RCAAP Repository

Objectivo: O nosso objectivo consistiu em avaliar a bilirrubina não conjugada (UCB) como candidato a biomarcador no espectro e na gravidade da esquizofrenia e perturbação esquizoafectiva. Para esse fim preparámos dois estudos: um estudo transversal restrospetivo de cinco anos, e um estudo longitudinal prospectivo com um ano de duração. Métodos: No nosso estudo transversal retrospetivo de 5 anos procurámos correlação entre os níveis médios de UCB, duração média de internamento e o diagnóstico em 255 indivíduos, incluindo 56 controlos saudáveis e 199 doentes (nomeadamente 44 com esquizofrenia, 99 com perturbação esquizoafectiva e 56 com perturbação bipolar). No nosso estudo longitudinal prospetivo 88 doentes psicóticos completaram a primeira avaliação durante internamento psiquiátrico em contexto de episódio psicótico agudo, metade com esquizofrenia, metade com perturbação esquizoafectiva. Nesta fase 44 doentes com perturbação bipolar, foram usados como controlos. Aos doze meses de seguimento completámos o protocolo longitudinal (dados laboratoriais, psicopatológicos e psicossociais) de 60 doentes, metade com esquizofrenia, metade com perturbação esquizoafectiva. Resultados: No nosso estudo transversal retrospetivo de 5 anos encontrámos diferença estatisticamente significativa, após teste de multicomparações de Bonferroni, entre os níveis médios de UCB dos doentes com esquizofrenia (0.41 mg/dL) e os doentes com perturbação esquizoafectiva (0.34 mg/dL) (p<0.03), os doentes com perturbação bipolar (0.28mg/dL) (p<0.001) e também os controlos saudáveis (0.28) (p<0.001). Encontrámos ainda diferença estatisticamente significativa entre os níveis médios de UCB de doentes com perturbação esquizoafectiva (0.34 mg/dL) e perturbação bipolar (0.28 mg/dL) (p<0.04). Relativamente à duração média de internamentos durante o período dos 5 anos, ao diagnóstico e ao nível de UCB, quando aplicámos um modelo de regressão linear ANOVA, com a duração média dos internamentos como variável dependente, encontrámos uma diferença estatisticamente significativa (p<0.001) entre os doentes com esquizofrenia (29 dias) e os doentes com perturbação bipolar (16 dias) mas não com os doentes com perturbação esquizoafectiva. No nosso estudo longitudinal prospetivo de 1 ano, encontrámos, aquando da primeira avaliação durante o episódio psicótico agudo (N=88): uma diferença estatisticamente significativa (ANOVA; p=0.002), confirmada após teste de multicomparações de Bonferroni, entre doentes com esquizofrenia (N=44), quer com os doentes com perturbação esquizoafectiva (N=44) (p=0.05), quer com os controlos com perturbação bipolar (N=44) (p=0.05). Encontrámos também uma correlação positiva (Pearson’s r=0.314) entre os valores médios de UCB e o item d) “disturbing and aggressive behavior” da escala de Personal and Social Performance; e ainda uma correlação positiva (R2=0.223), com significância estatística (p=0.008), entre os níveis médios de UCB e a duração média de internamento psiquiátrico dos doentes com perturbação esquizoafectiva que completaram o protocolo (N=30). Durante a remissão parcial, aquando da segunda avaliação (N=60), encontrámos: uma diferença estatisticamente significativa (ANOVA; p=0.006), confirmada após teste de multicomparações de Bonferroni (p=0.05) entre os doentes com esquizofrenia (N=30) e perturbação esquizoafectiva (N=30); bem como uma correlação negativa (Pearson’s r=−0.399) entre os níveis médios de UCB e o item G7 “psychomotor retardation” da Positive And Negative Syndrome Scale. Ao compararmos a primeira com a segunda avaliação (teste T Student para amostras emparelhadas) encontrámos uma diferença estatisticamente significativa (p=0.034) nos valores médios apenas entre doentes com perturbação esquizoafectiva (N=30). Conclusão: existe um interessante potencial na investigação da UCB como biomarcador das psicoses e respectiva gravidade, quer no espectro da esquizofrenia, quer no da perturbação esquizoafectiva, seja na recaída ou na remissão

