Concentration of tumor biomarkers using aqueous biphasic systems

According to the World Health Organization, around 8.2 million people die each year with cancer. Most patients do not perform routine diagnoses and the symptoms, in most situations, occur when the patient is already at an advanced stage of the disease, consequently resulting in a high cancer mortality. Currently, prostate cancer is the second leading cause of death among males worldwide. In Portugal, this is the most diagnosed type of cancer and the third that causes more deaths. Taking into account that there is no cure for advanced stages of prostate cancer, the main strategy comprises an early diagnosis to increase the successful rate of the treatment. The prostate specific antigen (PSA) is an important biomarker of prostate cancer that can be detected in biological fluids, including blood, urine and semen. However, the commercial kits available are addressed for blood samples and the commonly used analytical methods for their detection and quantification requires specialized staff, specific equipment and extensive sample processing, resulting in an expensive process. Thus, the aim of this MSc thesis consisted on the development of a simple, efficient and less expensive method for the extraction and concentration of PSA from urine samples using aqueous biphasic systems (ABS) composed of ionic liquids. Initially, the phase diagrams of a set of aqueous biphasic systems composed of an organic salt and ionic liquids were determined. Then, their ability to extract PSA was ascertained. The obtained results reveal that in the tested systems the prostate specific antigen is completely extracted to the ionic-liquid-rich phase in a single step. Subsequently, the applicability of the investigated ABS for the concentration of PSA was addressed, either from aqueous solutions or urine samples. The low concentration of this biomarker in urine (clinically significant below 150 ng/mL) usually hinders its detection by conventional analytical techniques. The obtained results showed that it is possible to extract and concentrate PSA, up to 250 times in a single-step, so that it can be identified and quantified using less expensive techniques.

Concentration of tumor biomarkers using aqueous biphasic systems

According to the World Health Organization, around 8.2 million people die each year with cancer. Most patients do not perform routine diagnoses and the symptoms, in most situations, occur when the patient is already at an advanced stage of the disease, consequently resulting in a high cancer mortality. Currently, prostate cancer is the second leading cause of death among males worldwide. In Portugal, this is the most diagnosed type of cancer and the third that causes more deaths. Taking into account that there is no cure for advanced stages of prostate cancer, the main strategy comprises an early diagnosis to increase the successful rate of the treatment. The prostate specific antigen (PSA) is an important biomarker of prostate cancer that can be detected in biological fluids, including blood, urine and semen. However, the commercial kits available are addressed for blood samples and the commonly used analytical methods for their detection and quantification requires specialized staff, specific equipment and extensive sample processing, resulting in an expensive process. Thus, the aim of this MSc thesis consisted on the development of a simple, efficient and less expensive method for the extraction and concentration of PSA from urine samples using aqueous biphasic systems (ABS) composed of ionic liquids. Initially, the phase diagrams of a set of aqueous biphasic systems composed of an organic salt and ionic liquids were determined. Then, their ability to extract PSA was ascertained. The obtained results reveal that in the tested systems the prostate specific antigen is completely extracted to the ionic-liquid-rich phase in a single step. Subsequently, the applicability of the investigated ABS for the concentration of PSA was addressed, either from aqueous solutions or urine samples. The low concentration of this biomarker in urine (clinically significant below 150 ng/mL) usually hinders its detection by conventional analytical techniques. The obtained results showed that it is possible to extract and concentrate PSA, up to 250 times in a single-step, so that it can be identified and quantified using less expensive techniques.

