Genetic Testing in Familial Hypercholesterolemia: Is It for Everyone?

ReviewPurpose of Review: Lipid measurements and genetic testing are the main diagnostic tools for FH screening that are available in many countries. A lipid profile is widely accessible, and genetic testing, although available worldwide, in some countries is only performed in a research context. Still FH is diagnosed late, showing lack of early screening programs worldwide. Recent Findings: Pediatric screening of FH was recently recognized by the European Commission Public Health Best Practice Portal as one on the best practices in non-communicable disease prevention. The early diagnosis of FH and the lowering of LDL-C values over lifespan can reduce the risk of coronary artery disease and offer health and socioeconomic gains. Summary: Current knowledge about FH shows that early detection through appropriate screening needs to become a priority in healthcare systems worldwide. Governmental programs for FH identification should be implemented to unify the diagnosis and increase patient identification.info:eu-repo/semantics/publishedVersio

Portuguese Familial Hypercholesterolaemia Study

A Hipercolesterolemia Familiar (FH) é uma doença genética associada a um elevado risco cardiovascular. Doentes com FH possuem valores muito elevados de colesterol no plasma, desde o nascimento. Até à data 3 genes foram associados à FH: LDLR (85-90%), APOB (5-8%) e PCSK9 (1-2%). Em 1999 foi estabelecido, no Instituto Nacional de Saúde Doutor Ricardo Jorge, o Estudo Português de Hipercolesterolemia Familiar (EPHF). Este estudo de investigação tem como objetivo principal identificar a causa genética da dislipidemia em doentes com critérios clínicos de FH. O EPHF identificou molecularmente 718 doentes heterozigotos com uma variante patogénica ou provavelmente patogénica (segundo as diretrizes da ACMG) num dos 3 genes associados à FH. Adicionalmente, 90 indivíduos possuem uma das 35 variantes de significado incerto cuja patogenicidade necessita de ser avaliada através de estudos funcionais. Foram também identificados 10 homozigotos (3 homozigotos verdadeiros e 7 heterozigotos compostos), com variantes patogénicas nos genes LDLR e PCSK9. O EPHF conseguiu identificar 3,8% dos portugueses que se calcula terem FH, colocando Portugal entre os dez países com mais doentes identificados. O risco cardiovascular dos doentes com FH é determinado pelos valores elevados de colesterol que os doentes apresentam desde o nascimento, mas também pela patogenicidade da variante identificada. A identificação precoce dos doentes com FH, através do diagnóstico genético, permite ao clínico implementar medidas terapêuticas adequadas e mais agressivas, de modo a diminuir o risco cardiovascular inerente a estes doentes.Familial Hypercholesterolemia (FH) is a genetic disorder associated with a high cardiovascular risk. Patients with FH have very high plasma cholesterol values since birth. Until now three genes have been associated to FH: LDLR (85-90%), APOB (5-8%) and PCSK9 (1-2%). In 1999 the Portuguese FH (PFHS) Study was established at the National Institute of Health. This study aims to identify the genetic cause of dyslipidemia in patients with clinical diagnosis of FH. The Portuguese FH Study genetically identified 718 heterozygous patients with a pathogenic or probably pathogenic variant (according to ACMG guidelines) in one of the 3 genes associated to FH. In addition, 90 individuals have one of 35 variants of uncertain significance whose pathogenicity needs to be assessed through functional studies. Ten homozygotes (3 true homozygotes and 7 heterozygous compounds) were also identified, with pathogenic variants in the LDLR and PCSK9 genes. The PFHS was able to identify 3.8% of the Portuguese that are estimated to have FH, placing Portugal among the ten countries with the most patients identified. The cardiovascular risk of FH patients is determined by the high cholesterol values present since birth, but also by the pathogenicity of the identified variant. The early identification of these patients through genetic diagnosis allows the clinician to implement appropriate and more aggressive therapeutic measures in order to reduce the cardiovascular risk inherent to these patients.info:eu-repo/semantics/publishedVersio

Comparative study on the performance of different classification algorithms, combined with pre- and post-processing techniques to handle imbalanced data, in the diagnosis of adult patients with familial hypercholesterolemia

