Untargeted metabolomics for plasma biomarker discovery for early chronic kidney disease diagnosis in pediatric patients using LC-QTOF-MS.

Pediatric chronic kidney disease (CKD) is a clinical syndrome characterized by renal hypofunction occurring due to gradual and irreversible kidney damage that can further progress over time. New biomarkers may help early diagnosis of pediatric patients suffering from CKD and improve the outcome. Untargeted metabolomics based on LC-QTOF-MS has been used to find new biomarkers for the early diagnosis of CKD in plasma from pediatric patients. In order to avoid any bias in the determination of statistically significant entities as a consequence of the data analysis method followed, two different chemometric approaches have been used, Mass Profiler Professional (MPP) software and Matlab R2015a software. Metabolic fingerprints of control and CKD pediatric patients were compared and five metabolites which showed a significant change common to both data analysis procedures were identified. Sphingosine-1-phosphate, n-butyrylcarnitine, cis-4-decenoylcarnitine and an unidentified feature with 126.0930 m/z were found to be increased in plasma from pediatric patients with CKD, whereas bilirubin was significantly decreased. A partial least squares discriminant analysis model built with these 5 entities classified correctly 96% of the samples. In addition, when considering only early CKD patients against controls, a performance of 97% was obtained. Thus, these promising metabolites could be suitable biomarkers for the early diagnosis of pediatric CKD in a clinical setting.

Plasma biomarker discovery for early chronic kidney disease diagnosis based on chemometric approaches using LC-QTOF targeted metabolomics data.

Chronic kidney disease (CKD) is a progressive pathological condition in which renal function deteriorates in time. The first diagnosis of CKD is often carried out in general care attention by general practitioners by means of serum creatinine (CNN) levels. However, it lacks sensitivity and thus, there is a need for new robust biomarkers to allow the detection of kidney damage particularly in early stages. Multivariate data analysis of plasma concentrations obtained from LC-QTOF targeted metabolomics method may reveal metabolites suspicious of being either up-regulated or down-regulated from urea cycle, arginine methylation and arginine-creatine metabolic pathways in CKD pediatrics and controls. The results show that citrulline (CIT), symmetric dimethylarginine (SDMA) and S-adenosylmethionine (SAM) are interesting biomarkers to support diagnosis by CNN: early CKD samples and controls were classified with an increase in classification accuracy of 18% when using these 4 metabolites compared to CNN alone. These metabolites together allow classification of the samples into a definite stage of the disease with an accuracy of 74%, being the 90% of the misclassifications one level above or below the CKD stage set by the nephrologists. Finally, sex-related, age-related and treatment-related effects were studied, to evaluate whether changes in metabolite concentration could be attributable to these factors, and to correct them in case a new equation is developed with these potential biomarkers for the diagnosis and monitoring of pediatric CKD.

Limited beneficial effects of piceatannol supplementation on obesity complications in the obese Zucker rat: gut microbiota, metabolic, endocrine, and cardiac aspects

Resveratrol is beneficial in obese and diabetic rodents. However, its low bioavailability raises questions about its therapeutic relevance for treating or preventing obesity complications. In this context, many related natural polyphenols are being tested for their putative antidiabetic and anti-obesity effects. This prompted us to study the influence of piceatannol, a polyhydroxylated stilbene, on the prevention of obesity complications in Zucker obese rats. A 6-week supplementation was followed by the determination of various markers in plasma, liver, adipose tissue and heart, together with a large-scale analysis of gut microbiota composition. When given in doses of 15 or 45 mg/kg body weight/day, piceatannol did not reduce either hyperphagia or fat accumulation. It did not modify the profusion of the most abundant phyla in gut, though slight changes were observed in the abundance of several Lactobacillus, Clostridium, and Bacteroides species belonging to Firmicutes and Bacteroidetes. This was accompanied by a tendency to reduce plasma lipopolysaccharides by 30 %, and by a decrease of circulating non-esterified fatty acids, LDL-cholesterol, and lactate. While piceatannol tended to improve lipid handling, it did not mitigate hyperinsulinemia and cardiac hypertrophy. However, it increased cardiac expression of ephrin-B1, a membrane protein that contributes to maintaining cardiomyocyte architecture. Lastly, ascorbyl radical plasma levels and hydrogen peroxide release by adipose tissue were similar in control and treated groups. Thus, piceatannol did not exhibit strong slimming capacities but did limit several obesity complications (AU)

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Limited beneficial effects of piceatannol supplementation on obesity complications in the obese Zucker rat: gut microbiota, metabolic, endocrine, and cardiac aspects.

