Human Breast Milk microRNAs, Potential Players in the Regulation of Nervous System.

Human milk is the biological fluid with the highest exosome amount and is rich in microRNAs (miRNAs). These are key regulators of gene expression networks in both normal physiologic and disease contexts, miRNAs can influence many biological processes and have also shown promise as biomarkers for disease. One of the key aspects in the regeneration of the nervous system is that there are practically no molecules that can be used as potential drugs. In the first weeks of lactation, we know that human breast milk must contain the mechanisms to transmit molecular and biological information for brain development. For this reason, our objective is to identify new modulators of the nervous system that can be used to investigate neurodevelopmental functions based on miRNAs. To do this, we collected human breast milk samples according to the time of delivery and milk states: mature milk and colostrum at term; moderate and very preterm mature milk and colostrum; and late preterm mature milk. We extracted exosomes and miRNAs and realized the miRNA functional assays and target prediction. Our results demonstrate that miRNAs are abundant in human milk and likely play significant roles in neurodevelopment and normal function. We found 132 different miRNAs were identified across all samples. Sixty-nine miRNAs had significant differential expression after paired group comparison. These miRNAs are implicated in gene regulation of dopaminergic/glutamatergic synapses and neurotransmitter secretion and are related to the biological process that regulates neuron projection morphogenesis and synaptic vesicle transport. We observed differences according to the delivery time and with less clarity according to the milk type. Our data demonstrate that miRNAs are abundant in human milk and likely play significant roles in neurodevelopment and normal function.

[Influence of donor profile on pre and post-pasteurization bacteriology of donated human milk]. / Influencia del perfil de las donantes en la bacteriología pre- y pospasteurización de la leche humana donada.

Introduction: Introduction: breastfeeding is the gold standard for infant's nutrition. Human milk bank is an essential tool to guarantee availability and safety in those situations when breastfeeding is not an option. The manipulation during the extraction of the human milk by the donors is a critical point to ensure an adequate microbiological safety. Therefore, knowing the bacterial flora that prevails in donated milk is essential to draw conclusions that can lead to taking measures in the management of the bank. Objectives: to analyze the prevalence of microorganisms in milk samples donated to the milk bank and how the bacterial flora behaves according to the profile of the donors and the donor milk circuits. Methods: the present work is a retrospective descriptive observational study that analyzes characteristics and the positive microbiological results within our milk bank samples from June 1, 2016 to December 31, 2020. All milk bank donors voluntarily signed an informed consent that authorizes the use of data to investigation. Results: during the study period, a total of 1,587 liters of raw milk were donated by 266 women, highlighting that, despite the SARS-CoV-2 virus pandemic, 2020 has been the year in which more volume of milk has been dispensed. The results show that 221 batches had at least one positive microbiological isolation (149 of them were before and 46 after pasteurization). Pre and post-pasteurization rate varies over the years with a decrease in the pre-pasteurization discard rate (3,9%) and increase in the post-pasteurization rate (5,3%). The most frequently isolated germs found in pre-pasteurization cultures were gram positive cocci and Enterobacter and a decrease in the positivity to S. aureus after establishing an eradication protocol. In post-pasteurization cultures, the most commonly found germs are Bacillus. Regarding the donor's profile, it was found a non statistically significant way that those women with the highest donation volume and/or admission of their offspring in Neonatology were associated with higher frequency of positive cultures. Conclusions: bacteriological analysis of milk samples is an essential part of quality control for a milk bank. Our results reflect a good extraction system and transportation, as well as good training from the bank's professionals. Nevertheless, improvement of processes is necessary to reduce the rate of contamination and the amount of discarded milk.