Síncope, a forma mais comum de perda temporária de consciência é responsável por até 5% das idas aos serviços de emergência e até 3% dos internamentos hospitalares. É um problema médico frequente, com múltiplos gatilhos, incapacitante, potencialmente perigoso e desafiante em termos diagnósticos e terapêuticos. Assim, é necessária uma anamnese detalhada para primeiro estabelecer a natureza da perda de consciência, mas, após o diagnóstico, as medidas terapêuticas existentes são pouco eficazes. Embora a fisiopatologia da síncope vasovagal ainda não tenha sido completamente esclarecida, alguns mecanismos subjacentes foram já desvendados. Em última análise, a síncope depende de uma falha transitória na perfusão cerebral pelo que qualquer factor que afecte a circulação sanguínea cerebral pode determinar a ocorrência de síncope. Assim, o objectivo do presente estudo é caracterizar o impacto hemodinâmico e autonómico nos mecanismos subjacentes à síncope reflexa, para melhorar o diagnóstico, o prognóstico e a qualidade de vida dos doentes e dos seus cuidadores. Para isso, desenhámos e implementámos novas ferramentas matemáticas e computacionais que permitem uma avaliação autonómica e hemodinâmica integrada, de forma a aprofundar a compreensão do seu envolvimento nos mecanismos de síncope reflexa. Além disso, refinando a precisão do diagnóstico, a sensibilidade e a especificidade do teste de mesa de inclinação (“tilt test”), estabelecemos uma ferramenta preditiva do episódio iminente de síncope. Isso permitiu-nos estabelecer alternativas de tratamento eficazes e personalizadas para os doentes refractários às opções convencionais, sob a forma de um programa de treino de ortostatismo (“tilt training”), contribuindo para o aumento da sua qualidade de vida e para a redução dos custos directos e indirectos da sua assistência médica. Assim, num estudo verdadeiramente multidisciplinar envolvendo doentes com síncope reflexa refractária à terapêutica, conseguimos demonstrar uma assincronia funcional das respostas reflexas autonómicas e hemodinâmicas, expressas por um desajuste temporal entre o débito cardíaco e as adaptações de resistência total periférica, uma resposta baroreflexa atrasada e um desequilíbrio incremental do tónus autonómico que, em conjunto, poderão resultar de uma disfunção do sistema nervoso autónomo que se traduz por uma reserva simpática diminuída. Igualmente, desenhámos, testámos e implementámos uma plataforma computacional e respectivo software associado - a plataforma FisioSinal –incluindo novas formas, mais dinâmicas, de avaliação integrada autonómica e hemodinâmica, que levaram ao desenvolvimento de algoritmos preditivos para a estratificação de doentes com síncope. Além disso, na aplicação dessas ferramentas, comprovámos a eficácia de um tratamento não invasivo, não disruptivo e integrado, focado na neuromodulação das variáveis autonómicas e cardiovasculares envolvidas nos mecanismos de síncope. Esta terapêutica complementar levou a um aumento substancial da qualidade de vida dos doentes e à abolição dos eventos sincopais na grande maioria dos doentes envolvidos. Em conclusão, o nosso trabalho contribuiu para preencher a lacuna entre a melhor informação científica disponível e sua aplicação na prática clínica, sustentando-se nos três pilares da medicina translacional: investigação básica, clínica e comunidade.

