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Embryonic development is a complex and dynamic process that unfolds over time and involves the production and diversification of increasing numbers of cells. The impact of developmental time on the formation of the central nervous system is well documented, with evidence showing that time plays a crucial role in establishing the identity of neuronal subtypes. However, the study of how time translates into genetic instructions driving cell fate is limited by the scarcity of suitable experimental tools. We introduce BirthSeq, a new method for isolating and analyzing cells based on their birth date. This innovative technique allows for in vivo labeling of cells, isolation via fluorescence-activated cell sorting, and analysis using high-throughput techniques. We calibrated the BirthSeq method for developmental organs across three vertebrate species (mouse, chick and gecko), and utilized it for single-cell RNA sequencing and novel spatially resolved transcriptomic approaches in mouse and chick, respectively. Overall, BirthSeq provides a versatile tool for studying virtually any tissue in different vertebrate organisms, aiding developmental biology research by targeting cells and their temporal cues.
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Análise de Célula Única , Animais , Camundongos , Análise de Célula Única/métodos , Embrião de Galinha , Lagartos/genética , Lagartos/embriologia , Desenvolvimento Embrionário/genética , Transcriptoma/genética , Citometria de Fluxo/métodos , Vertebrados/genética , Separação Celular/métodos , Galinhas , Análise de Sequência de RNA/métodosRESUMO
Sound classification plays a crucial role in enhancing the interpretation, analysis, and use of acoustic data, leading to a wide range of practical applications, of which environmental sound analysis is one of the most important. In this paper, we explore the representation of audio data as graphs in the context of sound classification. We propose a methodology that leverages pre-trained audio models to extract deep features from audio files, which are then employed as node information to build graphs. Subsequently, we train various graph neural networks (GNNs), specifically graph convolutional networks (GCNs), GraphSAGE, and graph attention networks (GATs), to solve multi-class audio classification problems. Our findings underscore the effectiveness of employing graphs to represent audio data. Moreover, they highlight the competitive performance of GNNs in sound classification endeavors, with the GAT model emerging as the top performer, achieving a mean accuracy of 83% in classifying environmental sounds and 91% in identifying the land cover of a site based on its audio recording. In conclusion, this study provides novel insights into the potential of graph representation learning techniques for analyzing audio data.
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Primary distal renal tubular acidosis (dRTA) is a rare tubulopathy characterised by the presence of hyperchloremic metabolic acidosis. It is caused by the existence of a defect in the function of the H+ -ATPase located on the luminal side of the α-intercalated cells or the Cl - HCO3- (AE1) anion exchanger located on the basolateral side. Patients do not acidify the urine after acid overload (NH4Cl) or after stimulating H+ secretion by obtaining a high intratubular concentration of an anion such as chlorine (pH is measured) or HCO3- (urinary pCO2 is measured). We present a family with autosomal dominant dRTA produced by a heterozygous mutation in the SLC4A1 gene in which the two paediatric members showed a test of normal maximum urinary pCO2. Our hypothesis is that since the H + -ATPase is intact, at least initially, the stimulation induced by intratubular electronegativity to secrete H + could be effective, which would allow the maximum urinary pCO2 to be paradoxically normal, which could explain the onset, moderate presentation of symptoms and late diagnosis in patients with this mutation. This is the first documented case of a dominant dRTA in Mexico.
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Acidose Tubular Renal , Humanos , Criança , Acidose Tubular Renal/diagnóstico , Acidose Tubular Renal/genética , Proteína 1 de Troca de Ânion do Eritrócito/genética , Mutação , Ânions/metabolismo , Adenosina Trifosfatases/genética , Adenosina Trifosfatases/metabolismoRESUMO
Polyhedrins are viral proteins present in a large family of baculoviruses that form occlusion bodies (polyhedra). These structures protect the virus particles from the outside environment until they are ingested by susceptible insects. Occluded viruses can sustain inclement weather for long periods of time. Therefore, the polyhedra is a natural preservative that keeps the viral structure intact at ambient temperature for years. In a previous study we identified the first 110 amino acids from polyhedrin (PH(1-110)) as a good candidate to carry antigens of interest. As a proof of concept, we produced a fusion protein with PH(1-110) and the green fluorescent protein (PH(1-110)GFP). The fusion protein associates spontaneously during its synthesis resulting in the formation of nanoparticles. Nasal immunization with these nanoparticles and in the absence of any adjuvant, results in a robust immune response with the production of IgG immunoglobulins that remained elevated for months and that selectively recognize the GFP but not PH(1-110). These results indicate that PH(1-110) is poorly immunogenic but capable of enhancing the immune response to GFP.
