Esta dissertação centra-se na inter-relação entre texto literário (João Miguel Fernandes Jorge) e fotografia (Jorge Molder) e pretende mostrar como a liminaridade das artes pode ser um factor determinante nesta que consideramos ser uma relação de contaminação e contiguidade. Há diversas possibilidades de análise deste cruzamento interartes, mas a que mais nos interessa relaciona-se com as formas de representação do corpo que, como veremos, deixa de ter como objectivo último a representação de um projecto de corpo uno para descobrir novas formas de representação plurais e descentradas. Trata-se, assim, de entender o texto literário e a fotografia como espaços de reconfiguração do corpo. Pelo uso da palavra reconfiguração procura evitar-se uma redução da literatura e da fotografia à representação, entendendo-as antes como modos de fazer e pensar (mostrar e figurar) a construção do corpo. Desta perspectiva, interessa-nos pensar de que forma configuração e representação se equacionam face à liminaridade das artes e como se escreve o corpo e se dá a ver, no texto e na fotografia. Corpo, fragmento e texto/fotografia: é uma dinâmica a quatro termos que aqui equacionaremos. Tirando partido reflexivo do que o pensamento filosófico e antropológico contemporâneo tem avançado sobre uma possível teoria do corpo pós-moderno, pretendemos analisar as obras de João Miguel Fernandes Jorge e Jorge Molder à luz daquilo que se tem designado como um corpo em crise. Defenderemos que estamos perante um novo conceito de corpo que se nos revela através da sua natureza fragmentária. Constitui, assim, nosso propósito analisar, por um lado, os efeitos de dispersão temática e formal em João Miguel Fernandes Jorge (no poema e livro) e em Jorge Molder (na fotografia e série) e, por outro, as várias estratégias de reconfiguração do corpo (corpo-testemunho e corpo-em-cena).

Stroke is a significant cause of death and disability in developed countries. It is a multifactorial disease, resulting from the interplay between genes and well-known life-style/environmental risk factors. Numerous studies have attempted to identify the genetic risk factors predisposing to stroke, but few have investigated the genetic factors involved in stroke outcome. This work aimed at the identification of genes contributing to stroke and influencing patient’s outcome after three months. Four inflammatory genes (IL1B, IL6, MPO and TNF) and two genes involved in the nitric oxide metabolism (NOS1 and NOS3) were tested for association with stroke. The results suggest that the IL6 and MPO genes influence stroke susceptibility through independent effects and non-additive interactions. Furthermore, they provided novel evidence for the involvement of the NOS1 gene in stroke susceptibility. Several studies have shown the important impact of oxidative stress, inflammation, angiogenesis, neurogenesis, neurovascular damage and neurovascular remodeling for stroke-associated brain damage and/or stroke recovery. Association analyses were thus carried out to assess the role of candidate genes involved in inflammatory processes (IL1B, IL6, MPO and TNF) and oxidative stress (NOS1 and NOS3), as well as matrix metalloproteinase genes (MMP2 and MMP9) and growth factor genes (BDNF, FGF2 and VEGFA) in patient’s outcome at three months. MMP2 genetic variants were found associated with patient’s outcome, and the results also indicate that two epistatic interactions between the BDNF and FGF2 genes and between the FGF2 and VEGFA genes influence this phenotype. A genome-wide association study was performed in stroke outcome using DNA pooled samples to provide novel insights into the mechanisms involved in stroke recovery. The BBS9 and GLIS3 genes were found associated with patient’s outcome at three months. Taken together, these results suggest that stroke susceptibility and outcome are modulated by a combination of main gene effects and gene-gene interactions, independently of stroke risk factors and/or severity parameters, highlighting the complexity of mechanisms predisposing to stroke and influencing recovery afterwards.

A identificação dos mecanismos que permitem promover a emergência e a manutenção de comportamentos de cooperação é um problema com relevância em diversas disciplinas científicas tais como a Biologia, a Sociologia, a Economia e a Informática. Numa população de indivíduos que interagem entre si, estes decidem, de vez em quando, alterar a forma como agem nas interacções em que participam. Potencialmente, a relação temporal entre os processos de actualização que ocorrem na população pode influenciar a proliferação de comportamentos cooperativos uma vez que condiciona a informação disponível da qual estes processos dependem. Esta relação estabelece uma dinâmica que denominamos por dinâmica de actualização. A actualização diz-se síncrona se os agentes actualizam a sua estratégia em simultâneo e assíncrona em caso contrário. Neste trabalho investigamos a influência da dinâmica de actualização na evolução da cooperação utilizando como plataforma de trabalho a Teoria dos Jogos Evolucionária. Os resultados mostram que, em geral, quando a dinâmica de actualização tem alguma influência, o assincronismo é benéfico à evolução da cooperação, ou seja, que é um mecanismo de promoção de comportamentos de cooperação numa população de indivíduos. O resultados são suportados pela utilização de uma grande variedade de condições, a qual permite proceder a uma caracterização da forma como a influência da dinâmica de actualização varia com as condições do sistema e a uma identificação dos aspectos que desempenham um papel relevante nessa influência.