Familial Hypercholesterolemia (FH) is an inherited disorder of cholesterol metabolism. Current criteria for FH diagnosis, like Simon Broome (SB) criteria, lead to high false positive rates. The aim of this work was to explore alternative classification procedures for FH diagnosis, based on different biological and biochemical indicators. For this purpose, logistic regression (LR), naive Bayes classifier (NB), random forest (RF) and extreme gradient boosting (XGB) algorithms were combined with Synthetic Minority Oversampling Technique (SMOTE), or threshold adjustment by maximizing Youden index (YI), and compared. Data was tested through a 10 x 10 repeated k-fold cross validation design. The LR model presented an overall better performance, as assessed by the areas under the receiver operating characteristics (AUROC) and precision-recall (AUPRC) curves, and several operating characteristics (OC), regardless of the strategy to cope with class imbalance. When adopting either data processing technique, significantly higher accuracy (Acc), G-mean and F-1 score values were found for all classification algorithms, compared to SB criteria (p < 0.01), revealing a more balanced predictive ability for both classes, and higher effectiveness in classifying FH patients. Adjustment of the cut-off values through pre or post-processing methods revealed a considerable gain in sensitivity (Sens) values (p < 0.01). Although the performance of pre and post-processing strategies was similar, SMOTE does not cause model's parameters to loose interpretability. These results suggest a LR model combined with SMOTE can be an optimal approach to be used as a widespread screening tool.The current work was supported by the programme Norte2020 (operação NORTE-08-5369-FSE-000018), awarded to JA, and by Fundação para a Ciência e Tecnologia (FCT), under the projects UID/MAT/00006/2019 and PTDC/SAU-SER/29180/2017.info:eu-repo/semantics/publishedVersio

Ciência com letras: uma experiência interdisciplinar na formação de professores/educadores

A construção da profissionalidade docente surge como um desafio para a supervisão pedagógica quando em contexto do Mestrado em Educação Pré-Escolar (PE) e Ensino do 1.º Ciclo do Ensino Básico (CEB). Pretende-se que os estudantes, no âmbito deste perfil formativo, desenvolvam a capacidade de problematizar e uma atitude empreendedora e criativa, que lhes permitam assumir com maior autonomia as responsabilidades que se atribuem ao educador/professor. Apresentam-se neste artigo os resultados da supervisão de uma experiência interdisciplinar planificada e desenvolvida numa perspetiva de trabalho colaborativo por quatro estudantes em contexto de estágio na mesma instituição, com crianças de três turmas do PE (3 a 5 anos) e de oito turmas do 1.º CEB (1.º ao 4.º ano). No âmbito da Semana da Ciência, os estudantes foram desafiados a planificar/delinear e realizar uma atividade interdisciplinar que motivasse as crianças para o Conhecimento do Mundo/Estudo do Meio e a refletir individualmente e em grupo sobre a atividade desenvolvida e as potencialidades do saber experiencial na construção da profissionalidade docente. Os estudantes estagiários organizaram uma exposição/workshop intitulado "Ciência com Letras" como oportunidade para abordar a História da Ciência, dando a conhecer cientistas que também se notabilizaram no domínio das Letras – Egas Moniz, Rómulo de Carvalho/António Gedeão e Abel Salazar –, referindo a época e os espaços em que viveram e a atividade científica, literária e/ou artística que desenvolveram. Proporcionou-se às crianças, a realização de experiências – Mensagens secretas (1.º/2.º ano), Polímeros (3.º/4.º ano) e Explosão de cores (3/4/5 anos) – e o desenvolvimento de atividades de escrita e desenho. As atividades foram planeadas tendo em conta a adequação da linguagem científica ao grupo etário e a abordagem dos conteúdos desenvolvidos nessa semana com os alunos de 1.º CEB, na área curricular do Português. Após a realização das atividades observadas pelos supervisores, os estudantes recolheram as perceções dos professores cooperantes e das crianças/alunos e elaboraram as suas narrativas. A análise destes registos permitiu perceber que o facto de terem planeado e desenvolvido atividades envolvendo diferentes áreas do conhecimento possibilitou aos estudantes a compreensão do potencial do trabalho interdisciplinar com as crianças/alunos do PE e do 1º CEB e do papel desempenhado pelo saber experiencial na construção da sua profissionalidade.The construction of teacher professionalism emerges as a challenge for pedagogical supervision in the context of the master’s degree in Pre-School Education and Teaching of the 1st Cycle of Basic Education (CBE). It is intended that trainee students, within the scope of this training profile, develop the capacity to problematize and improve responsible, entrepreneurial and creative attitudes, that allow them to assume with greater autonomy the responsibilities attributed to the educator/teacher. This paper presents the results of supervising an interdisciplinary experience planned collaboratively by four trainee students at the same institution, with children from three classes of the Pre-School Education (3-5 years) and students from eight groups of the 1st CBE (1st to 4th year).During the Science Week, trainee students were challenged to plan and carry out an interdisciplinary activity that motivated children/students to learn Knowledge of the World/Study of the Environment and to reflect individually and in group on the activity developed and the potentialities of experiential knowledge in the construction of teaching professionalism. The trainee students organized a workshop entitled "Science with Letters" as an opportunity to approach the History of Science, knowing Portuguese scientists that, at the same time, were recognized men of letters – Egas Moniz, Rómulo de Carvalho/António Gedeão and Abel Salazar –, referring to the time and spaces in which they lived and the scientific, literary and/or artistic activity they have developed. Students were given the opportunity to carry out experiments – Secret Messages (1st/2nd year), Polymers (3rd/4th year) and Color Explosion (3/4/5 years) – and to develop writing and drawing activities. The activities were planned, considering the adequacy of the scientific language to the age group and the approach of the contents developed this week with the students of 1st CBE, in the curricular area of Portuguese. After performing the activities, the trainee students collected the insight of the cooperating teachers and the children/students and elaborated their narratives. The analysis of these records showed the fact that they planned activities involving different areas of knowledge enabled the trainee students to understand the potential of interdisciplinary work with the children of the Pre-School Education and the 1st CBE and the role played by experiential knowledge in the construction of their professionalism. Keywords: pedagogical supervision, interdisciplinarity, teacher training