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Nanopartículas , Vacinas , Temperatura , Antígenos , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismoRESUMO
Objetivo: Identificar los factores asociados a la intoxicación intencionada por plaguicidas en Colombia durante 2007-2017. Metodología: Se llevó a cabo un estudio analítico transversal en la población de intoxicados con intención suicida por plaguicidas de Colombia entre 2007 a 2017. Se calcularon prevalencias ajustadas por edad, expresadas por 100 000 personas contadas por sexo. Se realizó un modelo de regresión bivariado, en el cual la condición final se constituyó en la variable de interés (variable y; 0=vivo 1=muerto) y las variables explicativas fueron (variables x): sociodemográficas (sexo, edad, escolaridad y procedencia) y específicas del evento (vía de exposición). Resultados: el comportamiento de los Odds Ratio evidencia que, los hombres tienen un riesgo de fallecer de 1.39 (95 % IC: 1.238-1.567) más veces que las mujeres. Con respecto al régimen de afiliación en salud, las personas no aseguradas tienen un riesgo de fallecer por suicidio de 1.33 más veces con respecto a los que sobreviven (95 % IC: 1.131-1.555). Para la mortalidad por suicidio se identifican los siguientes factores asociados a la supervivencia (intentos de suicidio no consumados): en primer lugar, el área de residencia urbana y, en segundo lugar, los grupos de edad, adolescente y la adultez temprana. Conclusiones: Al analizar la mortalidad por suicidio asociada a la intoxicación por plaguicidas los hombres son identificados y caracterizados como el grupo de mayor riesgo, siguiendo tendencias generales (todos los métodos) similares a estudios realizados a nivel mundial.
Objective: To identify the social factors associated with intentional poisoning by pesticides in Colombia, during the 2007-2017 period. Methodology: A cross-sectional analytical study was carried out in the population of those intoxicated by pesticides with suicidal intent in Colombia between 2007 and 2017. Age-adjusted prevalences were calculated, expressed per 100,000 people, counted by sex. A bi-variate regression model was carried out, in which the final condition was constituted in the variable of interest (variable y; 0 = alive 1 = dead), and the explanatory variables were (variables x): sociodemographic (sex, age, education and origin) and specific to the event (route of exposure). Results: the behavior of the Odds Ratio shows that men have a risk of dying 1,393 (95% CI: 1,238-1,567) more times than women. Regarding the health affiliation scheme, uninsured people have a risk of dying by suicide 1.33 times more than those who survive (95% CI: 1.131-1.555). For mortality due to suicide, the following factors associated with survival (unsuccessful suicide attempts) are identified: firstly, the urban residence area and secondly, age groups: adolescents and early adulthood. Conclusions: When analyzing suicide mortality associated with pesticide poisoning, men are identified and characterized as the highest risk group, following general trends (all methods) similar to studies carried out worldwide.
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Hypothalamic arginine vasopressin (AVP)-containing magnocellular neurosecretory neurons (AVPMNN) emit collaterals to synaptically innervate limbic regions influencing learning, motivational behaviour, and fear responses. Here, we characterize the dynamics of expression changes of two key determinants for synaptic strength, the postsynaptic density (PSD) proteins AMPAR subunit GluA1 and PSD scaffolding protein 95 (PSD95), in response to in vivo manipulations of AVPMNN neuronal activation state, or exposure to exogenous AVP ex vivo. Both long-term water deprivation in vivo, which powerfully upregulates AVPMNN metabolic activity, and exogenous AVP application ex vivo, in brain slices, significantly increased GluA1 and PSD95 expression as measured by western blotting, in brain regions reportedly receiving direct ascending innervations from AVPMNN (i.e., ventral hippocampus, amygdala and lateral habenula). By contrast, the visual cortex, a region not observed to receive AVPMNN projections, showed no such changes. Ex vivo application of V1a and V1b antagonists to ventral hippocampal slices ablated the AVP stimulated increase in postsynaptic protein expression measured by western blotting. Using a modified expansion microscopy technique, we were able to quantitatively assess the significant augmentation of PSD95 and GLUA1 densities in subcellular compartments in locus coeruleus tyrosine hydroxylase immunopositive fibres, adjacent to AVP axon terminals. Our data strongly suggest that the AVPMNN ascending system plays a role in the regulation of the excitability of targeted neuronal circuits through upregulation of key postsynaptic density proteins corresponding to excitatory synapses.