Resveratrol is beneficial in obese and diabetic rodents. However, its low bioavailability raises questions about its therapeutic relevance for treating or preventing obesity complications. In this context, many related natural polyphenols are being tested for their putative antidiabetic and anti-obesity effects. This prompted us to study the influence of piceatannol, a polyhydroxylated stilbene, on the prevention of obesity complications in Zucker obese rats. A 6-week supplementation was followed by the determination of various markers in plasma, liver, adipose tissue and heart, together with a large-scale analysis of gut microbiota composition. When given in doses of 15 or 45 mg/kg body weight/day, piceatannol did not reduce either hyperphagia or fat accumulation. It did not modify the profusion of the most abundant phyla in gut, though slight changes were observed in the abundance of several Lactobacillus, Clostridium, and Bacteroides species belonging to Firmicutes and Bacteroidetes. This was accompanied by a tendency to reduce plasma lipopolysaccharides by 30 %, and by a decrease of circulating non-esterified fatty acids, LDL-cholesterol, and lactate. While piceatannol tended to improve lipid handling, it did not mitigate hyperinsulinemia and cardiac hypertrophy. However, it increased cardiac expression of ephrin-B1, a membrane protein that contributes to maintaining cardiomyocyte architecture. Lastly, ascorbyl radical plasma levels and hydrogen peroxide release by adipose tissue were similar in control and treated groups. Thus, piceatannol did not exhibit strong slimming capacities but did limit several obesity complications.

Recomendaciones y manejo de la tirosinemia hereditaria Tipo I o Tirosinemia hepatorrenal / Recommendations and management of Type I hereditary or hepatorenal Tyrosinemia

La Tirosinemia tipo I es una enfermedad potencialmente letal si no se diagnostica y trata adecuadamente. Los avances diagnóstico terapéuticos en los últimos años han mejorado ostensiblemente el pronóstico de estos pacientes. Por ello es importante que el pediatra disponga de una guía de práctica clínica con recomendaciones para el diagnóstico, y manejo terapéutico de esta enfermedad que contribuya a una adecuada actuación (AU)

Tyrosinemia type I is a potentially lethal disease if not diagnosed and treated properly. Diagnostic and therapeutic advances in recent years have significantly improved the prognosis for these patients. It is therefore important that the pediatrician has a clinical practice guideline with recommendations for diagnosis and treatment of this disease that leads to the appropriate intervention (AU)

[Recommendations and management of type I hereditary or hepatorenal tyrosinemia]. / Recomendaciones y manejo de la tirosinemia hereditaria Tipo I o Tirosinemia hepatorrenal.

Tyrosinemia type I is a potentially lethal disease if not diagnosed and treated properly. Diagnostic and therapeutic advances in recent years have significantly improved the prognosis for these patients. It is therefore important that the pediatrician has a clinical practice guideline with recommendations for diagnosis and treatment of this disease that leads to the appropriate intervention.

Secondary disorders of glycosylation in inborn errors of fructose metabolism.