Introducción: Introducción: la leche humana es el patrón oro en la nutrición de los neonatos. Por ello, los bancos de leche se convierten en elementos esenciales para garantizar su disponibilidad y seguridad cuando la leche materna no está disponible. La manipulación que se realiza de la leche es un punto crítico para asegurar la seguridad microbiológica de las muestras. Por ello, analizar la flora de la leche donada es fundamental para tomar medidas de mejora de los bancos de leche. Objetivos: analizar los resultados microbiológicos positivos en un banco de leche humana entre las muestras de leche cruda donadas y tras su pasteurización y evaluar si existe relación entre los aislamientos, el perfil de la donante y los circuitos de leche donada. Métodos: estudio observacional descriptivo que analiza las características de la leche donada y los resultados microbiológicos positivos de muestras de leche donadas en nuestro banco desde junio de 2016 hasta diciembre de 2020. Todas las donantes firmaron un consentimiento informado. Resultados: durante el período de estudio fueron donados 1587 litros de leche cruda por 266 mujeres destacando que, a pesar de la pandemia, 2020 ha sido el año en el que más volumen se ha dispensado (280 L). Se obtuvieron 221 lotes de leche con al menos un aislamiento microbiológico positivo (14,2 % total), de ellos 149 previos y 46 posteriores a la pasteurización. La tasa de descarte pre y pospasteurización es variable a lo largo de los años con descenso en 2020 prepasteurización (3,9%) e incremento pospasteurización (5,3%). Los gérmenes más frecuentemente aislados fueron cocos grampositivos, seguidos por Enterobacterias prepasteurización detectándose un descenso en la positividad a S. aureus tras establecerse un protocolo de erradicación. En las muestras pospasteurización predomina el género Bacillus. Se ha encontrado una relación aunque no estadísticamente significativa (p > 0,05) entre mujeres con mayor volumen de donación y/o ingreso de su hijo/a en Neonatología y una mayor frecuencia cultivos positivos. Conclusiones: el análisis bacteriológico de las muestras es parte fundamental del control de calidad. Nuestros resultados traducen una buena sistemática de extracción y transporte, así como un buen entrenamiento de los profesionales del banco. No obstante, es necesario mejorar los procesos para reducir la tasa de contaminación y la cantidad de leche desechada.

Influencia del perfil de las donantes en la bacteriología pre- y pospasteurización de la leche humana donada / Influence of donor profile on pre and post-pasteurization bacteriology of donated human milk

Introducción: la leche humana es el patrón oro en la nutrición de los neonatos. Por ello, los bancos de leche se convierten en elementos esenciales para garantizar su disponibilidad y seguridad cuando la leche materna no está disponible. La manipulación que se realiza de la leche es un punto crítico para asegurar la seguridad microbiológica de las muestras. Por ello, analizar la flora de la leche donada es fundamental para tomar medidas de mejora de los bancos de leche. Objetivos: analizar los resultados microbiológicos positivos en un banco de leche humana entre las muestras de leche cruda donadas y tras su pasteurización y evaluar si existe relación entre los aislamientos, el perfil de la donante y los circuitos de leche donada. Métodos: estudio observacional descriptivo que analiza las características de la leche donada y los resultados microbiológicos positivos de muestras de leche donadas en nuestro banco desde junio de 2016 hasta diciembre de 2020. Todas las donantes firmaron un consentimiento informado. (AU)

Introduction: breastfeeding is the gold standard for infant’s nutrition. Human milk bank is an essential tool to guarantee availability and safety in those situations when breastfeeding is not an option. The manipulation during the extraction of the human milk by the donors is a critical point to ensure an adequate microbiological safety. Therefore, knowing the bacterial flora that prevails in donated milk is essential to draw conclusions that can lead to taking measures in the management of the bank. Objectives: to analyze the prevalence of microorganisms in milk samples donated to the milk bank and how the bacterial flora behaves according to the profile of the donors and the donor milk circuits. Methods: the present work is a retrospective descriptive observational study that analyzes characteristics and the positive microbiological results within our milk bank samples from June 1, 2016 to December 31, 2020. All milk bank donors voluntarily signed an informed consent that authorizes the use of data to investigation. (AU)

Lung Ultrasound Scores Progress Differently in Extreme and Very Preterm Infants after Birth: A Multicentre Prospective Study.

INTRODUCTION: The lung ultrasound score (LUS) has been suggested to predict moderate-severe bronchopulmonary dysplasia (msBPD) in preterm infants. We aimed to assess LUS evolution after birth in preterm infants and the effect of gestational age. METHODS: This multicentre prospective observational study was performed with newborns born before 33 weeks of gestation. We created two groups: group 1 (23-27 weeks) and group 2 (28-32 weeks). We compared LUSs between the groups from birth until 36 weeks of postmenstrual age, and we estimated the LUS evolution in each group with a linear multilevel mixed-effects regression model. The effects of the need for surfactant or an msBPD diagnosis were also studied. RESULTS: We included 339 patients: 122 (36%) in group 1 and 217 (64%) in group 2. The infants in group 1 showed a steady progression in the LUS from birth until 4 weeks of age and a subsequent decrease; the infants in group 2 showed a progressive decrease in the LUS throughout the study. This progression varied significantly in the first weeks of life in infants who required surfactant at birth and after the first week of life in the patients diagnosed with msBPD. DISCUSSION/CONCLUSIONS: Extremely preterm infants showed persistently high LUSs during the first weeks of life, regardless of the progression to msBPD. In this group, the infants who did not require surfactant at birth exhibited an increase in their LUSs after the first week until their values were equal to the remaining infants in their group.