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Sinapses , Tirosina 3-Mono-Oxigenase , Arginina Vasopressina/metabolismo , Hipocampo/metabolismo , Hipotálamo/metabolismo , Sinapses/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismoRESUMO
Resumen Objetivo: analizar el comportamiento epidemiológico del intento de suicidio, examinando las diferencias existentes entre el sexo, variables sociodemográficas, psicosociales y específicas durante 2012-2017. Materiales y métodos: un estudio analítico transversal; se incluyeron 524 casos reportados al Sistema Nacional de Vigilancia en Salud Pública (SIVIGILA) entre 2012 y 2017 en el municipio de Sogamoso. La variable de interés se fundó en el género, los datos fueron procesados y se construyó un modelo de regresión logística binaria con un nivel de confianza de 95% y una precisión del 5%. Resultados: con el modelo de regresión logística bivariada se encontró que la adolescencia y adultez temprana (p<0,05) se asocian significativamente con el género mujer, y con respecto al posible desencadenante, el conflicto de pareja (p 0,05; OR= 2,32). Lo anterior evidencia una interacción con la variable violencia (P<0,05). Por otro lado, el consumo de alcohol se constituyó en una variable asociada significativamente con el género hombre (p<0,05) y un OR= 3,6 con respecto a las mujeres. Conclusiones: se evidencia que las prevalencias ajustadas por edad no muestran una reducción significativa desde el inicio hasta el final del periodo, siendo evidente la concentración en edades tempranas del desarrollo. Los resultados observados en mujeres y hombres revelan una necesidad de capacitación de agentes comunitarios o guardianes (gatekeepers) y el fortalecimiento de centros de escucha y demás estrategias de intervención comunitaria.
Abstract Objective: To analyze the epidemiological behavior of the suicide attempt, examining the differences between gender, sociodemographic, psychosocial and specific variables during 2012-2017. Materials and methods: A cross-sectional analytical study including 524 cases reported to the National Public Health Surveillance System (SIVIGILA for its acronym in Spanish) between 2012 and 2017 in the municipality of Sogamoso. The variable of interest was based on gender, the data were processed, and a binary logistic regression model was built with a confidence level of 95% and an accuracy of 5%. Results: With the bivariate logistic regression model it was found that adolescence and early adulthood (p<0.05) are significantly associated with the female gender, and partner conflict (p<0.05; OR= 2.32); regarding the possible trigger thus evidencing an interaction with the violence variable (P<0.05). On the other hand, alcohol consumption became a variable significantly associated with the male gender (p<0.05) and an OR= 3.6 for females. Conclusions: it is evidenced that the age-adjusted impacts do not show a significant reduction from the beginning to the end of the period, being evident the concentration in early developmental ages. The results observed in women and men reveal a need for training community agents or guardians (gatekeepers) and the strengthening of listening centers and other community intervention strategies.
Resumo Objetivo: analisar o comportamento epidemiológico do intento de suicídio, examinando as diferenças existentes entre o sexo, variáveis sócio demográficas, psicossocial e específicas durante 2012-2017. Materiais e métodos: um estudo analítico transversal; se incluíram 524 casos reportados ao Sistema Nacional de Vigilância em Saúde Pública (SIVIGILA) entre 2012 e 2017 no município de Sogamoso. A variável de interesse se fundou no gênero, os dados foram processados e se construiu um modelo de regressão logística binaria com um nível de confiança de 95% e uma precisão o 5%. Resultados: com o modelo de regressão logística bivariada se encontrou que a adolescência e adultos precoces (p<0,05) se associam significativamente com o gênero mulher, e com respeito ao possível desencadeante, o conflito de casais (p<0,05; OR= 2,32). O anterior evidencia uma interação com a variável violência (P<0,05). Por outro lado, o consumo de álcool se constituiu em uma variável associada significativamente com o gênero home (p<0,05) e um OR= 3,6 com respeito às mulheres. Conclusões: Evidenciase que as prevalências ajustadas por idade não amostram uma redução significativa desde o início até o final do período, sendo evidente a concentração em idades precoces do desenvolvimento. Os resultados observados em mulheres e homens revelam uma necessidade de capacitação de agentes comunitários o guardiães (gatekeepers) e o fortalecimento de centros de escuta e também estratégias de intervenção comunitária.
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Childhood and adolescence represent critical periods where beverage and food consumption behaviors are learned and developed. Mexican mothers' presence and influence are instrumental in shaping such behaviors. The aim of this study was to estimate the prevalence and risk associations of maternal factors for unhealthy patterns of beverage intake. This study analyzed data from a population-based cross-sectional study of healthy children and adolescents from Mexico City. Data of subject's total water intake (TWI) and its' sources were collected using two 24-h recall surveys. Patterns of beverage intake were constructed based on the guidance system of beverage consumption in the US. Maternal factors of interest included age, body mass index (BMI), mother's educational level (MEL), socioeconomic status (SES), and belongingness to the paid workforce (BPW). Data of 1532 subject-mother dyads informed that 47% of subjects did not meet the Institute of Medicine (IOM) recommendations for TWI, and 94.6% showed an unhealthy beverage intake pattern, mainly consisting in a lower intake of water and a higher intake of caloric beverages with some nutrients; and calorically sweetened beverages. The major sources of hydration were caloric beverages with some nutrients (i.e., whole milk, fruit water, and flavored milk). The highest risk association for an unhealthy beverage intake pattern was seen in those subjects with mothers in the cluster with lower SES, lower MEL, lower proportion of BPW, higher BMI, and younger age (OR = 9.3, 95% CI 1.2-72.8, P = 0.03). Thus, there is a remarkably high prevalence of an unhealthy pattern of beverage intake, and specific maternal factors may be implicated as enablers of such behaviors, which is also addressable for future interventions.