Amyotrophic Lateral Sclerosis (ALS) belongs to a group of neurological disorders known as motor neuron diseases, which are caused by gradual degeneration and consequent death of motor neurons. In general, the disease begins in one of two fundamental ways: with involvement of the muscles of the bulbar region; or with loss of muscle strength of the upper or lower limbs - spinal form. Although other rare forms of manifestation are also cited in the literature, such as: respiratory (when respiratory muscles are initially affected); axial (initially affecting the cervical and paraspinal muscles) and the diffuse form (generalized onset of the disease). These forms of presentation determine the initial symptoms. Patients may initially develop muscle weakness in the limbs resulting in various clinical conditions with paresis, speech problems with dysarthria, dysphagia, and respiratory symptoms with dyspnea, and evolve to complete loss of body movements control. The degree of functional disability and dependence resulting from ALS lead the patient to gradually needing a caregiver for all their activities of daily life. The mean survival for ALS is around 3 - 5 years from first symptoms, and the fatal event usually occurs due to respiratory failure or infection. Although there is no effective treatment to halt disease progression, the clinical management has evolved positively in last years. The technological advance of medical interventions has contributed to a longer survival and higher quality of life. The monitoring of non-invasive ventilation has been very helpful for the clinical follow up and to decrease the anxiety experienced by caregivers. Unfortunately, the aerobic exercise is not a usual therapeutic option for ALS patients in the clinical management yet. Physical exercise has been suggested to promote growth factor delivery in experimental animal models of ALS. However, the aerobic exercise is understudied in ALS patients due the suspicious that the exercise could be harmful for this population. Meeting the recommendations from the last Cochrane review about Therapeutic exercise in ALS, we have analyzed the impact of aerobic exercise in the ALS progression. This thesis has 2 main contributions: 1 - To study the efficacy of moderated and accurate defined exercise program on the evolution and survival of ALS, and, 2 - to assess the feasibility to performing exercise monitored remotely from home in ALS patients. In addition, this thesis includes results from additional contributions, which are related to relevant issues always present during disease progression such as management of the respiratory failure (sub-chapters 5.2 and 5.3), the support for ALS caregivers (Chapter 6), and the potential impact of clinical management on disease progression in an environment with lack of resources (Chapter 7). In the sub-chapter 5.2 we present a work which describe the lack of consensus for the ideal timing to start NIV and about the use of alternative respiratory support. We address the impact of NIV and tracheostomy on family and / or informal caregivers, especially how it can affect quality of life. The importance of assessing the emotional, physical, social and psychological capacity of the caregiver is reinforced in order to cope with the increasing care needs of these patients. The sub-chapter 5.3 presents results on the importance of a careful management in the use and configuration of Non-Invasive Ventilation (NIV) parameters, in particular the role of ventilation and adherence adjustments in functional decline and survival of ventilated patients. A wide range of data recorded from the software used in ventilators, nocturnal pulse oximetry measurements and respiratory function tests were analyzed. Our results suggest that the variables that affect the respiratory comfort of the patient are relevant for adherence to NIV and positively affect survival in ALS. The Chapter 6 presents results of a feasibility study about a training program for caregivers. The uncertain progression of the disease and long-term care, as well as the insufficient number of skilled health professionals, determine that the admission of these patients to hospitals or to continuing care units is a complex option. Initiatives that allow better management of the disease at home can be an alternative solution. The functional disability of patients can promote significant financial constraints, and exposes their families to high levels of stress, which can compromise the provision of adequate health care, leading to the hospitalization of these patients. In this project, we identified an excellent level of participation as well as a good result in the evaluation of learning (above 70%). The main limiting factor for participation in the training program was the absence of a secondary caregiver. The Chapter 7 describes results from2 studies conducted with ALS patients in the African continent, which present data on disease progression in an environment with limited resources for clinical management. The Chapter 8 present a general discussion and conclusion of all works included in this Thesis. All the papers presented in this thesis aim to contribute to a broader view of the clinical management of ALS, where the role of exercise associated with more careful respiratory support, and the presence of a well-informed and trained caregiver, can together be an important contribution for the survival and quality of life of the patient with ALS. We hope our work presented in this thesis may contribute to a wider understanding on the clinical management of ALS. In particular demonstrating that controlled exercise associated with careful respiratory support, and the presence of a well-informed and trained caregiver, may be an added value in the survival and quality of life for ALS patient.

