Browsing by Issue Date, starting with "2023-10-30"
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- Estudo da influência do Bisfenol A nas células musculares lisas vasculares da artéria cerebral média e a possível influência no acidente vascular cerebralPublication . Costa, Henrique Elói Albino da; Oliveira, Maria Elisa Cairrão RodriguesO Bisfenol A (BPA) é um disruptor endócrino amplamente produzido mundialmente. O BPA está presente em plásticos policarbonatos e resinas epóxi, usados na produção de biberões para bebés, recipientes de armazenamento de alimentos e bebidas, materiais industriais, médicos, entre outros. O BPA possui propriedades estrogénicas e androgénicas, tendo sido encontrado a sua acumulação em tecidos cerebrais, sendo posteriormente associado a efeitos prejudiciais a nível neurológico e vascular, nomeadamente na Acidente Vascular Cerebral (AVC). Este tipo de lesão cerebral afeta consideravelmente as artérias e vasos sanguíneos, nomeadamente a artéria cerebral média (MCA), prejudicando a viabilidade das células musculares lisas (SMC). As SMC desempenham um papel fundamental na regulação do tónus vascular e consequentemente na integridade vascular e homeostasia cerebral. Neste sentido, este trabalho pretende analisar os mecanismos pelos quais o BPA altera na função contrátil das células musculares lisas da artéria cerebral média (SMC-MCA) de rato. Deste modo, foram isolados explantes da MCA de ratos Wistar e aderidos a placas de culturas previamente revestidas com colagénio, de modo a obter culturas puras de SMC. A partir destas culturas foram realizados três ensaios. O ensaio de MTT, de modo a testar a viabilidade celular, proliferação e citotoxicidade das SMC-MCA em resposta ao BPA. O ensaio de contractilidade por Planar Cell Surface Area, para analisar a resposta vasoativa das SMC-MCA em resposta ao agente contrátil, noradrenalina (NA), e ao agente relaxante, nitroprussiato de sódio (SNP). E em último por Real time-PCR, para avaliar a expressão das subunidades de canais iónicos e proteínas envolvidas na resposta contrátil da MCA. Os resultados deste estudo demonstraram que a exposição a BPA pode modificar a resposta vasorelaxante das SMC-MCA. Os efeitos genómicos dependeram da concentração de incubação com o BPA, induzindo uma alteração da resposta contrátil, devido a alterações na expressão da subunidade a do BKCa 1.1, bem como a proteína sGC. Em suma, estas observações sugerem que a exposição ao BPA altera a homeostasia vascular da SMC-MCA, podendo estar, por isso, implicado na ocorrência de AVC isquémico, evidenciando a necessidade urgente de compreender esta ligação e os mecanismos associados, sendo por isso necessários mais estudos.
- Formulation and characterization of taxifolin delivery systems as an anti-Human Papillomavirus drugPublication . Neto, Miguel Leite; Sousa, Ângela Maria Almeida de; Costa, Diana Rita BarataCancer is a leading cause of mortality worldwide, with over 19.3 million cases reported in 2020, resulting in approximately 10 million deaths. Various risk factors have been associated with the development of cancer, including age, genetic predisposition, ethnicity, environmental exposure, lifestyle, and infections caused by bacteria, parasites, or viruses. Human papillomavirus (HPV) is the primary causative agent of cervical cancer, which is the fourth most common cancer in women worldwide, responsible for over 340,000 deaths in 2020. This cancer is characterized by the overexpression of oncoproteins E6 and E7, which disrupt cell cycle regulation and proliferation, compromising the functions of tumor suppressor proteins p53 and pRb, respectively. Despite the availability of preventive HPV vaccines, their administration is not universally carried out. Consequently, efforts continue to develop effective therapies for this type of cancer to achieve a successful cure. In this regard, flavonoids have demonstrated remarkable therapeutic potential, offering an accessible and effective approach, allowing their use in less developed countries with limited healthcare resources. Taxifolin is a flavonoid with several anticancer properties, having shown particular relevance in cervical cancer therapy due to its ability to enhance the inhibition of oncoprotein E6, thereby increasing p53 expression and inducing apoptosis. However, the use of taxifolin is limited due to its very low aqueous solubility and low stability. Consequently, its poor bioavailability restricts its applications in cancer therapies. Thus, this master's dissertation aims to develop delivery systems for the encapsulation of taxifolin to improve its bioavailability and effectiveness in HPV-positive cells. To achieve this, various delivery systems consisting of chitosan, gelatin, and taxifolin were formulated. Initially, several ratios of systems composed solely of low molecular weight chitosan and gellan gum were tested. The most favourable ratio exhibited a size of 238.07 ± 31.18 nm, a PdI of 0.29 ± 0.08, and a zeta potential of +22.56 ± 2.83 mV and was chosen to proceed with the studies and attempt taxifolin encapsulation. For the incorporation of taxifolin into the delivery systems, various types of chitosan (5 kDa, low molecular weight, and high molecular weight) were tested at various taxifolin ratios. Systems composed of low molecular weight chitosan (LMW CH/GG/TAX) had a size of 276.23 ± 30.68 nm, a PdI of 0.36 ± 0.06, and a zeta potential of +30.80 ± 5.66 mV. Delivery systems formed by high molecular weight chitosan (HMW CH/GG/TAX) showed a size of 272.82 ± 53.58 nm, a PdI of 0.33 ± 0.10, and a zeta potential of +23.59 ± 5.94 mV, while systems composed of 5 kDa chitosan (5 kDa CH/GG/TAX) had a size of 249.00 ± 12.58 nm, a PdI of 0.35 ± 0.10, and a zeta potential of +17.35 ± 5.64 mV. Regarding encapsulation efficiency, LMW CH/GG/TAX exhibited an efficiency of 65%, while the remaining systems encapsulated only 55%. Scanning electron microscopy (SEM) was also performed for all formulated samples, and it was possible to observe that all systems exhibited a spherical and uniform morphology. In addition, Fourier-transform infrared spectroscopy (FTIR) and UV/vis spectroscopy were conducted to verify the presence and interactions of compounds in the delivery systems. Finally, in vitro release assays were performed at two different pH levels, one representative of the tumor microenvironment (pH 5.8) and one representative of physiological pH (pH 7.4). The results showed that HMW CH/GG/TAX systems released approximately 45% and 80% of taxifolin at pH 5.8 and pH 7.4, respectively. LMW CH/GG/TAX systems released 25% at pH 7.4 and approximately 70% at pH 5.8. The 5 kDa CH/GG/TAX systems released between 20% and 40% at the two different pH levels. Based on all the gathered information regarding the characterization of the delivery systems, only the most favourable formulation, LMW CH/GG/TAX, proceeded to cellular assays, where internalization assays were conducted on healthy cells and HPV-positive cells. The results demonstrated that the delivery systems were capable of internalizing into both cell lines; however, it is expected that they will solely exert an effect on HPV-positive cells due to their specific action against oncoprotein E6.
- Impact of the endocrine disruptor methoxychlor on the survival and glycolytic metabolism of Sertoli cellsPublication . Bordadágua, Mariana Costa; Correia, Sara Carina de Lima; Socorro, Sílvia Cristina da Cruz Marques; Feijó, Mariana PombalEndocrine-disrupting chemicals (EDCs) are manufactured or natural compounds that are capable of interfering with normal endocrine function and, consequently, with the body homeostasis. The EDC methoxychlor (MXC) is an organochloride pesticide and a xenoestrogen with moderate persistence in the environment, capable of adversely affecting spermatogenesis, deregulating the development of germ cells, and interfering with sperm function. However, a knowledge gap exists regarding the impact of MXC on other testicular cell types, namely, Sertoli cells (SCs), the somatic testicular cells within the seminiferous tubules. SCs physically and metabolically sustain germ cell survival, and any dysregulation induced by EDC action affects the spermatogenic output. The present dissertation investigated the effect of the EDC MXC on the survival and glycolytic metabolism of SCs, as well as their fatty acid uptake. For this purpose, primary SCs were isolated from 4-month-old rats and maintained in culture for 48 h in the presence (25 and 100 µM) or in the absence (control) of MXC. SCs’ survival and apoptotic rate were evaluated by trypan blue exclusion and spectrophotometrically, respectively. Glucose and fatty acid consumption, lactate production, and the expression of key regulators of the glycolytic pathway were measured by spectrophotometric assays and Western blot analyses. The results obtained demonstrated that both 25 and 100 µM MXC decreased the viability of SCs, accompanied by increased caspase-3-like activity, in a concentrationdependent manner. Enhanced glucose consumption and lactate production were also observed upon MXC exposure (25 and 100 µM). Given these results, we focused our metabolic intervenients analysis on the lowest MXC concentration. These observations were underpinned by the augmented expression levels of glucose transporter 3, and phosphofructokinase 1, and expression levels and activity of lactate dehydrogenase. In addition, 25 µM MXC increased fatty acid consumption by SCs. These findings demonstrated that MXC has an impact on the metabolic characteristics of SCs, which is a novel perspective on the effects of EDCs leading to male infertility. Further studies should be performed to identify other dysregulated molecules or susceptible metabolic pathways, which could help develop therapeutical and/or preventive strategies for male infertility.
- O paradoxo atual dos humanos a se tornarem máquinas versus as máquinas a se tornarem humanasPublication . Aragão, Marcia Dayane Maciel; Souza, José António Menezes Felippe de; Magalhães, Bianca GonçalvesEstamos, cada vez mais, a usar dispositivos tecnológicos para realizar tarefas e obter informações. Graças à tecnologia estamos a assistir a um aumento da saúde e bem-estar, com a criação de dispositivos como os já comuns rastreadores de exercícios físicos que ajudam as pessoas a monitorar as suas atividades diárias e manter-se saudáveis. A tecnologia também nos permite aproveitar o que a vida tem para nos oferecer, proporcionando-nos a possibilidade de viajar a qualquer lugar do mundo. A tecnologia veio a tornar a vida mais simples, tornando a comunicação mais acessível e permitindo-nos ficar conectados com o mundo. Enfim, a tecnologia veio para melhorar a qualidade de vida dos seres humanos. Portanto, os pesquisadores estão com desafios de desenvolver máquinas melhores e inteligentes, fazendo com que a interação Humanos-Máquinas fique cada vez mais natural. É evidente que a tecnologia nos permite evoluir, permitindo-nos usufruir de vidas mais longas e saudáveis, aproveitar o que a vida tem para oferecer e melhorar a nossa comunicação entre máquinas e humanos.