Adamowicz and colleagues raised the alert in 2007 about patients with atypical hereditary fructose intolerance (HFI) primarily misdiagnosed as CDG Ix. We describe a girl with neonatal hypertonia, facial trismus, absent swallowing and coughing reflexes, gastro-oesophageal reflux and sporadically elevated Krebs cycle metabolites and lactate. At 14 months microcephaly and hepatomegaly were noted, with hypertransaminasaemia but normal blood coagulation, glucose, phosphate, and absent urinary reducing substances. Neurological impairment persisted. Because of hepatic and neurological abnormalities with developmental delay, Tf IEF was performed and showed a severe type 1 pattern, resulting in a wrong diagnosis of CDG. Subsequently, an aversion to fruits suggested HFI, confirmed by the finding of ALDOB mutations (p.A150P/p.N335K). The girl improved with fructose-free diet, but liver cirrhosis led to hepatic transplantation. She is now 7 years old with good evolution; facial trismus and hypertonia reversed, but microcephaly persists. Transferrin MALDI-TOF MS characterization revealed underoccupation of glycosylation sites and glycan abnormalities, which reversed with dietary treatment. High maternal fructose concentrations might have caused neonatal abnormalities. Although in our patient's mother there is no fructose accumulation at present, it is possible that increased ingestion of fruits and vegetables during pregnancy, together with her heterozygosity, caused an accumulation of fructose that finally affected the fetus. We also describe slightly abnormal transferrin isoelectric focusing and MALDI-TOF MS patterns of intact transferrin and N-glycans in a fructose-1,6-bisphosphatase (FBP1)-deficient patient. While HFI is a well-known cause of secondary CDG, we found no reports of abnormal transferrin isoelectric focusing patterns in FBP1 deficiency and we introduce this condition as a possible secondary cause for altered transferrin isoelectric focusing.

[On-line resources in dealing with rare diseases]. / Recursos on-line en el manejo de enfermedades raras.

The increase in scientific knowledge and the need for its transmission to health professionals and patients has resulted in the creation of websites as a useful tool. In low prevalence diseases, such as rare diseases (RDs), sites are being created by scientific societies, institutions and patients. This section reviews the most important websites dealing with RDs both nationally and internationally.

Methylmalonic acidaemia: examination of genotype and biochemical data in 32 patients belonging to mut, cblA or cblB complementation group.

Methylmalonic acidaemia (MMA) is a genetic disorder caused by defects in methylmalonyl-CoA mutase or in any of the different proteins involved in the synthesis of adenosylcobalamin. The aim of this work was to examine the biochemical and clinical phenotype of 32 MMA patients according to their genotype, and to study the mutant mRNA stability by real-time PCR analysis. Using cellular and biochemical methods, we classified our patient cohort as having the MMA forms mut (n = 19), cblA (n = 9) and cblB (n = 4). All the mut (0) and some of the cblB patients had the most severe clinical and biochemical manifestations, displaying non-inducible propionate incorporation in the presence of hydroxocobalamin (OHCbl) in vitro and high plasma odd-numbered long-chain fatty acid (OLCFA) concentrations under dietary therapy. In contrast, mut (-) and cblA patients exhibited a milder phenotype with propionate incorporation enhanced by OHCbl and normal OLCFA levels under dietary therapy. No missense mutations identified in the MUT gene, including mut (0) and mut (-) changes, affected mRNA stability. A new sequence variation (c.562G>C) in the MMAA gene was identified. Most of the cblA patients carried premature termination codons (PTC) in both alleles. Interestingly, the transcripts containing the PTC mutations were insensitive to nonsense-mediated decay (NMD).

Recursos on-line en el manejo de enfermedades raras / On-line resources in dealing with rare diseases

El incremento del conocimiento científico y lanecesidad de su transmisión a los responsables de lasalud y a los pacientes, ha propiciado que una herramientaútil sea la creación de portales web. En enfermedadespoco prevalentes, como las enfermedadesraras (ER), se asiste a la creación de portales que surgendesde el área de las sociedades científicas, desdelos pacientes y las instituciones. En este capítulo seintenta reseñar los portales más significativos en ERtanto a nivel nacional como internacional

The increase in scientific knowledge and the needfor its transmission to health professionals andpatients has resulted in the creation of websites as auseful tool. In low prevalence diseases, such as rarediseases (RDs), sites are being created by scientificsocieties, institutions and patients. This sectionreviews the most important websites dealing with RDsboth nationally and internationally

[A randomized single-blind trial of the effects of vitamins C and E in familial hypercholesterolemia]. / Ensayo aleatorizado ciego-sencillo sobre los efectos de las vitaminas C y E en la hipercolesterolemia familiar.

