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We study the abundance patterns and the radial gradients of s-process elements (Y, Zr, Ba, La and Ce), r-process elements (Eu) and mixed-process elements (Mo, Nd and Pr) in the Galactic thin disc by means of a detailed two-infall chemical evolution model for the Milky Way with state-of-the-art nucleosynthesis prescriptions. We consider r-process nucleosynthesis from merging neutron stars (MNS), magneto-rotational supernovae (MR-SNe) and s-process synthesis from low- and intermediate- mass stars (LIMS) and rotating massive stars. The predictions of our model are compared with data from the sixth data release of the Gaia-ESO survey, from which we consider 62 open clusters with age > 0.1 Gyr and 1300 Milky Way disc field stars. We conclude that: i) the [Eu/Fe] vs. [Fe/H] is reproduced by both a prompt and a delayed source, but the quick source completely dominates the Eu production; ii) rotation in massive stars contribute substantially to the s-process elements of the first peak, but MNS and MR-SNe are necessary in order to reproduce the observations; iii) due to the adopted yields, our model overpredicts Pr and underpredicts Nd, while the [Mo/Fe] vs. [Fe/H] is nicely reproduced. For the radial gradients, we conclude that: i) our predicted slope of the [Fe/H] gradient is in agreement with the one observed in open clusters by Gaia-ESO and other high-resolution spectroscopic surveys. ii) The predicted slope of the [Eu/H] radial gradient is steeper than the observed one, independently on how quick the production of Eu is. We discuss the possible causes of this discrepancy in terms of both different Galaxy formation scenarios and stellar radial migration effects. iii) For all the elements belonging to the second s-process peak (Ba, La, Ce) as well as for Pr, we predict a plateau at low Galactocentric distances, which is probably due to the enhanced enrichment from LIMS in the inner regions., 17 pages, 11 figures. Submitted to MNRAS

We study the formation and evolution of elliptical galaxies and how they suppress star formation and maintain it quenched. A one-zone chemical model which follows in detail the time evolution of gas mass and its chemical abundances during the active and passive evolution, is adopted. The model includes both gas infall and outflow as well as detailed stellar nucleosynthesis. Elliptical galaxies with different infall masses, following a down-sizing in star formation scenario, are considered. In the chemical evolution simulation we include a novel calculation of the feedback processes. We include heating by stellar wind, core-collapse SNe, Type Ia SNe (usually not highlighted in galaxy formation simulations) and AGN feedback. The AGN feedback is a novelty in this kind of models and is computed by considering a Bondi-Eddington limited accretion onto the central supermassive black hole. We successfully reproduce several observational features, such as the [$\alpha$/Fe] ratios increasing with galaxy mass, mass-metallicity, $\rm M_{BH}-\sigma$ and $\rm M_{BH}-M_{*}$ relations. Moreover, we show that stellar feedback and in particular Type Ia SNe, has a main role in maintaining quenched the star formation after the occurrence of the main galactic wind, especially in low-mass ellipticals. For larger systems, the contribution from AGN to thermal energy of gas appears to be necessary. However, the effect of the AGN on the development of the main galactic wind is negligible, unless an unreasonable high AGN efficiency or an extremely low stellar feedback are assumed. We emphasize the important role played by Type Ia SNe in the energy budget of early-type galaxies., Comment: 15 pages, 10 figures, Accepted on MNRAS

RCAN proteins are endogenous regulators of the calcineurin-cytosolic nuclear factor of activated T cells (CN-NFATc) pathway that bind CN through similar conserved motifs PxIxIT and LxVP of the NFATc family. RCAN1 and RCAN3 protein levels were reported to correlate with overall survival of breast cancer patients. We additionally provided supporting results about RCAN3 role on cancer showing that overexpression of the native PxIxIT sequence of RCAN3-derived R3 peptide (PSVVVH, EGFP-R3178–210) dramatically inhibits tumor growth and tumor angiogenesis in an orthotopic mouse model of Triple Negative Breast Cancer (TNBC) in nude mice. On the other hand, RCAN3 protein and its derived peptide EGFP-R3178–210 bind to CN and inhibit NFAT-mediated cytokine gene expression without affecting CN phosphatase activity suggesting that RCAN3 and EGFP-R3178–210 peptide have tumor suppressor and immunosuppressant activity. Due to the known relationship between tumor development and immune system, as well as the relevance of CN-NFATc in the regulation of the immune system, in the present study we decided to assess the effect of EGFP-R3178–210 peptide in an orthotopic syngeneic TNBC mouse model, in order to ensure that the role of RCAN3 as immunosuppressant do not override its tumor suppressor activity. Our results evidence that EGFP-R3178–210 peptide displays an inhibitory potential on tumor growth and tumor angiogenesis similar to those obtained in the previous orthotopic TNBC model. These results highlight the importance of the RCAN3 peptide as a tumor suppressor protein and totally complement our previous results, indicating that this antitumor activity role is maintained in the presence of a complete functional immune system.

Background Enterovirus-A71 causes outbreaks of brainstem encephalitis, ranging from self-limited disease to acute flaccid paralysis. The aim of this study was to assess the role of cerebrospinal fluid (CSF) neopterin as a biomarker of disease severity in children with enterovirus-related brainstem encephalitis. Methods A descriptive, prospective cohort study was conducted from April 2016 to March 2017 in a tertiary hospital. Pediatric patients with a diagnosis of brainstem encephalitis with or without myelitis due to enterovirus infection were enrolled. The final study group comprised a convenience sample including all patients with sufficient CSF volume for neopterin determination. The major variables considered in estimating the severity were the diagnosis of encephalomyelitis, the presence of lesions and extensive lesions on brain and spinal magnetic resonance imaging (MRI), hospital stay length greater than seven days, and sequelae at day 30. Results Of 60 patients, CSF neopterin could be measured in 36. Median age was 26 months (interquartile range: 19 to 32). Thirty-three were diagnosed with brainstem encephalitis and three with encephalomyelitis. Enterovirus-A71 was the only identified genotype (25 of 25). CSF neopterin levels were elevated (>61 nmol/L) in 33 of 36 (92%), with a median of 347 nmol/L (interquartile range: 204 to 525). CSF neopterin was useful to distinguish patients with lesions on MRI (area under the receiver operating characteristic curve = 0.76; P = 0.02) and extensive lesions (area under the receiver operating characteristic curve = 0.76; P = 0.04). Conclusions This study suggests an association between CSF neopterin levels and the presence of inflammatory lesions on MRI.