Učinci ekstrakta Cytinus hypocistis (L.) L. u životinjskom modelu neoplazije inducirane papiloma virusom

Infections with certain types of papillomavirus, such as the human papillomavirus 16 (HPV16), are associated with the development of preneoplastic lesions and cancers of the anogenital, and head and neck regions. Cytinus hypocistis (L.) L. extracts are composed of substances presenting antiproliferative, antioxidant, anti-inflammatory and antibacterial properties, which might be promising as new therapeutic compounds. This study analysed the influence of topical application of an extract obtained from C. hypocistis (CH) on K14-HPV16 and FVB/n mice to evaluate its therapeutic and toxicological properties. To achieve the study goals, 30 female mice, 33–37 weeks old, were divided into six groups (n=5/group): I (HPV+CH3.1); II (HPV+CH6.2); III (HPV+CH12.4); IV (FVB/n+CH12.4); V (HPV+control) and VI (FVB/n+-control). CH was applied topically to both ears for 28 days. After this period, all animals were sacrificed for samples collection. Skin lesions were classified histologically. Toxicological parameters included haematological and biochemical blood markers, and hepatic oxidative stress analysis. Transgenic animals showed a decrease in mean body weight regardless of the concentration of extract applied. The extract had no influence on physiological parameters, organ weight, or biochemical and oxidative stress parameters. Histology demonstrated the presence of proliferative epithelial lesions in the skin and oral mucosa of K14-HPV16 mice, with no association with the application of this extract. Overall, the application of CH extract had no influence on the skin lesions and was well tolerated by the animals in these concentrations.Infekcije određenim vrstama papiloma virusa, poput humanog papiloma virusa 16 (HPV16), povezane su s razvojem preneoplastičnih lezija i karcinoma anogenitalnog područja i područja glave i vrata. Ekstrakti Cytinus hypocistis (CH) sadrže tvari koje pokazuju antiproliferativna, antioksidativna, protuupalna i antibakterijska svojstva te bi stoga mogle predstavljati nove, obećavajuće terapijske spojeve. Cilj je ovog rada bio analizirati utjecaj topikalne primjene ekstrakta dobivenog iz C. hypocistis (L.) L. na K14-HPV16 i FVB/n miševima za procjenu njegovih terapijskih i toksikoloških svojstava. Za postizanje ciljeva istraživanja, trideset ženki miševa starosti 33-37 tjedana podijeljeno je u šest skupina (n=5/ skupini): I (HPV+CH3,1); II (HPV+CH6,2); III (HPV+CH12,4); IV (FVB/n+CH12,4); V (HPV+kontrola) i VI (FVB/n+kontrola). CH je tijekom 28 dana topikalno primijenjen na oba uha. Nakon tog razdoblja sve životinje su žrtvovane u svrhu prikupljanja rezulata. Lezije kože su histološki klasificirane. Toksikološki parametri uključivali su hematološke i biokemijske markere krvi te analizu oksidativnog stresa jetre. Transgenične životinje pokazale su smanjenje srednje tjelesne mase, bez obzira na primijenjenu koncentraciju ekstrakta. Ekstrakt nije utjecao na fiziološke parametre, masu organa ili parametre biokemijskog i oksidativnog stresa. Histološki je dokazana prisutnost proliferativnih epitelnih lezija na koži i oralnoj sluznici K14-HPV16 miševa, bez povezanosti s primjenom ovog ekstrakta. Općenito, primjena CH ekstrakta nije utjecala na lezije kože te su ga životinje dobro podnosile u primijenjenim koncentracijama