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OBJECTIVE: In Spain, the number of persons that are in a surgery waiting list as well as the available surgery resources, differ across autonomous communities. The pandemic generated by COVID-19 has increased these waiting lists. In this study two objectives were pursued: on the one hand, to determine which are the resources that are determining the number of persons that are in a surgery waiting list per 1,000 inhabitants; on the other hand, to estimate the impact that the current pandemic has on the latter. METHODS: To estimate which are the resources that are having a greater impact on the waiting lists and to forecast the effect that the COVID-19 has on them, we use dynamic panel data models. The data on the surgery resources and on the waiting lists by autonomous communities is obtained from the Surveys on Health, Hospital Statistics and reports on waiting lists of the Ministry of Health, Consumption and Social Well Being and the Counsels. The sample period is 2012-2017 (last published year for surgery resources). In addition, a literature review is conducted and it shows the important and complexity of waiting list like a gestion tool of health system (Science, SciELO and Dialnet web data bases). RESULTS: COVID-19 will increase the waiting lists by approximately 7.6% to 19.14%, depending on the autonomous community. Not all the available surgery resources have the same relevance nor an equal effect on the reduction of the waiting lists. The most significant resources are the beds and operating rooms per 1,000 inhabitants. The hospital expenditure is not so relevant. CONCLUSIONS: The panel data models estimate the relation between the surgery resources and the waiting list. The latter is deemed complex and different across autonomous communities. In addition, these models allow to predict the expected increase in the waiting lists and are, thus, a useful instrument for their management.
OBJETIVO: En España, tanto el número de personas que están en espera de intervención quirúrgica, como los recursos quirúrgicos disponibles difieren entre comunidades autónomas. La pandemia ocasionada por la COVID-19 ha incrementado estas listas de espera. En este trabajo se persiguió un doble objetivo, por un lado, determinar cuáles son los recursos que tiene mayor influencia en el número de personas por mil habitantes que se encuentran en espera quirúrgica y, por otro, estimar el impacto que la pandemia actual ha generado. METODOS: Para estimar cuales son los recursos que más están repercutiendo en las listas de espera y predecir el efecto que la COVID-19 ejerce en ellas, utilizamos un modelo de datos de panel dinámico. Los datos de recursos quirúrgicos y listas de espera por comunidades autónomas se han obtenido de Encuestas de Salud, Estadísticas Hospitalarias e informes de listas de espera del Ministerio de Sanidad Consumo y Bienestar Social y de las Consejerías. El periodo muestral es 2012-2017 (último año publicado para recursos quirúrgicos). Además, se llevó a cabo una revisión de la literatura que permite corroborar la importancia y complejidad de las listas de espera como instrumento de gestión del sistema sanitario (bases de datos Web of Science, SciELO y Dialnet). RESULTADOS: La COVID-19 incrementará las listas de espera, aproximadamente entre el 7,6% y el 19,4%, dependiendo de la comunidad. No todos los recursos quirúrgicos disponibles tienen la misma importancia ni influyen de la misma manera en la reducción de las personas en espera. Los recursos más significativos son las camas y quirófanos por mil habitantes y no tanto el gasto en hospitales del Sistema Nacional de Salud. CONCLUSIONES: Los modelos de datos de panel permiten conocer la relación entre recursos quirúrgicos y lista de espera, la cual parece ser compleja y diferente entre comunidades. Además, estos modelos ayudan a predecir el incremento esperable y, por lo tanto, son un instrumento útil en la gestión de listas de espera.
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COVID-19/epidemiologia , Cirurgia Geral/estatística & dados numéricos , Listas de Espera , Coleta de Dados , Número de Leitos em Hospital , Humanos , Internet , Salas Cirúrgicas , EspanhaRESUMO
G protein-activated inward-rectifying potassium (K+ ) channels (Kir3/GIRK) participate in cell excitability. The GIRK5 channel is present in Xenopus laevis oocytes. In an attempt to investigate the physiological role of GIRK5, we identified a noncanonical di-arginine endoplasmic reticulum (ER) retention motif (KRXY). This retention motif is located at the N-terminal region of GIRK5, coded by two small exons found only in X. laevis and X. tropicalis. These novel exons are expressed through use of an alternative transcription start site. Mutations in the sequence KRXY produced functional channels and induced progesterone-independent oocyte meiotic progression. The chimeric proteins enhanced green fluorescent protein (EGFP)-GIRK5-WT and the EGFP-GIRK5K13AR14A double mutant, were localized to the ER and the plasma membrane of the vegetal pole of the oocyte, respectively. Silencing of GIRK5 or blocking of this channel by external barium prevented progesterone-induced meiotic progression. The endogenous level of GIRK5 protein decreased through oocyte stages in prophase I augmenting by progesterone. In conclusion, we have identified a unique mechanism by which the expression pattern of a K+ channel evolved to control Xenopus oocyte maturation.