Summary Background Factors affecting outcomes of SARS-CoV-2 infection in people living with HIV are unclear. We assessed the factors associated with SARS-CoV-2 diagnosis and severe outcomes among people living with HIV. Methods We did a retrospective cohort study using data from the PISCIS cohort of people with HIV in Catalonia (Spain) between March 1 and Dec 15, 2020. We linked PISCIS data with integrated health-care, clinical, and surveillance registries through the Public Data Analysis for Health Research and Innovation Program of Catalonia (PADRIS) to obtain data on SARS-CoV-2 diagnosis, chronic comorbidities, as well as clinical and mortality outcomes. Participants were aged at least 16 years in care at 16 hospitals in Catalonia. Factors associated with SARS-CoV-2 diagnoses and severe outcomes were assessed using univariable and multivariable Cox regression models. We estimated the effect of immunosuppression on severe outcomes (hospital admission for >24 h with dyspnoea, tachypnoea, hypoxaemia, asphyxia, or hyperventilation; or death) using Kaplan-Meier survival analysis. Findings We linked 20 847 (72·8%) of 28 666 participants in the PISCIS cohort with PADRIS data; 13 142 people had HIV. 749 (5·7%) people with HIV were diagnosed with SARS-CoV-2: their median age was 43·5 years (IQR 37·0–52·7), 131 (17·5%) were female, and 618 (82·5%) were male. 103 people with HIV (13·8%) were hospitalised, seven (0·9%) admitted to intensive care, and 13 (1·7%) died. SARS-CoV-2 diagnosis was more common among migrants (adjusted hazard ratio 1·55, 95% CI 1·31–1·83), men who have sex with men (1·42, 1·09–1·86), and those with four or more chronic comorbidities (1·46, 1·09–1·97). Age at least 75 years (5·2, 1·8–15·3), non-Spanish origin (2·1, 1·3–3·4), and neuropsychiatric (1·69, 1·07–2·69), autoimmune disease (1·92, 1·14–3·23), respiratory disease (1·84, 1·09–3·09), and metabolic disease (2·59, 1·59–4·23) chronic comorbidities were associated with increased risk of severe outcomes. A Kaplan-Meier estimator showed differences in the risk of severe outcomes according to CD4 cell count in patients with detectable HIV RNA (p=0·039) but no differences were observed in patients with undetectable HIV RNA (p=0·15). Interpretation People living with HIV with detectable HIV viraemia, chronic comorbidities, and some subpopulations could be at increased risk of severe outcomes from COVID-19. These groups should be prioritised in clinical management and SARS-CoV-2 vaccination programmes. Funding Fundacio "la Caixa". Translations For the Catalan, Spanish and Russian translations of the Summary see Supplementary Materials section.

BACKGROUND: Genetic defects of monoamine neurotransmitters are rare neurological diseases amenable to treatment with variable response. They are major causes of early parkinsonism and other spectrum of movement disorders including dopa-responsive dystonia. OBJECTIVES: The objective of this study was to conduct proteomic studies in cerebrospinal fluid (CSF) samples of patients with monoamine defects to detect biomarkers involved in pathophysiology, clinical phenotypes, and treatment response. METHODS: A total of 90 patients from diverse centers of the International Working Group on Neurotransmitter Related Disorders were included in the study (37 untreated before CSF collection, 48 treated and 5 unknown at the collection time). Clinical and molecular metadata were related to the protein abundances in the CSF. RESULTS: Concentrations of 4 proteins were significantly altered, detected by mass spectrometry, and confirmed by immunoassays. First, decreased levels of apolipoprotein D were found in severe cases of aromatic L-amino acid decarboxylase deficiency. Second, low levels of apolipoprotein H were observed in patients with the severe phenotype of tyrosine hydroxylase deficiency, whereas increased concentrations of oligodendrocyte myelin glycoprotein were found in the same subset of patients with tyrosine hydroxylase deficiency. Third, decreased levels of collagen6A3 were observed in treated patients with tetrahydrobiopterin deficiency. CONCLUSION: This study with the largest cohort of patients with monoamine defects studied so far reports the proteomic characterization of CSF and identifies 4 novel biomarkers that bring new insights into the consequences of early dopaminergic deprivation in the developing brain. They open new possibilities to understand their role in the pathophysiology of these disorders, and they may serve as potential predictors of disease severity and therapies. © 2020 International Parkinson and Movement Disorder Society.

The cosmological {7Li problem consists in explaining why the primordial Li abundance, as predicted by the standard Big Bang nucleosynthesis theory with constraints from WMAP and Planck, is a factor of 3 larger than the Li abundance measured in the stars of the Spite plateau defined by old, warm dwarf stars of the Milky Way halo. Several explanations have been proposed to explain this difference, including various Li depletion processes as well as non standard Big Bang nucleosynthesis, but the main question remains unanswered. In this paper, we present detailed chemical evolution models for dwarf spheroidal and ultra faint galaxies, compute the galactic evolution of 7Li abundance in these objects and compare it with observations of similar objects. In our models, Li is mainly produced by novae and cosmic rays and to a minor extent by low and intermediate mass stars. We adopt the yield combination which best fits the Li abundances in the Milky Way stars. It is evident that the observations of dwarf objects define a Spite plateau, identical to that observed in the Milky Way, thus suggesting that the Spite plateau could be a universal feature and its meaning should be discussed. The predictions of our models for dwarf galaxies, are obtained by assuming as Li primordial abundance either the one detected in the atmospheres of the oldest halo stars (Spite plateau; A(Li)=2.2 dex), or the one from cosmological observations (WMAP; A(Li)=2.66 dex). Finally, we discuss the implications of the universality of the Spite plateau results., 7 pages, 4 figures

