Assessment of Notch1 ligands production - key protein targets in breast cancer

Cell-to-cell communication is required for many biological processes in development and adult life. One of the most common systems utilized by a wide range of eukaryotes is the Notch signalling pathway. Four Notch receptors and five ligands have been identified in mammals that interact via their extracellular domains leading to transcription activation. Studies have shown that the Notch ligands expression is undetectable in normal breast tissues, but moderate to high expression has been detected in breast cancer. Thus, any of the Notch1 ligands can be studied as possible therapeutic targets for breast cancer. To study Notch pathway proteins there is the need to obtain stable protein solutions. E. coli is the host of excellence for recombinant proteins for the ease of use, fast growth and high cell densities. However, the expression of mammalian proteins in such systems may overwhelm the bacterial cellular machinery, which does not possess the ability for post-translational modifications, or dedicated compartments for protein synthesis. Mammalian cells are therefore preferred, despite their technical and financial increased demands. We aim to determine the best expression and purification conditions for the different ligand protein constructs, to develop specific function-blocking antibodies using the Phage Display technology. Moreover, we propose to crystallize the Notch1 ligands alone and in complex with the phage display selected antibodies, unveiling molecular details. hJag2DE3 and hDll1DE6 proteins were purified from refolded inclusion bodies or mammalian cell culture supernatants, respectively, and purity was confirmed by SDS-PAGE (>95%). Protein produced in mammalian cells showed to be more stable, apparently with the physiological disulfide pattern, contrary to what was observed in the refolded protein. Several nano-scale crystallization experiments were set up in 96-well plates, but no positive result was obtained. We will continue to pursue for the best expression for the Notch ligand constructs in both expression systems.

Res Publica: Citizenship and Political Representation in Portugal, 1820-1926

The term res publica (literally “thing of the people”) was coined by the Romans to translate the Greek word politeia, which, as we know, referred to a political community organised in accordance with certain principles, amongst which the notion of the “good life” (as against exclusively private interests) was paramount. This ideal also came to be known as political virtue. To achieve it, it was necessary to combine the best of each “constitutional” type and avoid their worst aspects (tyranny, oligarchy and ochlocracy). Hence, the term acquired from the Greeks a sense of being a “mixed” and “balanced” system. Anyone that was entitled to citizenship could participate in the governance of the “public thing”. This implied the institutionalization of open debate and confrontation between interested parties as a way of achieving the consensus necessary to ensure that man the political animal, who fought with words and reason, prevailed over his “natural” counterpart. These premises lie at the heart of the project which is now being presented under the title of Res Publica: Citizenship and Political Representation in Portugal, 1820-1926. The fact that it is integrated into the centenary commemorations of the establishment of the Republic in Portugal is significant, as it was the idea of revolution – with its promise of rupture and change – that inspired it. However, it has also sought to explore events that could be considered the precursor of democratization in the history of Portugal, namely the vintista, setembrista and patuleia revolutions. It is true that the republican regime was opposed to the monarchic. However, although the thesis that monarchy would inevitably lead to tyranny had held sway for centuries, it had also been long believed that the monarchic system could be as “politically virtuous” as a republic (in the strict sense of the word) provided that power was not concentrated in the hands of a single individual. Moreover, various historical experiments had shown that republics could also degenerate into Caesarism and different kinds of despotism. Thus, when absolutism began to be overturned in continental Europe in the name of the natural rights of man and the new social pact theories, initiating the difficult process of (written) constitutionalization, the monarchic principle began to be qualified as a “monarchy hedged by republican institutions”, a situation in which not even the king was exempt from isonomy. This context justifies the time frame chosen here, as it captures the various changes and continuities that run through it. Having rejected the imperative mandate and the reinstatement of the model of corporative representation (which did not mean that, in new contexts, this might not be revived, or that the second chamber established by the Constitutional Charter of 1826 might not be given another lease of life), a new power base was convened: national sovereignty, a precept that would be shared by the monarchic constitutions of 1822 and 1838, and by the republican one of 1911. This followed the French example (manifested in the monarchic constitution of 1791 and in the Spanish constitution of 1812), as not even republicans entertained a tradition of republicanism based upon popular sovereignty. This enables us to better understand the rejection of direct democracy and universal suffrage, and also the long incapacitation (concerning voting and standing for office) of the vast body of “passive” citizens, justified by “enlightened”, property- and gender-based criteria. Although the republicans had promised in the propaganda phase to alter this situation, they ultimately failed to do so. Indeed, throughout the whole period under analysis, the realisation of the potential of national sovereignty was mediated above all by the individual citizen through his choice of representatives. However, this representation was indirect and took place at national level, in the hope that action would be motivated not by particular local interests but by the common good, as dictated by reason. This was considered the only way for the law to be virtuous, a requirement that was also manifested in the separation and balance of powers. As sovereignty was postulated as single and indivisible, so would be the nation that gave it soul and the State that embodied it. Although these characteristics were common to foreign paradigms of reference, in Portugal, the constitutionalization process also sought to nationalise the idea of Empire. Indeed, this had been the overriding purpose of the 1822 Constitution, and it persisted, even after the loss of Brazil, until decolonization. Then, the dream of a single nation stretching from the Minho to Timor finally came to an end.

