Gold nanoparticles for the detection of DNA adducts as biomarkers of exposure to acrylamide

The main objective of this thesis was the development of a gold nanoparticle-based methodology for detection of DNA adducts as biomarkers, to try and overcome existing drawbacks in currently employed techniques. For this objective to be achieved, the experimental work was divided in three components: sample preparation, method of detection and development of a model for exposure to acrylamide. Different techniques were employed and combined for de-complexation and purification of DNA samples (including ultrasonic energy, nuclease digestion and chromatography), resulting in a complete protocol for sample treatment, prior to detection. The detection of alkylated nucleotides using gold nanoparticles was performed by two distinct methodologies: mass spectrometry and colorimetric detection. In mass spectrometry, gold nanoparticles were employed for laser desorption/ionisation instead of the organic matrix. Identification of nucleotides was possible by fingerprint, however no specific mass signals were denoted when using gold nanoparticles to analyse biological samples. An alternate method using the colorimetric properties of gold nanoparticles was employed for detection. This method inspired in the non-cross-linking assay allowed the identification of glycidamide-guanine adducts and DNA adducts generated in vitro. For the development of a model of exposure, two different aquatic organisms were studies: a goldfish and a mussel. Organisms were exposed to waterborne acrylamide, after which mortality was recorded and effect concentrations were estimated. In goldfish, both genotoxicity and metabolic alterations were assessed and revealed dose-effect relationships of acrylamide. Histopathological alterations were verified primarily in pancreatic cells, but also in hepatocytes. Mussels showed higher effect concentrations than goldfish. Biomarkers of oxidative stress, biotransformation and neurotoxicity were analysed after prolonged exposure, showing mild oxidative stress in mussel cells, and induction of enzymes involved in detoxification of oxygen radicals. A qualitative histopathological screening revealed gonadotoxicity in female mussels, which may present some risk to population equilibrium.

Synthesis of Polymeric Nanoparticles for biomedical delivery applications

Polymeric nanoparticles (PNPs) have attracted considerable interest over the last few years due to their unique properties and behaviors provided by their small size. Such materials could be used in a wide range of applications such as diagnostics and drug delivery. Advantages of PNPs include controlled release, protection of drug molecules and its specific targeting, with concomitant increasing of the therapeutic index. In this work, novel sucrose and cholic acid based PNPs were prepared from different polymers, namely polyethylene glycol (PEG), poly(D,L-lactic-co-glycolic acid) (PLGA) and PLGA-co-PEG copolymer. In these PNP carriers, cholic acid will act as a drug incorporation site and the carbohydrate as targeting moiety. The uptake of nanoparticles into cells usually involves endocytotic processes, which depend primarily on their size and surface characteristics. These properties can be tuned by the nanoparticle preparation method. Therefore, the nanoprecipitation and the emulsion-solvent evaporation method were applied to prepare the PNPs. The influence of various parameters, such as concentration of the starting solution, evaporation method and solvent properties on the nanoparticle size, size distribution and morphology were studied. The PNPs were characterized by using atomic force microscopy (AFM), scanning electron microscopy (SEM) and dynamic light scattering (DLS) to assess their size distribution and morphology. The PNPs obtained by nanoprecipitation ranged in size between 90 nm and 130 nm with a very low polydispersity index (PDI < 0.3). On the other hand, the PNPs produced by the emulsion-solvent evaporation method revealed particle sizes around 300 nm with a high PDI value. More detailed information was found in AFM and SEM images, which demonstrated that all these PNPs were regularly spherical. ζ-potential measurements were satisfactory and evidenced the importance of sucrose moiety on the polymeric system, which was responsible for the obtained negative surface charge, providing colloidal stability. The results of this study show that sucrose and cholic acid based polymeric conjugates can be successfully used to prepare PNPs with tunable physicochemical characteristics. In addition, it provides novel information about the materials used and the methods applied. It is hoped that this work will be useful for the development of novel carbohydrate based nanoparticles for biomedical applications, specifically for targeted drug delivery.