In this work, we compute rates of merging neutron stars (MNS) in galaxies of different morphological type, as well as the cosmic MNS rate in a unitary volume of the Universe adopting different cosmological scenarios. Our aim is to provide predictions of kilonova rates for future observations both at low and high redshift. In the adopted galaxy models, we take into account the production of r-process elements either by MNS or core-collapse supernovae. In computing the MNS rates we adopt either a constant total time delay for merging (10 Myr) or a distribution function of such delays. Our main conclusions are: i) the observed present time MNS rate in our Galaxy is well reproduced either with a constant time delay or a distribution function $\propto t^{-1}$. The [Eu/Fe] vs. [Fe/H] relation in the Milky Way can be well reproduced with only MNS, if the time delay is short and constant. If the distribution function of delays is adopted, core-collapse supernovae as are also required. ii) The present time cosmic MNS rate can be well reproduced in any cosmological scenario, either pure luminosity evolution or a typical hierarchical one, and spirals are the main contributors to it. iii) The spirals are the major contributors to the cosmic MNS at all redshifts in hierarchical scenarios. In the pure luminosity evolution scenario, the spirals are the major contributors locally, whereas at high redshift ellipticals dominate. iv) The predicted cosmic MNS rate well agrees with the cosmic rate of short Gamma Ray Bursts if the distribution function of delays is adopted, in a cosmological hierarchical scenario observationally derived. v) Future observations of Kilonovae in ellipticals will allow to disentangle among constant or a distribution of time delays as well as among different cosmological scenarios.

Aim Gamma-aminobutyric acid (GABA) is a major modulator in brain maturation and its role in many different neurodevelopmental disorders has been widely reported. Although the involvement of GABA in different disorders has been related to its regulatory function as an inhibitory neurotransmitter in the mature brain, co-transmitter, and signalling molecule, little is known about its role as a clinical biomarker in neuropaediatric disorders. The aim of this study is to report the cerebrospinal fluid (CSF) free-GABA concentrations in a large cohort of patients (n=85) with different neurological disorders. Method GABA was measured in the CSF of neuropaediatric patients using capillary electrophoresis with laser-induced fluorescence detection. Other neurotransmitters (amino acids and monoamines) were also analysed. Results GABA concentrations in CSF were abnormal, with a greater frequency (44%) than monoamines (20%) in neuropaediatric patients compared with our reference values. Although we included a few patients with inborn errors of metabolism, GABA levels in CSF were more frequently abnormal in metabolic disorders than in other nosological groups. Interpretation Our work suggests further research into brain GABAergic status in neuropaediatric disorders, which could also lead to new therapeutic strategies. What this paper adds Homeostasis of GABA seems more vulnerable than that of monoamines in the developing brain. The highest GABA levels are found in the primary GABA neurotransmitter disorder SSADH deficiency. GABA alterations are not specific for any clinical or neuroimaging presentation.

The presence of monoamines and their cofactors (the pterins and vitamin B6 (pyridoxal phosphate (PLP))) in human cerebrospinal fluid (CSF) can be used as indicators of the biosynthesis and turnover of dopamine and serotonin in the brain. In addition, abnormalities in the CSF levels of these molecules are associated with various neurological diseases, including genetic diseases leading to dopamine and serotonin deficiency. Here, we provide a set of quantitative high-performance liquid-chromatography (HPLC) approaches to determine CSF levels of monoamines and their cofactors. This protocol describes step-by-step procedures for CSF sample preparation for the analysis of different molecules, HPLC calibration and analysis, and data quantification and interpretation. Unlike plasma/tissue/blood samples, CSF requires minimal sample preparation: in this protocol, only the analysis of PLP requires mixing with trichloroacetic acid to release the protein-bound vitamin, centrifugation, and mixing of the supernatant with phosphate buffer and sodium cyanide for derivatization in alkaline conditions. Monoamines are analyzed by HPLC with coulometric electrochemical detection (ED), pterins are analyzed by HPLC with coupled coulometric electrochemical and fluorescence detection, and PLP is analyzed by HPLC with fluorescence detection. The quantification of all compounds is achieved by external calibration procedures, and internal quality control and standards are analyzed in each run. We anticipate that investigation of dopamine and serotonin disturbances will be facilitated by measurements of these compounds in human CSF and other biological samples. The estimated time for the different procedures primarily depends on the electrochemical detector stabilization. Overnight stabilization of this detector is advised, and, after that step, preanalytical equilibration rarely exceeds 3 h.

Primary and secondary conditions leading to thiamine deficiency have overlapping features in children, presenting with acute episodes of encephalopathy, bilateral symmetric brain lesions, and high excretion of organic acids that are specific of thiamine-dependent mitochondrial enzymes, mainly lactate, alpha-ketoglutarate, and branched chain keto-acids. Undiagnosed and untreated thiamine deficiencies are often fatal or lead to severe sequelae. Herein, we describe the clinical and genetic characterization of 79 patients with inherited thiamine defects causing encephalopathy in childhood, identifying outcome predictors in patients with pathogenic SLC19A3 variants, the most common genetic etiology. We propose diagnostic criteria that will aid clinicians to establish a faster and accurate diagnosis so that early vitamin supplementation is considered. Ann Neurol 2017;82:317-330.

The elevation of neopterin in cerebrospinal fluid (CSF) has been reported in several neuroinflammatory disorders. However, it is not expected that neopterin alone can discriminate among different neuroinflammatory etiologies. We conducted an observational retrospective and case–control study to analyze the CSF biomarkers neopterin, total proteins, and leukocytes in a large cohort of pediatric patients with neuroinflammatory disorders. CSF samples from 277 subjects were included and classified into four groups: Viral meningoencephalitis, bacterial meningitis, acquired immune-mediated disorders, and patients with no-immune diseases (control group). CSF neopterin was analyzed with high-performance liquid chromatography. Microbiological diagnosis included bacterial CSF cultures and several specific real-time polymerase chain reactions. Molecular testing for multiple respiratory pathogens was also included. Antibodies against neuronal and glial proteins were tested. Canonical discriminant analysis of the three biomarkers was conducted to establish the best discriminant functions for the classification of the different clinical groups. Model validation was done by biomarker analyses in a new cohort of 95 pediatric patients. CSF neopterin displayed the highest values in the viral and bacterial infection groups. By applying canonical discriminant analysis, it was possible to classify the patients into the different groups. Validation analyses displayed good results for neuropediatric patients with no-immune diseases and for viral meningitis patients, followed by the other groups. This study provides initial evidence of a more efficient approach to promote the timely classification of patients with viral and bacterial infections and acquired autoimmune disorders. Through canonical equations, we have validated a new tool that aids in the early and differential diagnosis of these neuroinflammatory conditions.