Parkinson’s disease (PD) is a progressive age-dependent neurodegenerative disease. Life expectancy increasing and a better knowledge in PD treatment management, including the advent of device-aided therapies, are likely to increase the number of patients who can reach an advanced disease stage and eventually enter the late stage (LS) of the disease in the next decades. LSPD is a recently recognized disease stage, in which patients are severely disable and dependent on activities of daily life (ADLs) due to the presence of poor treatment responsive motor and non-motor symptoms (NMS) thus highly impacting caregiver’s burden and social/health care system. Hence an operational clinical criteria to identify LSPD patients has been recently proposed suggesting adopt a Schwab and England activity of daily life score (S&E) < 50 in the MED ON condition. LSPD patients’ treatment management is challenging. Treatment-related adverse effects (AEs) are frequent and few evidence in terms of phamacological and non-pharmacological treatment efficacy are available as they are barely included in clinical or research studies and even the participation into routine hospital-based visits can be an unsurmountable limit. At the same time, even if general PD disease severity milestones have been described, we do not know how LSPD patients specifically progress, if they do evolve and if there are clinical markers or biomarkers of poor outcome that could be useful to focus specific therapeutic interventions for this specific disease stage. We aimed to deeply characterize the clinical phenotype, needs along with clinical markers or biomarkers of poor outcome of LSPD patients. As levodopa (L-dopa) is the mainstay of PD treatment and a simplification of treatment regimen in later disease stages has been suggested, we also aimed to investigate the real effect of L-dopa on motor symptoms and NMS among LSPD patients, if compared to advanced stage patients. Among NMS, we focused our work particularly on speech impairment, exploring speech response to L-dopa among LSPD patients and to fine stimulation parameters adjustment, in combination with L-dopa, in advanced PD patients submitted to deep brain stimulation (DBS). Participants were LSPD (Schwab and England ADL Scale [S&E] <50 or Hoehn Yahr Stage [HY] >3 in “MED ON” state) and advanced stage PD patients previously submitted to DBS. Cross-sectional data were obtained by means of a comprehensive clinical assessment including a L-dopa challenge test with a suprathreshold dose. A subgroup of thirteen LSPD patients underwent a neuroimaging study in order to study neuromelanin (NM) substantia nigra (SN) area changes in the latest disease stage if compared to previous ones. Automated analysis of speech were used to study the effect of a supramaximal L-dopa dose in twenty-four LSPD patients as well as L-dopa and frequency stimulation adjustment in twenty deep brain stimulated patients. Longitudinal data were collected only for LSPD patients. Descriptive, regression and survival curves analysis were performed. Fifty LSPD patients (female 46%) were included. Mean age was 77.5 ± 5.9 years and mean disease duration was 15.5± 6.5 years. At baseline, 76% had L-dopa-induced motor complications (MCs), mainly non-troublesome, 68%were demented, 54% had psychosis and 68% depression. Caregiver distress was high. L-dopa responsiveness was mild (18% ± 12 of improvement on MDS-UPDRS-III) and present only for appendicular signs, being tremor and rigidity the most responsive ones, while axial signs did not change. The clinical significance of this better motor response was marginal according to the Clinical Global Improvement Scale and the change in the S&E between OFF and ON state. The magnitude of L-dopa response correlated with the acute appearance of dyskinesias and the severity of MCs. After one-year, 20% of the patients were dead, 18% institutionalized in nursing home and 6% passed to a HY 5. MDS-UPDRS-motor mean score worsened 7.2 ± 10.3 points, corresponding to a 15.7% (±23.0) increase, with no difference between tremor-dominant versus akinetic-rigid phenotype or PD patients with/without dementia (PDD/non-PDD) at baseline. However, there was heterogeneity between patients in terms of disease progression, as 12 patients (37.5%) had a motor deterioration ≤ 3 points and 14 (43%) ≤ 5 points with concomitant worsening of the MDS-UPDRS-II (Motor Aspects of Experiences of Daily Living), of 2.1±4.1. Conversely, eleven cases (32%) did not deteriorate and, in fact, 10 of these improved between 1-6 points at the MDS-UPDRS-III. Overall NMS worsened, mostly in cognition/mood, urinary and gastrointestinal domains. Conversely, MCs improved despite similar L-dopa equivalent dose. Functional independence and quality of life worsened. Dysphagia severity at baseline predicted a poor combined outcome (death, being institutionalized or developing HY 5) (Hazard ratio 2.3, 95% CI 1.12- 4.4; p = 0.01) or death alone (Hazard ratio of 2.9, 95% CI 1.12- 8.6, p=0.04), whereas magnitude of L-dopa response of LSPD patients did not. SN area evaluated by NM-sensitive magnetic resonance imaging (MRI), resulted able to differentiate LSPD patients from both de novo PD patients and controls, though not founding statistical differences between LSPD patients and patients with two-five year disease duration. Performing an indirect comparison of the effect of L-dopa on motor symptoms and NMS among twenty LSPD patients and twenty-two, not-matched, advanced PD patients, a