INTRODUCTION: The aim of this study was to evaluate the effect of antioxidant vitamin C and E administration on dyslipidemia, plasma fatty acid composition, and biochemical inflammatory markers in children with heterozygous familial hypercholesterolemia (FH). PATIENTS: Forty girls and boys with heterozygous FH, aged between 2 and 18 years, and with plasma low-density lipoprotein (LDL)-cholesterol levels higher than 160 mg/dl were studied. METHODS: We performed an open longitudinal randomized trial over a 1-year period. All children followed a dietary intervention according to the National Cholesterol Education Program (NCEP)-1 guidelines and were randomized into two groups. One group (n = 21) received therapy with vitamin C (500 mg twice a day) and vitamin E (400 IU per day). A second group (n = 19) did not receive vitamin therapy. RESULTS: In patients receiving antioxidant vitamins, plasma linoleic acid levels (18:2 omega-6) significantly increased and the essential fatty acid deficiency ratio significantly decreased (Mead/arachidonic acid: 20:4 omega-6/20:3 omega-9). No significant differences were observed in plasma lipid profile, adhesion molecules, or reactive C protein. CONCLUSIONS: Antioxidant vitamin therapy in children with heterozygous FH modifies the plasma fatty acid profile. These modifications are independent of the degree of dyslipidemia and may represent an indicator of reduced cardiovascular risk.

Ensayo aleatorizado ciego-sencillo sobre los efectos de las vitaminas C y E en la hipercolesterolemia familiar / A randomized single-blind trial of the effects of vitamins C and E in familial hypercholesterolemia

Introducción El objetivo de este trabajo ha sido evaluar el efecto de la administración de las vitaminas antioxidantes C y E sobre la dislipemia, la composición de los ácidos grasos en plasma y los marcadores bioquímicos de inflamación en niños afectados de hipercolesterolemia familiar (HF) heterozigótica. Pacientes Cuarenta niños de ambos sexos afectados de HF heterozigótica, de edades comprendidas entre 2 y 18 años, y con valores de colesterol de las lipoproteínas de baja densidad (c-LDL) en plasma superiores a 160 mg/dl. Métodos Estudio aleatorizado longitudinal abierto de un año de duración. Todos los niños siguieron una intervención dietaria de acuerdo con las guías National Cholesterol Education Program (NCEP-1) y fueron aleatorizados en 2 grupos. El primer grupo (n = 21), recibió una terapia con vitamina C (500 mg dos veces al día) y vitamina E (400 U/día). Un segundo grupo (n = 19) no recibió terapia alguna con vitaminas. Resultados Los pacientes que recibieron vitaminas antioxidantes presentaron un aumento significativo en los niveles de ácido linoleico (18:2 ω-6) en plasma y un descenso significativo en el índice de deficiencia de ácidos grasos esenciales (Mead/araquidónico: 20:3 ω-9/20:4 ω-6). No se observaron diferencias significativas en el perfil lipídico en plasma, moléculas de adhesión o proteína C reactiva. Conclusiones La terapia con vitaminas antioxidantes en niños con HF heterozigótica muestra modificaciones del perfil de ácidos grasos que son independientes del grado de dislipemia y podría representar un indicador de disminución de riesgo cardiovascular

Introduction The aim of this study was to evaluate the effect of antioxidant vitamin C and E administration on dyslipidemia, plasma fatty acid composition, and biochemical inflammatory markers in children with heterozygous familial hypercholesterolemia (FH). Patients Forty girls and boys with heterozygous FH, aged between 2 and 18 years, and with plasma low-density lipoprotein (LDL)-cholesterol levels higher than 160 mg/dl were studied. Methods We performed an open longitudinal randomized trial over a 1-year period. All children followed a dietary intervention according to the National Cholesterol Education Program (NCEP)-1 guidelines and were randomized into two groups. One group (n = 21) received therapy with vitamin C (500 mg twice a day) and vitamin E (400 IU per day). A second group (n = 19) did not receive vitamin therapy. Results In patients receiving antioxidant vitamins, plasma linoleic acid levels (18:2 ω-6) significantly increased and the essential fatty acid deficiency ratio significantly decreased (Mead/arachidonic acid: 20:4 ω-6/20:3 ω-9). No significant differences were observed in plasma lipid profile, adhesion molecules, or reactive C protein. Conclusions Antioxidant vitamin therapy in children with heterozygous FH modifies the plasma fatty acid profile. These modifications are independent of the degree of dyslipidemia and may represent an indicator of reduced cardiovascular risk

Effect of docosahexaenoic acid administration on plasma lipid profile and metabolic parameters of children with methylmalonic acidaemia.

