cromatografía Líquida de Alta Presión/métodos , Femenino , Humanos , Recién Nacido , Estándares de Referencia , Reproducibilidad de los Resultados
15.
Cancer Sci ; 111(7): 2248-2258, 2020 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-32426941

RESUMEN

Myeloid-derived suppressor cells (MDSCs) play a crucial role in immunosuppression in tumor-bearing hosts. MDSCs express arginase-I and indoleamine 2,3-dioxygenase; they suppress T-cell function by reducing the levels of l-arginine and l-tryptophan, respectively. We examined the anticancer effects of supplementation of these amino acids in CT26 colon carcinoma-bearing mice. Oral supplementation of l-arginine or l-tryptophan (30 mg/mouse) did not affect tumor growth, whereas oral supplementation of d-arginine was lethal. Supplementation of l-arginine showed a tendency to augment the efficacy of cyclophosphamide (CP). CP reduced the proportions of granulocytic MDSCs and increased the proportions of monocytic MDSCs in the spleen and tumor tissues of CT26-bearing mice. l-Arginine supplementation alone did not affect the MDSC subsets. CP treatment tended to reduce the plasma levels of l-arginine in CT26-bearing mice and significantly increased the number of tumor-infiltrating CD8+ T cells. In addition, l-arginine supplementation significantly increased the proportions of tumor peptide-specific CD8+ T cells in draining lymph nodes. Importantly, additional supplementation of l-arginine significantly increased the number of cured mice that were treated with CP and anti-PD-1 antibody. Totally, l-arginine supplementation shows promise for boosting the therapeutic efficacy of chemoimmunotherapy.


Asunto(s)
Antineoplásicos Inmunológicos/farmacología , Antineoplásicos/farmacología , Arginina/administración & dosificación , Suplementos Dietéticos , Aminoácidos/sangre , Animales , Antineoplásicos/uso terapéutico , Linfocitos T CD8-positivos/efectos de los fármacos , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Ciclofosfamida/farmacología , Relación Dosis-Respuesta a Droga , Sinergismo Farmacológico , Femenino , Citometría de Flujo , Ratones , Células Supresoras de Origen Mieloide/efectos de los fármacos , Células Supresoras de Origen Mieloide/inmunología , Células Supresoras de Origen Mieloide/metabolismo , Neoplasias/tratamiento farmacológico , Neoplasias/inmunología , Neoplasias/metabolismo , Neoplasias/patología , Subgrupos de Linfocitos T/efectos de los fármacos , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismo
16.
Poult Sci ; 99(3): 1421-1431, 2020 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-32115029

RESUMEN

The crude protein (CP) content of 4 iso-energetic, maize-based diets containing 11.00 g/kg digestible lysine was reduced in gradations from 200 to 156 g/kg with increasing inclusions of synthetic, or unbound, essential amino acids. A constant dietary electrolyte balance (DEB) of 230 mEq/kg was maintained, but a second 156 g/kg CP diet had a DEB of 120 mEq/kg, and energy densities of the 156 g/kg CP diet were reduced in the sixth and seventh treatments. Each of the 7 dietary treatments were offered to 7 replicate cages (6 birds/cage) or a total of 294 Ross 308 off-sex male broilers from 14 to 35 D posthatch. Reductions in CP from 200 to 156 g/kg did not influence weight gain but quadratically increased feed conversion ratio (FCR) and linearly increased relative abdominal fat-pad weights and feed intakes. The reduction in DEB did not influence growth performance but did adversely influence some amino acid digestibilities. Reducing energy density by 100 kcal/kg did not influence growth performance of birds offered the 156 g/kg CP diet but numerically reduced fat-pad weights. The transition from 200 to 156 g/kg CP diets generally enhanced jejunal and ileal amino acid digestibility coefficients but had diverse effects on free amino acid concentrations in systemic plasma with a remarkable 116% increase in threonine. Starch:protein disappearance rate ratios linearly increased in the jejunum and the ileum following the same transition, and these expanding ratios were related to heavier fat-pads and compromised FCR. This study indicates that reductions in dietary CP from 200 to 172 g/kg supported by inclusions of unbound essential amino acids do not compromise growth performance, but a further reduction to 156 g/kg CP significantly increased FCR. Both heavier relative fat-pad weights and inferior FCR were related to expanding starch:protein disappearance rate ratios, which suggests condensed dietary starch:protein ratios may advantage birds offered reduced CP diets.