milder response on motor symptoms (11% vs. 37% of improvement on MDS-UPDRS-III) and an absence of response on NMS, namely anxiety, fatigue and pain, were found among LSPD patients, with concomitant higher frequency of drug-related AEs. Indeed orthostatic hypotension (OH) or drowsiness occurred among 35% of LSPD patients versus 13% of advanced PD patients, who still presented a benefit from L-dopa intake on pain and anxiety, while fatigue did not change. Scales applicability and blood pressure assessment while standing resulted challenging among LSPD patients with consequent missing data on depression, anxiety, pain and OH identification and possible underestimation of those symptoms. No effect of L-dopa was found on speech and voice by means of both automated analysis and clinical evaluation in LSPD patients. Respiratory support for speech and voice stability were the most affected speech and voice features among LSPD patients. Among axial symptoms, speech seemed to be the most L-dopa unresponsive one. Speech unresponsiveness to L-dopa was confirmed also among subthalamic (STN)-DBS treated patients with both mild and severe dysarthria, at least in combination with stimulation. Conversely, PD patients with severe dysarthria under chronic STN-DBS treatment showed a benefit of lowering frequency of stimulation from 130 Hz (High frequency stimulation [HFS]) to 60Hz (low frequency stimulation [LFS]), with concomitant increment of voltage, in order to keep constant the total energy delivered. Indeed speech intelligibility and articulatory diadochokinesis presented an acute improvement passing from HFS to LFS, as assessed by automated speech analysis and such a benefit, when present and clinically meaningful, lasted during six months with no motor worsening, though requiring medication adjustment. The present study provides further evidence to better delineate a recently recognized and poorly described PD stage. An extensive cross-sectional and longitudinal observation is proposed. LSPD patients clearly differ from previous stages in terms of both clinical features, needs, therapeutic response and drugs’ tolerability profile. Over one year, a heterogeneous disease progression of motor symptoms is still present and it seems even steeper if compared to previous stages, while functional independence globally worsened. As well as mild motor improvements are still possible with treatment adjustment, it is also possible to identify a clinical phenotype of LSPD patients who are likely to have a better response to L-dopa if compared to the other ones. Clinical assessment and therapeutic interventions for swallowing problems should be a priority. PDD or living in a nursing home remain other indicators of poor outcome. In the next few years the number of LSPD patients who have been previously submitted to device-aided therapies is expected to increase, bringing new clinical scenarios, such as the fine parameters adjustment of invasive treatment for challenging motor and NMS and the difficult management or eventual interruption of those treatments among elderly and frail LSPD patients. Overall, future research and fund allocations should be specifically oriented on LSPD patients, usually not included or considered in clinical trials or research studies, and on L-dopa not-responsive aspects and caregivers’ needs

The availability of resting sites is known to influence the distribution and density of animals and they are key resources to nocturnal mammals that need to spend daylight in safe refuges. This is the reason why these structures have to be taken in account in the efforts of mammal species conservation. Aside from being simply a place to rest, these structures also provide natural protection against predators and thermal isolation in harsh environments. In the 1960’s Eurasian otters (Lutra lutra) crashed in many parts of its European distribution, this being the case of Austria. With the species recolonization, favored by an interconnection with the Czech and the Hungarian populations, a need emerged for a better understanding on how animals are using such resources in the Alpine landscape as the species is still considered endangered. The main goals of this study were to evaluate (assess) if: (1) there is a structural pattern of daytime resting site according to gender and age; (2) human disturbance affects the choice of resting site type; (3) snow cover has any influence on the type of resting site used; (4) rest-site choice can be predicted based on environmental variables and (5) there is a geographic pattern in resting site choice. Four otters (two adult females, one adult male and one juvenile female) were captured and radio-tagged to locate resting structures which were characterized according to a set of physical and environmental parameters. Daily climate data was obtained from local weather stations. In spite of sampling size constrains results revealed that underground structures were used more often than surface lairs and suggest a different use of resting site type between females and breeding females. No relation was found between rest-site type and human density. Snow cover influence could not be asserted due to environmental conditions during the sampling period, but temperature, vegetation cover and river width seem to play a significant role in predicting resting site choice. Concerning the geographical pattern, the results, framed by previous studies, imply its existence but it was not possible to identify the factors behind it.