Envelhecimento ativo e o recurso à medicina tradicional chinesa : entre a responsabilidade individual e os fatores sociais determinantes da saúde

Tese de doutoramento, Antropologia (Antropologia da Saúde), Universidade de Lisboa, Instituto de Ciências Sociais, 2015

Personal Exposure to Grass Pollen: Relating Inhaled Dose to Background Concentration

Background Very few studies on human exposure to allergenic pollen have been conducted using direct methods, with background concentrations measured at city center monitoring stations typically taken as a proxy for exposure despite the inhomogeneous nature of atmospheric pollen concentrations. A 2003 World Health Organization report highlighted the need for an improved understanding of the relation between monitoring station data and actual exposure. Objective To investigate the relation between grass pollen dose and background concentrations measured at a monitoring station, to assess the fidelity of monitoring station data as a qualitative proxy for dose, and to evaluate the ratio of dose rate to background concentration. Methods Grass pollen dose data were collected in Aarhus, Denmark, in an area where grass pollen sources were prevalent, using Nasal Air Samplers. Sample collection lasted for approximately 25 to 30 minutes and was performed at 2-hour intervals from noon to midevening under moderate exercise by 2 individuals. Results A median ratio of dose rate to background concentration of 0.018 was recorded, with higher ratio values frequently occurring at 12 to 2 pm, the time of day when grass species likely to be present in the area are expected to flower. From 4 to 8 pm, dose rate and background concentration data were found to be strongly and significantly correlated (rs = 0.81). Averaged dose rate and background concentration data showed opposing temporal trends. Conclusion Where local emissions are not a factor, background concentration data constitute a good quantitative proxy for inhaled dose. The present ratio of dose rate to background concentration may aid the study of dose–response relations.

Translating proteomics into environmental health : biomarkers discovery for tobacco smoke-induced biological damage

Tese de doutoramento, Biologia (Biologia Molecular), Universidade de Lisboa, Faculdade de Ciências, 2015

Activities of an aqueous plant extract against Herpes simplex virus type 2 (HSV 2): role in virus replication and progeny yield

Tese de mestrado, Biologia Molecular e Genética, Universidade de Lisboa, Faculdade de Ciências, 2015

Construção e validação da tabela nacional de funcionalidade para as doenças crónicas

© Ordem dos Médicos 2016

The risk of viral dispersal and aerosolization by different hand-drying methods

Background World Health Organization hand hygiene guidelines state that if electric hand dryers are used, they should not aerosolize pathogens. Previous studies have investigated the dispersal by different hand-drying devices of chemical indicators, fungi and bacteria on the hands. This study assessed the aerosolization and dispersal of virus on the hands to determine any differences between hand-drying devices in their potential to contaminate other occupants of public washrooms and the washroom environment. Methods A suspension of MS2, an Escherichia coli bacteriophage virus, was used to artificially contaminate the hands of participants prior to using three different handdrying devices: jet air dryer, warm air dryer, paper towel dispenser. Virus was detected by plaque formation on agar plates layered with the host bacterium. Vertical dispersal of virus was assessed at a fixed distance (0.4 m) and over a range of different heights (0.0 – 1.8 m) from the floor. Horizontal dispersal was assessed at different distances of up to three metres from the hand-drying devices. Virus aerosolization and dispersal was also assessed at different times up to 15 minutes after use by means of air sampling at two distances (0.1 and 1.0 m) and at a distance behind and offset from each of the hand-drying devices. Results Over a range of heights, the jet air dryer was shown to produce over 60 times greater vertical dispersal of virus from the hands than a warm air dryer and over 1300 times greater than paper towels; the maximum being detected between 0.6 and 1.2 metres from the floor. Horizontal dispersal of virus by the jet air dryer was over 20 times greater than a warm air dryer and over 190 times greater than paper towels; virus being detected at distances of up to three metres. Air sampling at three different positions from the hand-drying devices 15 minutes after use showed that the jet air dryer produced over 50-times greater viral contamination of the air than a warm air dryer and over 110-times greater than paper towels. Conclusions Due to their high air speed, jet air dryers aerosolize and disperse more virus over a range of heights, greater distances, and for longer times than other hand drying devices. If hands are inadequately washed, they have a greater potential to contaminate other occupants of a public washroom and the washroom environment. Main messages: Jet air dryers with claimed air speeds of over 600 kph have a greater potential than warm air dryers or paper towels to aerosolize and disperse viruses on the hands of users. The choice of hand-drying device should be carefully considered. Jet air dryers may increase the risk of transmission of human viruses, such as norovirus, particularly if hand washing is inadequate.