Adaptação cultural e contributo para a validação da pain attitudes and believes scale for physiotherapists (PABS-PT) para a população de fisioterapeutas portugueses

RESUMO: Introdução: Vários estudos têm encontrado evidência para a relação entre as crenças e atitudes dos profissionais de saúde, a sua abordagem de tratamento, referida como orientada segundo o modelo biomédico ou modelo biopsicossocial (Bishop, 2008; Mutsaers, 2012), e os resultados obtidos. É sugerido que, no tratamento da dor lombar crónica, os profissionais que orientam o seu raciocínio e prática segundo o modelo biomédico, tendem a obter piores resultados quando comparados com os obtidos pelos profissionais que orientam o seu raciocínio e prática segundo uma abordagem biopsicossocial. Esta área de estudo tem salientado a importância de desenhar instrumentos capazes de identificar a orientação preferencial dos profissionais de saúde no tratamento da dor crónica de natureza músculo-esquelética, entre os quais se encontra a “Pain Attitudes and Beliefs Scale for Physiotherapists” (PABS-PT). Objetivo: Adaptar culturalmente a PABS-PT para a população de fisioterapeutas portugueses que intervêm em utentes com dor (lombar) crónica, e contribuir para a sua validação Metodologia: A versão original (língua inglesa) do PABS-PT foi adaptada para a língua e cultura portuguesas, através das etapas definidas nas normas orientadoras estabelecidas para este processo (Beaton et al., 2002; MAPI Institute, 2001). A avaliação das propriedades psicométricas da versão portuguesa foi realizada com recurso a uma amostra de 202 fisioterapeutas e estudantes finalistas do curso de licenciatura em Fisioterapia. Inicialmente foi realizada a análise fatorial exploratória da escala através do método das componentes principais. Posteriormente avaliou-se a consistência interna das componentes obtidas com recurso ao alpha de Cronbach (α). Para analisar a validade de constructo foram correlacionadas as componentes obtidas com a versão portuguesa da “Health Care Providers’ Pain and Impairment Relationship Scale” (HC-PAIRS), recorrendo ao cálculo do coeficiente de Spearman. Resultados: O processo de adaptação cultural não revelou dificuldades importantes sendo considerado que a versão portuguesa da PABS-PT é de fácil compreensão e preenchimento, e os seus itens adequados para avaliar as crenças e atitudes dos fisioterapeutas portugueses relativas à intervenção na dor crónica músculo-esquelética. Os resultados revelaram uma estrutura fatorial de duas componentes, identificadas com as componentes da escala original que explicam 30,96% da variância total. A consistência interna encontrada é boa, para a componente biomédica (α de Cronbach = 0,826), mas muito fraca para a componente biopsicossocial (α de Cronbach= 0,589). Relativamente à validade convergente e discriminativa, foi encontrada uma associação estatisticamente significativa e positiva, entre as componentes 1 (biomédica) da versão nportuguesa da PABS e a HC-PAIRS (Rs = 0,481, p≤ 0,005), e negativa, fraca e significativa entre a pontuação total da componente 2 (biopsicossocial) e a HC-PAIRS (Rs = -0,038, p=0,612). Conclusão: A versão portuguesa do PABS-PT é de fácil compreensão e aparenta ser um instrumento válido para a medição da orientação preferencial dos fisioterapeutas, relativamente às suas atitudes e crenças na avaliação e tratamento de utentes com dor crónica de natureza músculo-esquelética. No entanto, a componente biopsicossocial requer uma análise mais aprofundada para que possa, com rigor, ser utilizada na definição de uma orientação preferencialmente biopsicossocial.--------------ABSTRACT: Introduction: Previous studies have found a relation between the beliefs and attitudes of health professionals, their treatment approach, which can follows a biomedical or a biopsychosocial orientation (Bishop, 2008; Mutsaers, 2012), and the outcomes obtained. Therefore, is suggested that the professionals who tend to approach chronic low back pain patients according to the biomedical model have worse outcomes than professionals who use a biopsychosocial approach in there clinical reasoning. This research field has highlighted the importance of developing measures capable of identifying the preferred orientation of health professionals in the treatment of chronic pain of musculoskeletal nature, including the “Pain Attitudes and Beliefs Scale for Physiotherapists” (PABS-PT). Objective: To cross culturally adapt and validate the PABS-PT for the Portuguese population of physiotherapists. Methodology: The original version (English version) of the PABS-PT was adapted to the Portuguese language and culture, through the guidelines established for these processes (Beaton et al., 2002; MAPI Institute, 2001). The psychometric evaluation of the Portuguese version was carried out on a sample of 202 p physiotherapist and final year students of the physiotherapy course. Initially, an exploratory factorial analysis was performed through the method of the main components. Then, the internal consistence of the main components was evaluated using the Cronbach’s alpha (α). The convergent construct validity was analysed through the correlation between the obtained components of PABS-PT and the Health Care Provider’s Pain and Impairment Relationship Scale (HC-PAIRS), using the Spearmen correlation coefficient. Results: No major difficulties were found during the cultural adaptation process of PABS-PT to Portugal, which means that the Portuguese version is easy to understand and fulfill, and items are appropriated to evaluate the beliefs and attitudes of the Portuguese physiotherapists who treat chronic pain of musculoskeletal origins. The results revealed a factorial structure of two components, as the original scale, explaining 30,96% of the total variance. Internal consistence results were good, for the biomedical component (Cronbach’s α = 0,826), but very weak for the biopsychosocial componente (Cronbach’s α = 0,589). Relatively to convergent and discriminative validity, a statistically significant association was found, between the components 1 (biomedical) of the Portuguese version of PABS-PT and the HC-PAIRS (Rs= 0,481, p≤ 0,005) and negative, weak and significant between the total score of component 2 (biopsychosocial) and the HC-PAIRS (Rs = -0,038, p=0,612). Conclusion: The Portuguese Version of PABS-PT is easy to understand and seems to be a valid instrument to measure the attitudes and beliefs of physiotherapists in the management of patients with chronic low back pain. However, the biopsychosocial component requires a further deep analysis to examine a preferable biopsychosocial orientation.