Collective cell migration is a widespread biologic process that is crucial not only during homeostasis, but also in the context of disease. Mechanistically, it is based on the establishment of a front-rear polarity axis, and on the maintenance and dynamic regulation of cell-cell adhesions, which are crucial for traction force generation and oriented cell movements. Sprouting angiogenesis, a biologic process that enables the expansion of the vascular network, relies on the collective migration of specialized strands of endothelial cells composed by tip and stalk cells. Wnt5a, a non-canonical Wnt ligand, has been described as a pro-angiogenic factor that promotes cell proliferation, migration and network formation. More recently, it was shown to be crucial during sprouting angiogenesis. Indeed, genetic deletion of Wnt5a in endothelial cells reduces the expansion of the vascular plexus in the mouse retina. However, it is still unclear how Wnt5a exerts these functions in the endothelium and which specific molecular components could be involved. Therefore, to investigate these questions, we used the in vitro wound-healing assay to study endothelial collective cell behavior. With this system we uncovered that loss of Wnt5a leads to randomized follower cell polarity and migration, dramatically impairing coordination of collective cell behavior. Moreover, we also showed that cells lacking Wnt5a had a significant decrease in cell-cell force transmission and displayed a specific downregulation of vinculin co-localization to VE-cadherin at cell junctions. Then, we showed that randomized cell polarity in Wnt5a-depleted cells could be rescued by forcing the association of vinculin to cell junctions by overexpressing the αCat-Vinc fusion protein, or a constitutive active form of vinculin (Vinc-T12). Mechanistically, we identified ROR2/Cdc42 as the most relevant transducers in the signaling cascade triggered by Wnt5a. Altogether, we propose a model where Wnt5a, via ROR2/Cdc42, promotes vinculin and Arp2/3 association to adherens junctions, enhancing actin polymerization and junction stabilization. These strengthened adherens junctions would then act as a platform to enable stable force transmission between adjacent cells and coordination of endothelial cell behaviors during collective migration. Given its implications in evolutionarily conserved morphogenetic processes that require cell-cell mechanocoupling, we believe the mechanotransduction function of Wnt5a we described here may be applicable to other tissues and contexts. In the context of vascular biology, our findings may be relevant to understand the mechanical and molecular mechanisms involved in the collective migration of vessel sprouts driven by tip cells in vivo. Moreover, as Wnt5a has been described as a potential key regulator of cancer, our results might be relevant to understand disease onset and progression and even contribute to the development of potential therapies.

Os objetivos gerais deste estudo foram: i) caracterizar, numa amostra de trabalhadores, fatores organizacionais (no trabalho e na conciliação trabalho-família), psicossociais e biológicos, com impacte no desempenho profissional desses trabalhadores, medido pelo presentismo; ii) identificar presentismo, absentismo e sofrimento psicológico; iii) determinar a relação entre fatores biopsicossociais, saúde mental e bem-estar, resiliência e presentismo. Para o efeito, entre novembro de 2012 e julho de 2013, realizou-se um inquérito observacional transversal, com uma amostra não aleatória (de conveniência), de 405 trabalhadores de uma associação bancária mutualista. O inquérito foi constituído por um questionário eletrónico de autopreenchimento abrangendo informação biográfica, perceções sobre o emprego, atitudes face à organização, saúde, e aplicando diversas escalas, como o ASSET (deteção de stress em contexto organizacional), MHI – 5 (Inventário de Saúde Mental), escala de Satisfação no Trabalho, escala de Felicidade Subjetiva, CAGE (deteção de problemas relacionados com consumo de álcool), escala de Suporte Social – OSLO 3, escala de Presentismo, e escala de Resiliência Connor Davidson - CD-RISC. Efetuaram-se ainda medições antropométricas (peso, altura e perímetro da cintura), da tensão arterial e colheita de sangue para parâmetros biológicos (p. ex., genéticos). Os 405 participantes no estudo responderam à componente psicossocial do inquérito. Destes, 395 completaram a escala da resiliência e 260 participaram na componente biológica do estudo. A introdução dos dados foi automática pelo autopreenchimento do questionário eletrónico com exportação para SPSS V.20. O estudo global foi aprovado por duas Comissões de Ética e autorizado pela Comissão Nacional de Proteção de Dados (CNPD). Sobre os dados recolhidos realizaram-se análises estatísticas descritivas, inferenciais, fatoriais exploratórias e confirmatórias, e com modelo de equações estruturais. Das oito hipóteses de investigação (HI) que se colocaram, quatro foram suportadas pelos resultados obtidos (HI.1 há preditores, psicossociais e biológicos, de maior e menor resiliência nos trabalhadores; HI.5 há diferenças na resiliência consoante o sexo e o grupo funcional; HI.6 há associação entre presentismo e absentismo; HI.7 há associação entre absentismo e sofrimento psicológico). Três HI foram parcialmente suportadas