Comparison of virus dispersal and aerosolization by different hand-drying devices

Background World Health Organization and EU hand hygiene guidelines state that if electric hand dryers are used, they should not aerosolize pathogens. Previous studies have investigated the dispersal by different hand-drying devices of chemical indicators, fungi and bacteria on the hands. This study assessed the aerosolization and dispersal of virus on the hands to determine any differences between hand-drying devices in their potential to contaminate other occupants of public washrooms and the washroom environment. Methods A suspension of MS2, an Escherichia coli bacteriophage virus, was used to artificially contaminate the hands of participants prior to using three different hand-drying devices: jet air dryer, warm air dryer, paper towel dispenser. Virus was detected by plaque formation on agar plates layered with the host bacterium. Vertical dispersal of virus was assessed at a fixed distance (0.4 m) and over a range of different heights (0.0 – 1.8 m) from the floor. Horizontal dispersal was assessed at different distances of up to three metres from the hand-drying devices. Virus aerosolization and dispersal was also assessed at different times up to 15 minutes after use by means of air sampling at two distances (0.1 and 1.0 m) and at a distance behind and offset from each of the hand-drying devices. Results Over a range of heights, the jet air dryer was shown to produce over 60 times greater vertical dispersal of virus from the hands than a warm air dryer and over 1300 times greater than paper towels; the maximum being detected between 0.6 and 1.2 metres from the floor. Horizontal dispersal of virus by the jet air dryer was over 20 times greater than a warm air dryer and over 190 times greater than paper towels; virus being detected at distances of up to three metres. Air sampling at three different positions from the hand-drying devices 15 minutes after use showed that the jet air dryer produced over 50-times greater viral contamination of the air than a warm air dryer and over 110-times greater than paper towels. Conclusions Due to their high air speed, jet air dryers aerosolize and disperse more virus over a range of heights, greater distances, and for longer times than other hand drying devices. If hands are inadequately washed, they have a greater potential to contaminate other occupants of a public washroom and the washroom environment. Main messages: Jet air dryers with claimed air speeds of over 600 kph have a greater potential than warm air dryers or paper towels to aerosolize and disperse viruses on the hands of users. The choice of hand-drying device should be carefully considered. Jet air dryers may increase the risk of transmission of human viruses, such as norovirus, particularly if hand washing is inadequate.

Segurança alimentar nos refeitórios de uma universidade portuguesa : aplicação da teoria do comportamento planeado

A Organização Mundial de Saúde estima que nos países mais industrializados uma em cada três pessoas sofra, por ano, de uma doença de origem alimentar. De acordo com os dados da Agência Europeia para a Segurança Alimentar foram relatados pelos 27 Estados Membros da União Europeia, no ano 2012, um total de 5.363 surtos de origem alimentar, assistindo-se a uma prevalência do setor da restauração, como o local de maior ocorrência dos surtos de doenças de origem alimentar. Para o mesmo ano, Portugal reportou 7 surtos de origem alimentar, envolvendo 135 pessoas com 42 hospitalizações. Neste contexto, a aplicação de boas práticas de higiene, nomeadamente no setor da restauração, é essencial para proteger o consumidor das doenças de origem alimentar. Neste estudo, pretendeu-se identificar os constructos do modelo da Teoria do Comportamento Planeado (Theory of Planned Behaviour – TPB, segundo a terminologia anglo-saxónica), de Icek Ajzen, que melhor explicam a intenção dos operadores de alimentos em adotarem os comportamentos de higiene, a saber: i) utilização de luvas e touca de proteção de cabelos, e ii) remoção de adornos pessoais, durante a manipulação de alimentos. Para o efeito, foi aplicado um questionário tendo por base a Teoria do Comportamento Planeado, a uma amostra de cento e vinte e três operadores dos vários refeitórios de uma universidade portuguesa, na sua grande maioria do sexo feminino (91,1%) e que manipulam alimentos numa base diária, recorrendo-se primeiramente a uma fase preliminar de estudo qualitativo, ou pré-inquérito, para melhor selecionar os temas essenciais e as principais categorias a considerar na construção deste inquérito. Os inquéritos foram tratados estatisticamente recorrendo-se à estatística descritiva, à análise fatorial e avaliação da consistência interna dos fatores resultantes, seguido da aplicação de regressão linear e metodologia de análise de trajetórias (path modeling) com vista à validação do TPB. Os resultados obtidos apontam para o fato de a Atitude ser o melhor preditor da Intenção em adotar os comportamentos em estudo. Verificou-se também que a motivação de cumprir resulta da pressão exercida pelos superiores hierárquicos ou colegas, influenciando positivamente a intenção, na medida em que as crenças normativas assumiram-se como sendo o segundo preditor que melhor previu a intenção.