O direito de propriedade e as demais formas de apropriação privada do território e do espaço urbano constituem o tema central da dissertação. A abordagem do tema desenvolve-se num quadro de unidade do Direito indispensável à sua compreensão, incorporando as perspectivas do Direito Romano, da História do Direito Português, do Direito Civil e do Direito Constitucional, mas a análise do direito de propriedade é feita essencialmente à luz das normas legais e regulamentares que regem o planeamento e a gestão das cidades, pelo que tem por objecto principal o seu regime urbanístico. É, por isso, uma dissertação sobre o estatuto jurídico da propriedade imobiliária urbana no Direito do Urbanismo. Nesse contexto, são duas as principais preocupações evidenciadas ao longo da investigação. Por um lado, compreender a natureza jurídica do direito de construir a partir do estudo das restrições e limitações de interesse público urbanístico ao direito de propriedade privada e, por outro, avaliar em que medida a estrutura e o objecto desse direito são adequados à satisfação das necessidades colectivas inerentes à construção da cidade do século XXI. Uma das principais linhas de força da investigação é a de que o objecto do direito de propriedade imobiliária urbana deve tendencialmente corresponder ao conteúdo do aproveitamento urbanístico definido pelos instrumentos de planeamento elaborados pela Administração. Mais do que o solo, destinado preferencialmente a ser partilhado em regime de propriedade pública ou comunitária, é o espaço construído que deve constituir objecto de apropriação privada. Quando seja legalmente possível estabelecer essa correspondência, o direito de propriedade imobiliária urbana confunde-se com o direito de construir, que assim adquire a natureza de um direito real autónomo de realizar ou manter uma construção urbana em conformidade com o aproveitamento urbanístico definido pelo plano.

Microvesicular steatosis is a chronic hepatocellular change potentially induced by valproic acid (VPA). This effect has often been associated with a disturbed fatty acid metabolism and an altered dynamic balance of the essential co-factor coenzyme A (CoA). Furthermore, the parent drug or its metabolites may interact with mitochondrial metabolism at several sites, thereby deranging cellular functions. The experimental work presented in this thesis aimed to 1.) elucidate the impact of valproate on CoA and carnitine homeostasis, 2.) shed new light on the mitochondrial oxidative biotransformation of the drug and 3.) understand the interactions of valproyl-CoA with different functions of mitochondria. Valproate-induced mitochondrial dysfunction is often related to a potential secondary carnitine deficiency. The study of valproate interference on mitochondrial metabolism and CoA homeostasis was first focused on the evaluation of carnitine status in animal tissues using electrospray tandem mass spectrometry, since both cofactors are strictly related. Our studies showed a significant dose-related increase of free carnitine and total acylcarnitine levels in the livers of rats treated with a single dose of sodium valproate. However, these levels appeared to recover after a subchronic treatment, except from specific acylcarnitines derived from branched-chain-2-ketoacids and straight medium-chain fatty acids that remain elevated. In addition, γ-butyrobetaine dioxygenase activity was induced and γ-butyrobetaine levels were increased in the liver. The inter-organ redistribution of carnitine and stimulation of endogenous carnitine biosynthesis seem to be crucial for the immediate rescue of hepatic free carnitine levels. Moreover, the disturbances in rat liver acylcarnitines were suggestive of an altered CoA metabolism. The hepatocellular levels of free and total CoA were quantified in the biological samples. A clear depletion of hepatic free CoA was observed at the onset of VPA