Understanding students’ academic achievement in public High School : evidence for Portugal

Dissertation presented as the partial requirement for obtaining a Master's degree in Information Management, specialization in Knowledge Management and Business IntelligenceSeveral papers and studies have been conducted to better understand what are the main factors that influence students’ academic achievement and what measures should be taken to improve it. Therefore, based on 383.560 students’ observations, evaluated on secondary Portuguese public schools in 2014/2015 academic year, the purpose of this study is to provide a new approach to the collected data by using Data Mining predictive models. The results show differences on the academic achievement among females and male students, where females got better academic results. Access to computer and Internet found to be powerful tools in education that students can explore to their benefit and show to have a positive influence on academic results. Students benefiting from financial social support prove to have a lower performance in academic achievement. Results also point to the fact that the number of reproves still has a great negative impact on students’ academic achievement. This is one of the first studies to the best of the authors knowledge to employ analytic techniques on such a large dataset on the context of academic achievement

Clinically Expired Platelet Concentrates as a Source of Extracellular Vesicles for Targeted Anti-Cancer Drug Delivery

Funding Information: This research was funded by the Fundação para a Ciência e Tecnologia funded projects PEFPlateletValue (PTDC/BTM-ORG/32187/2017) and EXCELERATE (2022.10467.PTDC), iNOVA4Health-(UIDB/04462/2020 and UIDP/04462/2020), a program financially supported by FCT/Ministério da Ciência, Tecnologia e Ensino Superior, through national funds. The Associate Laboratory LS4FUTURE (LA/P/0087/2020) is also acknowledge. Publisher Copyright: © 2023 by the authors.The short shelf life of platelet concentrates (PC) of up to 5–7 days leads to higher wastage due to expiry. To address this massive financial burden on the healthcare system, alternative applications for expired PC have emerged in recent years. Engineered nanocarriers functionalized with platelet membranes have shown excellent targeting abilities for tumor cells owing to their platelet membrane proteins. Nevertheless, synthetic drug delivery strategies have significant drawbacks that platelet-derived extracellular vesicles (pEV) can overcome. We investigated, for the first time, the use of pEV as a carrier of the anti-breast cancer drug paclitaxel, considering it as an appealing alternative to improve the therapeutic potential of expired PC. The pEV released during PC storage showed a typical EV size distribution profile (100–300 nm) with a cup-shaped morphology. Paclitaxel-loaded pEV showed significant anti-cancer effects in vitro, as demonstrated by their anti-migratory (>30%), anti-angiogenic (>30%), and anti-invasive (>70%) properties in distinct cells found in the breast tumor microenvironment. We provide evidence for a novel application for expired PC by suggesting that the field of tumor treatment research may be broadened by the use of natural carriers.publishersversionpublishe

Towards large-scale production of human-induced pluripotent stem cell-derived extracellular vesicles in stirred-tank bioreactors

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Are cellulose nanofibrils safe for biomedical applications?

Nanomedicine is based on the application of nanotechnologies into the medical field to advance and improve diagnostics, prevention and treatment of human disease. While its expansion has been enormous in the last years, its progress must go hand in hand with nanosafety, i.e., with the safety evaluation of nanomaterials in an early phase of its development or application into a product. Cellulose appears as a natural and readily available material, which fits within the supply-demand chain: ecological, abundant and low cost. Particularly, cellulose nanofibrils (CNF) show great mechanical strength and high water-uptake capability and have the ability to form translucent structures with high elasticity and selective permeability, which make them attractive e.g., as constituents of surgical dressings and membranes for bone regeneration. Bacterial nanocellulose is already being used, but CNF produced from plants are also finding potential to be applied in tissue engineering and regenerative medicine. However, CNF may bring more toxicological concerns than the bacterial type, due to impurities associated with the chemical and mechanical processes used to produce them or due to their different physicochemical properties that may underlie unforeseen biological effects. The main objective of this work was to evaluate the safety of two different CNFs, through the analysis of their cytotoxic, genotoxic and epigenetic effects in human osteoblasts. The CNFs were obtained from the same raw material – industrial bleached Eucalyptus globulus kraft pulp - by two different methods: TEMPO-mediated oxidation and enzymatic hydrolisis. The physicochemical properties of the CNF gels obtained, including fibrillation yield, degree of polymerization and size were evaluated. The CNF cytotoxicity was assessed by the MTT assay and the genotoxicity by the cytokinesis-block micronucleus assay; their epigenetic effects were evaluated through gene expression analysis of the DNA methytransferases genes, DNMT1 and DMNT3b, responsible for the cellular methylation pattern, using qRT-PCR. The results obtained for the several endpoints were integrated in order to contribute to the characterization of the potential toxic effects of these new CNF in an early phase of their lifecycle. This knowledge will be relevant to decide whether these CNF may be further developed for applications in the nanomedicine field, or shall be modified to give rise to safer CNF.ToxApp4NanoCELFI , 2018-2021 - PTDC/SAU-PUB/32587/2017; ToxOmics - Centro de Investigação em Toxicogenómica e Saúde Humana - UID/BIM/00009/2013N/

hiPSC and hiPSC-cardiomyocytes are alternative EV biofactories for cardiac regeneration