AIM: To evaluate the effect of administration of docosahexaenoic acid (DHA) on dyslipidaemia, plasma fatty acid composition and metabolic parameters of children with isolated methylmalonic acidaemia (MMA) (McKusick 25100). METHODS: Four children (3 male, 1 female) with MMA (mut(0)), participated in a crossover, randomized study of DHA administration (25 mg/kg per day, divided into three daily doses). The control group comprised 56 healthy children, aged 10+/- 2.7 years, (51 male, 5 female), who were followed in our clinic owing to possible familial risk of cardiovascular disease. RESULTS: The comparison of plasma fatty acid composition of children with MMA versus control children demonstrated that the patients had significantly higher values for oleic acid (p = 0.004) and linolenic acid (p = 0.008). No differences were observed in the levels of DHA and arachidonic acid. Plasma concentrations of insulin, glycine, ammonia, total cholesterol and cholesterol fractions did not change with DHA administration. No significant changes were observed in urinary excretion of methylmalonic acid. As expected, the percentage of DHA and n-3 fatty acids in plasma increased significantly after therapy (p = 0.005 and 0.014, respectively). The most remarkable result was a decrease of plasma levels of triglycerides after DHA therapy (p = 0.014). CONCLUSION: As previously found in normal children, dietary supplementation with DHA decreases the triglyceride levels, normalizing the hypertriglyceridaemia of these children without any evidence of short-term adverse effects.

Síntomas guía de las enfermedades lisosomales. Una orientación para el pediatra general / Symptoms indicative of lysosomal storage diseases: a guide for the general pediatrician

Las enfermedades lisosomales son un amplio grupo de entidades infrecuentes de manera aislada, que, a diferencia de otros grupos de enfermedades metabólicas, suelen alterar el fenotipo a lo largo de su evolución. Este hecho motiva que algunas de ellas puedan ser diagnosticadas por el pediatra de visu. Además, la posibilidad reciente de terapia enzimática de sustitución hace que algunas tengan ya posibilidad de tratamiento, y otras, próximamente. Estas dos características conllevan la necesidad de que el pediatra conozca su sintomatología y las características de los fenotipos para incrementar la sospecha diagnóstica

Lysosomal storage diseases comprise a broad group of rare diseases which, unlike other groups of metabolic disorders, tend to produce phenotypic changes over the course of time (facial dysmorphism and other skeletal abnormalities). This circumstance enables pediatricians to diagnose some of them on the basis of visual inspection. Moreover, the recent introduction of emzyme replacement therapy has made it possible to treat some of these diseases, and others will follow in the near future. Thus, since a high degree of clinical suspicion is the key to their diagnosis, the pediatrician must be familiar with the symptomatology and the most common phenotypic features associated with these metabolic disorders

Leucoencefalopatía mitocondrial del lactante. ¿Forma precoz de un síndrome de Leigh? / Mitochondrial leukoencephalopathy of infancy: is it an early expression of Leigh syndrome?