Sublineage A1 Drives Multi-Organ Carcinogenesis

by Fundação para a Ciência e Tecnologia/Ministério da Ciência, Tecnologia e Ensino Superior (FCT/MCTES, Portugal) through national funds to iNOVA4Health (UIDB/04462/2020 and UIDP/ 04462/2020); from the GenomePT project (POCI-01-0145-FEDER-022184), supported by COMPETE 2020—Operational Programme for Competitiveness and Internationalisation (POCI), Lisboa Portugal Regional Operational Programme (Lisboa2020), Algarve Portugal Regional Operational Pro- Int. J. Mol. Sci. 2022, 23, 12371 8 of 10 gramme (CRESC Algarve2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF), and by Fundação para a Ciência e a Tecnologia (FCT). This work was also supported by Fundos FEDER through the Programa Operacional Factores de Competitividade—COMPETE and by Fundos Nacionais through the Fundação para a Ciência e a Tecnologia within the scope of the project UID/BIM/00009/2019 (Centre for Toxicogenomics and Human Health-ToxOmics); and from LA/P/0045/2020 (ALiCE), UIDB/00511/2020 and UIDP/00511/2020 (LEPABE), funded by national funds through FCT/MCTES (PIDDAC); 2SMART (NORTE-01-0145- FEDER-000054), supported by Norte Portugal Regional Operational Programme (NORTE 2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF).The study of human papillomavirus (HPV)-induced carcinogenesis uses multiple in vivo mouse models, one of which relies on the cytokeratin 14 gene promoter to drive the expression of all HPV early oncogenes. This study aimed to determine the HPV16 variant and sublineage present in the K14HPV16 mouse model. This information can be considered of great importance to further enhance this K14HPV16 model as an essential research tool and optimize its use for basic and translational studies. Our study evaluated HPV DNA from 17 samples isolated from 4 animals, both wild-type (n = 2) and HPV16-transgenic mice (n = 2). Total DNA was extracted from tissues and the detection of HPV16 was performed using a qPCR multiplex. HPV16-positive samples were subsequently whole-genome sequenced by next-generation sequencing techniques. The phylogenetic positioning clearly shows K14HPV16 samples clustering together in the sub-lineage A1 (NC001526.4). A comparative genome analysis of K14HPV16 samples revealed three mutations to the human papillomaviruses type 16 sublineage A1 representative strain. Knowledge of the HPV 16 variant is fundamental, and these findings will allow the rational use of this animal model to explore the role of the A1 sublineage in HPV-driven cancer.publishersversionpublishe

MicroRNA-21 expression and susceptibility to HPV-induced carcinogenesis - role of microenvironment in K14-HPV16 mice model

Aims: Human papillomaviruses (HPVs) are responsible for several types of cancer. K14-HPV16 transgenic mice express the HPV16 early genes, developing multi-step carcinogenesis associated with marked inflammation, as observed in human patients. MicroRNAs (MiRNA) constitute a class of non-coding RNAs that regulate gene expression. In particular, miR-21 has been associated with carcinogenesis. However, little is known about this microRNA in the normal tissue microenvironment and its possible relationship with cancer predisposition. We hypothesized that miR-21 expression influences each tissue's susceptibility to HPV-induced carcinogenesis. Main methods: In order to test this hypothesis, we evaluated miR-21 expression by RT-qPCR in ear and chest skin samples from 24-28 weeks old, female K14-HPV16 transgenic and wild-type mice. Key findings: In wild-type mice (HPV-/-) miR-21 expression was lower in ear skin compared with that of chest skin (p = 0.036). Under the influence of HPV16 oncogenes, transgenic animals (HPV16+/-), developed in situ carcinoma in all ear samples and epidermal hyperplasia in chest samples. These results are consistent with the hypothesis that microRNA expression in the microenvironment of normal tissues may influence HPV-associated carcinogenesis. Furthermore, among transgenic animals, miR-21 expression was lower in in situ carcinoma samples compared with hyperplasia (p = 0043). Significance: This suggests that, despite the well-known role of miR-21 as an oncogene, its anti-inflammatory and immunomodulatory properties may modulate HPV-induced carcinogenesis in a tissue-dependent manner. Further studies are warranted in order to explore the role of microRNAs in tissue susceptibility to carcinogenesis