In this article, we review the state-of-the-art analysis of different biomarkers in the cerebrospinal fluid for the diagnosis of genetically conditioned, rare, neurometabolic diseases, including glucose transport defects, neurotransmitter (dopamine, serotonin, and gamma-aminobutyric acid) and pterin deficiencies, and vitamin defects (folate, vitamin B6, and thiamine) that affect the brain. The analysis of several key metabolites are detailed, which thus highlights the preanalytical and analytical factors that should be cautiously controlled to avoid misdiagnosis; moreover, these factors may facilitate an adequate interpretation of the biochemical profiles in the context of severe neuropediatric disorders. Secondary disturbances in these biomarkers, which are associated with other genetic or environmental conditions, are also detailed. Importantly, the early biochemical identification of biochemical disturbances in the cerebrospinal fluid may improve the clinical outcomes of a remarkable number of patients, who may exhibit good neurologic outcomes using the available therapies for these disorders.

Thiamine is a key cofactor for energy metabolism in brain tissue. There are four major genetic defects (SLC19A2, SLC19A3, SLC25A19 and TPK1) involved in the metabolism and transport of thiamine through cellular and mitochondrial membranes. Neurological involvement predominates in three of them (SLC19A3, SCL25A19 and TPK1), whereas patients with SLC19A2 mutations mainly present extra-neurological features (e.g. diabetes mellitus, megaloblastic anaemia and sensori-neural hearing loss). These genetic defects may be amenable to therapeutic intervention with vitamins supplementation and hence, constitutes a main area of research.We conducted a literature review of all reported cases with these genetic defects, and focused our paper on treatment efficacy and safety, adverse effects, dosing and treatment monitoring. Expert commentary: Doses of thiamine vary according to the genetic defect: for SLC19A2, the usual dose is 25-200 mg/day (1-4 mg/kg per day), for SLC19A3, 10-40 mg/kg per day, and for TPK1, 30 mg/kg per day. Thiamine supplementation in SLC19A3-mutated patients restores CSF and intracellular thiamine levels, resulting in successful clinical benefits. In conclusion, evidence collected so far suggests that the administration of thiamine improves outcome in SLC19A-2, SLC19A3- and TPK1-mutated patients, so most efforts should be aimed at early diagnosis of these disorders.

Mitochondrial diseases are a group of genetic disorders leading to the dysfunction of mitochondrial energy metabolism pathways. We aimed to assess the clinical phenotype and the biochemical cerebrospinal fluid (CSF) biogenic amine profiles of patients with different diagnoses of genetic mitochondrial diseases. We recruited 29 patients with genetically confirmed mitochondrial diseases harboring mutations in either nuclear or mitochondrial DNA (mtDNA) genes. Signs and symptoms of impaired neurotransmission and neuroradiological data were recorded. CSF monoamines, pterins, and 5-methyltetrahydrofolate (5MTHF) concentrations were analyzed using high-performance liquid chromatography with electrochemical and fluorescence detection procedures. The mtDNA mutations were studied by Sanger sequencing, Southern blot, and real-time PCR, and nuclear DNA was assessed either by Sanger or next-generation sequencing. Five out of 29 cases showed predominant dopaminergic signs not attributable to basal ganglia involvement, harboring mutations in different nuclear genes. A chi-square test showed a statistically significant association between high homovanillic acid (HVA) values and low CSF 5-MTHF values (chi-square = 10.916; p = 0.001). Seven out of the eight patients with high CSF HVA values showed cerebral folate deficiency. Five of them harbored mtDNA deletions associated with Kearns-Sayre syndrome (KSS), one had a mitochondrial point mutation at the mtDNA ATPase6 gene, and one had a POLG mutation. In conclusion, dopamine deficiency clinical signs were present in some patients with mitochondrial diseases with different genetic backgrounds. High CSF HVA values, together with a severe cerebral folate deficiency, were observed in KSS patients and in other mtDNA mutation syndromes.

Objective The mechanisms of the ketogenic diet remain unclear, but several predictors of response have been proposed. We aimed is to study the relationship between the etiology of epilepsy, cerebrospinal fluid neurotransmitters, pterins, and amino acids, and response to a ketogenic diet. Methods We studied 60 patients who began classic ketogenic diet treatment for refractory epilepsy. In 24 of 60 individuals, we analyzed cerebrospinal fluid neurotransmitters, pterins, and amino acids in baseline conditions. Mean age at epilepsy onset was 24 months, 83.3% were focal epilepsies, and in 51.7% the etiology of the epilepsy was unknown. Results Six months after initiating the ketogenic diet, it was effective (greater than a 50% reduction in seizure frequency) in 31.6% of patients. We did not find a link between rate of efficacy for the ketogenic diet and etiologies of epilepsy, nor did we find a link between the rate of efficacy for the ketogenic diet and cerebrospinal fluid pterins and biogenic amines concentrations. However, we found statistically significant differences for lysine and arginine values in the cerebrospinal fluid between ketogenic diet responders and nonresponders, but not for the other amino acids analyzed. Significance The values of some amino acids were significantly different in relationship with the ketogenic diet efficacy; however, the epilepsy etiology and the cerebrospinal fluid biogenic amine and pterin values were not.

Cellular folate metabolism is highly compartmentalized, with mitochondria folate transport and metabolism being distinct from the well-known cytosolic folate metabolism. There is evidence supporting the association between low folate status and mitochondrial DNA (mtDNA) instability, and cerebral folate deficiency is relatively frequent in mitochondrial disorders. Furthermore, folinic acid supplementation has been reported to be beneficial not only in some patients with mitochondrial disease, but also in patients with relatively common diseases where folate deficiency might be an important pathophysiological factor. In this review, we focus on the evidence that supports the potential involvement of impaired folate metabolism in the pathophysiology of mitochondrial disorders.