Introducción: La enfermedad de Leigh es, probablemente, la enfermedad metabólica más frecuente de presentación en la infancia. La forma clásica de la enfermedad afecta de forma principal a los ganglios basales y el troncoencéfalo. La afectación extensa de la sustancia blanca en ausencia de alteraciones de los ganglios basales es una manifestación poco habitual. Objetivo: Describir el caso de cuatro pacientes que debutaron antes del año de edad. Pacientes y métodos: Los cuatro pacientes presentaron una clínica estereotipada, consistente en regresión de las habilidades psicomotrices adquiridas y desarrollo progresivo de un síndrome piramidal. Las manifestaciones clínicas junto con el hallazgo en la neuroimagen sugerían, inicialmente, una leucodistrofia primaria. La determinación de cifras elevadas de ácido láctico y pirúvico orientaron hacia una enfermedad mitocondrial. Los estudios enzimáticos confirmaron un déficit de los complejos enzimáticos de la cadena respiratoria en dos de los pacientes y, en otras dos hermanas, del complejo piruvato deshidrogenasa. En estas dos últimas, el examen necrópsico mostró las lesiones elementales de la enfermedad de Leigh pero con una distribución inusual. Conclusión: Se debe tener en cuenta que el síndrome de Leigh puede manifestarse con la apariencia de una leucodistrofia en ausencia de las alteraciones típicas de los ganglios basales si afecta a un niño de pocos meses de edad (AU)

[Mitochondrial leukoencephalopathy of infancy: is it an early expression of Leigh syndrome?]. / Leucoencefalopatía mitocondrial del lactante. ¿Forma precoz de un síndrome de Leigh?

INTRODUCTION: Leigh syndrome is probably the most frequent metabolic disorder in infancy and childhood. The classic form of the disease is characterized by bilateral lesions of basal ganglia and brainstem. The extensive involvement of white matter, without radiological basal ganglia abnormalities, is an unusual manifestation of the disease. OBJECTIVE: Four patients who presented the disease during the first year of life are described. PATIENTS AND METHODS: The four patients presented a stereotyped clinical picture, consisting of regression of already acquired psychomotor abilities and very prominent pyramidal signs. These clinical manifestations and results of neuroimaging studies suggested a primary leukodystrophy. Increased values of lactic and piruvic acids suggested a mitochondrial disorder. Enzymatic studies confirmed a mitochondrial respiratory chain deficiency in two patients, and a pyruvate dehydrogenase complex defect in the remaining two patients. The pathological findings in the latter two sisters were consistent with the characteristic microscopic lesions of Leigh syndrome, but with atypical distribution. CONCLUSION: Diagnosis of Leigh syndrome must be taken into consideration in infants presenting with a leukodystrophic clinical and radiological pattern, despite the lack of basal ganglia involvement.

Polyunsaturated fatty acid deficiency during dietary treatment of very long-chain acyl-CoA dehydrogenase deficiency. Rescue with soybean oil.

Nutritional management of very long-chain acyl-CoA dehydrogenase (VLCAD) deficiency is based on the avoidance of fasting and substitution of medium-chain triglycerides for long- and very long-chain triglycerides. We report two cases of this disease, which developed omega-6 essential fatty acid deficiency after three and five months from the beginning of nutritional therapy (SHS product: Monogen). This alteration could be especially dangerous in these patients owing to their possible susceptibility to the development of pigmentary retinopathy. The incorporation of linoleic acid as 3-4% of total caloric intake supported as soybean oil ameliorates this deficiency. We wish to remark on this early complication in the nutritional management of VLCAD deficiency and the possibility of rescue by the incorporation of soybean oil into the diet.

Continuous versus cyclic parenteral nutrition during bone marrow transplantation: assessment and follow-up.

The aim of this prospective, randomized clinical trial was to compare the efficacy of parenteral nutrition (PN) in adult bone marrow transplant (BMT) patients on a continuous (CON) versus a cyclic (CYC) regimen. Twelve patients received CON PN and 12 CYC PN. The groups were homogeneous. CON PN received 27.2 +/- 3.7 kcal/kg/day and for CYC PN 25.9 +/- 4.2 kcal/kg/day (P= 0.45). The duration of PN was 20.4 +/- 7.9 days and 27.3 +/- 13.4 days respectively (P = 0.14). There were no differences between initial and final body weights, either within or between groups. The initial pre-albumin levels were 16.1 +/- 7.5 mg/dl and 20.1 +/- 4.9 mg/dl in CON PN and CYC PN, respectively (P= 0.22), and these were maintained throughout the study. Blood glucose levels did not differ between groups. Measures of liver function were moderately increased, but with no significant differences between groups. There were no significant differences regarding the efficacy of PN or the appearance of complications between CON PN and CYC PN in patients with BMT.