Establishment of Relational Model of Congenital Heart Disease Markers and GO Functional Analysis of the Association between Its Serum Markers and Susceptibility Genes

Purpose. The purpose of present study was to construct the best screening model of congenital heart disease serum markers and to provide reference for further prevention and treatment of the disease. Methods. Documents from 2006 to 2014 were collected and meta-analysis was used for screening susceptibility genes and serum markers closely related to the diagnosis of congenital heart disease. Data of serum markers were extracted from 80 congenital heart disease patients and 80 healthy controls, respectively, and then logistic regression analysis and support vector machine were utilized to establish prediction models of serum markers and Gene Ontology (GO) functional annotation. Results. Results showed that NKX2.5, GATA4, and FOG2 were susceptibility genes of congenital heart disease. CRP, BNP, and cTnI were risk factors of congenital heart disease (p<0.05); cTnI, hs-CRP, BNP, and Lp(a) were significantly close to congenital heart disease (p<0.01). ROC curve indicated that the accuracy rate of Lp(a) and cTnI, Lp(a) and BNP, and BNP and cTnI joint prediction was 93.4%, 87.1%, and 97.2%, respectively. But the detection accuracy rate of the markers’ relational model established by support vector machine was only 85%. GO analysis suggested that NKX2.5, GATA4, and FOG2 were functionally related to Lp(a) and BNP. Conclusions. The combined markers model of BNP and cTnI had the highest accuracy rate, providing a theoretical basis for the diagnosis of congenital heart disease

Genetics and Etiology of Down Syndrome

This book provides a concise yet comprehensive source of current information on Down syndrome. Research workers, scientists, medical graduates and paediatricians will find it an excellent source for reference and review. This book has been divided into four sections, beginning with the Genetics and Etiology and ending with Prenatal Diagnosis and Screening. Inside, you will find state-of-the-art information on: 1. Genetics and Etiology 2. Down syndrome Model 3. Neurologic, Urologic, Dental & Allergic disorders 4. Prenatal Diagnosis and Screening Whilst aimed primarily at research workers on Down syndrome, we hope that the appeal of this book will extend beyond the narrow confines of academic interest and be of interest to a wider audience, especially parents and relatives of Down syndrome patients

Optimisation of stem cell based approaches towards cardiac regeneration

Aberrant behaviour of resident cells causes several illnesses. Among them, cardiovascular diseases are the leading cause of death in the western world. The human heart possesses only negligible myocardial regeneration capacity after a myocardial infarction, thus, it cannot compensate the massive cell loss self-reliantly. The thesis addresses the optimization of different cell biological approaches for enrichment of cardiac subtypes. Moreover, the thesis demonstrates different genetic manipulation strategies (DNA-, mRNA- and microRNA-based) for an efficient modulation of cell fate.Abnormales Verhalten von körpereigenen Zellen führt zu zahlreichen Krankheitsbildern, unter diesen stellen kardiovaskuläre Krankheiten die Haupttodesursache in der westlichen Welt dar. Das menschliche Herz weist jedoch nur ein geringes Regenerationspotenzial nach einem Myokardinfarkt auf, wodurch der massive Zellverlust nicht selbstständig kompensiert werden kann. Diese Dissertation befasst sich mit der Optimierung verschiedener zellbiologischer Ansätze zur Anreicherung kardialer Subtypen sowie verschiedener Strategien der genetischen Manipulation zur Modulation des Zellschicksals

