Biossíntese da proteína Catecol-O-metiltransferase membranar humana: optimização com recurso a desenho experimental Plackett-Burman e composto central

Catechol-O-methyltransferase (COMT, EC 2.1.1.6) is an S-adenosyl-L-methionine-dependent methyltransferase enzyme that catalyzes the methylation of catechol substrates (catecholamines, catecholestrogens). Physiologically, it is responsible for the elimination of biologically active or toxic catechols, making it a protein with great clinical relevance as therapeutic target in serious disorders, like schizophrenia and Parkinson´s disease. To fulfill pharmaceuticals requirements, new strategies of optimization and large-scale production of COMT enzyme are crucial. Statistical optimization approaches have demonstrated their enormous value in laboratory and industrial scale, namely in biotechnological production processes, in which an incremental enhancement can be a perpetual improvement. In this work, we aimed the optimization of recombinant human membrane-bound COMT (hMBCOMT) enzymatic activity yields following a statistical optimization as a solving approach. Plackett-Burman design was used as a first optimization step to identify which factors have a significant effect in hMBCOMT productivity and activity levels, and Response Surface Methodology (RSM), by a Central Composite Design (CCD), to optimize the process. We applied Brevibacillus choshinensis cells for the biosynthesis of hMBCOMT and a semi-defined medium for cell growth. This medium was subjected to a first screening using the Plackett–Burman design to evaluate the influence of the culture parameters (chemicals and physicals) in hMBCOMT enzymatic activity levels. Enzymatic activity were measured in a high performance liquid chromatography (HPLC) coupled to a coulochemical detector. Among the eleven variables tested, polypeptone, ammonium sulfate, glucose and temperature were selected owing to their significant effect on human MBCOMT enzymatic activity. The biological human MBCOMT activity obtained with the semi-defined medium in Plackett-Burman design were very promising, while were higher than the obtained with 2SYNm medium, a traditional growth medium for Brevibacillus cells of this work. Typically, we obtained values of 93nmol/h for hMBCOMT total enzymatic activity and 30 nmol/h/mg of specific activity with protein in its native form, without the use of any kind of detergents on protein solubilization step. Based on the results of Plackett–Burman design, a CCD was adopted to define optimal components concentration and temperature in order to maximize our response. The CCD model presented a multiple correlation coefficient value of 0.635 and a significant lack of fit, showing the lack aptness of the model to the process optimization and the failure to attain the optimal concentration of each variable.Catecol-O-metiltransferase (COMT, CE 2.1.1.6) é uma enzima metiltransferase dependente de S-adenosil-L-metionina (SAM) que catalisa a metilação de substratos catecóis (catecolaminas, catecolestrogénios). Fisiologicamente, é responsável pela eliminação de catecóis biologicamente activos ou tóxicos, tornando-a uma proteína de elevado interesse clínico e utilizada como alvo terapêutico em doenças graves, como a esquizofrenia e a doença de Parkinson. Para suprir as necessidades farmacêuticas, novas estratégias de otimização e produção em larga escala desta enzima são fundamentais. Abordagens de otimização estatística têm demonstrado o seu enorme valor à escala laboratorial e industrial, nomeadamente nos processos de produção biotecnológicos, em que um pequeno detalhe melhorado pode significar um grande passo para o sucesso. Neste trabalho, objetivou-se a otimização do nível de atividade enzimática da proteína recombinante COMT, na sua forma membranar, através do recurso a modelos de otimização estatística como uma abordagem resolutiva. Numa primeira fase de otimização e de seleção dos fatores mais significativos para a atividade enzimática da proteína em estudo foi utilizada a técnica de desenho experimental Plackett-Burman. Após esta seleção foi aplicada a Metodologia de Superfície de Resposta (RSM), através de desenho composto central (DCC), para otimização da concentração dos fatores que revelaram ser mais significativos e, consequentemente, do processo. Foi utilizado o sistema de expressão Brevibacillus choshinensis para a biossíntese da proteína membranar COMT e um meio semi-definido para o seu crescimento. Este meio foi submetido a uma primeira triagem através do desenho experimental Plackett-Burman, avaliando-se desta forma a influência dos parâmetros de cultura (produtos químicos e físicos) nos níveis de actividade enzimática da COMT membranar. Os níveis de actividade enzimática foram medidos num sistema de cromatografia líquida de alta eficiência acoplado a um detector amperométrico. Entre as onze variáveis testadas, a polipeptona, sulfato de amónio, glucose e temperatura foram as variáveis selecionadas dado o seu significativo efeito na actividade enzimática da COMT membranar. Os níveis de atividade enzimática obtidos nesta primeira triagem revelaram-se bastante promissores, sendo mais elevados do que os obtidos com o meio 2SYNm, meio de crescimento mais comum para as células usadas neste trabalho. Foram obtidos valores de 93nmol/h para a actividade enzimática total e cerca 30 nmol/h/mg de actividade enzimática específica com a proteína na sua forma nativa, sem o uso de qualquer tipo de detergentes no processo de solubilização. Com base nos resultados do desenho Plackett Burman foi aplicado o desenho Composto Central para a otimização dos quatro fatores em causa a fim de maximizar a nossa resposta. Este apresentou um valor do coeficiente de correlação múltipla de 0,635 e uma falta de ajuste significativa, demonstrando a falta de adequação do modelo para a otimização do processo