pelos resultados (HI.2 os stressores organizacionais como ‘Recursos e Comunicação’, ‘Equilíbrio trabalho-família’, ‘Aspetos do trabalho’, ‘Segurança no emprego’, ‘Controlo’, ‘Relações no trabalho’ e ‘Sobrecarga laboral’ têm impacte no presentismo absoluto, através do efeito do bem-estar psicológico dos trabalhadores; HI.3 o efeito das variáveis ‘compromisso do colaborador face à organização’ e do ‘compromisso da organização face ao colaborador’, no presentismo absoluto, é mediado pela saúde mental; HI.8 há mais presentismo do que absentismo e as mulheres têm mais absentismo e mais sofrimento psicológico). Uma HI não foi suportada (HI.4 menos de metade da população da amostra tem uma autoperceção de saúde global ‘boa ou muito boa’). Com o trabalho desenvolvido, atingiram-se objetivos que permitiram: i) identificar preditores de presentismo e de resiliência que podem contribuir para novos modelos de stress laboral; ii) identificar determinantes psicossociais e biológicos, nomeadamente genéticos, na área da resiliência e do stress; iii) utilizar escalas que foram validadas e/ou traduzidas. Considera-se que a identificação e análise de determinantes biopsicossociais, com impacte no presentismo e absentismo, são bases importantes para o planeamento e implementação de ações em promoção da saúde mental no contexto laboral, visando maior produtividade da organização, mais equilíbrio trabalho-família e bem-estar do trabalhador.

Introduction: Cardiovascular disease (CVD) and mood disorders (MD) are two of the world’s leading health problems. Both conditions are chronic, debilitating, and extremely prevalent among the general population, often occurring together in the same individual. Epidemiological evidence has been growing suggesting that there is a two-way relationship between them. In fact, a significant proportion of people with no prior record of MD develop depression after a cardiovascular event. As well, physically healthy individuals who are diagnosed with depression are at increased risk of developing CVD compared to the general population. This link is particularly evident between depression and heart failure (HF). The HF population with depression is particularly known to have an increased risk of hospitalizations, emergency visits and re-admissions with higher mortality rates and worse health-related outcomes. The burden of these diseases is overwhelming when they occur individually and even greater when they occur in comorbidity, yet there is still no consensus or any particular guidelines for the best method to treat patients with comorbid MD and CVD. This may be partially due to the fact that MD remains poorly understood. To better clarify the pathophysiology behind this spectrum of disorders we should not be confined to certain conceptual boundaries. First, although there are clear clinical distinctions between depression and Bipolar Disorder (BD), experts still debate the possibility of some degree of continuum between these diseases. Second, the biological mechanisms behind MD are not yet fully understood; namely what separates them and what mechanisms they have in common. This is also true if we aim to identify common biological changes between CVD and MD, particularly between HF and depression. HF is one of the most common causes of hospitalization, mortality, and economic burden worldwide. It is crucial to develop new and improved models of care for this condition. A new program design at Health Sciences North (HSN) called the Heart Failure Disease Management Program (HFDMP) utilizes various outpatient strategies that aim to avoid emergency department (ED) visits, to decrease HF hospitalizations, to improve outcomes, and to decrease mortality and health care costs. The success of this outpatient program model needs to be evaluated. Furthermore, since the association of depression with HF seems to be extremely relevant the prevalence and impact of depression in an original program like the HFDMP needs to be clarified. The general aims of this work were: 1. To investigate pathophysiological mechanisms in mood disorders; 2. To investigate common pathophysiological mechanisms between mood disorders and cardiovascular disease, particularly depression and heart failure; 3. To further investigate the relationship between depression and heart failure from a clinical perspective. The following specific aims were established to address the general goals of this work: 1 To investigate pathophysiological changes in mood disorders, namely the role of mechanisms known to be relevant to the pathophysiology of cardiovascular disease. 1.1 To investigate oxidative stress changes across the life span of patients with MD, Specifically, to analyse the levels of oxidative damage in plasma from 185 subjects, consisting of 110 euthymic older BD patients (BD-I and BD-II with 35 years of evolution of disease) compared to 75 healthy controls; 1.2 To investigate oxidative changes in different mood episodes and mood polarities (depression, hypomania and euthymia) and their relationship with hippocampus changes in patients with MD; specifically, to analyse peripheral levels of oxidative stress markers and decreased hippocampal subvolume dentate gyrus–cornu ammonis (CA4) in correlation to depressive, hypomanic and euthymic episodes in 62 subjects consisting of 29 patients with BD-II and 33 healthy controls. 