Making the cake and eating it as well

Educação Médica 1991; 2(2): p.3-4.

A first analysis of the atmospheric measurements from TANSO/GOSAT

Dissertação apresentada na Faculdade de Ciências e Tecnologia da Universidade Nova de Lisboa para obtenção do grau em Mestre em Engenharia Física

Liaisons dangereuses, conservation of modern and contemporary art: a study of the synthetic binding media in Portugal

Dissertação apresentada para obtenção do Grau de Doutor em Conservação e Restauro, especialidade de Ciências da Conservação, pela Universidade Nova de Lisboa, Faculdade de Ciências e Tecnologia

Biological activity of well defined hydantoin derivatives on efflux pump systems of bacteria and cancer cells

A multi-resistência a antibióticos e medicamentos usados em quimioterapia é um dos grandes problemas com os quais as instituições de saúde se debatem hoje em dia. A acção provocada por bombas de efluxo é uma das suas causas. Estas bombas têm uma importância fundamental, uma vez que, ao expelirem todo o tipo de tóxicos para o exterior das células, também expelem medicamentos, fazendo com que estes não tenham o efeito desejado dentro delas. As bombas de efluxo são transportadores que se encontram nas membranas de todo o tipo de células. Existem dois grandes tipos de bombas de efluxo: as primárias e as secundárias. As primeiras conferem multi-resistência principalmente em células eucariotas, como as células do cancro em humanos, tendo como função a mediação da repulsa de substâncias tóxicas por intermédio da hidrólise de ATP. A primeira a ser descoberta e mais estudada destas bombas foi a ABCB1 que é o gene que codifica a glicoproteína-P (P de permeabilidade). Enquanto as secundárias, que são a maior fonte de multi-resistência em bactérias, promovem a extrusão de substâncias tóxicas através da força motriz de protões. Neste tipo de bombas são conhecidas quatro famílias principais, das quais uma das mais importantes é a superfamília RND, uma vez que inclui a bomba AcrAB-TolC, que é muito importante no metabolismo xenobiótico de bactérias Gramnegativas, nomeadamente a E.coli. Com o objectivo de reverter a multi-resistência, tanto em células eucariotas como procariotas, têm-se desenvolvido estratégias de combate que envolvem a descoberta de substâncias que inibam as bombas de efluxo. Assim sendo, ao longo dos tempos têm sido descobertas variadas substâncias que cumprem este objectivo. É o caso, por exemplo, dos derivados de fluoroquinolonas usados como inibidores de bombas de efluxo em bactérias ou do Tamoxifen, utilizado na terapia de pacientes com cancro da mama. Um dos grupos de substâncias estudados para o desenvolvimento de possíveis compostos que actuem como reversores de multi-resistência são os compostos derivados de hidantoínas. Estes, são conhecidos por possuírem uma grande variedade de propriedades bioquímicas e farmacológicas, sendo portanto usados para tratarem algumas doenças em humanos, como a epilepsia. Nestes, estão englobados compostos com actividade anti-convulsão que constitui a sua grande mais-valia e, dependente da substituição no anel que os constitui, uma grande variedade de outras propriedades farmacológicas como a anti-fungica, a anti-arritmica, a anti-viral, a anti-diabética ou por exemplo a antagonização de determinados receptores, como os da serotonina. Apesar de pouco usados em estudos experimentais para desenvolver substâncias anti-carcinogénicas, existem alguns estudos com este efeito. Objectivos: O presente projecto envolve o estudo de bombas de efluxo primárias e secundárias, em células eucariotas e procariotas, respectivamente. Em bactérias, foram usados quatro modelos experimentais: Staphylococcus aureus ATCC 25923, Enterococcus faecalis ATCC 29212, E. coli AG 100 e Salmonella Enteritidis NCTC 13349. Em