Problemática associada à utilização de diferentes tipos de mochila para transporte de material escolar

O transporte de cargas é uma tarefa comum para crianças, adolescentes e adultos, pela necessidade de transferência diária de objetos pessoais, livros e artigos de papelaria para os locais de trabalho ou escolas. Diversos autores apontam que o peso carregado durante transporte de material é o principal responsável pelo aparecimento de dor lombar. Deste modo é importante o constante estudo da temática para a definição recomendações e limites. O presente estudo teve como principais objetivos a caraterização da problemática associada à utilização de mochilas e a determinação do Peso Máximo Aceitável (PMA) e do Índice de Esforço Percebido (IEP) para a tarefa de transporte de mochilas, através da abordagem psicofísica. O estudo foi desenvolvido com estudantes do 7º, 8º e 9º ano de escolaridade e, foi dividido em duas fases. Na 1ª fase foram aplicados questionários para a análise da problemática associada à utilização de diferentes tipos de mochilas escolares. Nesta fase, foram incluídos aspetos associados à identificação do tipo de mochila mais utilizada, as rotinas e hábitos dos estudantes e as características da mochila utilizada. Verificou-se que os estudantes utilizam, maioritariamente, a mochila de duas alças para transporte de material escolar. Posteriormente foram efetuadas medições de peso da mochila, altura e peso aos 131 estudantes que constituíram a amostra da 1º fase. O principal objetivo deste ponto foi identificar o tipo de mochila habitualmente utilizada pelos estudantes assim como, o peso transportado nas mochilas. Na 2ª fase foi efetuado um estudo para a determinação do PMA e do IEP, através da abordagem psicofísica, para a tarefa de transporte de mochila, considerando-se uma amostra constituída por 10 estudantes. Para este estudo, apenas foi considerada a mochila mais frequentemente utilizada, identificada na 1º fase. A tarefa consistiu no transporte da mochila nos dois ombros e com as alças devidamente ajustadas ao corpo, num percurso pré-definido, de acordo com o procedimento experimental. Os resultados indicaram que nem todos os estudantes transportam mochilas com pesos dentro das recomendações da Organização Mundial de Saúde. O PMA determinado pelos estudantes foi de 6.8 kg para a mochila de duas alças e a região dos ombros foi identificada durante todo o estudo como sendo a que apresentava maior intensidade de dor durante o transporte da mochila.