Asunto(s)
Pollos/fisiología , Carbohidratos de la Dieta/metabolismo , Proteínas en la Dieta/metabolismo , Digestión , Metabolismo Energético , Equilibrio Hidroelectrolítico , Aminoácidos/sangre , Alimentación Animal/análisis , Fenómenos Fisiológicos Nutricionales de los Animales , Animales , Dieta/veterinaria , Relación Dosis-Respuesta a Droga , Íleon/fisiología , Yeyuno/fisiología , Masculino , Almidón/metabolismo
17.
Int J Mol Sci ; 21(5)2020 Mar 05.
Artículo en Inglés | MEDLINE | ID: mdl-32150890

RESUMEN

Traumatic brain injury (TBI) can cause biochemical and metabolomic alterations in the brain tissue and serum. These alterations can be used for diagnosis and prognosis of TBI. Here, the serum concentrations of seventeen amino acids (AA) were studied for their potential utility as biomarkers of TBI. Twenty-five female, 4-week-old piglets received diffuse (n = 13) or focal (n = 12) TBI. Blood samples were obtained both pre-injury and at either 24-h or 4-days post-TBI. To find a robust panel of biomarkers, the results of focal and diffuse TBIs were combined and multivariate logistic regression analysis, coupled with the best subset selection technique and repeated k-fold cross-validation method, was used to perform a thorough search of all possible subsets of AAs. The combination of serum glycine, taurine, and ornithine was optimal for TBI diagnosis, with 80% sensitivity and 86% overall prediction rate, and showed excellent TBI diagnostic performance, with 100% sensitivity and 78% overall prediction rate, on a separate validation dataset including four uninjured and five injured animals. We found that combinations of biomarkers outperformed any single biomarker. We propose this 3-AA serum biomarker panel to diagnose mild-to-moderate focal/diffuse TBI. The systematic approaches implemented herein can be used for combining parameters from various TBI assessments to develop/evaluate optimal multi-factorial diagnostic/prognostic TBI metrics.


Asunto(s)
Aminoácidos/sangre , Biomarcadores/sangre , Lesiones Traumáticas del Encéfalo/diagnóstico , Encéfalo/metabolismo , Modelos Animales de Enfermedad , Animales , Animales Recién Nacidos , Lesiones Traumáticas del Encéfalo/sangre , Femenino , Metabolómica , Porcinos
18.
Nutrients ; 12(2)2020 Jan 31.
Artículo en Inglés | MEDLINE | ID: mdl-32023931

RESUMEN

Cooking changes the texture and tenderness of red meat, which may influence its digestibility, circulatory amino acids (AA) and gastrointestinal (GI) hormonal responses in consumers. In a randomised crossover intervention, healthy males (n = 12) consumed a beef steak sandwich, in which the beef was cooked by either a pan-fried (PF) or sous-vide (SV) method. Plasma AA were measured by ultrahigh performance liquid chromatography (UPLC), while plasma GI hormones were measured using a flow cytometric multiplex array. Following meat ingestion, the circulatory concentrations of some of the essential AA (all the branched-chain AA: leucine, isoleucine and valine; and threonine), some of the nonessential AA (glycine, alanine, tyrosine and proline) and some of the nonproteogenic AA (taurine, citrulline and ornithine) were increased from fasting levels by 120 or 180 min (p < 0.05). There were no differences in circulating AA concentrations between cooking methods. Likewise, of the measured GI hormones, glucose-dependent insulinotropic peptide (GIP) and glucagon-like peptide-1 (GLP-1) concentrations increased from fasting levels after consumption of the steak sandwich (p < 0.05), with no differences between the cooking methods. In the healthy male adults, protein digestion and circulating GI hormone responses to a beef-steak breakfast were unaltered by the different cooking methods.


Asunto(s)
Aminoácidos/sangre , Culinaria/métodos , Proteínas en la Dieta/sangre , Hormonas Gastrointestinales/sangre , Carne Roja , Adolescente , Adulto , Estudios Cruzados , Ingestión de Alimentos , Ayuno/sangre , Voluntarios Sanos , Humanos , Masculino , Periodo Posprandial , Adulto Joven
19.
J Nutr ; 150(5): 1178-1185, 2020 05 01.
Artículo en Inglés | MEDLINE | ID: mdl-32006007