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Motivos de Aminoácidos , Sequência de Aminoácidos , Retículo Endoplasmático/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização Acoplados a Proteínas G/química , Canais de Potássio Corretores do Fluxo de Internalização Acoplados a Proteínas G/metabolismo , Oócitos/metabolismo , Domínios e Motivos de Interação entre Proteínas , Proteínas de Xenopus/química , Proteínas de Xenopus/metabolismo , Animais , Sequência Conservada , Canais de Potássio Corretores do Fluxo de Internalização Acoplados a Proteínas G/genética , Humanos , Oócitos/efeitos dos fármacos , Filogenia , Ligação Proteica , Proteínas de Xenopus/genética , Xenopus laevisRESUMO
The kidney controls body fluids, electrolyte and acid-base balance. Previously, we demonstrated that hyperpolarization-activated and cyclic nucleotide-gated (HCN) cation channels participate in ammonium excretion in the rat kidney. Since acid-base balance is closely linked to potassium metabolism, in the present work we aim to determine the effect of chronic metabolic acidosis (CMA) and hyperkalemia (HK) on protein abundance and localization of HCN3 in the rat kidney. CMA increased HCN3 protein level only in the outer medulla (2.74 ± 0.31) according to immunoblot analysis. However, immunofluorescence assays showed that HCN3 augmented in cortical proximal tubules (1.45 ± 0.11) and medullary thick ascending limb of Henle's loop (4.48 ± 0.45) from the inner stripe of outer medulla. HCN3 was detected in brush border membranes (BBM) and mitochondria of the proximal tubule by immunogold electron and confocal microscopy in control conditions. Acidosis did not alter HCN3 levels in BBM and mitochondria but augmented them in lysosomes. HCN3 was also immuno-detected in mitoautophagosomes. In the distal nephron, HCN3 was expressed in principal and intercalated cells from cortical to medullary collecting ducts. CMA did not change HCN3 abundance in these nephron segments. In contrast, HK doubled HCN3 level in cortical collecting ducts and favored its basolateral localization in principal cells from the inner medullary collecting ducts. These findings further support HCN channels contribution to renal acid-base and potassium balance.
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Acidose/etiologia , Acidose/metabolismo , Hiperpotassemia/etiologia , Hiperpotassemia/metabolismo , Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização/metabolismo , Néfrons/metabolismo , Canais de Potássio/metabolismo , Animais , Biomarcadores , Doença Crônica , Células Epiteliais/metabolismo , Imunofluorescência/métodos , Expressão Gênica , Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização/genética , Túbulos Renais Proximais/metabolismo , Alça do Néfron/metabolismo , Néfrons/ultraestrutura , Canais de Potássio/genética , RatosRESUMO
Bilingualism was implemented in the Community of Madrid (Spain) more than ten years ago, through the incorporation of the English language in the teaching methods of certain schools. Since that time, various research projects have been carried out, with the objective of comparing the academic performance of students in bilingual schools with those in non-bilingual schools. The present paper makes use of primary education data from the Department of Education and Research for the Community of Madrid in an effort to analyze whether or not bilingualism results in the relative improvement of educational outcomes achieved in primary schools in the Region of Madrid, in Spain. More specifically, the data used is from sixth grade classrooms, given that, generally speaking, in this grade all schools give a standardized test which measures academic performance in Math, Science and Technology, Spanish Language Arts and English Language Arts. Our assessment makes use of a multinomial logit model, and includes the most common variables found in the research on the determination of educational outcomes (variables related to whether or not schools are bilingual, which is the main focus of this paper), as well as other less common variables considered to be relevant. These include absenteeism, satisfaction levels among families and students, and the percentage of students in second chance programs. The results show that bilingualism does not lower children performance in the subjects taught in English or in the subjects taught in Spanish. Academic performance in Mathematics, Science and Technology, and Spanish Language Arts is similar with respect to those schools which are not bilingual. However, results in English are significantly higher in bilingual schools when compared to non-bilingual schools.
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Desempenho Acadêmico/normas , Sucesso Acadêmico , Multilinguismo , Criança , Humanos , Estudos de Linguagem/normas , Instituições Acadêmicas/normas , Espanha , EstudantesRESUMO
Overweight (OW) and obesity (OB) during childhood/adolescence are major public health problems in Mexico. Several obesogenic lifestyle (OL) risk factors have been identified, but the burden and consequences of them in Mexican children/adolescents remain unclear. The objective of this study was to estimate the prevalence of OL components and describe their relationships with adiposity, and OW/OB. A population-based cross-sectional study of Mexican children/adolescents with nutritional assessment, data collection on daily habits and adiposity as fat-mass index (FMI) by dual-energy X-ray absorptiometry was performed. Individual OL-components: "inactivity," "excessive screen time," "insufficient sleep," "unhealthy-diet", were defined according to non-adherence to previously published healthy recommendations. Results: 1449 subjects were assessed between March 2015 to April 2018. Sixteen percent of subjects had all four OL-components, 40% had three, 35% had two, 9% had one, and 0.5% had none. A cumulative OL score showed a significant dose-response effect with FMI. The combination of inactivity, excessive screen time, and insufficient sleep showed the highest risk association to OW/OB and higher values of FMI. Conclusions: The prevalence of OL-components was extremely high and associated with increased adiposity and OW/OB. Several interventions are needed to revert this major public health threat.