Utilidad de las prostaglandinas intravenosas en el espasmo vascular neonatal / Use of intravenous prostaglandins in neonatal vasospasm

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Effects of Hypothermia and Allopurinol on Oxidative Status in a Rat Model of Hypoxic Ischemic Encephalopathy.

Hypoxic ischemic encephalopathy (HIE) is one of the main causes of morbidity and mortality during the neonatal period, despite treatment with hypothermia. There is evidence that oxidative damage plays an important role in the pathophysiology of hypoxic-ischemic (HI) brain injury. Our aim was to investigate whether postnatal allopurinol administration in combination with hypothermia would reduce oxidative stress (OS) biomarkers in an animal model of HIE. Postnatal 10-day rat pups underwent unilateral HI of moderate severity. Pups were randomized into: Sham operated, hypoxic-ischemic (HI), HI + allopurinol (HIA), HI + hypothermia (HIH), and HI + hypothermia + allopurinol (HIHA). Biomarkers of OS and antioxidants were evaluated: GSH/GSSG ratio and carbonyl groups were tested in plasma. Total antioxidant capacity (TAC) was analyzed in plasma and cerebrospinal fluid, and 8-iso-prostaglandin F2α was measured in brain tissue. Plasma 2,2'-azinobis-(3-ethyl-benzothiazoline-6-sulfonic acid) (ABTS) levels were preserved in those groups that received allopurinol and dual therapy. In cerebrospinal fluid, only the HIA group presented normal ferric reducing ability of plasma (FRAP) levels. Protein oxidation and lipid peroxidation were significantly reduced in all groups treated with hypothermia and allopurinol, thus enhancing neuroprotection in HIE.

The Practicality of Feeding Defatted Human Milk in the Treatment of Congenital Chylothorax.

Introduction: Congenital chylothorax (CC) is a rare and life-threating condition. Since its treatment is founded on the elimination of long-chain fatty acids from the diet, breastfeeding has been traditionally contraindicated. However, breast milk could be very beneficial due to its immunological and nutritional benefits. Only limited research has been published about the usage of modified-fat breast milk (MBM) in chylothorax treatment. Methods and Results: Systematic review methods were used by two independent reviewers. Only a few case report studies (quality assessment on the domains of the GRADE approach), two small controlled studies, a retrospective study, and some test-tube-based laboratory research met the inclusion criteria. Despite this, we have observed a widespread clinical adoption of this novel treatment in health institutions. Data suggest that modified-fat breast milk does facilitate the resolution of chylothoraces. Refrigerated centrifuge (2°C, 3,000 rpm for 15 minutes) and syringe fat removal methods were the most efficient options in terms of fat reduction. Conclusions: Feeding of human milk is advisable in CC and feasible by means of a simple milk defatting procedure. Open questions remain, related to length and degree of fat restriction and need for individualized fortification of defatted breast milk.

Neuroprotection with hypothermia and allopurinol in an animal model of hypoxic-ischemic injury: Is it a gender question?

BACKGROUND: Hypoxic-ischemic encephalopathy (HIE) is one of the most important causes of neonatal brain injury. Therapeutic hypothermia (TH) is the standard treatment for term newborns after perinatal hypoxic ischemic injury (HI). Despite this, TH does not provide complete neuroprotection. Allopurinol seems to be a good neuroprotector in several animal studies, but it has never been tested in combination with hypothermia. Clinical findings show that male infants with (HI) fare more poorly than matched females in cognitive outcomes. However, there are few studies about neuroprotection taking gender into account in the results. The aim of the present study was to evaluate the potential additive neuroprotective effect of allopurinol when administrated in association with TH in a rodent model of moderate HI. Gender differences in neuroprotection were also evaluated. METHODS: P10 male and female rat pups were subjected to HI (Vannucci model) and randomized into five groups: sham intervention (Control), no treatment (HI), hypothermia (HIH), allopurinol (HIA), and dual therapy (hypothermia and allopurinol) (HIHA). To evaluate a treatment's neuroprotective efficiency, 24 hours after the HI event caspase3 activation was measured. Damaged area and hippocampal volume were also measured 72 hours after the HI event. Negative geotaxis test was performed to evaluate early neurobehavioral reflexes. Learning and spatial memory were assessed via Morris Water Maze (MWM) test at 25 days of life. RESULTS: Damaged area and hippocampal volume were different among treatment groups (p = 0.001). The largest tissue lesion was observed in the HI group, followed by HIA. There were no differences between control, HIH, and HIHA. When learning process was analyzed, no differences were found. Females from the HIA group had similar results to the HIH and HIHA groups. Cleaved caspase 3 expression was increased in both HI and HIA. Despite this, in females cleaved caspase-3 was only differently increased in the HI group. All treated animals present an improvement in short-term (Negative geotaxis) and long-term (WMT) functional tests. Despite this, treated females present better long-term outcome. In short-term outcome no sex differences were observed. CONCLUSIONS: Our results suggest that dual therapy confers great neuroprotection after an HI event. There were functional, histological, and molecular improvements in all treated groups. These differences were more important in females than in males. No statistically significant differences were found between HIHA and HIH; both of them present a great improvement. Our results support the idea of different regulation mechanisms and pathways of cell death, depending on gender.