Cerebral folate deficiency is defined as any neurological condition associated with low cerebrospinal fluid folate concentrations. It is becoming increasingly associated with several neurological diseases, either genetic or environmental. Treatment of cerebral folate deficiency by folate supplementation is generally effective, improving the neurological outcome of some patients. However, to treat cerebral folate deficiency, the proper choice of one of the available folate forms is essential. The distinct brain folate metabolism features compared with peripheral folate metabolic pathways strongly suggest the investigation of different folate forms, such as the biologically active folinic acid and 5-methyltetrahydrofolate, since they are efficiently transported to the brain. Regarding the oral doses of the different folate forms, despite the fact that there are some recommendations, there is no general consensus. Further investigation and designing clinical trials are advisable to elucidate these aspects.

Altres ajuts: This work is funded by the European Regional Development Fund (FEDER) and the Fundació La Marató TV3 (20,143,130 to BPD). LMS has a grant from Fundació Sant Joan de Déu. MAK holds a Wellcome Intermediate Clinical Fellowship. The SLC39A14, SLC30A10 and SLC39A8 are considered to be key genes involved in manganese (Mn) homeostasis in humans. Mn levels in plasma and urine are useful tools for early recognition of these disorders. We aimed to explore further biomarkers of Mn deposition in the central nervous system in two siblings presenting with acute dystonia and hypermanganesemia due to mutations in SLC39A14. These biomarkers may help clinicians to establish faster and accurate diagnosis and to monitor disease progression after chelation therapy is administered. A customized gene panel for movement disorders revealed a novel missense variant (c.311G > T; p.Ser104Ile) in SLC39A14 gene in two siblings presenting at the age of 10 months with acute dystonia and motor regression. Mn concentrations were analyzed using inductively coupled mass spectrometry in plasma and cerebrospinal fluid, disclosing elevated Mn levels in the index case compared to control patients. Surprisingly, Mn values were 3-fold higher in CSF than in plasma. We quantified the pallidal index, defined as the ratio between the signal intensity in the globus pallidus and the subcortical frontal white matter in axial T1-weighted MRI, and found significantly higher values in the SLC39A14 patient than in controls. These values increased over a period of 10 years, suggesting the relentless pallidal accumulation of Mn. Following genetic confirmation, a trial with the Mn chelator NaCaEDTA led to a reduction in plasma Mn, zinc and selenium levels. However, parents reported worsening of cervical dystonia, irritability and sleep difficulties and chelation therapy was discontinued. Our study expands the very few descriptions of patients with SLC39A14 mutations. We report for the first time the elevation of Mn in CSF of SLC39A14 mutated patients, supporting the hypothesis that brain is an important organ of Mn deposition in SLC39A14-related disease. The pallidal index is an indirect and non-invasive method that can be used to rate disease progression on follow-up MRIs. Finally, we propose that patients with inherited defects of manganese transport should be initially treated with low doses of NaCaEDTA followed by gradual dose escalation, together with a close monitoring of blood trace elements in order to avoid side effects.

Pyridoxal phosphate (PLP) is the active form of vitamin B6 and a cofactor in many enzyme reactions including neurotransmitter metabolism. PLP metabolism disturbances may mostly lead to refractory seizures. In this report, we review the main pathophysiological factors related with PLP deficiency and our experience in PLP treatment in pediatric patients with low-normal cerebrospinal fluid PLP values who presented epilepsy. Only one case had a definite diagnosis (Phelan-McDermid syndrome). The results of extensive metabolic workups and targeted genetic studies were normal for all patients. In 5 cases, the response to PLP supplementation (10-30mg/kg/d) was initially positive. PLP adverse reactions were noticed in 4 patients and PLP was discontinued; however, one of the most noticeable symptoms was an asymptomatic increase in liver enzymes. These negative results with PLP supplementation are worth reporting, to improve the information we use to treat our patients.

Aim To determine the prevalence of dopaminergic abnormalities in 1388 children with neurological disorders, and to analyse their clinical, neuroradiological, and electrophysiological characteristics. Method We studied biogenic amines in 1388 cerebrospinal fluid (CSF) samples from children with neurological disorders (mean age 3y 10mo, SD 4y 5mo; 712 males, 676 females. Correlations among CSF homovanillic acid (HVA) values and other biochemical, clinical, neuroradiological, and electrophysiological parameters were analysed. Results Twenty-one patients with primary dopaminergic deficiencies were identified. Of the whole sample, 20% showed altered HVA. We report neurological diseases with abnormal CSF HVA values such as pontocerebellar hypoplasia, perinatal asphyxia, central nervous system infections, mitochondrial disorders, and other genetic diseases. Overlapping HVA levels between primary and secondary dopamine deficiencies were observed. Prevalence of low CSF HVA levels was significantly higher in neonatal patients (χ2=84.8, p

Melatonin is synthesized from serotonin and it is excreted as sulphatoxymelatonin in urine. We aim to evaluate urinary sulphatoxymelatonin as a biomarker of brain serotonin status in a cohort of patients with mutations in genes related to serotonin biosynthesis. We analized urinary sulphatoxymelatonin from 65 healthy subjects and from 28 patients with genetic defects. A total of 18 patients were studied: 14 with autosomal dominant and recessive guanosine triphosphate cyclohydrolase-I deficiency; 3 with sepiapterin reductase deficiency; and 1 with aromatic L-amino acid decarboxylase deficiency. Further 11 patients were studied after receiving serotoninergic treatment (serotonin precursors, monoamine oxidase inhibitors, selective serotonin re-uptake inhibitors): 5 with aromatic L-amino acid decarboxylase deficiency; 1 with sepiapterin reductase deficiency; 3 with dihydropteridine reductase deficiency; and 2 with 6-pyruvoyltetrahydropterin synthase deficiency. Among the patients without therapy, 6 presented low urinary sulphatoxymelatonin values, while most of the patients with guanosine triphosphate cyclohydrolase-I deficiency showed normal values. 5 of 11 patients under treatment presented low urine sulphatoxymelatonin values. Thus, decreased excretion of sulphatoxymelatonin is frequently observed in cases with severe genetic disorders affecting serotonin biosynthesis. In conclusion, sulphatoxymelatonin can be a good biomarker to estimate serotonin status in the brain, especially for treatment monitoring purposes.