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Adiposidade , Estilo de Vida , Obesidade Infantil/epidemiologia , Obesidade Infantil/etiologia , Adolescente , Composição Corporal , Criança , Feminino , Humanos , Masculino , México/epidemiologia , Sobrepeso/epidemiologia , Sobrepeso/etiologia , Vigilância em Saúde PúblicaRESUMO
The hyperpolarization-activated cyclic nucleotide-gated (HCN) channels are encoded by a family of four genes (HCN1-4). All isoforms are expressed in the heart, HCN4 being the most abundant in the sinoatrial node (SAN). HCN channels are responsible for the "funny" current (If) associated with the generation and autonomic control of the diastolic depolarization phase of cardiac action potential. In this work we performed a proteomic analysis of HCN4 transfected in HEK293 cells. Most of the identified proteins in the HCN4 network belonged to mitochondria. The subcellular localization of HCN channels was predicted in plasma membrane, mitochondria and nucleus. Experimentally, HCN2 (full-length, truncated), HCN3 (full-length, truncated) and HCN4 (truncated) were detected in rat heart mitochondria by immunoblotting. If sensitive to ZD7288, was recorded by patch-clamp in mitoplasts from cardiomyocytes. Mitochondrial membrane potential (ΔΨm) assessment in H9c2 cells revealed that ZD7288 induced almost 50% higher hyperpolarization respect to control at 30 min. Furthermore, ZD7288 reduced oxygen consumption attributed to ATP synthesis in H9c2 cells. In conclusion, we identify for the first time functional HCN channels in mammalian cardiac mitochondria and demonstrate their impact on ΔΨm and respiration.
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Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização/metabolismo , Potencial da Membrana Mitocondrial , Mitocôndrias Cardíacas/metabolismo , Consumo de Oxigênio , Animais , Linhagem Celular , Células HEK293 , Humanos , Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização/análise , Camundongos Endogâmicos BALB C , Miócitos Cardíacos/metabolismo , Ratos WistarRESUMO
Rosmarinus officinalis L. (Lamiaceae) is an aromatic plant widely popular mainly due to its uses in traditional medicine as an anti-inflammatory, diuretic and antimicrobial, as well as in the prevention and treatment of diseases. These biological activities are mainly related to the presence of phenolic and terpenic compounds. This work reports a chemical profile analysis of extracts from leaves and calli of rosemary obtained by both pressurized liquid extraction and maceration. Chemical profiles were determined on calli extracts of 3, 6, 9, and 15 days of culture; chemical characterization and quantification of compounds was carried out using ultrahigh performance liquid chromatography-mass spectrometry. A total of 53 metabolites were identified in callus and 47 compounds in leaf extracts, of which 25 correspond to phenolic compounds, mainly flavonoids and flavones, 13 terpenes that include phenolic terpenes and one diterpenolactone, two glycosides which correspond to 6-O-caffeoyl-ß-D-fructofuranosil-(2â1)-α-D-glucopyranoside and primulaverin, an aromatic compound identified as fenantrenone and a growth regulator 12-hydroxy jasmonic acid. These results showed that undifferentiated rosemary cells accumulate the same compounds identified mainly in highly specialized tissues such as leaves. The plant cell culture supply the possibility of developing biotechnological processes to obtain compounds of commercial interest.
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Cromatografia Líquida de Alta Pressão/métodos , Espectrometria de Massas/métodos , Extratos Vegetais/química , Folhas de Planta/química , Rosmarinus/química , Cinamatos/análise , Cinamatos/química , Depsídeos/análise , Depsídeos/química , Flavonoides/análise , Flavonoides/química , Glicosídeos/análise , Glicosídeos/química , Terpenos/análise , Terpenos/química , Ácido RosmarínicoRESUMO
INTRODUCTION: The development of motor skills influences the capacity of the child to interact with the environment. Thus, several instruments have been created for their assessment. OBJECTIVE: To evaluate the internal consistency, reproducibility, and agreement level of age band 1 of the Movement Assessment Battery for Children - 2 in a preschool group. PATIENTS AND METHOD: Assessment study of diagnostic tests with 29 preschoolers, selected by convenience, enrollments in an educational ins titution in Bucaramanga city, Colombia. For the inter-evaluators reproducibility assessment, three evaluators watched each video independently. In the intra-evaluator reproducibility assessment, each evaluator watched the same video on two different occasions. The internal consistency, the intra- and inter-evaluator reproducibility, and the agreement level were determined using Cronbach's alpha co efficient, the Intraclass Correlation Coefficient (ICC), and the Bland and Altman limits of agreement method, respectively. RESULTS: Internal consistency of the total test for each of the three evaluators was higher than 0.60. Very good reliability was found for all items, domains, and total score of age band 1 of MABC-2 (ICC > 0.85), as well as good limits of agreement. CONCLUSIONS: age band 1 of MABC-2 Spanish version is an instrument with adequate reliability psychometric properties that can be used for the motor skills development evaluation in preschoolers.
