Familial amyloid polyneuropathy: TTR sequencing and "in silico" analysis

Familial amyloid polyneuropathy (FAP) or paramiloidosis is an autosomal dominant neurodegenerative disease with onset on adult age that is characterized by mutated protein deposition in the form of amyloid substance. FAP is due to a point alteration in the transthyretin (TTR) gene and until now more than 100 amyloidogenic mutations have been described in TTR gene. FAP shows a wide variation in age-at-onset (AO) (19-82 years, in Portuguese cases) and the V30M mutation often runs through several generation of asymptomatic carriers, before expressing in a proband, but the protective effect disappear in a single generation, with offspring of late-onset cases having early onset. V30M mutation does not explain alone the symptoms and AO variability of the disease observed in the same family. Our aim in this study was to identify genetic factors associated with AO variability and reduced penetrance which can have important clinical implications. To accomplish this we genotyped 230 individuals, using a directautomated sequencing approach in order to identify possible genetic modifiers within the TTR locus. After genotyping, we assessed a putative association of the SNPs found with AO and an intensive in silico analysis was performed in order to understand a possible regulation of gene expression. Although we did not find any significant association between SNPs and AO, we found very interesting and unreported results in the in silico analysis since we observed some alterations in the mechanism of splicing, transcription factors binding and miRNAs binding. All of these mechanisms when altered can lead to dysregulation of gene expression, which can have an impact in AO and phenotypic variability. These putative mechanisms of regulation of gene expression within the TTR gene could be used in the future as potential therapeutical targets, and could improve genetic counselling and follow-up of mutation carriers.

Cannabinoids impact on pregnancy: effects in trophoblast cells

Cannabinoids (CBs) can be classified as: phytocannabinoids, the constituents of the Cannabis sativa plant; synthetic cannabinoids lab-synthesized and the endocannabinoids that are endogenous lipid mediators. Cannabinoid compounds activate cannabinoid receptors – CB1 and CB2. The most prevalent psychoactive phytocannabinoid is Δ9tetrahydrocannabinol (THC), but more than 60 different CBs were already identified in the plant. The best characterized endocannabinoids (eCBs) are anandamide (AEA) and 2arachidonoylglycerol (2-AG), that are involved in several physiological processes including synaptic plasticity, pain modulation, energy homeostasis and reproduction. On the other hand, some synthetic cannabinoids that were initially designed for medical research, are now used as drugs of abuse. During the period of placental development, highly dynamic processes of remodeling occur, involving proliferation, apoptosis, differentiation and invasion of trophoblasts. It is known that a tight control of eCBs levels is required for normal pregnancy progression and that eCBs are involved in trophoblast cells turnover. Therefore, by sharing activation of the same receptors, exposure to exocannabinoids either by recreational or medicinal use may lead to alterations in the eCBs levels and in the endocannabinoid system homeostasis In this work, it was studied the impact of CBs in BeWo trophoblastic cells and in primary cultures of human cytotrophoblasts. Cells were treated for 24 hours with different concentrations of THC, the synthetic cannabinoid WIN‐55,212 (WIN) and 2-AG. Treatment with THC did not affect BeWo cells viability while WIN and 2-AG caused a dose-dependent viability loss. Morphological studies together with biochemical markers indicate that 2-AG is able to induce apoptosis in cytotrophoblasts. On the other hand, morphological studies after acridine orange staining suggest that autophagy may take part in WIN-induced loss of cell viability. All cannabinoids caused a decrease in mitochondrial membrane potential (Δψm) but only 2-AG led to ROS/RNS generation, though no changes in glutathione levels were observed. In addition, ER-stress may be involved in the 2-AG induced-oxidative stress, as preliminary results point to an increase in CCAAT-enhancer-binding protein homologous protein (CHOP) expression. Besides the decrease in cell viability, alterations in cell cycle progression were observed. WIN treatment induced a cell cycle arrest in G0/G1 phase, whereas 2-AG induced a cell cycle arrest in G2/M phase. Here it is reinforced the relevance of cannabinoid signaling in fundamental processes of cell proliferation and cell death in trophoblast cells. Since cannabis-based drugs are the most consumed illicit drugs worldwide and some of the most consumed recreational drugs by pregnant women, this study may contribute to the understanding of the impact of such substances in human reproduction.

Abordagem da temática das plantas num contexto EDS orientado para o pensamento crítico no 1º CEB

Numa sociedade ocidental que se apresenta em constante mutação é impossível dominar todo o conhecimento, daí que haja a necessidade de se dotar os jovens alunos de ferramentas que lhes permitam enfrentar a incerteza do futuro. Uma ferramenta imprescindível para os ajudar a ultrapassar dificuldades com que se confrontem são as suas capacidades de Pensamento Crítico (PC), porquanto podem concorrer para tomarem decisões mais racionais. Neste sentido, este estudo, desenvolvido numa turma de 2.º ano do 1.º CEB com 18 alunos, tem como finalidade desenvolver (adaptar, conceber, produzir, implementar e avaliar) atividades promotoras de PC num contexto de Educação para o Desenvolvimento Sustentável (EDS), tendo por base a temática das plantas. Decorrente da finalidade, formularam-se as questões de investigação: Qual o contributo das atividades promotoras de PC na promoção deste tipo de pensamento dos alunos? Qual o contributo das atividades promotoras de PC para a construção/mobilização de conhecimentos dos alunos? Qual a opinião dos alunos acerca das sessões que envolveram atividades promotoras de PC? Quais as representações dos alunos acerca das atividades promotoras de PC? Neste estudo, optou-se por uma metodologia orientada para a prática, assente num plano de investigação-ação. Os alunos realizaram atividades promotoras de PC, num contexto EDS, relacionadas com a temática das plantas abordada na área disciplinar de Estudo do Meio. Recolheram-se dados através de vários instrumentos no âmbito de diferentes técnicas de recolha de dados, incluindo um instrumento de análise das produções dos alunos. Na análise de dados, a técnica privilegiada foi a análise de conteúdo. Os resultados obtidos sugerem que as atividades promotores de PC desenvolvidas no âmbito do estudo contribuíram para a mobilização/desenvolvimento de capacidades de PC dos alunos e para a mobilização/construção de conhecimentos científicos. Na opinião dos alunos, as aulas em que se implementaram atividades promotoras de PC foram mais motivadoras e mais produtivas que as outras aulas de Estudo do Meio. Pela perspetiva dos alunos, as sessões em que realizaram as atividades promotoras de PC contribuíram para a mobilização/desenvolvimento de capacidades de PC e para a mobilização/construção de conhecimentos científicos.