In this chapter, current knowledge about biomarkers for the study of genetic diseases leading to monoamine deficiencies (dopamine, adrenaline, noradrenaline, and serotonin) is reviewed. Twelve genetic diseases will be discussed as causes of primary disturbances in monoamine metabolism, including deficiencies in tyrosine hydroxylase, dopamine beta hydroxylase, L-aromatic amino acid decarboxylase, monoamine oxidase A, tetrahydrobiopterin biosynthesis, dopamine transporter 1, and vesicular monoamine transporter type 2. Different biomarkers are available for the diagnosis, treatment, and monitoring of these diseases, which can be divided into peripheral markers (that can be analyzed in blood or urine) and central markers (cerebrospinal fluid analysis is required). Among these biomarkers, biogenic amines (homovanillic and 5-hydroxyindoleacetic acids) are the most informative to confirm the clinical suspicion. Biochemical procedures for the study of these conditions are stated. Genetic analysis is mandatory for the definitive diagnosis of monoamine diseases. Next-generation sequencing techniques are becoming available in clinical ­laboratories, expanding our knowledge in this field.

Primary and secondary conditions leading to thiamine deficiency have overlapping features in children, presenting with acute episodes of encephalopathy, bilateral symmetric brain lesions, and high excretion of organic acids that are specific of thiamine-dependent mitochondrial enzymes, mainly lactate, alpha-ketoglutarate, and branched chain keto-acids. Undiagnosed and untreated thiamine deficiencies are often fatal or lead to severe sequelae. Herein, we describe the clinical and genetic characterization of 79 patients with inherited thiamine defects causing encephalopathy in childhood, identifying outcome predictors in patients with pathogenic SLC19A3 variants, the most common genetic etiology. We propose diagnostic criteria that will aid clinicians to establish a faster and accurate diagnosis so that early vitamin supplementation is considered. Ann Neurol 2017;82:317-330.

Resumen Introduccion La enfermedad celiaca (EC) y la diabetes mellitus tipo 1 son alteraciones cronicas que comparten susceptibilidad genetica, presencia de anticuerpos organo especificos e influencia de factores ambientales. Segun un estudio transversal realizado el ano 2001 en nuestro centro, el 8,3% de los pacientes diabeticos tiene anticuerpos especificos de la EC y el 4,9% presenta una lesion intestinal Marsh III. Con estos antecedentes, se instaura el cribado serologico sistematico para la EC en los pacientes que debutan diabetes mellitus de tipo 1 (DM1). El objetivo de este estudio es valorar el resultado de esta estrategia y caracterizar a los pacientes celiacos detectados. Material y metodos Se estudian los 202 pacientes diabeticos que debutan durante el periodo 2002–2007 controlados en la Unidad de Diabetes de nuestro hospital. Se determina la concentracion serica de anticuerpos antitransglutaminasa en el debut diabetico con controles anuales. Resultados y discusion El 6,4% (13/202) de los pacientes diabeticos debutantes tiene EC, siendo mas frecuente entre los que la debutan precozmente (p=0,016) y mostrando un orden de aparicion aleatorizado de ambas enfermedades. En estos pacientes, la EC es mayoritariamente asintomatica. La mitad de ellos muestra una respuesta serologica parcial a la dieta sin gluten, lo que sugiere una falta de motivacion para el tratamiento dietetico. Segun nuestra experiencia, el control metabolico de los pacientes diabeticos con EC puede ser complicado y una elevada proporcion de casos (p=0,013) requiere el uso de bombas de infusion continua de insulina.

The aim of this work was to establish the effect of the cis9,trans11 conjugated linoleic acid (CLA) isomer on mucosal immunity during early life in rats, a period when mucosal immunoglobulin production is poorly developed, as is also the case in humans. CLA supplementation was performed during three life periods: gestation, suckling, and early infancy. The immune status of supplemented animals was evaluated at two time points: at the end of the suckling period (21-day-old rats) and 1 week after weaning (28-day-old rats). Secretory IgA was quantified in intestinal washes from 28-day-old rats by ELISA technique. IgA, TGFbeta, and PPARgamma mRNA expression was measured in small intestine and colon by real time PCR, using Taqman specific probes and primers. IgA mucosal production was enhanced in animals supplemented with CLA during suckling and early infancy: in 28-day-old rats, IgA mRNA expression was increased in small intestine and colon by approximately 6- and 4-fold, respectively, and intestinal IgA protein by approximately 2-fold. TGFbeta gene expression was independent of age and type of tissue considered, and was not modified by dietary CLA. Gene expression of PPARgamma, a possible mediator of CLA's effects was also upregulated in animals receiving CLA during early life. In conclusion, dietary supplementation with CLA during suckling and extended to early infancy enhances development of the intestinal immune response in rats.

Thiamine transporter-2 deficiency is caused by mutations in the SLC19A3 gene. As opposed to other causes of Leigh syndrome, early administration of thiamine and biotin has a dramatic and immediate clinical effect. New biochemical markers are needed to aid in early diagnosis and timely therapeutic intervention. Thiamine derivatives were analysed by high performance liquid chromatography in 106 whole blood and 38 cerebrospinal fluid samples from paediatric controls, 16 cerebrospinal fluid samples from patients with Leigh syndrome, six of whom harboured mutations in the SLC19A3 gene, and 49 patients with other neurological disorders. Free-thiamine was remarkably reduced in the cerebrospinal fluid of five SLC19A3 patients before treatment. In contrast, free-thiamine was slightly decreased in 15.2% of patients with other neurological conditions, and above the reference range in one SLC19A3 patient on thiamine supplementation. We also observed a severe deficiency of free-thiamine and low levels of thiamine diphosphate in fibroblasts from SLC19A3 patients. Surprisingly, pyruvate dehydrogenase activity and mitochondrial substrate oxidation rates were within the control range. Thiamine derivatives normalized after the addition of thiamine to the culture medium. In conclusion, we found a profound deficiency of free-thiamine in the CSF and fibroblasts of patients with thiamine transporter-2 deficiency. Thiamine supplementation led to clinical improvement in patients early treated and restored thiamine values in fibroblasts and cerebrospinal fluid.