RESUMEN

BACKGROUND: Legumes are an excellent plant source of the limiting indispensable amino acid (IAA) lysine in vegetarian, cereal-based diets. However, their digestibility is poor largely because of their antiprotease content. Extrusion can enhance digestibility by inactivating trypsin inhibitors and thus potentially improve the protein quality of legumes. OBJECTIVE: We measured the digestibility of extruded chickpea and yellow pea protein with use of a dual stable isotope method in moderately stunted South Indian primary school children. METHODS: Twenty-eight moderately stunted children (height-for-age z scores <-2.0 SD and >-3.0 SD) aged 6-11 y from low to middle socioeconomic status were randomly assigned to receive a test protein (extruded intrinsically [2H]-labeled chickpea or yellow pea) along with a standard of U-[13C]-spirulina protein to measure amino acid (AA) digestibility with use of a dual stable isotope method. Individual AA digestibility in the test protein was calculated by the ratios of AA enrichments in the test protein to the standard protein in the food and their appearance in blood plasma collected at 6 and 6.5 h during the experiment, representing a plateau state. RESULTS: The mean AA digestibility of extruded chickpea and yellow pea protein in moderately stunted children (HAZ; -2.86 to -1.2) was high and similar in both extruded test proteins (89.0% and 88.0%, respectively, P = 0.83). However, lysine and proline digestibilities were higher in extruded chickpea than yellow pea (79.2% compared with 76.5% and 75.0% compared with 72.0%, respectively, P < 0.02). CONCLUSION: Extruded chickpea and yellow pea protein had good IAA digestibility in moderately stunted children, which was 20% higher than an earlier report of their digestibility when pressure-cooked, measured by the same method in adults. Higher digestibility of lysine and proline highlights better retention of these AA in chickpea during extrusion-based processing. Extrusion might be useful for developing high-quality protein foods from legumes. This trial was registered at www.ctri.nic.in as CTRI/2018/03/012439.


Asunto(s)
Aminoácidos/metabolismo , Cicer/química , Digestión , Manipulación de Alimentos/métodos , Trastornos del Crecimiento/metabolismo , Guisantes/química , Aminoácidos/sangre , Isótopos de Carbono , Niño , Deuterio , Femenino , Humanos , India , Lisina/metabolismo , Masculino , Proteínas de Plantas/metabolismo , Prolina/metabolismo , Inhibidores de Tripsina
20.
J Nutr ; 150(5): 1160-1166, 2020 05 01.
Artículo en Inglés | MEDLINE | ID: mdl-32006029

RESUMEN

BACKGROUND: Poor nutritional status is frequently observed in end-stage renal disease patients and associated with adverse clinical outcomes and increased mortality. Loss of amino acids (AAs) during hemodialysis (HD) may contribute to protein malnutrition in these patients. OBJECTIVE: We aimed to assess the extent of AA loss during HD in end-stage renal disease patients consuming their habitual diet. METHODS: Ten anuric chronic HD patients (mean ± SD age: 67.9 ± 19.3 y, BMI: 23.2 ± 3.5 kg/m2), undergoing HD 3 times per week, were selected to participate in this study. Spent dialysate was collected continuously and plasma samples were obtained directly before and after a single HD session in each participant. AA profiles in spent dialysate and in pre-HD and post-HD plasma were measured through ultra-performance liquid chromatography to determine AA concentrations and, as such, net loss of AAs. In addition, dietary intake before and throughout HD was assessed using a 24-h food recall questionnaire during HD. Paired-sample t tests were conducted to compare pre-HD and post-HD plasma AA concentrations. RESULTS: During an HD session, 11.95 ± 0.69 g AAs were lost via the dialysate, of which 8.26 ± 0.46 g were nonessential AAs, 3.69 ± 0.31 g were essential AAs, and 1.64 ± 0.17 g were branched-chain AAs. As a consequence, plasma total and essential AA concentrations declined significantly from 2.88 ± 0.15 and 0.80 ± 0.05 mmol/L to 2.27 ± 0.11 and 0.66 ± 0.05 mmol/L, respectively (P < 0.05). AA profiles of pre-HD plasma and spent dialysate were similar. Moreover, AA concentrations in pre-HD plasma and spent dialysate were strongly correlated (Spearman's ρ = 0.92, P < 0.001). CONCLUSIONS: During a single HD session, ∼12 g AAs are lost into the dialysate, causing a significant decline in plasma AA concentrations. AA loss during HD can contribute substantially to protein malnutrition in end-stage renal disease patients. This study was registered at the Netherlands Trial Registry (NTR7101).


Asunto(s)
Aminoácidos/sangre , Soluciones para Diálisis/análisis , Fallo Renal Crónico/terapia , Desnutrición Proteico-Calórica/etiología , Diálisis Renal/efectos adversos , Anciano , Anciano de 80 o más Años , Aminoácidos/análisis , Dieta , Proteínas en la Dieta/administración & dosificación , Femenino , Humanos , Masculino , Persona de Mediana Edad , Estado Nutricional
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