células de cancro foram usadas, células T de linfoma de rato parentais e células T de linfoma de rato transfectadas com o gene humano MDR-1. O principal objectivo deste estudo foi a pesquisa de novos moduladores de bombas de efluxo presentes em bactérias e células do cancro, tentando assim contribuir para o desenvolvimento de novos agentes farmacológicos que consigam reverter a multi-resistência a medicamentos. Assim sendo foram testados trinta compostos derivados de hidantoínas: SZ-2, SZ-7, LL-9, BS-1, JH-63, MN-3, TD-7k, GG-5k, P3, P7, P10, P11, RW-15b, AD-26, RW-13, AD-29, KF-2, PDPH-3, Mor-1, KK-XV, Thioam-1, JHF-1, JHC-2, JHP-1, Fur-2, GL-1, GL-7, GL-14, GL-16, GL-18. Como forma de atingir estes objectivos, a actividade biológica dos trinta compostos derivados de hidantoínas foi avaliada nas quatro estirpes de bactérias da seguinte forma: foram determinadas as concentrações mínimas inibitórias dos trinta compostos como forma de definir as concentrações em que os compostos seriam utilizados. Os compostos foram posteriormente testadas com um método fluorométrico de acumulação de brometo de etídeo, que é um substrato comum em bombas de efluxo bacterianas, desenvolvido por Viveiros et al. A actividade biológica dos compostos derivados de hidantoínas nas células de cancro foi demonstrada por diferentes métodos. O efeito anti-proliferativo e citotóxico dos trinta compostos foi avaliado nas células T de linfoma de rato transfectadas com o gene humano MDR-1 pelo método de thiazolyl de tetrazólio (MTT). Como o brometo de etídeo também é expelido pelos transportadores ABC, estes compostos foram posteriormente testados com um método fluorométrico de acumulação de brometo de etídeo desenvolvido por Spengler et al nos dois diferentes tipos de células eucariotas. Resultados: A maioria dos compostos derivados de hidantoínas foi eficaz na modulação de bombas de efluxo, nas duas estirpes de bactérias Gram-negativas e nos dois diferentes tipos de células T de linfoma. Em contraste com estes resultados, nas duas estirpes de células Gram-positivas, a maioria dos compostos tiveram pouco efeito na inibição de bombas de efluxo ou até nenhum, em muitos dos casos. De uma maneira geral os melhores compostos nas diferentes estirpes de bactérias foram: Thioam-1, SZ-2, P3, Rw-15b, AD-26, AD-29, GL-18, GL-7, KF-2, SZ-7, MN-3, GL-16 e GL- 14. Foram portanto estes os compostos que provocaram maior acumulação de brometo de etídeo, inibindo assim com maior eficácia as bombas de efluxo. No presente estudo, a maioria dos compostos conseguiu inibir a resistência provocada pela bomba de efluxo ABCB1, tanto nas células parentais bem como nas células que sobre-expressam esta bomba, causando a acumulação de brometo de etídeo dentro das células. As células que sobreexpressam a bomba ABCB1 foram posteriormente testadas com citometria de fluxo que é a técnica padrão para pesquisa de inibidores de bombas de efluxo. Os compostos que foram mais efectivos na inibição da bomba ABCB1, causando assim maior acumulação de brometo de etídeo nas células que sobre-expressam esta bomba foram: PDPH-3, GL-7, KK-XV, AD-29, Thioam-1, SZ-7, KF-2, MN-3, RW-13, LL-9, P3, AD-26, JH-63 e RW- 15b. Este facto não corroborou totalmente os resultados da citometria de fluxo uma vez que os moduladores que provocaram maior inibição da bomba ABCB1 foram o MN-3, JH-63 e o BS-1, sendo que o último não foi seleccionado como um bom composto usando o método fluorométrico de acumulação de brometo de etídeo. Conclusão: Os compostos derivados de hidantoínas testados tiveram maior efeito nas estirpes de bactérias Gram-negativas do que nas Gram-positivas. Relativamente às células eucariotas, as estruturas mais activas apresentam substituintes aromáticos bem como alguns fragmentos aminicos terciários.