BACKGROUND: Inherited neurometabolic disorders (iNMDs) represent a group of almost seven hundred rare diseases whose common manifestations are clinical neurologic or cognitive symptoms that can appear at any time, in the first months/years of age or even later in adulthood. Early diagnosis and timely treatments are often pivotal for the favorable course of the disease. Thus, the elaboration of new evidence-based recommendations for iNMD diagnosis and management is increasingly requested by health care professionals and patients, even though the methodological quality of existing guidelines is largely unclear. InNerMeD-I-Network is the first European network on iNMDs that was created with the aim of sharing and increasing validated information about diagnosis and management of neurometabolic disorders. One of the goals of the project was to determine the number and the methodological quality of existing guidelines and recommendations for iNMDs. ----- METHODS: We performed a systematic search on PubMed, the National Guideline Clearinghouse (NGC), the Guidelines International Network (G-I-N), the Scottish Intercollegiate Guideline Network (SIGN) and the National Institute for Health and Care Excellence (NICE) to identify all the published guidelines and recommendations for iNMDs from January 2000 to June 2015. The methodological quality of the selected documents was determined using the AGREE II instrument, an appraisal tool composed of 6 domains covering 23 key items. ----- RESULTS: A total of 55 records met the inclusion criteria, 11 % were about groups of disorders, whereas the majority encompassed only one disorder. Lysosomal disorders, and in particular Fabry, Gaucher disease and mucopolysaccharidoses where the most studied. The overall methodological quality of the recommendation was acceptable and increased over time, with 25 % of the identified guidelines strongly recommended by the appraisers, 64 % recommended, and 11 % not recommended. However, heterogeneity in the obtained scores for each domain was observed among documents covering different groups of disorders and some domains like 'stakeholder involvement' and 'applicability' were generally scarcely addressed. ----- CONCLUSIONS: Greater efforts should be devoted to improve the methodological quality of guidelines and recommendations for iNMDs and AGREE II instrument seems advisable for new guideline development. The elaboration of new guidelines encompassing still uncovered disorders is badly needed.

To determine whether glutamine (Gln) supplementation would have a role modifying both the oxidative stress and the inflammatory response of critically ill children.Prospective, randomized, double-blind, interventional clinical trial. Selection criteria were children requiring parenteral nutrition for at least 5 days diagnosed with severe sepsis or post major surgery. Patients were randomly assigned to standard parenteral nutrition (SPN, 49 subjects) or standard parenteral nutrition with glutamine supplementation (SPN + Gln, 49 subjects).Glutamine levels failed to show statistical differences between groups. At day 5, patients in the SPN + Gln group had significantly higher levels of HSP-70 (heat shock protein 70) as compared with the SPN group (68.6 vs 5.4, p = 0.014). In both groups, IL-6 (interleukine 6) levels showed a remarkable descent from baseline and day 2 (SPN: 42.24 vs 9.39, p0.001; SPN + Gln: 35.20 vs 13.80, p0.001) but only the treatment group showed a statistically significant decrease between day 2 and day 5 (13.80 vs 10.55, p = 0.013). Levels of IL-10 (interleukine 10) did not vary among visits except in the SPN between baseline and day 2 (9.55 vs 5.356, p0.001). At the end of the study, no significant differences between groups for PICU and hospital stay were observed. No adverse events were detected in any group.Glutamine supplementation in critically-ill children contributed to maintain high HSP-70 levels for longer. Glutamine supplementation had no influence on IL-10 and failed to show a significant reduction of IL-6 levels.

[Background] The clinical characteristics distinguishing treatable thiamine transporter-2 deficiency (ThTR2) due to SLC19A3 genetic defects from the other devastating causes of Leigh syndrome are sparse., [Methods] We report the clinical follow-up after thiamine and biotin supplementation in four children with ThTR2 deficiency presenting with Leigh and biotin-thiamine-responsive basal ganglia disease phenotypes. We established whole-blood thiamine reference values in 106 non-neurological affected children and monitored thiamine levels in SLC19A3 patients after the initiation of treatment. We compared our results with those of 69 patients with ThTR2 deficiency after a review of the literature., [Results] At diagnosis, the patients were aged 1 month to 17 years, and all of them showed signs of acute encephalopathy, generalized dystonia, and brain lesions affecting the dorsal striatum and medial thalami. One patient died of septicemia, while the remaining patients evidenced clinical and radiological improvements shortly after the initiation of thiamine. Upon follow-up, the patients received a combination of thiamine (10–40 mg/kg/day) and biotin (1–2 mg/kg/day) and remained stable with residual dystonia and speech difficulties. After establishing reference values for the different age groups, whole-blood thiamine quantification was a useful method for treatment monitoring., [Conclusions] ThTR2 deficiency is a reversible cause of acute dystonia and Leigh encephalopathy in the pediatric years. Brain lesions affecting the dorsal striatum and medial thalami may be useful in the differential diagnosis of other causes of Leigh syndrome. Further studies are needed to validate the therapeutic doses of thiamine and how to monitor them in these patients., Supported by Fondo de Investigación Sanitaria Grant PI12/02010 and PI12/02078; Centre for Biomedical Research on Rare Diseases, an initiative of the Instituto de Salud Carlos III, Barcelona, Spain; Agència de Gestio’ d’Ajuts Universitaris i de Recerca-Agaur FI-DGR 2014 (JD Ortigoza-Escobar).

