Departamento de Promoção da Saúde e Prevenção de Doenças Não Transmissíveis
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Browsing Departamento de Promoção da Saúde e Prevenção de Doenças Não Transmissíveis by advisor "Alves, Ana Catarina"
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- APOB Variants Spectrum and Functional Characterization in Portuguese Patients with Familial Hypercholesterolaemia PhenotypePublication . Ferreira, Maria Rafael Simões do Carmo; Alves, Ana Catarina; Rebelo, Maria TeresaFamilial hypercholesterolaemia (FH) is an autosomal semi dominant disorder of lipid metabolism clinically characterized by increased levels of circulating LDL cholesterol and associated with elevated cardiovascular risk. The genetic diagnosis is usually based on the analysis of LDLR, APOB, and PCSK9 genes. APOB variants are responsible for 5-10% of FH cases, and the variant spectrum of APOB has increased due to sequencing of the whole gene through Next Generation Sequencing, consequently increasing the number of variants that need to be functionally assessed. This dissertation aimed to verify the correlation between phenotype and genotype in individuals from the Portuguese FH Study, as well as create a database including all APOB variants found up to date in this study. Moreover, it was intended to characterize two APOB variants identified in subjects from this cohort. Graphics regarding LDL cholesterol levels were designed for index cases FH positive and negative and relatives FH positive. The variants previously detected by NGS were confirmed by PCR and Sanger sequencing, and cascade screening was carried out in families. All APOB variants with MAF <1% were gathered into a database. LDL from index cases and relatives was separated using sequential ultracentrifugation and labelled with FITC for uptake assessment by flow cytometry in CHO-ldlA7 cells, and proliferation assays were performed with U937 cells. A definite diagnosis was possible for 4 individuals carrying known pathogenic variants, and c.6639_6641del/p.(Asp2213del) and c.10121T>C/p.(Ile3374Thr) alterations from exon 26 were functionally assessed. In vitro studies showed a neutral effect on the apoB function for these variants. Furthermore, 143 different variants were discovered located throughout the whole gene, of which more than 90% were variants of uncertain significance. Functional studies, combined with the association between phenotype and genotype, allow a better and more personalized treatment according to the needs of each individual.
- Estudos Funcionais de Variantes no Gene APOB em Doentes com Diagnóstico Clínico de Hipercolesterolemia FamiliarPublication . Ramos, Diana Catarina Fernandes; Bourbon, Mafalda; Alves, Ana CatarinaA hipercolesterolemia familiar (FH) é uma doença caracterizada por níveis elevados de colesterol LDL, que se acumulam nas artérias, promovendo o desenvolvimento precoce de eventos cardiovasculares. Geneticamente, a FH é transmitida de forma autossómica semi-dominante, sendo causada por variantes nos genes LDLR, APOB e PCSK9. As variantes no gene APOB afetam a ligação do colesterol LDL ao seu recetor, representando 5 a 10% dos casos de FH. No entanto, para a maioria das variantes identificadas, o efeito funcional na proteína ainda não está determinado. Esta dissertação teve como objetivo principal investigar a correlação entre fenótipo e genótipo, além de caracterizar funcionalmente variantes identificadas no gene APOB em indivíduos com diagnóstico clínico de FH na população portuguesa. Como objetivo secundário, foi desenvolvida uma base de dados que reúne todas as variantes deste gene com estudos funcionais realizados até ao momento. A identificação das variantes foi realizada por sequenciação de nova geração (NGS) e confirmada por PCR e sequenciação de Sanger, seguida da pesquisa da variante nos familiares dos casos index. As variantes em estudo foram identificadas no âmbito do projeto EPHF. O LDL dos participantes foi isolado por ultracentrifugação a partir do soro de casos index e familiares com variantes no gene APOB, bem como de indivíduos normolipidémicos. Este LDL foi marcado com fluorescência e incubado em células CHO-ldlΔ7 para avaliar a capacidade de ligação e internalização do LDL. A caracterização funcional, realizada por citometria de fluxo com LDL fluorescente, revelou que as variantes p.(Asp1456Asn), p.(Val4295Leu) e p.(Arg4519Thr) apresentavam ligação e internalização normais do LDL, indicando que estas variantes não comprometem a função da proteína. Este trabalho destaca a importância de caracterizar funcionalmente as variantes no gene APOB para melhorar o diagnóstico dos indivíduos com FH, possibilitando tratamentos mais adequados a cada caso e contribuindo para a redução do risco cardiovascular.