Maternal thyroid hormones role in Zebrafish neural development

Thyroid hormones (TH) are essential for proper embryonic development of the central nervous system. During this period maternal supply of TH is the only source of these hormones to the embryo. Using a zebrafish MCT8 knockdown model, with consequent inhibition of maternal thyroid hormones (MTH) uptake to the target cells, the aim of this thesis is to start a comprehensive understanding of the role of MTH during embryonic neural development. We characterised the transcriptome in 25hpf CTRL and MCT8MO zebrafish embryos and found 4,343 differentially expressed genes. Reactome analysis show that MTH regulate the expression of core developmental pathways such as NOTCH, SHH and WNT. The cellular distribution of neural MTH-target genes demonstrated their cell specific action on neural stem cells and differentiated neuron classes. We identified a series of genes involved in several key neurogenic processes to be modulated by MTH. By analysing these genes by qPCR in a temporal series, from the start of segmentation through hatching, we determined the developmental time-window where MTH are required for appropriate CNS development. We show MTH are involved in the regulation of NOTCH pathway components such as notch1a, dla, dld, her2 and her4 during neurogenesis, whereas neuroectodermal genes are not affected. Response to MTH begins at 12hpf, and the time window between 22-25hpf is particularly sensitive to MTH action. Overall, these results, show that MTH is not involved in neuroectoderm specification nor CNS compartmentalisation but stress the involvement of MTH in the early stages of neurogenesis by promoting the maintenance of specific neural progenitor populations. Analyzing the cytoarchitecture of the spinal cord we found that by the end of embryogenesis cells populating the spinal cord of control and MCT8 MO zebrafish are substantially different. Lack of thyroid hormone uptake leads to a generalized disorganization of the neural tissue, together with a decrease in: neural stem cells population, subpopulations of neuron progenitor cells, radial glial cells, mature glial cells and oligodendrocyte precursors, while the primary motor neuron domain was maintained. Colocalization analysis of neural progenitors with thraa, thrab and mct8 allowed identifying cells under the regulation of MTH via MCT8. Survival and proliferation of neural progenitor cells are compromised in MCT8MO, which could later impact on the diversity of neural cell populations obtained in the end of embryogenesis. Analysis of cell autonomous Notch activation showed it cannot rescue the phenotype induced by the lack of MTH demonstrating the niche importance in the regulation of TH action. Given that MTH regulate several important morphogenetic pathways it is likely that its action occurs as an integrator enabling an adequate equilibrium between all these signals in a time a context dependent manner. MTH actions are reflected on the timely development of neurons and glial cells. It is of great interest to continue to explore the significance of these findings to further clarify the genetic and cellular causes underlying human AHDS syndrome. In conclusion with this work, we show that thyroid hormone transferred from the mother to the embryo allows the enrichment of neural progenitor pools and the generation of cell diversity necessary to produce a fully functional central nervous tissue.As hormonas da tiróide (TH) são essenciais para o correto desenvolvimento embrionário do sistema nervoso central (CNS). Durante este período o fornecimento de hormona da tiróide pela via materna é a única fonte destas hormonas para o embrião, uma vez que a produção endógena desta hormona é iniciada apenas numa fase posterior do desenvolvimento. Evidências mostram que mesmo níveis baixos de deficiência de hormona da tiróide na mãe estão associados a desordens neurológicas e psiquiátricas nos filhos. No entanto os mecanismos moleculares envolvidos na ação desta hormona no embrião e feto continuam amplamente desconhecidos. Em humanos, mutações no principal transportador celular de TH, MCT8, causa a síndrome de Herndon-Dudley (AHDS). Esta síndrome é caracterizada pelo atraso mental, atraso global no desenvolvimento, impossibilidade de falar e uma deficiência neuromotora severa. O MCT8 é o principal transportador de hormona da tiróide presente no embrião. No presente trabalho utilizamos o “knockdown” deste transportador, inibindo a sua tradução, no modelo de peixe zebra. Este modelo foi previamente estabelecido e possui características semelhantes à síndrome AHDS humana. Neste modelo o transporte da hormona da tiróide materna (MTH) para as células alvo é bloqueado, uma vez que o MCT8 está ausente, inibindo a ação da hormona. O objetivo desta tese é começar a compreender o papel destas hormonas durante o desenvolvimento neural embrionário. Para isso caracterizamos o transcriptoma de peixe-zebra CTRL e MCT8MO às 25 horas pósfertilização (hpf) por RNA-seq. Foram encontrados 4343 genes diferencialmente expressos. A análise utilizando o Reactome revelou que a MTH está envolvida direta ou indiretamente na expressão de genes pertencentes a importantes vias de