Influence of catch up growth on spatial learning and memory in a mouse model of intrauterine growth restriction.

BACKGROUND: Intrauterine growth restriction (IUGR) and rapid postnatal weight gain or catch up growth (CUG) increase the susceptibility to metabolic syndrome during adult life. Longitudinal studies have also revealed a high incidence of learning difficulties in children with IUGR. The aim of the present study was to investigate the effect of nutrition and CUG on learning memory in an IUGR animal model. We hypothesized that synaptic protein expression and transcription, an essential mechanism for memory consolidation, might be affected by intrauterine undernutrition. METHODS: IUGR was induced by 50% maternal caloric undernutrition throughout late gestation. During the suckling period, dams were either fed ad libitum or food restricted. The pups were divided into: Normal prenatal diet-Normal postnatal diet (NN), Restricted prenatal diet- Normal postnatal diet + catch up growth (RN+), Normal prenatal diet-Restricted postnatal diet (NR) and Restricted prenatal diet-Restricted postnatal diet (RR). At 4 weeks of age, memory was assessed via a water maze test. To evaluate synaptic function, 2 specific synaptic proteins (postsynaptic density-95 [PSD95], synaptophysin) as well as insulin receptors (IR) were tested by Western Blot and quantitative polymerase chain reaction (qPCR). Brain-derived neurotrophic factor and serum insulin levels were also studied. RESULTS AND CONCLUSIONS: The RN+ group presented a learning curve similar to the NN animals. The RR animals without CUG showed learning disabilities. PSD95 was lower in the RR group than in the NN and RN+ mice. In contrast, synaptophysin was similar in all groups. IR showed an inverse expression pattern to that of the PSD95. In conclusion, perinatal nutrition plays an important role in learning. CUG after a period of prenatal malnutrition seems to improve learning skills. The functional alterations observed might be related to lower PSD95 activity and a possible dysfunction in the hormone regulation of synaptic plasticity.

Learning and memory disabilities in IUGR babies: Functional and molecular analysis in a rat model.

INTRODUCTION: 1Intrauterine growth restriction (IUGR) is the failure of the fetus to achieve its inherent growth potential, and it has frequently been associated with neurodevelopmental problems in childhood. Neurological disorders are mostly associated with IUGR babies with an abnormally high cephalization index (CI) and a brain sparing effect. However, a similar correlation has never been demonstrated in an animal model. The aim of this study was to determine the correlations between CI, functional deficits in learning and memory and alterations in synaptic proteins in a rat model of IUGR. METHODS: 2Utero-placental insufficiency was induced by meso-ovarian vessel cauterization (CMO) in pregnant rats at embryonic day 17 (E17). Learning performance in an aquatic learning test was evaluated 25 days after birth and during 10 days. Some synaptic proteins were analyzed (PSD95, Synaptophysin) by Western blot and immunohistochemistry. RESULTS: 3Placental insufficiency in CMO pups was associated with spatial memory deficits, which are correlated with a CI above the normal range. CMO pups presented altered levels of synaptic proteins PSD95 and synaptophysin in the hippocampus. CONCLUSIONS: 4The results of this study suggest that learning disabilities may be associated with altered development of excitatory neurotransmission and synaptic plasticity. Although interspecific differences in fetal response to placental insufficiency should be taken into account, the translation of these data to humans suggest that both IUGR babies and babies with a normal birth weight but with intrauterine Doppler alterations and abnormal CI should be closely followed to detect neurodevelopmental alterations during the postnatal period.

