Bridging metabolomics with mathematical tools to study virus-host interactions

The increasing demand of baculovirus- based biopharmaceuticals raises the interest in developing high-titer production processes. Baculovirus infection boosts the host biosynthetic activity towards the production of viral components and the recombinant protein of interest, hyper-productive phenotypes being the result of a successful adaptation of the cellular network to that scenario. Thus, the comprehensive knowledge of the host cell metabolism, mainly in the post-infection phase, can provide clues on the metabolic adaptations that occur and, ultimately, aid bioprocess engineers in developing rational and targeted optimization strategies.(...

Enhancing enveloped viral particles production by targeted supplementation design: Releasing bottlenecks in IC-BEVS

The increasing demand of Insect Cell-Baculovirus Expression Vector System (IC-BEVS) based biopharmaceuticals raises the interest in developing high-titer production processes. Previously, we addressed the impact of the baculovirus infection on the physiology of High Five and Sf9 host cell lines, stressing out key cellular features that support higher productivities. This information was applied to design tailored supplementation schemes to boost IC-BEVS production yields of three targets with increasing complexity: recombinant influenza neuraminidase (rNeur); enveloped influenza VLPs (Inf-VLP) and the baculovirus itself (BV). Higher rNeur productivities were achieved when supplementing High Five cultures with cholesterol. For Sf9 cells, GSH, antioxidants combined with polyamines and cholesterol yielded the best outputs during Inf-VLPs and infectious BVs production. The results also show that the viral load influence the cellular responsiveness to the supplements, with lower MOIs retrieving higher improvements of IC-BEVS specific productivity. The careful selection of the MOI in a batch infection process, along with the supplementation of culture medium with compounds altering cellular redox state and cholesterol, yielded an improvement of the systems’ specific productivity up to 6 fold. The correlation between systems productivity, host cell line and target product was extensively analyzed. The resulting implications for the development of rational strategies for increased productivity are discussed

Redes de apoio social: um estudo de caso sobre pessoas submetidas a artroplastia da anca

A presente proposta de investigação tem como temática principal a análise de redes sociais, mais precisamente, as redes de apoio social na prestação de cuidados de saúde. O objectivo será conhecer as dinâmicas/lógicas da rede de apoio social dos doentes submetidos a artroplastia da anca e investigar as regularidades. Conhecer as dimensões da rede, tais como o tamanho e a densidade; estrutura da rede, tendo como exemplo a medida de centralidade identificar dinâmicas na prestação do apoio social, estabelecendo comparações entre a rede de cada indivíduo. E, por último, identificar dinâmicas na relação hospital - rede de apoio social do doente (família, amigos, vizinhos, outras solidariedades). A pertinência do estudo prende-se com o fenómeno social do envelhecimento da população e o que isso acarreta em termos de co morbilidades. O diagnóstico desta situação poderá revelar pistas sobre como contribuir para a acção, no que toca à apresentação de propostas de intervenção socio-organizacional; Abstract: The main focus of this research proposal is the analysis of the social networks, specifically those concerning social support in the area of health care. The objective is to ascertain the dynamics/logic of the social support network for patients who have undergone hip replacement surgery and investigate the regularities. To learn the dimension of the network, such as its size and concentration; the strength of the network, to identify dynamics in the supply of social support, taking as example the measure of centrality to establish comparisons between the network of each individual. Lastly, to identify dynamics in the relationship hospital-patient's social support network (family, friends, neighbors, other groups). The relevance of the study relates to the social phenomenon of an aging population and what that entails in terms of co-morbidities. The diagnosis of this could reveal clues about how to contribute to the action, with regard to proposals for socio-organizational intervention

U.dream goes to market: an analysis of the financial impacts of commercialising u.purpose & crescer com consciência and project wrap-up

U.Dream is a social enterprise targeted at developing social leadership skills. It recently launched U.Purpose (service) and Crescer com Consciência (product) aiming to ensure future financial sustainability. However, these were launched without conducting a thorough market research and clear guiding strategy. The focus of this project is to assess the attractiveness of the CSR and children’s book markets and U.Dream’s current strategy, providing a strategic revision. From a financial standpoint, U.Purpose has the highest potential, with evident future opportunities. Both projects showcase potential to financially sustain U.Dream, namely through diversification benefits and a strong alignment with the social value delivered