- Health inequalities in cerebro and cardiovascular health – an analysis of the e_COR studyPublication . Afonso dos Santos, Carolina Sofia; Antunes, Marília Cristina de Sousa; Alves, Ana CatarinaAccording to the World Health Organization (WHO), health inequalities are systematic differences in the health status of different population groups with significant social and economic costs for individuals and societies. These differences are unfair and can be mitigated through the action of government policies, so it is necessary to understand their impact. Cardiovascular diseases (CVD) are a group of disorders of the brain, heart and blood vessels and include coronary heart disease, cerebro-cardiovascular disease, rheumatic heart disease and other conditions. More than four out of five CVD deaths are due to heart attacks and strokes, and one third of these deaths occur prematurely in people under 70 years of age (WHO). The events of interest considered in this work were stroke, acute myocardial infarction, and peripheral arterial disease. In Portugal in 2019, strokes were the number one cause of death representing 9.8% of the total mortality and the deaths by myocardial infarction represented 3.8% of the total mortality, according to INE. It is estimated that about 10 670 potential years of life were lost due to cerebro-cardiovascular diseases in Portugal in 2019. In addition to being interesting for being very prevalent across Europe, cerebrovascular diseases are impacted by the lifestyle and behavior of individuals. Several studies have concluded that a substantial part of the risk of developing cerebro-cardiovascular disease can be reduced through personal choices and preventive approaches, such as adapting healthy lifestyles. An individual’s socioeconomic position has a potential effect on their health status and also on the health care they receive, which means that socioeconomic inequalities are determinants of health. The European Deprivation Index (EDI) is a way of quantifying the level of deprivation of individuals and, therefore, the version adapted to the Portuguese reality, developed in 2016 by a multinational and multidisciplinary team of researchers, was incorporated in this work. The main objective of this work is to evaluate the effect of health inequalities, namely those associated with low education and levels of deprivation in cerebro-cardiovascular disease. To achieve this objective, the main set of data used resulted from the e_COR study, developed by the Instituto Nacional de Saúde Doutor Ricardo Jorge, between 2012 and 2014, within the scope of cardiovascular prevention and with the objective of estimating the prevalence of cardiovascular risk factors. In the present work, the past occurrence of at least one cerebro-cardiovascular event and the number of risk factors that each of the participants in the e_COR study had were studied. The variables under study are some of the variables collected during the e_COR study, referring to demographic, physical, metabolic, medical history, and lifestyle characteristics of the participants. Some other variables were collected such as variables that characterize access to healthcare, extracted from INE, and variables related to socioeconomic status, that resulted from the EDI adapted to the Portuguese context. To model the occurrence of a cerebro-cardiovascular event, the logistic regression method was used, as it suits the binary nature of the response variable. Before modeling the data, possible correlations between the variables under study were evaluated. The variables that were included in the Generalized Linear Model were selected through the Stepwise Selection process, considering an inclusion p-value of 0.20 and an exclusion p-value of 0.25. The confounding variables, sex and age, were the first to be introduced in the model and, regardless of their p-value, they always remained in the model. After the Stepwise process, variables related to levels of education and deprivation were introduced. The random effect of the region of residence was added to the final model. Since there was a large imbalance between cases and non-cases of cerebro-cardiovascular disease, the SMOTE technique was used to create a more balanced dataset and the same modeling process was applied. For the validation of the models obtained, the ROC curve with the respective AUC was calculated and cross validation was also performed. Poisson regression was used to model the number of risk factors, since it is one of the methods indicated when the response variable is a count variable. In this model, once again, confounding variables, sex and age were included, as well as the variables related to inequalities, education levels and deprivation levels. Multiple comparisons were performed to assess how variables associated with inequalities affect the number of risk factors that each individual has. The recorded cerebro-cardiovascular events that occurred happened in the past and the risk factors that individuals have are in the present, so the interpretation of the models is not the most conventional and some of the variables would have different values if they had been evaluated before the event occurred. Both logistic regression models suggest that in the presence of two individuals with similar physical and biological characteristics, the influence of their levels of education or deprivation is not significant, and these are not good indicators to distinguish individuals who have already suffered a cerebro-cardiovascular event from those who have not. The random effect of the region is also not statistically significant. Since the event has already occurred, this model is useful mainly to distinguish individuals who have already suffered the event from those who have not suffered, through their characteristics. The Poisson regression model suggests that an individual's education levels affect the number of risk factors that an individual has, i.e., it was found that individuals with more advanced education had a lower number of risk factors.