Terapia prenatal con sulfato de magnesio: evolución clínica de los recién nacidos pretérmino menores de 29 semanas y correlación con la magnesemia neonatal / Prenatal treatment with magnesium sulphate: initial clinical outcomes in pre-term infants less than 29 weeks and correlation with neonatal magnesium levels

INTRODUCCIÓN: La administración prenatal de MgSO4ha mostrado su eficacia en reducir la parálisis cerebral y la disfunción motora severa a los 2 años de edad. El objetivo de este trabajo es estudiar la evolución clínica inicial de los neonatos menores de 29 semanas, que han recibido prenatalmente MgS04 con indicación neuroprotectora y dilucidar la asociación entre la dosis de magnesio administrada a la madre y las concentraciones de magnesio en suero neonatal. MATERIAL Y MÉTODOS: Estudio prospectivo de cohortes en el que se incluyó a los neonatos menores de 29 semanas ingresados en la Unidad de Cuidados Intensivos Neonatales del Hospital Universitario de Vigo desde diciembre del 2012 hasta julio del 2015. Análisis comparativo de resultados perinatales, de morbimortalidad neonatal y magnesemia entre el grupo expuesto prenatalmente al sulfato de magnesio y un grupo control. RESULTADOS: Se incluyó a un total de 42 recién nacidos, en 28 de los cuales sus madres habían recibido MgSO4. Se encontró significación estadística en la variable mortalidad. No hubo diferencias significativas en el resto de las variables estudiadas. Se obtuvo una correlación significativa entre la dosis total de MgSO4 recibida por la madre y los niveles de magnesio del recién nacido en las primeras 24 h de vida (r2 0,436; p < 0,001). CONCLUSIONES: Se ha obtenido una menor mortalidad en el grupo expuesto a MgS04. No se han encontrado efectos secundarios significativos derivados de la administración de MgS04. La dosis de MgSO4 recibida por las madres tiene una relación lineal con los niveles de magnesio obtenidos en los recién nacidos

INTRODUCTION: Antenatal magnesium sulphate (MgSO4) administration has shown to be effective in minimising cerebral palsy and severe motor dysfunction at the age of 2 years. The aim of this study is to analyse the initial clinical outcome of preterm neonates less than 29 weeks who have received prenatal MgSO4, as well as to determine the relationship between the magnesium dose delivered to the mother and the magnesium concentration in the neonates. MATERIAL AND METHODS: A prospective cohort study was conducted on neonates of less than 29 weeks gestation admitted to the Neonatal Intensive Care Unit (NICU) of Hospital Universitario de Vigo from December 2012 to July 2015. Comparative analysis was performed on the perinatal outcomes, neonatal morbidity, mortality, and magnesium levels between the groups of neonates exposed to magnesium sulphate and the control group. RESULTS: A total of 42 neonates were included in the study. The mothers of 28 of them had received MgSO4 as a neuroprotective agent. Statistical significance was obtained in the mortality variable. There were no significant differences in the rest of studied variables. There was a significant correlation between the full dose of MgSO4 received by the mother and the levels of magnesium in the neonate in the first 24hours of life (r2 0.436; P<.001). CONCLUSIONS: A lower mortality was observed in the group that had been exposed to MgS04. No significant side effects were found as a result of administering of MgS04. The MgS04 dose received by mother has a linear relationship with the magnesium levels obtained in neonates

[Prenatal treatment with magnesium sulphate: Initial clinical outcomes in pre-term infants less than 29 weeks and correlation with neonatal magnesium levels]. / Terapia prenatal con sulfato de magnesio: evolución clínica de los recién nacidos pretérmino menores de 29 semanas y correlación con la magnesemia neonatal.

INTRODUCTION: Antenatal magnesium sulphate (MgSO4) administration has shown to be effective in minimising cerebral palsy and severe motor dysfunction at the age of 2 years. The aim of this study is to analyse the initial clinical outcome of preterm neonates less than 29 weeks who have received prenatal MgSO4, as well as to determine the relationship between the magnesium dose delivered to the mother and the magnesium concentration in the neonates. MATERIAL AND METHODS: A prospective cohort study was conducted on neonates of less than 29 weeks gestation admitted to the Neonatal Intensive Care Unit (NICU) of Hospital Universitario de Vigo from December 2012 to July 2015. Comparative analysis was performed on the perinatal outcomes, neonatal morbidity, mortality, and magnesium levels between the groups of neonates exposed to magnesium sulphate and the control group. RESULTS: A total of 42 neonates were included in the study. The mothers of 28 of them had received MgSO4 as a neuroprotective agent. Statistical significance was obtained in the mortality variable. There were no significant differences in the rest of studied variables. There was a significant correlation between the full dose of MgSO4 received by the mother and the levels of magnesium in the neonate in the first 24hours of life (r2 0.436; P<.001). CONCLUSIONS: A lower mortality was observed in the group that had been exposed to MgS04. No significant side effects were found as a result of administering of MgS04. The MgS04 dose received by mother has a linear relationship with the magnesium levels obtained in neonates.