Development of new biomaterials based on liquid crystalline phases of cellulose derivatives

Dissertação apresentada na Faculdade de Ciências e Tecnologia da Universidade Nova de Lisboa para obtenção do Grau de Mestre em Engenharia Biomédica

Negative non-ground queries in well founded semantics

Dissertação apresentada na Faculdade de Ciências e Tecnologia da Universidade Nova de Lisboa para obtenção do grau de Mestre em Computational Logic

Three transducers for one photodetector: essays for optical communications

Dissertation presented to obtain the PhD degree in Electrical and Computer Engineering - Electronics

Stratigraphic interpretation of Well-Log data of the Athabasca Oil Sands of Alberta Canada through Pattern recognition and Artificial Intelligence

Dissertation submitted in partial fulfillment of the requirements for the Degree of Master of Science in Geospatial Technologies.

Pre-treatment of different types of lignocellulosic biomass using ionic liquids

Dissertação apresentada na Faculdade de Ciências e Tecnologia da Universidade Nova de Lisboa para obtenção do grau Mestre em Biotecnologia

Preparation and characterization of microfiltration flat polymeric membranes for biomedical applications

Dissertation presented to Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa for obtaining the master degree in Membrane Engineering

Fate of nitrogen in bioreactor landfills: lab-scale in situ aeration in well decomposed MSW

Dissertação para obtenção do Grau de Mestre em Engenharia do ambiente, perfil de engenharia sanitária

Fiber functionalization for biomedical applications

Deep-eutectic solvents (DES) are considered novel renewable and biodegradable solvents, with a cheap and easy synthesis, without waste production. Later it was discovered a new subclass of DES that even can be biocompatible, since their synthesis uses primary metabolites such as amino acids, organic acids and sugars, from organisms. This subclass was named natural deep-eutectic solvents (NADES). Due to their properties it was tried to study the interaction between these solvents and biopolymers, in order to produce functionalized fibers for biomedical applications. In this way, fibers were produced by using the electrospinning technique. However, it was first necessary to study some physical properties of NADES, as well as the influence of water in their properties. It has been concluded that the water has a high influence on NADES properties, which can be seen on the results obtained from the rheology and viscosity studies. The fluid dynamics had changed, as well as the viscosity. Afterwards, it was tested the viability of using a starch blend. First it was tested the dissolution of these biopolymers into NADES, in order to study the viability of their application in electrospinning. However the results obtained were not satisfactory, since the starch polymers studied did not presented any dissolution in any NADES, or even in organic solvents. In this way it was changed the approach, and it was used other biocompatible polymers. Poly(ethylene oxide), poly(vinyl alcohol) and gelatin were the others biopolymers tested for the electrospinning, with NADES. All polymers show good results, since it was possible to obtain fibers. However for gelatin it was used only eutectic mixtures, containing active pharmaceutical ingredients (API’s), instead of NADES. For this case it was used mandelic acid (antimicrobial properties), choline chloride, ibuprofen (anti-inflammatory properties) and menthol (analgesic properties). The polymers and the produced fibers were characterized by scanning electron microscope (SEM), Transmission electron microscopy (TEM) and Fourier transform infrared spectroscopy (FTIR). With the help of these techniques it was possible to conclude that it was possible to encapsulate NADES within the fibers. Rheology it was also study for poly(ethylene oxide) and poly(vinyl alcohol), in a way to understand the influence of polymer concentration, on the electrospinning technique. For the gelatin, among the characterization techniques, it was also performed cytotoxicity and drug release studies. The gelatin membranes did not show any toxicity for the cells, since their viability was maintained. Regarding the controlled release profile experiment no conclusion could be drawn from the experiments, due to the rapid and complete dissolution of the gelatin in the buffer solution. However it was possible to quantify the mixture of choline chloride with mandelic acid, allowing thus to complete, and confirm, the information already obtained for the others characterization technique.

Thermal performance of cool facades: evaluation by Infrared Thermography

High reflective paints (cool paints) are used on flat roofs to reduce heat gains from the incidence of solar radiation and thus improve the thermal comfort and energy efficiency of buildings, especially in summer periods. Given the application potential of these paints on vertical surfaces, a research study has been developed to evaluate the thermal performance of reflective paints on walls under real exposure conditions. Accordingly, different reflective paints have been applied as the final coating of an ETICS type solution, on the facades of a full scale experimental cell built at LNEC campus. For being applied in an ETICS system a paint has to fulfill several requirements, whether aesthetic or functional (such as the adhesion between the coating layers or the durability of the insulation), essential for its efficient performance. Since this construction coating system is subject to a prolonged sun exposure, various problems may arise, such as paint degradation or deterioration of the thermal insulation properties, particularly when dark colors are applied. To evaluate the thermal performance of the chosen paints, the method of non-destructive analysis by Infrared Thermography was used. Thermography allows knowing the temperature distribution of facades by measuring the radiation emitted by their surfaces. To complement the thermographic diagnosis, thermocouples were placed between the insulation and the paint system of the experimental cell. Additional laboratory tests allowed the characterization of the optical properties (reflectance and emittance) of the different reflective paints used in this study. The comparative analysis of the thermal performance of reflective and conventional paints revealed that the reflective paint allows a reduction of the facade surface temperature, reducing the risk of loss of insulating properties of the ETICS system and thus ensuring its longevity and functionality. The color of the paint used affects, naturally, the reflective ability of the surface and may have an important role in energy balance of the building. This paper also showed the potential of infrared thermography in the evaluation of the thermal performance of reflective paints.