In cardiac regenerative medicine, there is a growing interest in using EV as cell-mimetic therapeutics due to their potential superior efficacy and overall advantages over cell transplantation: i) absence of oncogenic risk, ii) low immunogenicity [1], iii) easier large-scale manufacturing and iv) consistent product profile. However, most studies have focused particularly on the potential of either mesenchymal or cardiac progenitor cell-derived EV to promote cardiac repair [2]. Here, we study the potential of human induced pluripotent stem cells (hiPSC) and hiPSC-derived cardiomyocytes (hiPSC-CM) as alternative cell factories for the production of a high yield of therapeutic EV for cardiac regeneration. Due to their high self-renewal ability and capacity to differentiate into functional cardiomyocytes, these cells can provide an unlimited source of EV for application in cardiac regeneration. We generated and characterized EV derived from key stages of hiPSC-CM differentiation and maturation, i.e. from hiPSC (hiPSC-EV), cardiac progenitors (CPC-EV), immature (CMi-EV) and mature (CMm-EV) cardiomyocytes, with the goal of studying their potential role as therapeutics, and whether their yield and function was influenced by the state of their parent cell. Two hiPSC lines were differentiated into hiPSC-CM and cultured as 3D spheroids in a fatty acid supplemented medium to improve CM maturation [3,4]. EV isolation was performed based on density separation on an iodixanol discontinuous gradient, and EV were characterized in terms of particle size and particle size distribution, presence of EV-specific markers, and imaging through transmission electron microscopy. Functional studies were performed using human umbilical vein endothelial cells (HUVECs) to evaluate EV-uptake, migration and angiogenesis. EV yield varied along CM differentiation stages, with a minimum for CPC, for both cell lines. Bioactivity assays with HUVECs showed that uptake of PKH26-labelled EV could be blocked by dynasore, an inhibitor of dynamin-2, a GTPase that plays a crucial role in clathrin and caveolin-dependent endocytosis. Increased migration was observed in HUVECs treated with hiPSC, CPC and CM-derived EV (92.25 ± 14.69% wound closure at 24h for hiPSC, 77.13 ± 13.64 % for CPC, 74.71 ± 19.86% for CMi, 69.2 ± 19.12% for CMm versus 45.65 ± 7.26% for control), but angiogenic properties were found only for hiPSC-EV (fold change of 11.2 ± 4.59 in total segment length vs. control, p\u3c0.001). Current efforts towards the characterization of EV small RNA cargo aim at understanding the correlation between cargo composition and in vitro activity, to identify the optimal cell factory for scalable therapeutic EV production. Funding: FCT PhD fellowship PD/BD/139078/2018; IC&TD Projects MetaCardio” (PTDC/BTM-SAL/32566/2017) and NETDIAMOND (SAICTPAC/0047/2015), and iNOVA4Health Research Unit (UIDB/04462/2020). 1. Zhu, X., et al. Comprehensive toxicity and immunogenicity studies reveal minimal effects in mice following sustained dosing of extracellular vesicles derived from HEK293T cells. Journal of Extracellular Vesicles 6, 1, 2017. 2. El Harane, et al. Acellular therapeutic approach for heart failure: in vitro production of extracellular vesicles from human cardiovascular progenitors. European Heart Journal 39, 20, 2018. 3. Correia, C., et al. Distinct carbon sources affect structural and functional maturation of cardiomyocytes derived from human pluripotent stem cells. Scientific reports 7, 1, 2017. 4. Correia, C., et al. 3D aggregate culture improves metabolic maturation of human pluripotent stem cell derived cardiomyocytes. Biotechnology and Bioengineering 115, 3, 2018

Investigation of potential respiratory adverse effects of micro/nanofibrillated cellulose and cellulose nanocrystals using human lung cell lines.