Rett Syndrome is a neurodevelopmental autism spectrum disorder caused by mutations in the gene coding for methyl CpG-binding protein (MeCP2). The disease is characterized by abnormal motor, respiratory, cognitive impairment, and autistic-like behaviors. No effective treatment of the disorder is available. Mecp2 knockout mice have a range of physiological and neurological abnormalities that resemble the human syndrome and can be used as a model to interrogate new therapies. Herein, we show that the combined administration of Levodopa and a Dopa-decarboxylase inhibitor in RTT mouse models is well tolerated, diminishes RTT-associated symptoms, and increases life span. The amelioration of RTT symptomatology is particularly significant in those features controlled by the dopaminergic pathway in the nigrostratium, such as mobility, tremor, and breathing. Most important, the improvement of the RTT phenotype upon use of the combined treatment is reflected at the cellular level by the development of neuronal dendritic growth. However, much work is required to extend the duration of the benefit of the described preclinical treatment.

Objectives Thiamine transporter-2 (hTHTR2) deficiency due to SLC19A3 mutations is a potentially reversible cause of Leigh syndrome for which no biochemical markers are currently available. Our aim was to assess the sensitivity of thiamine quantification in cerebrospinal fluid (CSF) and fibroblasts from patients with Leigh encephalopathy and SLC19A3 defects as compared with other causes of Leigh syndrome. Methods Thiamine vitamers (free-thiamine, thiamine monophosphate (TMP) and thiamine diphosphate (TDP)) were analyzed by HPLC-fluorescence detection in whole-blood and cerebrospinal fluid (CSF) samples from 106 and 38 paediatric controls, respectively. Results were compared with patients with Leigh syndrome due to SLC19A3 defects (N=6) and mitochondrial respiratory chain defects (N=9). In all but one SLC19A3 patient, samples were collected before thiamine supplementation. Thiamine vitamers were also analyzed by HPLC in fibroblasts from SLC19A3 patients (N=3) and patients with other metabolic defects (N=6). Results A negative correlation between thiamine isoforms and age was detected in whole-blood and CSF, thus three reference intervals were established for free-thiamine and two intervals for TMP and TDP. Free-thiamine was severely reduced in five non-treated SLC19A3 patients CSF, but not TMP and TPP. The SLC19A3 patient under thiamine supplementation showed thiamine values above reference range. Nine Leigh patients with mitochondrial defects showed normal o slightly reduced values for CSF thiamine. In SLC19A3 patient's fibroblasts, a reduction in free-thiamine was detected as compared with control sample. These values normalized after thiamine was added to the culture medium. Conclusion SLC19A3 patients show a profound deficiency of free-thiamine in the CSF that allows their identification from other causes of Leigh syndrome. SLC19A3 is essential to maintain CSF thiamine homeostasis and to prevent brain damage. Thiamine overload can supply the SLC19A3 defect and restore thiamine values in fibroblasts and CSF, probably by an alternative transport system.

The measurement of γ-aminobutyric acid (GABA) is suitable for investigating various neurological disorders. In this study, a sensitive and selective method for free GABA quantification in cerebrospinal fluid (CSF) has been standardised. This method is based on CE with LIF detection using 4-fluoro-7-nitrobenzo-2-oxa-1,3-diazole (NBD-F) as a derivatisating agent. The reaction conditions (NBD-F concentration, pH, temperature and reaction time) and the electrophoretic parameters (run buffer composition and pH and separation voltage) were optimised to obtain the maximum derivatisation efficiency and electrophoretic resolution. The best resolution was obtained using 200 mM sodium borate, 10 mM SDS, 8.5 mM β-CD, pH 10 and 20 kV voltage. The method was linear in the concentration range of 2.5-1000 nM with good inter- and intra-assay precision values. The effects of CSF handling on free GABA concentrations were also evaluated. Our results show that the time delay between CSF collection and freezing strongly increases the CSF GABA values. Age-related reference values were established in 55 paediatric controls. The influence of antiepileptic therapy on free CSF GABA was studied in 38 neuropaediatric patients. Significantly, higher GABA values were obtained in patients taking valproic acid or vigabatrin therapy, which are antiepileptic drugs that modulate GABA metabolism.

Epilepsy is detected in about 23% of patients with fragile X syndrome (FXS). Absence or reduced levels of the fragile X mental retardation protein (FMRP), a global regulator of translation in neurons and an important factor in synaptic plasticity, produce the observed epileptic patterns. The brain-derived neurotrophic factor (BDNF) gene is a specific regulator of synaptic plasticity, and disturbances in its function cause dendrite abnormalities similar to those observed in FXS. A putative reciprocal regulation of FMRP and BDNF has been hypothesized. The Val66Met polymorphism in the BDNF gene may be involved in the alteration of normal secretion of the mature peptide and may modulate the epileptic phenotype observed in some patients with FXS. We investigated the relationship of this Met66 allele to the prevalence of epilepsy in 77 patients with FXS. No association was observed between this polymorphism and epilepsy in our group of patients. Therefore, it should not be considered a biomarker for developing epilepsy in patients with FXS.

We study the evolution of europium (Eu) and barium (Ba) abundances in Local Group dwarf spheroidal and ultrafaint dwarf galaxies by means of detailed chemical evolution models and compare our results with new sets of homogeneous abundances. The adopted models include gas infall and outflow and have been previously tested. We investigate several production scenarios for r-process elements: merging neutron stars and magnetorotational-driven supernovae. Production of Ba through the main s-process acting in low- and intermediate-mass stars is considered as well. We also test different sets of nucleosynthesis yields. For merging neutron stars we adopt either a constant and short delay time for merging or a delay time distribution function. Our simulations show that (i) if r-process elements are produced only by a quick source, it is possible to reproduce the [Eu/Fe] versus [Fe/H], but those models fail in reproducing the [Ba/Fe] versus [Fe/H]. (ii) If r-process elements are produced only with longer delays the opposite happens. (iii) If both a quick source and a delayed one are adopted, such as magnetorotational-driven supernovae and merging neutron stars with a delay time distribution, the [Eu/Fe] abundance pattern is successfully reproduced, but models still fail in reproducing the [Ba/Fe]. (iv) On the other hand, the characteristic abundances of Reticulum II can be reproduced only if both the Eu and the r-process fraction of Ba are produced on short and constant time delays during a single merging event. We discuss also other possible interpretations, including an inhomogeneous mixing of gas that might characterize this galaxy.