treatment in rats. The levels of CoA normalized after prolonged treatment with VPA probably due to active adaptive cellular mechanisms to valproate-induced stress on mitochondrial metabolism. However, the decreased availability of this cofactor may contribute to the adverse effects associated with VPA, often observed at the start of VPA therapy or in cases of overdose. The structural similarity between the intermediates derived from the β-oxidation of VPA and from the oxidative metabolism of branched-chain amino acids (BCAAs) suggested that VPA might use key enzymes from the BCAAs catabolic pathways for its own metabolism. We have demonstrated that two enzymes from the isoleucine and leucine catabolism, namely short-branched chain acyl-CoA dehydrogenase (SBCAD) and isovaleryl-CoA dehydrogenase (IVD), are involved in the first step of the β-oxidation of VPA, the dehydrogenation of valproyl-CoA into Δ2-valproyl-CoA. We have further demonstrated that short-chain enoyl-CoA hydratase (ECHS1) and 2-methyl-3-hydroxybutyryl-CoA dehydrogenase (MHBD) are the unique enzymes hydrating Δ2-valproyl-CoA and oxidizing 3-hydroxyvalproyl-CoA, respectively. Moreover, the activity of IVD and SBCAD were inhibited in the presence of the tested VPA Summary xiv metabolites, valproyl-CoA and valproyl-dephosphoCoA. The drug metabolite 3-hydroxyvalproyl-CoA also inhibited the activity of MHBD. This result is in accordance with the increased urinary excretion of the isoleucine metabolite 2-methyl-3-hydroxybutyric acid in patients under valproate therapy. It has been reported that patients treated with valproate reveal an increased excretion of 3-hydroxyisovaleric acid in urine, a secondary metabolite of the leucine oxidative pathway. This finding has not been explained and its biochemical basis remains elusive. Our results suggest that valproyl-CoA inhibits the activity of 3-methylcrotonyl-CoA carboxylase (3MCC) through a direct effect on the enzyme, or through the inhibition of biotinidase. This is in agreement with the significant increased levels of 3-hydroxyisovaleric acid observed in urine of patients treated with valproate as well as with the skin rash and hair loss, which are side effects often reported in VPA-treated patients. To elucidate the underlying mechanisms of VPA-induced interferences on oxidative phosphorylation (OXPHOS), the effects of VPA and specific metabolites were studied both on the mitochondrial respiration and on the formation of ATP. Our results show that oxygen consumption and ATP synthesis were inhibited by both valproate and Δ4-VPA, when 2-oxoglutarate or glutamate, were used as respiratory substrates. Accordingly, the results demonstrated a selective and mild inhibition of dihydrolipoyl dehydrogenase (DLDH) (E3 subunit) activity by valproyl-CoA, which suggest that valproate might limit energy production in mitochondria. In addition, the measurement of the GTP- and ATP-specific succinate:CoA ligases (G-SUCL and A-SUCL) activity revealed that valproyl-CoA was found to inhibit the activity of SUCL, in particular the ATP-specific enzyme. Since SUCL is a crucial enzyme in the Krebs cycle, interference with its activity might derange the cellular energy status. Furthermore, valproate might induce an imbalance of nucleotides in mitochondria explaining the potential valproate-induced liver failure in patients with deficiencies within the mitochondrial DNA replicase such as polymerase γ 1. The elucidation of the interference of valproate with mitochondrial metabolism promoted by the present studies, may explain the potencial idiosyncratic reaction reported in patients treated with the drug. The prescription of valproate should be carefully considered in patients with inborn errors of metabolism affecting not only fatty acid oxidation and urea cycle but also the oxidative pathways of branched-chain amino acids, the krebs cycle and the DNA replicase system.