Internamentos hospitalares durante a primeira vaga de COVID-19 em Portugal

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Epidemiology of Psoriasis in Portugal: A Population-Based Study

Introduction: Psoriasis is a common, chronic, and inflammatory skin disorder with a high personal, social and economic burden and important implications for healthcare systems. The aim of this study was to provide an epidemiological characterization of individuals with psoriasis in Portugal. Material and Methods: A large observational, cross-sectional, nationwide, population-based survey study developed by the Portuguese Psoriasis Group of the Portuguese Society of Dermatology and Venereology (GPP-SPDV). A structured questionnaire was designed and applied by experienced interviewers to a random, representative sample of Portuguese individuals with psoriasis and/or psoriatic arthritis. Patients were considered to have psoriasis if they replied positively to one of the following questions: “Does any physician have ever diagnosed you with psoriasis?” or “Do you have a skin disorder characterized by scaling, reddish skin lesions located in the elbows/knees/scalp?”. Results: A total of 6381 individuals were interviewed, of which 283 met the criteria for psoriasis, corresponding to a prevalence rate of 4.4% (95% CI 3.95 – 4.98). Out of the participants that met psoriasis criteria, 24% had suggestive signs/symptoms but did not have a clinical diagnosis established and were not being monitored by a physician. Although more than 70% of participants had active disease (scaling, erythema, or pruritus) and one third had joint symptoms, only 12% were on systemic treatment. Fifty percent of participants with psoriasis (n = 139) had relevant comorbidities (most frequently depression/anxiety and cardiometabolic diseases). Sixteen percent of participants with psoriasis (n = 46) reported that psoriasis interfered with their daily activities (median impact of 5 in a 0 – 10 scale) and 12% mentioned the disease had an impact in their sexual life (median impact of 5 in a 0 – 10 scale). Conclusion: The results of this study suggest that the prevalence rate of psoriasis is likely to be high in Portugal, and several gaps exist at different levels of healthcare delivery to these patients, from diagnosis to treatment. This study provides important data for the future planning of interventions targeting the improvement of psoriasis care in Portugal

Nationwide access to endovascular treatment for acute ischemic stroke in portugal

Publisher Copyright: Copyright Ordem dos M dicos 2021.Introduction: Since the publication of endovascular treatment trials and European Stroke Guidelines, Portugal has re-organized stroke healthcare. The nine centers performing endovascular treatment are not equally distributed within the country, which may lead to differential access to endovascular treatment. Our main aim was to perform a descriptive analysis of the main treatment metrics regarding endovascular treatment in mainland Portugal and its administrative districts. Material and Methods: A retrospective national multicentric cohort study was conducted, including all ischemic stroke patients treated with endovascular treatment in mainland Portugal over two years (July 2015 to June 2017). All endovascular treatment centers contributed to an anonymized database. Demographic, stroke-related and procedure-related variables were collected. Crude endovascular treatment rates were calculated per 100 000 inhabitants for mainland Portugal, and each district and endovascular treatment standardized ratios (indirect age-sex standardization) were also calculated. Patient time metrics were computed as the median time between stroke onset, first-door, and puncture. Results: A total of 1625 endovascular treatment procedures were registered. The endovascular treatment rate was 8.27/100 000 inhabitants/year. We found regional heterogeneity in endovascular treatment rates (1.58 to 16.53/100 000/year), with higher rates in districts closer to endovascular treatment centers. When analyzed by district, the median time from stroke onset to puncture ranged from 212 to 432 minutes, reflecting regional heterogeneity. Discussion: Overall endovascular treatment rates and procedural times in Portugal are comparable to other international registries. We found geographic heterogeneity, with lower endovascular treatment rates and longer onset-to-puncture time in southern and inner regions. Conclusion: The overall national rate of EVT in the first two years after the organization of EVT-capable centers is one of the highest among European countries, however, significant regional disparities were documented. Moreover, stroke-onset-to-first-door times and in-hospital procedural times in the EVT centers were comparable to those reported in the randomized controlled trials performed in high-volume tertiary hospitalspublishersversionpublishe