Desenvolvimento de um Code Set para a escrita manual baseado na classificação internacional de funcionalidade versão crianças e jovens

Compreender a funcionalidade de uma criança é um desafio persistente em contextos de saúde e educação. Na tentativa de superar esse desafio, em 2007, a Organização Mundial de Saúde desenvolveu a Classificação Internacional de Funcionalidade, Incapacidade e Saúde para Crianças e Jovens (CIF-CJ) como o primeiro sistema de classificação universal para documentar a saúde e funcionalidade da criança. Apesar de a CIF-CJ não ser um instrumento de avaliação e intervenção, tem, no entanto, a capacidade de servir de enquadramento para o desenvolvimento de ferramentas adaptadas às necessidades dos seus utilizadores. Considerando que no contexto escolar, a escrita manual encontra-se entre as atividades mais requeridas para a participação plena de uma criança, parece ser pertinente a definição de um conjunto de códigos destinados a caracterizar o perfil de funcionalidade de uma criança, no que se refere à escrita manual. O objetivo deste estudo foi, pois, o desenvolvimento de um conjunto preliminar de códigos baseado na CIF-CJ que possa vir a constituir um code set para a escrita manual. Dada a complexidade do tema e atendendo a que se pretende alcançar consenso entre os especialistas sobre quais as categorias da CIF-CJ que devem ser consideradas, optou-se pela utilização da técnica de Delphi. A escolha da metodologia seguiu a orientação dos procedimentos adotados pelo projeto Core Set CIF. De dezoito profissionais contactados, obtiveram-se respostas de sete terapeutas ocupacionais com experiência em pediatria, que participaram em todas as rondas. No total, três rondas de questionários foram realizadas para atingir um consenso, com um nível de concordância, previamente definido, de 70%. Deste estudo resultou um conjunto preliminar de códigos com 54 categorias da CIF-CJ (16 categorias de segundo nível, 14 categorias de terceiro nível e uma categoria de quarto nível), das quais 31 são categorias das funções do corpo, uma categoria das estruturas do corpo, 12 categorias de atividades e participação e 10 categorias de fatores ambientais. Este estudo é um primeiro passo para o desenvolvimento de um code set para a escrita manual baseado na CIF-CJ , sendo claramente necessário a realização de mais pesquisas no contexto do desenvolvimento e da validação deste code set.

The role of the efflux mechanisms in multidrug resistance in Mycobacterium tuberculosis