Proper ion transport is crucial for maintenance, hydration and also protection against infection of epithelial tissues. The dysfunction of the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Cl- channel results in Cystic Fibrosis (CF), the most common genetic disease among Caucasians. CFTR and Ca2+-activated Cl- channels (CaCCs) are main Cl- channels present in the respiratory epithelia and are stimulated by cAMP and Ca2+, respectively. Data demonstrating upregulated Ca2+ activated Cl- currents (ICaCC) when CFTR is absent or defective have been well described for long and are of remarkable interest for further studies and potential therapeutic approaches to CF. In the first part of this doctoral work, we used Anoctamin 1 (Ano 1) null mice to look into the role of CaCCs in the airways, so as to elucidate the potential contribution of this alternative Cl- channel to tissue homeostasis. Ussing chamber measurements were made on freshly isolated tracheas from newborn Ano -/- and Ano +/+ mice. Application of the muscarinic M3 receptor agonist carbachol (CCH) or apical stimulation of purinergic receptors through UTP and ATP was performed to elicit Ca2+-activated Cl- secretion. In Ano 1 +/+ mice, CCH induced Cl- secretion was sensitive to the CaCCs inhibitor niflumic acid and also to Ca2+ depletion from the endoplasmic reticulum (ER). On the other hand, in Ano 1 -/- animals Cl- secretion was completely abolished when using CCH as an agonist. Additionally, both UTP and ATP evoked a transient Cl- secretion in Ano 1 +/+ tracheas, but had only small effects on Ano 1 -/- tracheas. We also indirectly evaluated the efficiency of mucociliary transport in the airways upon cholinergic stimulation by measuring the movement of polystyrene microspheres over time on fleshly isolated tracheas. Particle transport was activated in Ano 1 +/+ tracheas by stimulation with CCH, while on Ano 1 -/- tissues a lack of cholinergic mucociliary clearance was observed. Altogether, our results demonstrate that in the absence of Ano 1 both Cl- secretion and mucociliary clearance are impaired, pointing towards a critical role of Ano 1 in the airways. Furthermore, we speculate that the novel insights in the function of Ano 1 provide the basis for novel treatments in CF (the so-called "bypass therapies"), being Ano 1 an alternative conductor of Cl- when CFTR is absent or defective. In the second part of this work we aimed to investigate the link between CFTR and CaCCs. We first compared the mRNA levels of membrane localized Ano 1, as well as those of Bestrophin 1, in freshly isolated nasal cells from F508del-homozygous CF patients and non-CF healthy controls. We also compared the mRNA levels of these channels in cell lines stably expressing wild type (wt) or F508del-CFTR. In both cases no increase in transcript levels of neither Ano 1 nor Best 1 was found in the F508del-CFTR cells. It is thus unlikely that changes in ion channel expression account for enhanced CaCC activity in native CF airway epithelial cells. Exposure to lipopolysaccharides (LPS) only slightly increased Ano 1 and Best 1 mRNA levels, both in wt- and F508del-CFTR cells. Taken together, the enhanced Ca2+-activated Cl- conductance observed in airway epithelial cells expressing F508del-CFTR is not explained neither by an increase in transcripts of known CaCCs nor exposure to proinflammatory agents such as LPS. We next examined whether F508del-CFTR has an effect on receptor-mediated Ca2+ signalling. Our results showed that the presence of F508del-CFTR augments receptor mediated Ca2+ signalling, comparatively to the effects observed in the presence of wt-CFTR. To elucidate the possible mechanism by which ER-localized F508del-CFTR augments intracellular Ca2+ signals, we hypothesized that F508del-CFTR trapped in the ER could affect receptor-mediated Ca2+ release from ER Ca2+ stores. Our results showed that not only UTP-induced increase in intracellular calcium concentration ([Ca2+]i) was largely Cl- dependent but also that the presence of F508del-CFTR in the ER may affect intracellular Ca2+ signalling by acting as a Cl- counter-ion channel, thereby facilitating Ca2+ movement across the ER membrane. Altogether, these results give new insights into the fine tuning of ion transport in the airways. Moreover, they contribute to a better understanding of the pathophysiological basis of CF disease, giving new perspectives to bypass therapies for Cystic Fibrosis.

As Redes de transportes sempre estruturaram o território. Desde as estradas e aquedutos romanos, passando pelos canais de navegação, os caminhos-de-ferro, a electricidade, as estradas de macadame, até à internet de banda larga, que as redes de transporte configuram o território dilatando-o ou comprimindo-o, determinando a forma como as actividades se distribuem, o território se organiza, as fronteiras se definem e criando uma nova tipologia de espaços: espaços de fluxos e de conexões. Mas não são apenas novas tipologias de espaços que surgem à medida que as redes de transportes se reconfiguram ao longo do tempo. Também a topologia das redes sofrem modificações, mais propriamente a estrutura de nós e arcos sofre transformações. Para tentar medir e avaliar as características das redes e da influência que exercem no território, e em particular nas relações entre os lugares, esta dissertação propõe um novo modelo de análise de redes, de base matemática e geográfica, com métricas provenientes da Teoria dos Grafos (medidas de conectividade e medidas de acessibilidade) e das Redes Complexas (Mundo pequeno, coeficiente de clustering e índice de Freeman ou grau de intermediação). Face à crescente complexidade dos sistemas e aos padrões territoriais emergentes, as teorias, os modelos e as técnicas clássicas e analógicas deixaram de conseguir dar uma resposta adequada, quer pela excessiva simplicidade da realidade que consagram (Wilson, 2000), quer pela ausência da dinâmica e da anisotropia do espaço nos processos transformativos dos sistemas. Para respostas mais eficazes são precisos modelos que incluam métodos mais evoluídos e para isso é fundamental o recurso à geocomputação (Openshaw, 1995/97). Os recentes avanços na capacidade de processamento dos computadores e nas técnicas de modelação computacional forneceram, os meios para modelar e compreender os problemas mais difíceis de complexidade organizada (Longley, 1999). O modelo criado nesta dissertação é um produto da geocomputação. Trata-se de um modelo Sistema de Informação Geográfica para Transportes (SIG-T), que se designou de geo-grafo e que usa como hospedeiro o interface gráfico do mais popular e comercial programa de Sistemas de Informação Geográfica (SIG) – ArcGIS -, mas que pode ‘correr’ em qualquer outro programa SIG que aceite a linguagem de programação (LP) Visual Basic for Applications (VBA)

Drosophila teashirt (tsh) encodes a zinc finger protein essential during Drosophila development. My PhD aimed to clarify the tsh role in cell proliferation studying tsh function in three Drosophila systems: follicle stem cells (FSC), neuroblasts and S2 cells. I found that Tsh and Hh were co-expressed in the Drosophila terminal filament and cap cells. A new tsh female sterile mutation exhibiting early oogenesis arrest allowed me to show that Tsh acts via Hh-mediated signalling pathway to control FSC proliferation; tsh overexpression mimics the hh overexpression associated phenotype. Additionally, in tsh background, the hsGal4 driven expression of tsh or hh partially rescues the abnormal proliferation phenotype allowing oogenesis progression. These results suggest that Tsh is critical to control hh expression contributing for the regulation of FSC proliferation and specification of somatic cell identity. In neuroblasts, hypomorphic tsh mutation leads to a metaphase-like arrest with highly condensed chromosomes associated with apparently normal mitotic spindles and centrosomes. Additionally, checkpoint proteins Bub1 and ROD were localised normally and CID imunodetection excluded an eventual premature sister chromatid separation. Furthermore, cyclin-A appears normally degraded, whereas cyclin-B remains detectable. Double tsh rod and tsh bub1 mutants phenotypes resemble the tsh single mutant. Finally, tsh depletion in S2 cells mimics the tsh mutant phenotype in neuroblasts. These results suggest that Tsh is critical during mitosis progression and I propose that Tsh loss maintains active the spindle checkpoint and/or unable the APC/C activity. We proposed a tsh acts through Hh signalling in cell proliferation. During oogenesis, tsh affects FSC proliferation mediating Hh expression. In neuroblasts, tsh could acts through Hh signalling, since expression of interference hedgehog, a positive mediator of Hh signal, was severely downregulated in microarrays-based analysis.

Human adult mesenchymal stem cells (hMSC) represent a potential source for cellbased therapies in Diabetes Mellitus. Yet, till now, no definitive data exist for clinical application. Aims 1. Evaluation of the pancreatic endocrine differentiation potential of humanmesenchymal stem-cells, immortalized with telomerase (hMSC-TERT); 2. Comparative analysis of the plasticity and molecular signature of human-islet-derived pancreatic cells (hIPC) and bone-marrow-derived hMSC (hBM-MSC). Research strategies: 1. In vitro differentiation of hMSC-TERT into insulin producing cells through: a) Endocrine-promoting culture conditions; b) Ectopic expression of two key regulatory genes of human endocrine pancreas, neurogenin3 (hNGN3) and pancreatic-duodenal homeobox 1 (/hPDX-1) genes in hMSC-TERT. 2. Comparison of the functional signature of hIPC and hBM-MSC by: a) Immunophenotype characterization; b) In vitro mesenchymal and endocrine differentiation of primary hBM-MSC and hIPC; c) Transcriptome analysis for detection of mesenchymal and adult stem-cell related gene markers. Identification of target genes, related to the origin and function of hBM-MSC and hIPC. Results We could demonstrate that hMSC-TERT can be differentiated in vitro into pancreatic endocrine phenotypes with the ability to synthesise and store insulin. Similar to pancreatic development, in hMSC-TERT, NGN3 contributes to the activation of PDX-1 promoter. Insulin secretion in a glucose-sensing manner was not detected in our cell system, which indicates that a mature state of beta-cell was not attained in these phenotypes; hIPC and hBM-MSC display very similar MSC phenotypes. However, our data suggest distinct functions of these populations, which are related to a “mesodermal tissue specification” and “positional memory” of these cells. In hIPC, we confirmed a state of epithelial-mesenchymal transition. Conclusions MSC populations are adult stem cells with distinct levels of plasticity and commitment. Future studies will help to identify those MSC populations with greater potential to replace or regenerate beta cells in type 1 Diabetes.