sinalização, incluindo Notch, Shh e Wnt. A análise da distribuição celular de genes alvo da MTH por hibridação in-situ revelou uma ação celular específica em células estaminais neurais, mas também sobre várias classes de neurónios diferenciados. A análise transcriptómica revelou também que uma série de genes envolvidos em vários passos de processos-chave da neurogénese são modulados pela MTH. Analisando alguns destes genes numa série temporal desde a fase da segmentação até à eclosão do embrião de peixe-zebra por qPCR, determinamos a janela temporal na qual a MTH é necessária para um correto desenvolvimento do sistema nervoso central no peixezebra. Mostramos que a MTH está envolvida na regulação de componentes da sinalização Notch, tais como notch1a, dla, dld, her2 e her4 durante o processo de neurogénese, enquanto que genes envolvidos na formação e manutenção da neuroectoderme não são regulados pela MTH. A resposta à ausência de MTH inicia-se às 12hpf, sendo que a janela de desenvolvimento entre as 22 e 25hpf parece ser particularmente sensível à ação da MTH. Globalmente estes resultados mostram que a MTH não está envolvida na indução neural, nem na compartimentalização do sistema nervoso central. No entanto é demonstrada a importância da MTH na fase inicial da neurogénese pela promoção da manutenção de populações de células progenitoras neurais. A análise à citoarquitectura da medula espinhal revelou que na fase final do desenvolvimento embrionário as células neurais presentes em embriões CTRLMO e MCT8MO diferem substancialmente. A diminuição de transporte de MTH para as células alvo leva a uma desorganização geral do tecido neural, para além duma redução em células estaminais neurais, subpopulações de células progenitoras de neurónios, células radiais gliais, células da glia maduras e progenitoras de oligodendrócitos; enquanto que a população de neurónios motores primários é mantida na ausência de MTH. Análise de colocalização de células progenitoras neurais her2(+), dla(+) e fabp7a(+) com RNAm de componentes do metabolismo da hormona da tiroide, nomeadamente recetores (thraa e thrab) e o transportador mct8, permitiu a identificação de células que estão sob a regulação da MTH. Um dos papeis clássicos da TH envolve a sobrevivência a proliferação celulares, no entanto a sua função varia com o tipo celular, o estadio de desenvolvimento e contexto celular. Verificámos que em embriões MCT8MO a sobrevivência e proliferação de células progenitoras neurais her2(+) e dla(+) estava comprometida, tanto na medula espinhal como no romboencéfalo. Este facto pode impactar a diversidade das células neurais obtidas no final da embriogénese. Na verdade, a divisão assimétrica de células progenitoras originando células da glia ou progenitoras intermediárias está diminuída nos embriões MCT8MO, mostrando um possível papel da MTH na sua formação. Com vista a esclarecer o papel da MTH sobre a sinalização Notch procedeu-se à ativação da sinalização Notch em células neurais da espinhal medula de embriões MCT8MO e CTRLMO. Esta sobre-expressão celular isolada da sinalização Notch em células individuais mostrou ser insuficiente para recuperar o fenótipo induzido pela falta de MTH, demonstrando a importância do nicho celular na regulação da ação da MTH. A ação da MTH parece estar envolvida no memanismo de divisão assimétrica de células neuroepiteliais. Este tipo de divisão está na origem da diversidade de células neurais obtidas. Um reflexo da perda deste tipo de divisões na espinhal medula pode ser a perda de progenitores intermediários que expressam neurogenina 1 e neurod6b. Dado que a MTH regula várias vias morfogenéticas é provável que a sua ação seja a de integradora entre estas vias, permitindo o equilíbrio entre estes sinais num determinado tempo e contexto celular específicos. As ações da MTH refletem-se no desenvolvimento correto e atempado de neurónios e células da glia. É de grande interesse continuar a explorar o significado destas descobertas, nomeadamente para esclarecer as causas genéticas e celulares que causam a síndrome de Herdnon-Dudley (AHDS). Foi desenvolvido um modelo de ação da MTH durante o desenvolvimento no peixe zebra, em que em subpopulações de células progenitoras contendo MCT8 e recetores da hormona da tiroide (sensíveis a TH) na ausência desta sofrem apoptose e/ou redução na proliferação em momentos distintos do desenvolvimento neural. As células progenitoras não responsivas à TH prosseguem o seu desenvolvimento. O balanço final será uma diminuição da diversidade de progenitores neurais e consequente perda na variedade de neurónios e células da glia maduros. Este efeito a nível celular terá efeito na função do CNS. Em conclusão este trabalho mostra que a hormona da tiroide transmitida da mãe para o embrião e que dá entrada nas células alvo através do MCT8 permite o desenvolvimento de populações de progenitores neurais e o consequente enriquecimento da diversidade celular, necessária à formação de um sistema nervoso central funcional.This study received Portuguese national funds from operational programmes CRESC Algarve 2020 and COMPETE 2020 through project EMBRC.PT ALG-01-0145-FEDER- 022121. The author was a recipient of a FCT PhD grant SFRH/BD/111226/201