Estudo do efeito da densidade celular na interacção baculovírus - célula de insecto: perspectiva metabólica

Tese de mestrado, Biologia (Microbiologia Aplicada), Universidade de Lisboa, Faculdade de Ciências, 2009O Sistema de Expressão Baculovírus-Célula de Insecto (BEVS-IC) apresenta um grande potencial na indústria biotecnológica e farmacêutica. As suas aplicações são vastas passando pela produção de proteínas recombinantes, vacinas e VLPs e, mais recentemente, vectores para terapia génica. Não obstante, o nível de conhecimento fundamental sobre este sistema é ainda reduzido. Neste trabalho, propusemo-nos a potenciar a capacidade produtiva do sistema BEVS-IC, explorando as alterações metabólicas que decorrem da interacção vírus-célula no cenário pós-infecção. Foram investigados os passos limitantes do metabolismo celular e respectivas enzimas reguladoras. Os resultados obtidos foram complementados com dados de análise de fluxo metabólico, numa abordagem integrativa de Biologia de Sistemas. Dois pontos-chave no controlo do metabolismo oxidativo foram identificados: a regulação do fluxo glicolítico é exercida pela hexokinase, enquanto que a isocitrato desidrogenase é responsável pela actividade do ciclo dos ATCs. Estas enzimas foram portanto identificadas como alvos promissores para engenharia metabólica. A infecção com baculovírus provocou um aumento na actividade da enzima glutamato desidrogenase, estando associado à manutenção do status energético celular e assegurando assim melhores condições para a replicação viral. Este resultado evidencia a capacidade de manipulação da maquinaria celular e utilização o metabolismo celular por parte do vírus. Este trabalho permitiu-nos identificar os pontos-chave de controlo do metabolismo de células Sf9. De um ponto de vista fundamental, foram estudados e racionalizados os cenários metabólicos mais marcantes que se estabelecem no período pós-infecção, decorrentes da interacção vírus-célula. De um ponto de vista tecnológico, os resultados obtidos servem agora de plataforma para o desenvolvimento de novas estratégias biomoleculares e de bioprocesso em vista à optimização da produtividade do sistema.The Insect Cells-Baculovirus Vector Expression System (IC-BEVS) has a great potential in the biotechnological and pharmaceutical industry. It has been recognized regarding its numerous applications, such as the production of heterologous proteins, vaccines and virus like particles (VLPs) and, more recently, vectors for gene therapy. Nevertheless, a more fundamental understanding of this system is still needed. In the work herein presented we intended to improve the productivity of the IC-BEVS, by exploiting the major metabolic alterations that take place during the post-infection scenario as the result of the interactions between the host cell and the virus itself. Within this scope, the main metabolic pathways and the flux controlling enzymes involved were investigated. The results obtained were complemented with metabolic flux analysis data, with the main aim of integrating them in a Systems Biology perspective. Two key-points regarding the oxidative metabolism were identified: hexokinase is responsible for the regulation of the glycolytic flux, and isocitrate dehydrogenase limits the flux through the TCA cycle. Therefore both enzymes were selected as promising targets for metabolic engineering. The activities of glutamate dehydrogenases increased after infection with baculovirus, and this happened within the maintenance of the cellular energetic state to guarantee better conditions for viral replication. This result demonstrates the capacity of the virus to manipulate the cellular metabolism for its own profit, providing better conditions for viral replication. This work allowed us to unveil the metabolic regulatory key-points of Sf9 cells. From a fundamental point of view, the most important metabolic scenarios that occur during the post-infection period were addressed. From a technological point of view, the results herein presented can be exploited for further development of new strategies in terms of bioprocess engineering to attain the ultimate goal: improvement of systems productivity

Metabolic drivers of IC-BEVS productivity: Tackling the production of enveloped viral particles