Monotherapy and combined therapy of new potential antitumor compounds: antiproliferative activities and biological targets

Review article Martins, P., Marques, M., Coito, L., Pombeiro, A.J.L., Baptista, P.V., Fernandes, A.R. 2014. Organometallic Compounds in Cancer Therapy: Past Lessons and Future Directions. Anti-cancer Agents in Medicinal Chemistry 14. PMID: 25173559

Immunohistochemistry detection of putative miR-200c and miR-203 targets in breast cancer patients

Part of this thesis will be published in the following: Gomes, B.C., Santos, B. 2015. Methods for studying microRNAs expression and their targets in formalin-fixed, paraffin-embedded (FFPE) breast cancer tissues. In Methods in Molecular Biology: Cancer Drug Resistance (Rueff, J. & Rodrigues, A.S. eds), Springer Protocols.

The Vase’s Representation (Cantharus, Crater) on the Roman Mosaic in Portugal: A Significant Formal and Iconographic Path from Classic Antiquity to Late Antiquity

11TH INTERNATIONAL COLLOQUIUM ON ANCIENT MOSAICS OCTOBER 16TH  20TH, 2009, BURSA TURKEY Mosaics of Turkey and Parallel Developments in the Rest of the Ancient and Medieval World: Questions of Iconography, Style and Technique from the Beginnings of Mosaic until the Late Byzantine Era

Popular music exhibitions in Portugal: practices of representation from 2007 to 2013

This thesis focuses on the representation of Popular Music in museums by mapping, analyzing, and characterizing its practices in Portugal at the beginning of the 21st century. Now that museums' ability to shape public discourse is acknowledged, the examination of popular music's discourses in museums is of the utmost importance for Ethnomusicology and Popular Music Studies as well as for Museum Studies. The concept of 'heritage' is at the heart of this processes. The study was designed with the aim of moving the exhibiting of popular music in museums forward through a qualitative inquiry of case studies. Data collection involved surveying pop-rock music exhibitions as a qualitative sampling of popular music exhibitions in Portugal from 2007 to 2013. Two of these exhibitions were selected as case studies: No Tempo do Gira-Discos: Um Percurso pela Produção Fonográfica Portuguesa at the Museu da Música in Lisbon in 2007 (also Faculdade de Letras, 2009), and A Magia do Vinil, a Música que Mudou a Sociedade at the Oficina da Cultura in Almada in 2008 (and several other venues, from 2009 to 2013). Two specific domains were observed: popular music exhibitions as instances of museum practice and museum professionals. The first domain encompasses analyzing the types of objects selected for exhibition; the interactive museum practices fostered by the exhibitions; the concepts and narratives used to address popular music discursively, as well as the interpretative practices they allow. The second domain, focuses museum professionals and curators of popular music exhibitions as members of a group, namely their goals, motivations and perspectives. The theoretical frameworks adopted were drawn from the fields of ethnomusicology, popular music studies, and museum studies. The written materials of the exhibitions were subjected of methods of discourse analysis methods. Semi-structured interviews with curators and museum professional were also conducted and analysed. From the museum studies perspective, the study research suggests that the practice adopted by popular music museums largely matches that of conventional museums. From the ethnomusicological and popular music studies stand point, the two case studies reveal two distinct conceptual worlds: the first exhibition, curated by an academic and an independent researcher, points to a mental configuration where popular music is explained through a framework of genres supported by different musical practices. Moreover, it is industry actors such as decision makers and gatekeepers that govern popular music, which implies that the visitors' romantic conception of the musician is to some extent dismantled; the second exhibition, curated by a record collector and specialist, is based on a more conventional process of the everyday historical speech that encodes a mismatch between “good” and “bad music”. Data generated by a survey shows that only one curator, in fact that of my first case study, has an academic background. The backgrounds of all the others are in some way similar to the curator of the second case study. Therefore, I conclude that the second case study best conveys the current practice of exhibiting Popular Music in Portugal.

Tool for spatial and dynamic representation of artistic performances

This proposal aims to explore the use of available technologies for video representation of sets and performers in order to serve as support for composition processes and artistic performer rehearsals, while focusing in representing the performer’s body and its movements, and its relation with objects belonging to the three-dimensional space of their performances. This project’s main goal is to design and develop a system that can spatially represent the performer and its movements, by means of capturing processes and reconstruction using a camera device, as well as enhance the three-dimensional space where the performance occurs by allowing interaction with virtual objects and by adding a video component, either for documentary purposes, or for live performances effects (for example, using video mapping video techniques in captured video or projection during a performance).