Engineering of metal oxide nanoparticles for application in electrochemical devices

The growing demand for materials and devices with new functionalities led to the increased inter-est in the field of nanomaterials and nanotechnologies. Nanoparticles, not only present a reduced size as well as high reactivity, which allows the development of electronic and electrochemical devices with exclusive properties, when compared with thin films. This dissertation aims to explore the development of several nanostructured metal oxides by sol-vothermal synthesis and its application in different electrochemical devices. Within this broad theme, this study has a specific number of objectives: a) research of the influence of the synthesis parameters to the structure and morphology of the nanoparticles; b) improvement of the perfor-mance of the electrochromic devices with the application of the nanoparticles as electrode; c) application of the nanoparticles as probes to sensing devices; and d) production of solution-pro-cessed transistors with a nanostructured metal oxide semiconductor. Regarding the results, several conclusions can be exposed. Solvothermal synthesis shows to be a very versatile method to control the growth and morphology of the nanoparticles. The electrochromic device performance is influenced by the different structures and morphologies of WO3 nanoparticles, mainly due to the surface area and conductivity of the materials. The dep-osition of the electrochromic layer by inkjet printing allows the patterning of the electrodes without wasting material and without any additional steps. Nanostructured WO3 probes were produced by electrodeposition and drop casting and applied as pH sensor and biosensor, respectively. The good performance and sensitivity of the devices is explained by the high number of electrochemical reactions occurring at the surface of the na-noparticles. GIZO nanoparticles were deposited by spin coating and used in electrolyte-gated transistors, which promotes a good interface between the semiconductor and the dielectric. The produced transistors work at low potential and with improved ON-OFF current ratio, up to 6 orders of mag-nitude. To summarize, the low temperatures used in the production of the devices are compatible with flexible substrates and additionally, the low cost of the techniques involved can be adapted for disposable devices.

The impact of online promotional efforts on customer acquisitiom and retention at continente.pt

Continente Online (COL) is the market leader of online grocery retailing in Portugal. Aiming at sustaining this position, it has been focusing its promotional efforts in acquiring and retaining customers. This report intakes an extensive study on the impact of COL’s exclusive promotional efforts (free delivery offers and ten percent discounts on loyalty card) on customer acquisition and retention, as well as, a cost analysis on these offers. Two econometric models were developed and the results were interesting. It was concluded that free delivery offers have a significant impact both in terms of customer acquisition and retention, having also a lower cost in comparison with ten percent on loyalty card discounts.