The Insect Cell-Baculovirus Expression System (IC-BEVS) has a major track record for the production of recombinant proteins and vaccines. Although its widespread use, the physiological aspects that contribute to systems productivity are still to be fully disclosed. In the present work, the metabolic features of the two main insect host cell lines, Sf9 and High Five, were analyzed during cellular growth and after baculovirus infection for the production of enveloped Influenza VLPs (Inf-VLPs). The gathered data were contextualized in a metabolic network representative of central carbon and nitrogen metabolism. Metabolic Flux Analysis (MFA) was performed to have a quantitative overview of the cellular fluxome dynamics that followed infection. In addition, the main carbon sources that contributed most to flux activity were identified. The impact of baculovirus infection on the physiology of High Five and Sf9 host cell lines was assessed by metabolomics, aiming at the identification of metabolic markers of productivity. The information herein generated was used to design tailored supplementation schemes that could boost IC-BEVS production yields of two enveloped viral particles: influenza VLPs (Inf-VLP) as a vaccine candidate and the recombinant baculovirus (BV). The strong correlation observed between the metabolic state of the host cell and baculovirus infection highlights the capacity of this virus to act as a metabolic engineer, re-directing the cellular fluxome to support virus replication and production. The results also show that the viral load influence the cellular responsiveness to the supplements, with lower MOIs retrieving higher improvements in specific productivity. The careful selection of the MOI, along with the supplementation of culture medium with compounds altering cellular redox state and cholesterol metabolism, yielded a 6-fold improvement of specific productivity. These results pave the way to deepen our knowledge on the relationship between host cell and virus, contributing to the disclosure of the metabolic determinants that contribute to productivity

Verificação da previsão meteorológica obtida com o modelo WRF no contexto de aproveitamento eólico

O presente relatório resulta do estágio curricular desenvolvido na empresa Smartwatt, particularmente no vetor Intelligence. O estudo realizado visa a verificação da qualidade das previsões meteorológicas do modelo WRF, relativamente à variável meteorológica intensidade do vento a 100 metros, efetuada operacionalmente na empresa, para o contexto de aproveitamento eólico. Neste caso de estudo, foram realizadas múltiplas simulações para se verificar qual o melhor conjunto de parametrizações a usar no modelo de modo a melhorar o desempenho deste sistema de previsão efetuado na empresa. Esta verificação foi realizada a partir de uma comparação ideal entre os parâmetros simulados e os observados de intensidade de vento para períodos e localizações específicas nas diferentes simulações do modelo. Para este exercício de comparação aferiu-se e quantificou-se um conjunto de métricas de erro, diretamente simples e diretas, que permitiram avaliar os erros associados a cada simulação efetuada. Os resultados obtidos a partir destas análises permitiram retirar algumas conclusões no que diz respeito ao desempenho do modelo WRF para a previsão da intensidade do vento a 100 metros e, consequentemente, tomar decisões relativamente ao que pode ser empregue para melhorar ou tornar mais eficiente o sistema de previsão WRF efetuado operacionalmente na empresa.This report follows the curricular internship at the company Smartwatt, more particularly the study conducted on the vector Intelligence. This study aims to verify the quality of the weather forecasts as described by the WRF model, concerning the meteorological variable wind intensity to 100 meters, conducted operationally in the company, as an input towards wind energy. In this case study, multiple simulations were carried out to determine the best set of settings to use in order to improve the performance of this system of prediction made in the company. This verification was performed from a comparison between the simulated and observed parameters of wind intensity for periods and specific locations in the various model simulations. This comparison exercise showed and quantified a set of error metrics directly simple and direct that allowed to evaluate the errors associated with each simulation. The results obtained from these analyses made it possible to draw some conclusions regarding the performance of the WRF model for the prediction of wind strength at the 100 meters and, consequently, to reach some conclusions on how to enhance and increase the efficacy of the company’s forecast system WRF.Mestrado em Engenharia do Ambient

Introductory programming in higher education: A systematic literature review

A systematic literature review was performed on 33 papers obtained from the ACM, IEEE and Sciencedirect databases, in order to understand in depth, the introductory programming discipline (CS1) in higher education. Recently published works have been covered, providing an overview of the teaching-learning process of introductory programming and enabling to find out whether the research developed by universities worldwide is in line with the proposals made by ACM/IEEE-CS group for computer courses, regarding the transition to the competency-based model. The results show that the new techniques/technologies currently used in software development, as an example of agile methodology, has influenced the teaching-learning process of CS1 together with methods such as visual programming and e-learning. The analyzed papers discuss the importance of developing not only technical, but also social skills, corroborating that methodologies used in introductory programming courses need to focus on preparing students for an increasingly competitive market, associating new skills with technical aspects.This work is supported by CIEd – Research Centre on Education, Institute of Education, University of Minh