- Prevalence of Thrombogenic Risk Factors in the Portuguese population and Identification of High-risk Individuals in PortugalPublication . Gomes dos Santos, Micaela Patrícia; Alves, Ana Catarina; Rivera, Isabel AntolinIntrodução: A trombose é uma patologia comum e complexa, referente à formação de um coágulo que dificulta o normal fluxo sanguíneo. Sob condições normais, os mecanismos procoagulantes, anticoagulantes e fibrinolíticos regulam a hemostasia para evitar a formação patológica destes coágulos. No entanto, alterações nos genes de proteínas envolvidas direta ou indiretamente na regulação hemostática podem levar a um aumento anómalo da coagulação e a uma maior predisposição da formação de coágulos obstrutivos. Esta condição é conhecida como trombofília e descreve um estado de hipercoagulabilidade responsável pelo desenvolvimento de eventos trombóticos que comprometem a vida (como o enfarte do miocárdio, acidente vascular cerebral e embolia pulmonar) e que geralmente se desenvolvem na presença de um ou mais fatores de risco herdados (variantes genéticas) e ambientais (adquiridos). A trombose tem, portanto, uma origem multifatorial causada pela interação destes fatores de risco herdados e/ou outros adquiridos (tabagismo, gravidez, uso de contracetivos orais e terapia hormonal), que podem coexistir no mesmo indivíduo e conferir um risco contínuo ao longo da vida. Devido ao seu estado adquirido de hipercoagulabilidade, as grávidas apresentam ainda um risco acrescido à manifestação de tromboses, traduzido pela ocorrência de abortos espontâneos recorrentes. Através da eliminação de fatores de risco adquiridos é possível prevenir (até certo ponto) o desenvolvimento de trombose. É assim importante conhecer o perfil genético da população de forma a reunir dados que permitam a identificação de indivíduos de alto risco e antecipar a ocorrência destes eventos, bem como desenvolver medidas preventivas dirigidas aos mesmos. Os objetivos deste projeto são então estimar a prevalência das variantes trombogénicas FV G1691A (FV Leiden), FII G20210A, AT Cambridge II, PAI-1 4G/5G, MTHFR C677T e MTHFR A1298C na população portuguesa, estabelecer associações entre a presença destas variantes genéticas e outros fatores de risco adquiridos no desenvolvimento de eventos trombóticos e identificar grupos de indivíduos de alto risco em Portugal. Introduction: Thrombosis is a pathology in which a blood clot (thrombus) hampers the normal blood flow to a tissue. Under normal conditions, procoagulant, anticoagulant and fibrinolytic pathways regulate hemostasis to avoid pathological clot formation. However, changes in the genes encoding proteins, directly or indirectly involved in hemostatic regulation, can lead to enhanced abnormal blood coagulation, resulting in an increased disposition to the formation of obstructive clots, known as thrombophilia. This state of hypercoagulability is responsible for the development of life-threatening thromboembolic events (like myocardial infarction, ischemic stroke and pulmonary embolism), which usually develop when one or more of both genetic (inherited) and environmental (acquired) risk factors come into play. Pregnant women are also at major risk to manifest thrombosis by recurrent spontaneous abortion episodes. Thus, thrombosis results from the interplay of inherited DNA changes and/or acquired risk factors (cigarette smoking, pregnancy and oral contraceptive or hormone therapy use) that may coexist in the same individual; however, it is preventable. Hence, it is important to recognize the genetic thrombogenic profile of the population to assemble data for identification of high-risk individuals and to develop local guidelines for prevention. With this in mind, this project aims to estimate the prevalence of the FV G1691A (FV Leiden), FII G20210A, AT Cambridge II, PAI-1 4G/5G and MTHFR C677T and A1298C thrombogenic variants in the Portuguese population, to establish associations between the presence of these genetic variants and the presence of other increasing risk factors in the development of thrombotic-related events and to identify groups of high-risk individuals in Portugal.