Acesso a Tratamento Endovascular para Acidente Vascular Cerebral Isquémico em Portugal

Introduction: Since the publication of endovascular treatment trials and European Stroke Guidelines, Portugal has re-organized stroke healthcare. The nine centers performing endovascular treatment are not equally distributed within the country, which may lead to differential access to endovascular treatment. Our main aim was to perform a descriptive analysis of the main treatment metrics regarding endovascular treatment in mainland Portugal and its administrative districts. Material and Methods: A retrospective national multicentric cohort study was conducted, including all ischemic stroke patients treated with endovascular treatment in mainland Portugal over two years (July 2015 to June 2017). All endovascular treatment centers contributed to an anonymized database. Demographic, stroke-related and procedure-related variables were collected. Crude endovascular treatment rates were calculated per 100 000 inhabitants for mainland Portugal, and each district and endovascular treatment standardized ratios (indirect age-sex standardization) were also calculated. Patient time metrics were computed as the median time between stroke onset, first-door, and puncture. Results: A total of 1625 endovascular treatment procedures were registered. The endovascular treatment rate was 8.27/100 000 inhabitants/year. We found regional heterogeneity in endovascular treatment rates (1.58 to 16.53/100 000/year), with higher rates in districts closer to endovascular treatment centers. When analyzed by district, the median time from stroke onset to puncture ranged from 212 to 432 minutes, reflecting regional heterogeneity. Conclusion: The overall national rate of EVT in the first two years after the organization of EVT-capable centers is one of the highest among European countries, however, significant regional disparities were documented. Moreover, stroke-onset-to-first-door times and in-hospital procedural times in the EVT centers were comparable to those reported in the randomized controlled trials performed in high-volume tertiary hospitals.Introdução: A aprovação do tratamento endovascular para o acidente vascular cerebral isquémico obrigou à reorganização dos cuidados de saúde em Portugal. Os nove centros que realizam tratamento endovascular não estão distribuídos equitativamente pelo território, o que poderá causar acesso diferencial a tratamento. O principal objetivo deste estudo é realizar uma análise descritiva da frequência e métricas temporais do tratamento endovascular em Portugal continental e seus distritos. Material e Métodos: Estudo de coorte nacional multicêntrico, incluindo todos os doentes com acidente vascular cerebral isquémico submetidos a tratamento endovascular em Portugal continental durante um período de dois anos (julho 2015 a junho 2017). Foram colhidos dados demográficos, relacionados com o acidente vascular cerebral e variáveis do procedimento. Taxas de tratamento endovascular brutas e ajustadas (ajuste indireto a idade e sexo) foram calculadas por 100 000 habitantes/ano para Portugal continental e cada distrito. Métricas de procedimento como tempo entre instalação, primeira porta e punção foram também analisadas. Resultados: Foram registados 1625 tratamentos endovasculares, indicando uma taxa bruta nacional de tratamento endovascular de 8,27/100 000 habitantes/ano. As taxas de tratamento endovascular entre distritos variaram entre 1,58 e 16,53/100 000/ano, com taxas mais elevadas nos distritos próximos a hospitais com tratamento endovascular. O tempo entre sintomas e punção femural entre distritos variou entre 212 e 432 minutos. Conclusão: Portugal continental apresenta uma taxa nacional de tratamento endovascular elevada, apresentando, contudo, assimetrias regionais no acesso. As métricas temporais foram comparáveis com as observadas nos ensaios clínicos piloto