Immunological aspects of glycosylation: from aberrant to defective glycosylation

Glycosylation is crucial in many biological processes, like cell recognition, signaling and development. Many diseases present altered glycosylation and two extremes are cancer and congenital disorders of glycosylation (CDG), with aberrant and defective glycosylation, respectively. Sialic acids are glycans’ terminal sugars with an immunomodulatory role and when decreased, typically activate immune cells, as dendritic cells. Interestingly, both ST6Gal-I and its derived α2,6 sialylation are overexpressed in cancer. Here, we hypothesized that cancer cells secret functional ST6Gal-I that modulates immune cells’ glycosylation and their activity as a cancer immune evasion mechanism. Also interestingly, patients with PMM2-CDG (the most frequent CDG type) present immunological affectation. Here, we hypothesized that the PMM2-CDG-defective glycosylation observed also influences the function of immune cells. Therefore, the main goals of this study comprised the assessment of the immunological aspects of cancer cells and CDG glycosylation. Specifically, we intended to (1) study the expression and secretion of ST6Gal-I by colorectal cancer (CRC) cells and test its function in modulating immune cells activity; (2) develop a PMM2-CDG leukocyte cell line as a model to unravel patients’ immunity and to evaluate their response to mitogenic stimulation. Moreover, as PMM2-CDG have a profound impact in patients’ quality of life (QoL), patient and observer reported outcomes measures (PROMs and ObsROMs) were reviewed. These may integrate primary endpoints in clinical trials to find treatment to PMM2-CDG. Our data demonstrated that (1) CRC cells secret ST6Gal-I enzyme, however further work is needed to evaluate its role in immune modulation; (2) PMM2-CDG T cells have higher proliferation capacity and IFN-γ cytokine expression, in response to a mitogen as compared to the healthy control and (3) there are significant numbers of tools for future evaluation of PMM2-CDG patients’ and caregivers’ QoL. This study may contribute to better understand the glycan-related pathological mechanisms

Investigation of histone lysine methylation in stem cell differentiation using inhibitor peptide

In an effort to expand the histone code, we examine a novel site of methylation on lysine 43 of histone H2B. In mouse embryonic stem cells (ESCs), KDM5b acts as the lysine demethylase for H2BK43me2, diminishing this histone mark as cells differentiate. We utilize a synthetic peptide mimetic corresponding to amino acids 37-49 of histone H2B in order to sterically inactivate KDM5b enzyme. The addition of inhibitor peptide into culture enhanced stem cell differentiation, upregulating cell cycle and neural-specific markers while downregulating the expression of pluripotency genes. Global gene analysis patterns of peptide-treated ESCs were representative of differentiated cell populations. Applying a novel inhibition method, we reveal an accelerated rate of stem cell differentiation through the upregulation of KDM5b targets. Our investigation serves to elucidate the role of histone H2BK43 methylation in an epigenetically regulated model of development