Storage of water molecules into biomimetic heterostructures: the role of roughness

The development of devices based on heterostructured thin films of biomolecules conveys a huge contribution on biomedical field. However, to achieve high efficiency of these devices, the storage of water molecules into these heterostructures, in order to maintain the biological molecules hydrated, is mandatory. Such hydrated environment may be achieved with lipids molecules which have the ability to rearrange spontaneously into vesicles creating a stable barrier between two aqueous compartments. Yet it is necessary to find conditions that lead to the immobilization of whole vesicles on the heterostructures. In this work, the conditions that govern the deposition of open and closed liposomes of 1.2-dipalmitoyl-sn-Glycero-3-[Phospho-rac-(1-glycerol)] (sodium Salt) (DPPG) onto polyelectrolytes cushions prepared by the layer-by-layer (LbL) method were analyzed. Electronic transitions of DPPG molecules as well as absorption coefficients were obtained by vacuum ultraviolet spectroscopy, while the elemental composition of the heterostructures was characterized by x-ray photoelectron spectroscopy (XPS). The presence of water molecules in the films was inferred by XPS and infrared spectroscopy. Quartz crystal microbalance (QCM) data analysis allowed to conclude that, in certain cases, the DPPG adsorbed amount is dependent of the bilayers number already adsorbed. Moreover, the adsorption kinetics curves of both adsorbed amount and surface roughness allowed to determine the kinetics parameters that are related with adsorption processes namely, electrostatic forces, liposomes diffusion and lipids re-organization on surface. Scaling exponents attained from atomic force microscopy images statistical analysis demonstrate that DPPG vesicles adsorption mechanism is ruled by the diffusion Villain model confirming that adsorption is governed by electrostatic forces. The power spectral density treatment enabled a thorough description of the accessible surface of the samples as well as of its inner structural properties. These outcomes proved that surface roughness influences the adsorption of DPPG liposomes onto surfaces covered by a polyelectrolyte layer. Thus, low roughness was shown to induce liposome rupture creating a lipid bilayer while high roughness allows the adsorption of whole liposomes. In addition, the fraction of open liposomes calculated from the normalized maximum adsorbed amounts decreases with the cushion roughness increase, allowing us to conclude that the surface roughness is a crucial variable that governs the adsorption of open or whole liposomes. This conclusion is fundamental for the development of well-designed sensors based on functional biomolecules incorporated in liposomes. Indeed, LbL films composed of polyelectrolytes and liposomes with and without melanin encapsulated were successfully applied to sensors of olive oil.

Characterization of molecular damage induced by UV photons and carbon ions on biomimetic heterostructures

The study of the effect of radiation on living tissues is a rather complex task to address mainly because they are made of a set of complex functional biological structures and interfaces. Particularly if one is looking for where damage is taking place in a first stage and what are the underlying reaction mechanisms. In this work a new approach is addressed to study the effect of radiation by making use of well identified molecular hetero-structures samples which mimic the biological environment. These were obtained by assembling onto a solid support deoxyribonucleic acid (DNA) and phospholipids together with a soft water-containing polyelectrolyte precursor in layered structures and by producing lipid layers at liquid/air interface with DNA as subphase. The effects of both ultraviolet (UV) radiation and carbon ions beams were systematically investigated in these heterostructures, namely damage on DNA by means vacuum ultraviolet (VUV), infrared (IR), X-Ray Photoelectron (XPS) and impedance spectroscopy. Experimental results revealed that UV affects furanose, PO2-, thymines, cytosines and adenines groups. The XPS spectrometry carried out on the samples allowed validate the VUV and IR results and to conclude that ionized phosphate groups, surrounded by the sodium counterions, congregate hydration water molecules which play a role of UV protection. The ac electrical conductivity measurements revealed that the DNA electrical conduction is arising from DNA chain electron hopping between base-pairs and phosphate groups, with the hopping distance equal to the distance between DNA base-pairs and is strongly dependent on UV radiation exposure, due loss of phosphate groups. Characterization of DNA samples exposed to a 4 keV C3+ ions beam revealed also carbon-oxygen bonds break, phosphate groups damage and formation of new species. Results from radiation induced damage carried out on biomimetic heterostructures having different compositions revealed that damage is dependent on sample composition, with respect to functional targeted groups and extent of damage. Conversely, LbL films of 1,2-dipalmitoyl-sn-Glycero-3-[Phospho-rac-(1-glycerol)] (Sodium Salt) (DPPG) liposomes, alternated with poly(allylamine hydrochloride) (PAH) revealed to be unaffected, even by prolonged UV irradiation exposure, in the absence of water molecules. However, DPPG molecules were damaged by the UV radiation in presence of water with cleavage of C-O, C=O and –PO2- bonds. Finally, the study of DNA interaction with the ionic lipids at liquid/air interfaces revealed that electrical charge of the lipid influences the interaction of phospholipid with DNA. In the presence of DNA in the subphase, the effects from UV irrladiation were seen to be smaller, which means that ionic products from biomolecules degradation stabilize the intact DPPG molecules. This mechanism may explain why UV irradiation does not cause immediate cell collapse, thus providing time for the cellular machinery to repair elements damaged by UV.