Al-doped ZnO ceramic sputtering targets based on nanocrystalline powders produced by emulsion detonation synthesis – deposition and application as a transparent conductive oxide material

Transparent conducting oxides (TCOs) have been largely used in the optoelectronic industry due to their singular combination of low electrical resistivity and high optical transmittance. They are usually deposited by magnetron sputtering systems being applied in several devices, specifically thin film solar cells (TFSCs). Sputtering targets are crucial components of the sputtering process, with many of the sputtered films properties dependent on the targets characteristics. The present thesis focuses on the development of high quality conductive Al-doped ZnO (AZO) ceramic sputtering targets based on nanostructured powders produced by emulsion detonation synthesis method (EDSM), and their application as a TCO. In this sense, the influence of several processing parameters was investigated from the targets raw-materials synthesis to the application of sputtered films in optoelectronic devices. The optimized manufactured AZO targets present a final density above 99 % with controlled grain size, an homogeneous microstructure with a well dispersed ZnAl2O4 spinel phase, and electrical resistivities of ~4 × 10-4 Ωcm independently on the Al-doping level among 0.5 and 2.0 wt. % Al2O3. Sintering conditions proved to have a great influence on the properties of the targets and their performance as a sputtering target. It was demonstrated that both deposition process and final properties of the films are related with the targets characteristics, which in turn depends on the initial powder properties. In parallel, the influence of several deposition parameters in the film´s properties sputtered from these targets was investigated. The sputtered AZO TCOs showed electrical properties at room temperature that are superior to simple oxides and comparable to a reference TCO – indium tin oxide (ITO), namely low electrical resistivity of 5.45 × 10-4 Ωcm, high carrier mobility (29.4 cm2V-1s-1), and high charge carrier concentration (3.97 × 1020 cm-3), and also average transmittance in the visible region > 80 %. These superior properties allowed their successful application in different optoelectronic devices.

Ontology-based representation and generation of workflows for micro-task human-machine computation

Doctoral Program in Computer Science

Prediction of drug targets in human pathogens

The identification of new and druggable targets in bacteria is a critical endeavour in pharmaceutical research of novel antibiotics to fight infectious agents. The rapid emergence of resistant bacteria makes today's antibiotics more and more ineffective, consequently increasing the need for new pharmacological targets and novel classes of antibacterial drugs. A new model that combines the singular value decomposition technique with biological filters comprised of a set of protein properties associated with bacterial drug targets and similarity to protein-coding essential genes of E. coli has been developed to predict potential drug targets in the Enterobacteriaceae family [1]. This model identified 99 potential target proteins amongst the studied bacterial family, exhibiting eight different functions that suggest that the disruption of the activities of these proteins is critical for cells. Out of these candidates, one was selected for target confirmation. To find target modulators, receptor-based pharmacophore hypotheses were built and used in the screening of a virtual library of compounds. Postscreening filters were based on physicochemical and topological similarity to known Gram-negative antibiotics and applied to the retrieved compounds. Screening hits passing all filters were docked into the proteins catalytic groove and 15 of the most promising compounds were purchased from their chemical vendors to be experimentally tested in vitro. To the best of our knowledge, this is the first attempt to rationalize the search of compounds to probe the relevance of this candidate as a new pharmacological target.

3D representation of the urban evolution of Braga using the cityengine tool

The morphological evolution of the city of Braga has been the subject of several studies focusing on different urban areas in different periods. Using the accumulated knowledge provided by the available archaeological, historical and iconographic data of Braga, from the Roman times to the nineteenth century, we intend to present a working methodology for 3D representation of urban areas and its evolution, using the CityEngine ESRI tool. Different types of graphic and cartographic data will be integrated in an archaeological information system for the characterization of urban buildings. Linking this information system to the rules of characterization of urban spaces through the CityEngine tool, we can create the 3D urban spaces and their changes. The building characterization rules include several parameters of architectural elements that can be dynamically changed according the latest information. This methodology will be applied to the best known areas within of the city allowing the creation of different and dynamic layouts. Considerations about the concepts, challenges and constraints of using the CityEngine tool for recording and representing urban evolution knowledge will be discussed.

A glycolytic phenotype is associated with prostate cancer progression and aggressiveness: a role for Monocarboxylate Transporters as metabolic targets for therapy.