Business strategy and performance of a business unit

Relatório de Estágio do Mestrado em Gestão apresentado à Faculdade de EconomiaAtualmente, a globalização e o consequente aumento da concorrência nos mercados têm sujeitado as organizações a desafios estratégicos cada vez maiores. A par da inovação, o marketing é, hoje, um dos pilares da gestão do negócio. A estratégia de marketing visa a criação de uma resposta por parte da organização às condições competitivas e ao alcance dos seus objetivos organizacionais em mercados-alvo. Em mercados business-to-bussiness as organizações procuram promover os seus produtos perante outras organizações, e dadas as particularidades quando comparado com o mercado business-to-consumer, a estratégia de marketing tem necessariamente as suas particularidades.Este relatório, resultado do estágio realizado na Unidade de Negócio dos Transformadores da Efacec Energia, tem como principal objetivo responder ao desafio proposto pela empresa para a definição de uma estratégia comercial para a unidade de negócio com vista à melhoria da sua performance numa situação de constrangimento de mercado com a qual esta unidade de negócio se depara atualmente. Neste contexto, este relatório debruça-se em aspetos relacionados com a definição da estratégia de marketing e a sua relação com a estratégia organizacional. Foi proposta uma nova estratégia comercial para a unidade de negócio tendo sido definidas várias alavancas de melhoria comercial, das quais se destacam o aumento do pipeline comercial e a melhoria do aproveitamento comercial com vista à obtenção do objetivo primordial, o aumento do número de encomendas. Apoiado pela literatura, este trabalho constitui uma análise crítica que pretende, através da visão obtida durante os quatros meses de estágio curricular, entregar ao seu leitor uma noção prática da importância da definição de uma estratégia de marketing em mercados business-to-business, bem como a sua importância para a estratégia global do negócio.Globalization and the resulting increase in competition in the markets have now placed organizations under increasing strategic challenges. Alongside innovation, marketing is today one of the pillars of business management. The marketing strategy aims the construction of a response by the organization to the competitive conditions and the fulfillment of its organizational objectives in target markets. In business-to-business markets organizations seek to promote their products to other organizations, and given the specificities when compared to the business-to-consumer market, the marketing strategy necessarily has its particularities, too. This report, as a result of the internship carried out in Efacec Energia Transformers Business Unit, has as main objective to respond to the challenge proposed by the company to define a business strategy for the business unit with the purpose of improving its performance in a highly competitive market that this business unit is currently facing. In this context, this report focuses on aspects related to the definition of the marketing strategy and its relationship with the organizational strategy. A new commercial strategy was proposed for the business unit, with several levers of commercial improvement being defined, among which the increase of the commercial pipeline and the improvement of the commercial use in order to achieve the primary objective, the increase in the number of orders. Supported by the literature, this work constitutes a critical analysis that aims, through the vision obtained during the four months of curriculum internship, to give to its reader a practical notion of the importance of defining a marketing strategy in business-to-business markets, as well as its importance to the overall business strategy

Impact of an educational programme in Portuguese people with diabetes

This study, conducted in the north of Portugal, is the result of a group of nurses’ intent to develop a closer and meaningful person-centered care for patients with diabetes, empowering them with skills to better manage their therapeutic regimen. Through a participatory action research process, the group of nurses involved in this study, reflected on the current healthcare enabling them an in-depth analysis of the best scientific evidence, aiming to design context tailored solutions, which would contribute to patients’ empowerment. Through a longitudinal study developed over several stages, the nurses outlined the educational programme Living in harmony with diabetes, which was applied to 85 patients with diabetes mellitus type 2. The programme had a positive impact on the psychosocial variables, in self-care behaviour and in the metabolic control, helping patients to better cope with their disease and have greater control over their health condition. Throughout the process of participatory action research, the nurses were able to increase their self-efficacy and empowerment on person-centered care implementation.info:eu-repo/semantics/publishedVersio

Portuguese recommendations for the treatment of psoriasis with biologic therapy

Copyright © 2021 John Libbey EurotextPsoriasis is a highly prevalent chronic, inflammatory multisystem disease with a considerable impact on patients' quality of life and the healthcare system. This report presents the recommendations developed by the Portuguese Psoriasis Group of the Portuguese Society of Dermatology and Venereology that address several clinical questions arising during the management and care of psoriasis with biologic therapy, based on the available evidence. The recommendations were generated following thorough evaluation of existing guidelines on the treatment of psoriasis, publications concerning new biologic treatments that have not yet been considered in existing guidelines, as well as expert-based recommendations. Considerations regarding the severity of psoriasis, indications for initiating biologic therapy, parameters to be considered in treatment choice (in particular, treatment goals), as well as recommendations for using and monitoring therapy and screening programmes are also included.info:eu-repo/semantics/publishedVersio