Abstract publicado em: Environ Mol Mutagen. 2022 Aug;63(Suppl 1):72-73. (Abstracts from the 13th International Conference on Environmental Mutagens and 53rd Annual Meeting of the Environmental Mutagenesis and Genomics Society). https://onlinelibrary.wiley.com/toc/10982280/2022/63/S1Micro/nanofibrillated (CMF/CNF) and nanocrystalline (CNC) celluloses are innovative materials with enormous potential for industrial and biomedical applications. Their expanding production/application urges the investigation of their safety for human health. This study aimed at investigating the potential respiratory outcomes of two CMF/CNF and one CNC produced from bleached Eucalyptus globulus kraft pulp using human alveolar epithelial (A549) cells grown in monoculture or co-cultured with THP-1 monocyte-derived macrophages, by assessing their cellular uptake, cytotoxic, immunotoxic, genotoxic, and epigenetic effects. The nanocelluloses were characterized for their physicochemical properties: CMF displays a low percentage of nanofibrils while CNF comprises 100% fibrils with a diameter (D) circa 11 nm; CNC consists of nanorods with D of 4-5 nm and aspect ratio around 42. TEM analysis evidenced that CMF and CNF were internalised into A549 cells whereas CNC were not. Neither cytotoxicity (colorimetric and clonogenic assays) nor ROS induction was observed for any of the nanocelluloses. CMF caused chromosomal alterations (in vitro micronucleus assay) in A549 cells while negative results were obtained in co-culture and for the other micro/nanocelluloses in mono- or co-culture. Results in progress of DNA damage and gene mutation analyses will complement mutagenesis assessment. Additionally, potential inflammatory and epigenetic effects are being evaluated. These results contribute to the weight of evidence of nanocelluloses biological effects and knowledge of the underlying molecular mechanisms. Such information will drive the synthesis of the safest nanocelluloses,thus minimising potential negative impacts of their use on human and environmental health.FCT/MCTES through nacional funds (PTDC/SAU-PUB/32587/2017; UIDP/00009/2020; UIDB/00009/2020; 020.07168.BD)info:eu-repo/semantics/publishedVersio

Nanocellulose toxicity in vitro models: contributing to its safety assessment to human health

As nanoceluloses são nanomateriais inovadores com potencial para uma vasta gama de aplicações industriais e biomédicas. No entanto, a expansão da sua produção tem vindo a suscitar preocupações quanto aos possíveis efeitos, a longo prazo, na saúde humana. Este estudo teve como objetivo avaliar a segurança de algumas nanoceluloses produzidas a partir de matéria-prima nacional, através da caracterização da sua potencial toxicidade em células de mamífero. Para tal, testaram-se duas celuloses nano /microfibrilares (CNF e CMF ) e uma celulose nanocristalina (CNC). Analisou-se a citotoxicidade usando ensaios colorimétricos e o ensaio clonogénico, e a genotoxicidade pelo ensaio do micronúcleo in vitro em células pulmonares de mamífero (A549 e V79 ) e em osteoblastos humanos ( MG-63 ). A indução de espécies reativas de oxigénio (ROS) e a internalização celular foram também estudadas nas células A549. Observou- -se citotoxicidade no ensaio clonogénico, principalmente no caso da CNC, mas não nos restantes ensaios, não havendo também indução de ROS. Todas as nanoceluloses revelaram efeitos genotóxicos nalgumas concentrações, uma vez que induziram micronúcleos e /ou pontes nucleoplásmicas num dos modelos celulares. Para além disso, visualizou-se a internalização da CNF e CMF, mas não da CNC, em células A549. Esta primeira avaliação toxicológica veio contribuir para prevenir a exposição a materiais celulósicos potencialmente perigosos, procurando impulsionar o desenvolvimento de materiais inovadores e mais seguros.Nanocelluloses are innovative nanomaterials with potential for a wide range of industrial and biomedical applications. However, the expansion of its production has raised concerns about their possible long-term effects on human health. This study aimed to evaluate the safety of various nanocelluloses through the characterization of their potential toxicity in mammalian cells. Two samples of nano/microfibrillar celluloses with different pre-treatments (CNF and CMF) and a nanocrystalline cellulose (CNC) were tested. The cytotoxicity of the nanocelluloses was analyzed using colorimetric assays and the clonogenic assay, and genotoxicity by the in vitro micronucleus assay in human alveolar epithelial cells (A549), human osteoblasts (MG-63) and Chinese hamster fibroblasts (V79). Induction of reactive oxygen species (ROS) and cellular internalization were also studied in A549 cells. Cytotoxicity was observed through the clonogenic assay, mainly in the case of CNC, but not in the remaining assays, with no induction of ROS. All nanocelluloses, at some of the concentrations tested, induced micronuclei and/or nucleoplasmic bridges in one of the cellular models. Furthermore, the internalization of CNF and CMF, but not of CNC was visualized in A549 cells. These results aim to contribute to preventing exposure to potentially hazardous cellulosic materials, seeking to boost the development of innovative and safer materials.Projeto ToxApp4NanoCELFI – Uma abordagem de toxicologia preditiva para a caracterização dos potenciais efeitos respiratórios de fibras de nanocelulose funcionalizadas num sistema de co-cultura (PTDC/SAU-PUB/32587/2017).info:eu-repo/semantics/publishedVersio