Management consulting labs- Pt

The purpose of this project was to assist PT in identifying how the company could capture value with a new technology in the short-medium term in Portugal. The analysis performed consisted on identifying, evaluating and prioritizing the business opportunities that could have this new technology. Thus, a prioritization methodology was set to find the ones with greater value, and a deep analysis was conducted to its assessment. Subsequently, a strategy for the whole organization and for the priorities discovered was outlined. Based on these analyses, a set of recommendations was given to PT concerning its position regarding this new technolog

Engineered MRI nanoprobes based on superparamagnetic iron oxide nanoparticles

This project aimed to engineer new T2 MRI contrast agents for cell labeling based on formulations containing monodisperse iron oxide magnetic nanoparticles (MNP) coated with natural and synthetic polymers. Monodisperse MNP capped with hydrophobic ligands were synthesized by a thermal decomposition method, and further stabilized in aqueous media with citric acid or meso-2,3-dimercaptosuccinic acid (DMSA) through a ligand exchange reaction. Hydrophilic MNP-DMSA, with optimal hydrodynamic size distribution, colloidal stability and magnetic properties, were used for further functionalization with different coating materials. A covalent coupling strategy was devised to bind the biopolymer gum Arabic (GA) onto MNPDMSA and produce an efficient contrast agent, which enhanced cellular uptake in human colorectal carcinoma cells (HCT116 cell line) compared to uncoated MNP-DMSA. A similar protocol was employed to coat MNP-DMSA with a novel biopolymer produced by a biotechnological process, the exopolysaccharide (EPS) Fucopol. Similar to MNP-DMSA-GA, MNP-DMSA-EPS improved cellular uptake in HCT116 cells compared to MNP-DMSA. However, MNP-DMSA-EPS were particularly efficient towards the neural stem/progenitor cell line ReNcell VM, for which a better iron dose-dependent MRI contrast enhancement was obtained at low iron concentrations and short incubation times. A combination of synthetic and biological coating materials was also explored in this project, to design a dynamic tumortargeting nanoprobe activated by the acidic pH of tumors. The pH-dependent affinity pair neutravidin/iminobiotin, was combined in a multilayer architecture with the synthetic polymers poy-L-lysine and poly(ethylene glycol) and yielded an efficient MRI nanoprobe with ability to distinguish cells cultured in acidic pH conditions form cells cultured in physiological pH conditions.

Field-effect transistors based on Zinc oxide nanoparticles

This work reports the development of field-effect transistors (FETs), whose channel is based on zinc oxide (ZnO) nanoparticles (NPs). Using screen-printing as the primary deposition technique, different inks were developed, where the semiconducting ink is based on a ZnO NPs dispersion in ethyl cellulose (EC). These inks were used to print electrolyte-gated transistors (EGTs) in a staggered-top gate structure on glass substrates, using a lithium-based polymeric electrolyte. In another approach, FETs with a staggered-bottom gate structure on paper were developed using a sol-gel method to functionalize the paper’s surface with ZnO NPs, using zinc acetate dihydrate (ZnC4H6O4·2H2O) and sodium hydroxide (NaOH) as precursors. In this case, the paper itself was used as dielectric. The various layers of the two devices were characterized using X-ray diffraction (XRD), scanning electron microscopy (SEM), Fourier Transform Infrared spectroscopy (FTIR), thermogravimetric and differential scanning calorimetric analyses (TG-DSC). Electrochemical impedance spectroscopy (EIS) was used in order to evaluate the electric double-layer (EDL) formation, in the case of the EGTs. The ZnO NPs EGTs present electrical modulation for annealing temperatures equal or superior to 300 ºC and in terms of electrical properties they showed On/Off ratios in the order of 103, saturation mobilities (μSat) of 1.49x10-1 cm2(Vs)-1 and transconductance (gm) of 10-5 S. On the other hand, the ZnO NPs FETs on paper exhibited On/Off ratios in the order of 102, μSat of 4.83x10- 3 cm2(Vs)-1and gm around 10-8 S.