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Transtornos das Habilidades Motoras/diagnóstico , Destreza Motora/fisiologia , Criança , Pré-Escolar , Colômbia , Feminino , Humanos , Idioma , Masculino , Variações Dependentes do Observador , Psicometria , Reprodutibilidade dos TestesRESUMO
Hyperpolarization-activated cationic HCN channels comprise four members (HCN1-4) that control dendritic integration, synaptic transmission and action potential firing. In the kidney, HCN1, HCN2 and HCN3 are differentially expressed and contribute to the transport of sodium, potassium (K+) and ammonium into the nephrons. HCN3 is regulated by K+ diets in the kidney. In this work we performed a proteomic analysis of HCN3 expressed in human embryonic kidney cells (HEK293 cells). More than 50% of the interacting proteins belonged to mitochondria. Therefore, we explored the presence of HCN channels in kidney mitochondria. By immunoblotting and immunogold electron microscopy HCN3 protein expression was found in rat kidney mitochondria; it was also confirmed in human kidney. Patch-clamp recordings of renal mitochondria and mitochondria from HEK293 cells overexpressing HCN1, HCN2 and HCN3 channels, stained with MitoTracker Green FM, indicated that only HCN3 could produce inwardly K+ currents that were inhibited by ZD7288, a specific blocker of HCN channels. Furthermore, ZD7288 caused inhibition of the oxygen consumption coupled to ATP synthesis and hyperpolarization of the inner mitochondrial membrane. In conclusion, we show for the first time that pacemaker HCN channels contribute to K+ transport in mitochondria facilitating the activity of the respiratory chain and ATP synthesis by controlling the inner mitochondrial membrane potential.
Assuntos
Rim/metabolismo , Mitocôndrias/metabolismo , Canais de Potássio/metabolismo , Potenciais de Ação , Respiração Celular , Cromatografia Líquida , Ativação do Canal Iônico , Mitocôndrias/genética , Nucleotídeos Cíclicos/metabolismo , Proteoma , Proteômica/métodos , Espectrometria de Massas em TandemRESUMO
INTRODUCCIÓN: La adquisición de las destrezas motoras fundamentales influyen en la capacidad que tiene el niño para interactuar con el ambiente. Por lo anterior diversos instrumentos han sido creados para su evaluación. OBJETIVO: Evaluar la consistencia interna, reproducibilidad y el nivel de acuerdo de la Batería para la evaluación del Movimiento en Niños -2- banda 1 (MABC-2) en un grupo de preescolares. PACIENTES Y MÉTODO: Estudio de evaluación de pruebas diagnósticas con 29 preescolares, seleccionados por conveniencia, matriculados en una institución educativa de la ciudad de Bucaramanga, Colombia. Para la evaluación de la reproducibilidad inter-evaluadores, tres evaluadores observaron cada video de forma independiente. En la evaluación de la reproducibilidad intra-evaluador, cada evaluador observó el mismo video en dos oportunidades diferentes. Fue calculado el a de Cronbach para establecer la consistencia interna, la reproducibilidad intra y entre evaluadores fue calculada con el coeficiente de correlación intraclase (CCI) y el nivel de acuerdo fue determinado em pleando la metodología de Bland y Altman. RESULTADOS: La consistencia interna del total de la prueba para cada uno de los tres evaluadores fue superior a 0,60. Se encontró muy buena reproducibilidad para todos los ítems, atributos y el puntaje total del MABC-2 banda 1 (CCI > 0,85), así como buenos niveles de acuerdo. CONCLUSIÓN: La banda 1 del MABC-2 versión en español es un instrumento con adecuadas propiedades psicométricas de confiabilidad que puede ser utilizado para la evaluación del desarrollo motor en preescolares.