Handbook of Life Course Health Development

Health development science; Developmental origins of chronic illnesses; Community; Diabetes; Autism; Obesity; Nutrition; Health disparities across the lifespan; Fetal programmin

Effects of maternal high fat diet and pharmacological intervention on the developmental origins of metabolic &amp; cardiovascular disease

A high fat (HF) diet leads to hypercholesterolemia and predisposes the individual to developing cardiovascular disease (CVD). We hypothesised that mother‘s HF diet before and during pregnancy and lactation can also influence predisposition to CVD in offspring fed a similar diet. The thesis sets out to investigate whether (1) the effects of long-term consumption of a HF diet by the mother predisposes her offspring to developing a CVD/ metabolic syndrome in adult life and (2) pharmacological intervention using statin alleviates the detrimental effects of maternal HF diet on the health of the dams and their offspring. Female C57BL/6 mice were fed either a HF diet (45% kcal fat) or standard chow (C; 21% kcal fat) from weaning through pregnancy and lactation. Pregnant C57/BL6 mice on HF diet were further given pravastatin in the drinking water (5 mg/kg of body weight per day) either short-term (2nd half of pregnancy and during lactation) or long-term (from weaning through to pregnancy and lactation) to lower cholesterol and improve post-weaning maternal blood pressure. Weaned female offspring from each group were then fed either a HF or C diets to adulthood. Body weight, blood pressure, plasma cholesterol, C-reactive protein (CRP) and bone marrow derived endothelial progenitor cells (EPC) were measured at 24, 28 and 36 weeks post-weaning in different experiments. Histology of the liver and kidneys were performed. Offspring from hypercholesterolemic mothers on HF diet were significantly obese (bodyweight in grams; 17.2+4.2 vs. 13.8+4.7; P&lt;0.05), hypertensive (SBP mmHg; 134+4.2 vs. 117+3.4; P&lt;0.001), less active (distance in cm; 312 + 31 vs. 563 + 45; P&lt;0.001), demonstrated increased lipid laden vacuoles in liver and kidneys; and showed reduced expression of EPC (P&lt;0.05) than offspring from C dams independent of their postnatal nutrition respectively. Pravastatin therapy in HF mothers resulted in abrogation of these variables in offspring independent of post weaning nutrition (P&lt;0.05). The effects were more permanent when the dams were given long-term statin treatment. The study demonstrates that long-term maternal HF feeding from weaning through pregnancy and lactation predisposes offspring to hypertension, raised plasma lipids, fatty liver, kidney disorders, raised CRP and inhibition of EPC numbers and expression in offspring. Pravastatin treatment of these dams inhibits these effects on the offspring and may reduce their risk of later cardiovascular pathophysiology. The findings may have implications for understanding the effects of the ?nutritional transition‘ to higher dietary intake of fat which could lead to increased cardiovascular disease in many societies.<br/

Abstracts from the 50th European Society of Human Genetics Conference: Posters

International audienc

Personalized Nutrition

Awareness of the influence of our genetic variation to dietary response (nutrigenetics) and how nutrients may affect gene expression (nutrigenomics) is prompting a revolution in the field of nutrition. Nutrigenetics/Nutrigenomics provide powerful approaches to unravel the complex relationships among nutritional molecules, genetic variants and the biological system. This publication contains selected papers from the ‘3rd Congress of the International Society of Nutrigenetics/Nutrigenomics’ held in Bethesda, Md., in October 2009. The contributions address frontiers in nutrigenetics, nutrigenomics, epigenetics, transcriptomics as well as non-coding RNAs and posttranslational gene regulations in various diseases and conditions. In addition to scientific studies, the challenges and opportunities facing governments, academia and the industry are included