Portuguese Familial Hypercholesterolemia Study (1999-2021): phenotype-genotype characterization

A Hipercolesterolemia Familiar (FH) é uma condição genética comum do metabolismo dos lípidos, que se encontra subdiagnosticada. Existem três genes primários associados à FH (LDLR, APOB e PCSK9) e 5 genes fenocópias (LDLRAP1, LIPA, ABCG5, ABCG8 e APOE), que conferem um fenótipo semelhante à FH. Neste trabalho pretende-se apresentar a relação fenótipo-genótipo dos indivíduos com critérios clínicos de FH referenciados ao Estudo Português de Hipercolesterolemia Familiar. Até ao fim de 2021 foram estudados molecularmente 1005 indivíduos com critérios clínicos de FH. Destes, foram confirmados geneticamente com FH (FH positivos), 417 casos-índex (408 heterozigotos e 9 homozigotos). Com os estudos familiares identificaram- se adicionalmente 581 heterozigotos e 2 homozigotos. De entre os FH positivos, os casos-índex com variantes de alelo nulo apresentam um fenótipo mais severo do que os casos-índex com variantes de alelo defeituoso. Cerca de 1% dos casos-índex foram diagnosticados com outras causas monogénicas. Dos FH negativos, 34% apresenta hiper-Lp(a), 18% tem uma hipercolesterolemia de causa poligénica e 1% possui uma variante patogénica em heterozigotia nos genes fenocópias da FH. As diferentes causas genéticas contribuem para uma variedade de fenótipos que requerem diferentes formas de gestão da doença, terapias específicas e têm implicações na estratificação do risco cardiovascular e no rastreio dos familiares, sendo por esta razão essencial que seja identificada a etiologia da hipercolesterolemia o mais precocemente possível para melhorar o prognóstico dos indivíduos com FH.Familial Hypercholesterolemia (FH) is a common genetic condition of lipid metabolism that is underdiagnosed. There are three primary genes associated with FH (LDLR, APOB and PCSK9) and 5 phenocopy genes (LDLRAP1, LIPA, ABCG5, ABCG8 and APOE), which confer a phenotype FH-like. In this work, we present the phenotype – genotype characterization of individuals with clinical criteria of FH referred to the Portuguese FH Study. Until the end of 2021, 1005 individuals with clinical diagnosis of FH were molecularly studied. Of these, 417 index cases (408 heterozygotes and 9 homozygotes) were genetically confirmed with FH (FH positive). Cascade screening also identified 581 heterozygotes and 2 homozygotes. Among the FH positives, the index cases with null allele variants show a more severe phenotype than the index cases with defective allele variants. About 1% of the index cases were diagnosed with other monogenic causes. Of the FH negatives, 34% have hyper-Lp(a), 18% have a polygenic hypercholesterolemia and 1% have a pathogenic variant in heterozygosity in a FH phenocopy genes. The different genetic causes contribute to a variety of phenotypes that require different forms of disease management, specific therapies and have implications for cardiovascular risk stratification and family screening. Therefore, it is essential that the etiology of hypercholesterolemia is identified as early as possible to improve the prognosis of individuals with FH.info:eu-repo/semantics/publishedVersio

Efeito da aplicação repetida de Cytinus hypocistis em murganhos transgénicos (K14HPV16): avaliação de humane endpoints

Em, 1959 Russel e Burch propuseram os 3Rs (Reduce, Refine e Replacement) em experimentação animal e desde então as diversas entidades envolvidas na temática têm feito esforços conjuntos para os aplicar (1). Nesse sentido foi gradualmente introduzida a avaliação e registo dos humane endpoints (HE) dos animais durante procedimentos experimentais, para monitorizar o seu sofrimento e tomar decisões sobre a necessidade, ou não, de finalizar o ensaio antes da data prevista (2). Contudo, apesar das exigências legais, observa-se grande escassez de publicações sobre a avaliação de HE nos procedimentos com animais de laboratório. Assim, é nosso objetivo com este trabalho apresentar os HE durante a aplicação diária de um creme com Cytinus hypocistis numa estirpe de murganhos transgénicos.Financiamento nacional pela FCT, Fundação para a Ciência e a Tecnologia, no âmbito da bolsa de doutoramento 2020.07675.BD de Beatriz Medeiros Fonseca.info:eu-repo/semantics/publishedVersio