VO2 thin films and nanoparticles for energy-saving applications in architectural elements

Vanadium dioxide (VO2) is a promising material with large interest in construction industry and architecture, due to its thermochromic properties. This material may be used to create "smart" coatings that result in improvements in the buildings energy efficiency, by reducing heat exchanges and, consequently, the need for acclimatization. In this work, VO2 thin films and coatings were produced and tested in laboratory, to apply in architectural elements, such as glass, rooftop tiles and exterior paints. Thin films were produced by RF magnetron sputtering and VO2 nanoparticles were obtained through hydrothermal synthesis, aiming to create "smart" windows and tiles, respectively. These coatings have demonstrated the capability to modulate the transmittance of infrared radiation by around 20%. The VO2 nanoparticle coatings were successfully applied on ceramic tiles. The critical temperature was reduced to around 40ºC by tungsten doping. Ultimately, two identical house models were built, in order to test the VO2 coatings, in real atmospheric conditions during one of the hottest months of the year, in Portugal – August.

Chitosan-based magnetic nanoparticles for osteosarcoma theranostic

Cancer is a well-known disease with a significant impact in society not only due to its incidence, more evident in more developed countries, but also due to the expenses related to medical treat-ments. Cancer research is considered an increasingly logical science with great potential for the development of new treatment options. Advances in nanomedicine have resulted in rapid devel-opment of nanomaterials with considerable potential in cancer diagnostics and treatment. The combination of diagnosis and treatment in a single nano-platform is named theranostic. In this PhD thesis a theranostic system for osteosarcoma was proposed, composed by a magnetic core, a polymeric coating, and a chemotherapeutic drug. The presence of a specific targeting agent, in this case a monoclonal antibody, provides high specificity to the proposed theranostic system. For the core of the proposed theranostic system, stable aqueous suspensions of superparamagnetic iron oxide nanoparticles with an average diameter of 9 nm were produced. Chitosan-based poly-meric nanoparticles with a hydrodynamic diameter around 150 nm were successfully produced. Incorporation of iron oxide nanoparticles into the polymeric ones increased their hydrodynamic diameter to at least 250 nm. A monoclonal antibody specific for a transmembranar protein (car-bonic anhydrase IX) present in solid tumors was developed by hybridoma technology. Functional hybridomas producing the desired monoclonal antibodies were obtained. The proposed theranostic system functionality was evaluated in separated parts of its components. Uncoated and coated iron oxide nanoparticles with chitosan-based polymers generated heat under the application of an external alternating magnetic field. Uncoated iron oxide nanoparticles sta-bilized with oleic acid were able to enhance contrast in magnetic resonance imaging. Drug deliv-ery studies were conducted in chitosan-based polymeric nanoparticles without and with the in-corporation of iron oxide nanoparticles, demonstrating to be an effective drug delivery platform for doxorubicin. The theranostic system proposed in this PhD thesis is very promising for cancer theranostic, demonstrating to be applicable in solid tumors such as osteosarcoma.

Corporate governance failure at PT

The following case study depicts the bitter transfiguration of Portugal Telecom, SGPS (PT), a multinational telecommunications company that was once an honourable flag of innovation, corporate governance standards in Portugal and overseas. It scrutinises the controversial episodes that paved the way for the pitiful condition in which PT is nowadays: a company that carries the weight of a ruinous €897 million investment in a defaulted company and no more than a 25.6% stake in a heavily indebted Brazilian carrier. The free-fall is made, ironically, of a complete disregard for best corporate governance practices, allowing for PT’s major shareholders to take over the helm of the company, using it selfishly as a cash cow.

PT Multimedia

The purpose of the present case – and accompanying Teaching Notes – is to better understand the spin-off of PT Multimédia, by Portugal Telecom, after receiving a Public Takeover Offer from Sonaecom, in 2006. The Government and the Competition Authority had never looked in a serious way at PT’s dominant position and the lack of room for competition in the TMT sector – PT was the owner of both the cable and copper networks, having access to privileged information from its competitors with control over the wholesale and retail businesses. In 2006, the company received a takeover offer from Sonaecom, the TMT subsidiary from the Portuguese conglomerate Sonae. The offer was voted and rejected by a majority of PT shareholders, but the whole process triggered several recommendations from the regulatory bodies. As a result, PT divested its cable business with the spin-off of PT Multimédia, giving birth to a new competitor and a totally different landscape in the telecommunications sector in Portugal.