Abstract The emergence of multi and extensively drug resistant tuberculosis (MDRTB and XDRTB) has increased the concern of public health authorities around the world. The World Health Organization has defined MDRTB as tuberculosis (TB) caused by organisms resistant to at least isoniazid and rifampicin, the main first-line drugs used in TB therapy, whereas XDRTB refers to TB resistant not only to isoniazid and rifampicin, but also to a fluoroquinolone and to at least one of the three injectable second-line drugs, kanamycin, amikacin and capreomycin. Resistance in Mycobacterium tuberculosis is mainly due to the occurrence of spontaneous mutations and followed by selection of mutants by subsequent treatment. However, some resistant clinical isolates do not present mutations in any genes associated with resistance to a given antibiotic, which suggests that other mechanism(s) are involved in the development of drug resistance, namely the presence of efflux pump systems that extrude the drug to the exterior of the cell, preventing access to its target. Increased efflux activity can occur in response to prolonged exposure to subinhibitory concentrations of anti-TB drugs, a situation that may result from inadequate TB therapy. The inhibition of efflux activity with a non-antibiotic inhibitor may restore activity of an antibiotic subject to efflux and thus provide a way to enhance the activity of current anti-TB drugs. The work described in this thesis foccus on the study of efflux mechanisms in the development of multidrug resistance in M. tuberculosis and how phenotypic resistance, mediated by efflux pumps, correlates with genetic resistance. In order to accomplish this goal, several experimental protocols were developed using biological models such as Escherichia coli, the fast growing mycobacteria Mycobacterium smegmatis, and Mycobacterium avium, before their application to M. tuberculosis. This approach allowed the study of the mechanisms that result in the physiological adaptation of E. coli to subinhibitory concentrations of tetracycline (Chapter II), the development of a fluorometric method that allows the detection and quantification of efflux of ethidium bromide (Chapter III), the characterization of the ethidium bromide transport in M. smegmatis (Chapter IV) and the contribution of efflux activity to macrolide resistance in Mycobacterium avium complex (Chapter V). Finally, the methods developed allowed the study of the role of efflux pumps in M. tuberculosis strains induced to isoniazid resistance (Chapter VI). By this manner, in Chapter II it was possible to observe that the physiological adaptation of E. coli to tetracycline results from an interplay between events at the genetic level and protein folding that decrease permeability of the cell envelope and increase efflux pump activity. Furthermore, Chapter III describes the development of a semi-automated fluorometric method that allowed the correlation of this efflux activity with the transport kinetics of ethidium bromide (a known efflux pump substrate) in E. coli and the identification of efflux inhibitors. Concerning M. smegmatis, we have compared the wild-type M. smegmatis mc2155 with knockout mutants for LfrA and MspA for their ability to transport ethidium bromide. The results presented in Chapter IV showed that MspA, the major porin in M. smegmatis, plays an important role in the entrance of ethidium bromide and antibiotics into the cell and that efflux via the LfrA pump is involved in low-level resistance to these compounds in M. smegmatis. Chapter V describes the study of the contribution of efflux pumps to macrolide resistance in clinical M. avium complex isolates. It was demonstrated that resistance to clarithromycin was significantly reduced in the presence of efflux inhibitors such as thioridazine, chlorpromazine and verapamil. These same inhibitors decreased efflux of ethidium bromide and increased the retention of [14C]-erythromycin in these isolates. Finaly, the methods developed with the experimental models mentioned above allowed the study of the role of efflux pumps on M. tuberculosis strains induced to isoniazid resistance. This is described in Chapter VI of this Thesis, where it is demonstrated that induced resistance to isoniazid does not involve mutations in any of the genes known to be associated with isoniazid resistance, but an efflux system that is sensitive to efflux inhibitors. These inhibitors decreased the efflux of ethidium bromide and also reduced the minimum inhibitory concentration of isoniazid in these strains. Moreover, expression analysis showed overexpression of genes that code for efflux pumps in the induced strains relatively to the non-induced parental strains. In conclusion, the work described in this thesis demonstrates that efflux pumps play an important role in the development of drug resistance, namely in mycobacteria. A strategy to overcome efflux-mediated resistance may consist on the use of compounds that inhibit efflux activity, restoring the activity of antimicrobials that are efflux pump substrates, a useful approach particularly in TB where the most effective treatment regimens are becoming uneffective due to the increase of MDRTB/XDRTB.

Plano de Negócio para a aplicação mobile Nutri Consult

Trabalho de projeto apresentado à Escola Superior de Comunicação Social como parte dos requisitos para obtenção de grau de mestre em Publicidade e Marketing.

Quality of Life Predictors and Normative Data

This study identifies predictors and normative data for quality of life (QOL) in a sample of Portuguese adults from general population. A cross-sectional correlational study was undertaken with two hundred and fifty-five (N = 255) individuals from Portuguese general population (mean age 43 years, range 25–84 years; 148 females, 107 males). Participants completed the European Portuguese version of the World Health Organization Quality of Life short-form instrument and the European Portuguese version of the Center for Epidemiologic Studies Depression Scale. Demographic information was also collected. Portuguese adults reported their QOL as good. The physical, psychological and environmental domains predicted 44 % of the variance of QOL. The strongest predictor was the physical domain and the weakest was social relationships. Age, educational level, socioeconomic status and emotional status were significantly correlated with QOL and explained 25 % of the variance of QOL. The strongest predictor of QOL was emotional status followed by education and age. QOL was significantly different according to: marital status; living place (mainland or islands); type of cohabitants; occupation; health. The sample of adults from general Portuguese population reported high levels of QOL. The life domain that better explained QOL was the physical domain. Among other variables, emotional status best predicted QOL. Further variables influenced overall QOL. These findings inform our understanding on adults from Portuguese general population QOL and can be helpful for researchers and practitioners using this assessment tool to compare their results with normative data