INTRODUCTION: The development of motor skills influences the capacity of the child to interact with the environment. Thus, several instruments have been created for their assessment. OBJECTIVE: To evaluate the internal consistency, reproducibility, and agreement level of age band 1 of the Movement Assessment Battery for Children - 2 in a preschool group. PATIENTS AND METHOD: Assessment study of diagnostic tests with 29 preschoolers, selected by convenience, enrollments in an educational ins titution in Bucaramanga city, Colombia. For the inter-evaluators reproducibility assessment, three evaluators watched each video independently. In the intra-evaluator reproducibility assessment, each evaluator watched the same video on two different occasions. The internal consistency, the intra- and inter-evaluator reproducibility, and the agreement level were determined using Cronbach's alpha co efficient, the Intraclass Correlation Coefficient (ICC), and the Bland and Altman limits of agreement method, respectively. RESULTS: Internal consistency of the total test for each of the three evaluators was higher than 0.60. Very good reliability was found for all items, domains, and total score of age band 1 of MABC-2 (ICC > 0.85), as well as good limits of agreement. CONCLUSIONS: age band 1 of MABC-2 Spanish version is an instrument with adequate reliability psychometric properties that can be used for the motor skills development evaluation in preschoolers.
Assuntos
Humanos , Masculino , Feminino , Pré-Escolar , Criança , Transtornos das Habilidades Motoras/diagnóstico , Destreza Motora/fisiologia , Psicometria , Variações Dependentes do Observador , Reprodutibilidade dos Testes , Colômbia , IdiomaRESUMO
Management of hyper-virulent generalist pathogens is an emergent global challenge, yet for most disease systems we lack a basic understanding as to why some host species suffer mass mortalities, while others resist epizootics. We studied two sympatric species of frogs from the Colombian Andes, which coexist with the amphibian pathogen Batrachochytrium dendrobatidis (Bd), to understand why some species did not succumb to the infection. We found high Bd prevalence in juveniles for both species, yet infection intensities remained low. We also found that bacterial community composition and host defense peptides are specific to amphibian life stages. We detected abundant Bd-inhibitory skin bacteria across life stages and Bd-inhibitory defense peptides post-metamorphosis in both species. Bd-inhibitory bacteria were proportionally more abundant in adults of both species than in earlier developmental stages. We tested for activity of peptides against the skin microbiota and found that in general peptides did not negatively affect bacterial growth and in some instances facilitated growth. Our results suggest that symbiotic bacteria and antimicrobial peptides may be co-selected for, and that together they contribute to the ability of Andean amphibian species to coexist with the global pandemic lineage of Bd.
Assuntos
Anuros/microbiologia , Quitridiomicetos/isolamento & purificação , Microbiota , Peptídeos/farmacologia , Animais , Anuros/crescimento & desenvolvimento , Bactérias/efeitos dos fármacos , Bactérias/isolamento & purificação , Colômbia , Micoses/microbiologia , Micoses/veterinária , Peptídeos/análise , Pele/química , Pele/microbiologia , Simbiose , SimpatriaRESUMO
Symbiotic bacteria can produce secondary metabolites and volatile compounds that contribute to amphibian skin defense. Some of these symbionts have been used as probiotics to treat or prevent the emerging disease chytridiomycosis. We examined 20 amphibian cutaneous bacteria for the production of prodigiosin or violacein, brightly colored defense compounds that pigment the bacteria and have characteristic spectroscopic properties making them readily detectable, and evaluated the antifungal activity of these compounds. We detected violacein from all six isolates of Janthinobacterium lividum on frogs from the USA, Switzerland, and on captive frogs originally from Panama. We detected prodigiosin from five isolates of Serratia plymuthica or S. marcescens, but not from four isolates of S. fonticola or S. liquefaciens. All J. lividum isolates produced violacein when visibly purple, while prodigiosin was only detected on visibly red Serratia isolates. When applied to cultures of chytrid fungi Batrachochytrium dendrobatidis (Bd) and B. salamandrivorans (Bsal), prodigiosin caused significant growth inhibition, with minimal inhibitory concentrations (MIC) of 10 and 50 µM, respectively. Violacein showed a MIC of 15 µM against both fungi and was slightly more active against Bsal than Bd at lower concentrations. Although neither violacein nor prodigiosin showed aerosol activity and is not considered a volatile organic compound (VOC), J. lividum and several Serratia isolates did produce antifungal VOCs. White Serratia isolates with undetectable prodigiosin levels could still inhibit Bd growth indicating additional antifungal compounds in their chemical arsenals. Similarly, J. lividum can produce antifungal compounds such as indole-3-carboxaldehyde in addition to violacein, and isolates are not always purple, or turn purple under certain growth conditions. When Serratia isolates were grown in the presence of cell-free supernatant (CFS) from the fungi, CFS from Bd inhibited growth of the prodigiosin-producing isolates, perhaps indicative of an evolutionary arms race; Bsal CFS did not inhibit bacterial growth. In contrast, growth of one J. lividum isolate was facilitated by CFS from both fungi. Isolates that grow and continue to produce antifungal compounds in the presence of pathogens may represent promising probiotics for amphibians infected or at risk of chytridiomycosis. In a global analysis, 89% of tested Serratia isolates and 82% of J. lividum isolates were capable of inhibiting Bd and these have been reported from anurans and caudates from five continents, indicating their widespread distribution and potential for host benefit.