Deficiência na desidrogenase dos ésteres acil-coenzima a de cadeia média (MCAD): caracterização de uma nova proteína mutante (p.G377V)

Relatório de projecto no âmbito de Bolsa Universidade de Lisboa/Fundação Amadeu Dias (2010/2011). Universidade de Lisboa. Faculdade de Farmáci

Development of gel formulations for control of varroa destructor

Trabalho Final de Mestrado Integrado, Ciências Farmacêuticas, Universidade de Lisboa, Faculdade de Farmácia, 201

Exploring the toxicity of cellulose nanofibrils in a lung epithelial cell line

Nanotechnologies and nanomaterials (NMs) applications have been growing in recent years, bringing benefits to society but raising also some concerns about their safety to human health. Cellulose is a natural material that fits the global trend of sustainability: ecological, low cost, abundant and renewable nature. In particular, cellulose nanofibrils (CNF)1 are forest-derived products with advantageous mechanical, optical and rheological properties, assuming a high industrial potential, e.g., in paper, food, pharmaceutical and biomedical industries. With the innovative applications expanding, CNF synthesis and production has increasing, leading to concerns about occupational exposure, particularly by inhalation, or consumers exposure. The toxicity studies of other NMs, like MWCNT, have had a major impact on the understanding of the nanofibres health effects on humans. MWCNTs have been reported to cause adverse effects in vitro and in vivo, such as DNA damage and oxidative stress2. Because CNF show a high resemblance in terms of aspect ratio to MWCNT, our main focus is to identify if some of the CNF synthesized have a genotoxic or carcinogenic potential. This study aims to assess the safety of two types of CNF produced with different pre-treatments (TEMPO-mediated oxidation and enzymatic hydrolysis) of an industrial bleached Eucalyptus globulus kraft pulp, through the characterization of its cytotoxicity and genotoxicity in human cells. The CNF cytotoxicity was assessed using lung epithelial alveolar (A549) cells by two methodologies, the MTT and the clonogenic assay, whereas the genotoxicity was assessed by the cytokinesis-block micronucleus assay. Dose-range finding experiments were performed using the MTT (24h, 48h and 72h exposure) and the clonogenic (8 days exposure) assays, which revealed that both CNF were not cytotoxic at concentrations between 3,125 and 100 μg/ml. On the contrary, both CNFs were able to increase cell viability at the highest concentrations tested (50 and 100 μg/ml). This effect had been previously observed in the same cell line exposed to CNF produced by TEMPO-mediated oxidation, but at the lowest concentration level3. The potential of the CNF to induce chromosomal alterations, either chromosome breaks or loss is being analysed through the micronucleus assay and the results will be presented. Overall, this study is expected to uncover potential adverse outcomes of CNF to human health, in order to promote the design of safer CNF and CNF-based products that will allow a more sustainable and responsible industrial development. References: 1) Gamelas, J., Pedrosa, J., Lourenço, A., Mutjé, P., González, I., Chinga-Carrasco, G., Singh, G. and Ferreira, P. (2015). On the morphology of cellulose nanofibrils obtained by TEMPO-mediated oxidation and mechanical treatment. Micron, 72, 28-33. 2) Louro, H., Pinhão, M., Santos, J., Tavares, A., Vital, N. and Silva, M. (2016). Evaluation of the cytotoxic and genotoxic effects of benchmark multi-walled carbon nanotubes in relation to their physicochemical properties. Toxicology Letters, 262, 123-134. 3) Ventura, C., Lourenço, A., Sousa-Uva, A., Ferreira, P. and Silva, M. (2018). Evaluating the genotoxicity of cellulose nanofibrils in a co-culture of human lung epithelial cells and monocyte-derived macrophages. Toxicology Letters, 291, 173-183.FCT-MCTES (PTDC/SAU-PUB/32587/2017) through national funds (PIDDAC).info:eu-repo/semantics/publishedVersio