Metabolic adaptation is considered an emerging hallmark of cancer, whereby cancer cells exhibit high rates of glucose consumption with consequent lactate production. To ensure rapid efflux of lactate, most cancer cells express high levels of monocarboxylate transporters (MCTs), which therefore may constitute suitable therapeutic targets. The impact of MCT inhibition, along with the clinical impact of altered cellular metabolism during prostate cancer (PCa) initiation and progression, has not been described. Using a large cohort of human prostate tissues of different grades, in silico data, in vitro and ex vivo studies, we demonstrate the metabolic heterogeneity of PCa and its clinical relevance. We show an increased glycolytic phenotype in advanced stages of PCa and its correlation with poor prognosis. Finally, we present evidence supporting MCTs as suitable targets in PCa, affecting not only cancer cell proliferation and survival but also the expression of a number of hypoxia-inducible factor target genes associated with poor prognosis. Herein, we suggest that patients with highly glycolytic tumours have poorer outcome, supporting the notion of targeting glycolytic tumour cells in prostate cancer through the use of MCT inhibitors.

SMYD3 contributes to a more aggressive phenotype of prostate cancer and targets Cyclin D2 through H4K20me3

Prostate cancer (PCa) is one of the most incident cancers worldwide but clinical and pathological parameters have limited ability to discriminate between clinically significant and indolent PCa. Altered expression of histone methyltransferases and histone methylation patterns are involved in prostate carcinogenesis. SMYD3 transcript levels have prognostic value and discriminate among PCa with different clinical aggressiveness, so we decided to investigate its putative oncogenic role on PCa.We silenced SMYD3 and assess its impact through in vitro (cell viability, cell cycle, apoptosis, migration, invasion assays) and in vivo (tumor formation, angiogenesis). We evaluated SET domain's impact in PCa cells' phenotype. Histone marks deposition on SMYD3 putative target genes was assessed by ChIP analysis.Knockdown of SMYD3 attenuated malignant phenotype of LNCaP and PC3 cell lines. Deletions affecting the SET domain showed phenotypic impact similar to SMYD3 silencing, suggesting that tumorigenic effect is mediated through its histone methyltransferase activity. Moreover, CCND2 was identified as a putative target gene for SMYD3 transcriptional regulation, through trimethylation of H4K20.Our results support a proto-oncogenic role for SMYD3 in prostate carcinogenesis, mainly due to its methyltransferase enzymatic activity. Thus, SMYD3 overexpression is a potential biomarker for clinically aggressive disease and an attractive therapeutic target in PCa.

Ontological representation of tumor-node-metastasis classification and an ontology-driven classifier: a study on colorectal cancer

Dissertação de mestrado integrado em Engenharia Biomédica (área de especialização em Informática Médica)

Comprensión en resolución de problemas en Física: niveles de representación e inferencias basadas en el conocimiento. Comprehension in Physics Problem-Solving: levels of representation and knowledge-based inferences.

Se propone desarrollar e integrar estudios sobre Modelado y Resolución de Problemas en Física que asumen como factores explicativos: características de la situación planteada, conocimiento de la persona que resuelve y proceso puesto en juego durante la resolución. Interesa comprender cómo los estudiantes acceden al conocimiento previo, qué procedimientos usan para recuperar algunos conocimientos y desechar otros, cuáles son los criterios que dan coherencia a sus decisiones, cómo se relacionan estas decisiones con algunas características de la tarea, entre otras. Todo ello con miras a estudiar relaciones causales entre las dificultades encontradas y el retraso o abandono en las carreras.Se propone organizar el trabajo en tres ejes, los dos primeros de construcción teórica y un tercero de implementación y transferencia. Se pretende.1.-Estudiar los procesos de construcción de las representaciones mentales en resolución de problemas de física, tanto en expertos como en estudiantes de diferentes niveles académicos.2.-Analizar y clasificar las inferencias que se producen durante las tareas de comprensión en resolución de problemas de física. Asociar dichas inferencias con procesos de transición entre representaciones mentales de diferente naturaleza.3.-Desarrollar materiales y diseños instruccionales en la enseñanza de la Física, fundamentado en un conocimiento de los requerimientos psicológicos de los estudiantes en diversas tareas de aprendizaje.En términos generales se plantea un enfoque interpretativo a la luz de marcos de la psicología cognitiva y de los desarrollos propios del grupo. Se trabajará con muestras intencionales de alumnos y profesores de física. Se utilizarán protocolos verbales y registros escritos producidos durante la ejecución de las tareas con el fin de identificar indicadores de comprensión, inferencias, y diferentes niveles de representación. Se prevé analizar material escrito de circulación corriente sea comercial o preparado por los docentes de las carreras involucradas.Las características del objeto de estudio y el distinto nivel de desarrollo en que se encuentran los diferentes ojetivos específicos llevan a que el abordaje contemple -según consideracion de Juni y Urbano (2006)- tanto la lógica cualitativa como la cuantitativa.