2 To investigate common pathophysiological mechanisms between mood disorders and cardiovascular disease, particularly between depression and heart failure: 2.1 To review the role of inflammation as a pathophysiological mechanism shared between depression and CVD; specifically, to discuss the evidence of the benefit ofanti-inflammatory drugs in both conditions in support of the hypothesis of a shared pathophysiology between these diseases through inflammation. 2.2 To investigate further common biological pathways between MD and CVD, in HF patients with depression; specifically, to investigate whether NT-proBNP mediated reduced ejection fraction (EF) and depressive symptoms in 124 subjects with HF and depression compared to HF patients without depression. 3 To evaluate an original outpatient program for heart failure management and the relationship of depression with patients as well as the program’s outcomes: 3.1 To investigate the safety and effectiveness of the program HFDMP in managing HF in an outpatient setting; specifically, to investigate the ability of the program to reduce emergency department visits, to decrease HF admissions and readmissions, to improve HF outcome, and to decrease mortality in 138 patients enrolled and followed in the program over a period of 12 months; 3.2 To investigate the relationship between depression, quality of life (QoL) and HF outcomes in the HFDMP; specifically, to investigate the prevalence of depression in the HFDMP and the impact of depression in the QoL and clinical symptoms and outcomes of 124 patients enrolled and followed in the program over a period of 12-months. Results and discussion: The pathophysiological mechanisms responsible for MD are complex and multifaceted. The multiple studies conducted in this work and its respective findings support the idea of that complexity. The hypothesis of a central role for oxidative stress in MD is supported and highlighted by the findings (*1.1) of increased levels of an early component of the peroxidation chain, the lipid hydroperoxide (LPH), in euthymic patients with BD-I and BD-II into late life. This suggests a persistent effect of reactive species of oxygen throughout a BD patient’s life. Further proof for this hypothesis can be gleaned from the findings (*1.2) of increased peripheral levels of two lipid peroxidation markers, the 4-hydroxy-2-nonenal (4-HNE) and LPH, which is significantly correlated with depressive episodes and with decreased dentate gyrus–CA4 volume. It was shown that a larger number of depressive episodes predict a greater volume loss in patient’s hippocampus. Furthermore, it was shown that 4-HNE is negatively associated with left and right dentate gyrus–CA4 volumes. Altogether, these results are consistent in proving the by oxidative stress in the pathophysiology of MD. Furthermore, they suggest that depressive episodes and elevated oxidative stress might contribute to hippocampal volume loss in these patients. In addition, these findings provide further support for the hypothesis that peripheral lipid peroxidation markers may reflect brain alterations and may one day represent a biomarker for these disorders. Oxidative stress is a mechanism well established in CVD and in our findings it appears to also play a central role in MD pathophysiology. Therefore, oxidative stress is an extremely likely biological interface between mood and heart problems. Oxidative stress and inflammation are closely related pathophysiological mechanisms. One can be easily induced by the other, therefore both processes are simultaneously found in many pathological conditions. Similar to oxidative stress, inflammation also seems to be implicated in the pathophysiology of both depression and CVD. Inflammation also seems to have a central role in MD and to be a central candidate mediating the link between depression and CVD, as suggested by the results of our review (*2.1). Taking this hypothesis further, we have found original and seemingly important biological pathways shared between depression and HF, highlighted by the findings in our study (*2.2) that suggest that NT-proBNP is a potential mediator between reduced EF and depression in patients with HF. This indicates that NT-proBNP may be a potential biomarker for HF patients with reduced EF and depression who are at higher risk in terms of disease prognosis and mortality. In this work we have also investigated the original approach established at HSN for HF management named HFDMP. This program was shown (*3.1) to be a safe and effective way to manage HF and avoid ED visits and decrease HF hospitalizations and mortality, all while improving clinical symptoms and decreasing health care expenditures. Finally, we have shown that depression is extremely prevalent in this program’s HF population (*3.2) and is associated with worse QoL levels in patients and worse outcomes in the program. On the other hand, this program was shown to be highly effective in improving patients’ QoL over time, both physically and emotionally. Altogether, these studies provide further evidence of the strong association between HF and depression, from molecular to clinical perspectives. Conclusion: Altogether this work leads to several conclusions. Firstly, oxidative stress and inflammation appear to have a central role in the pathophysiology of MD. Oxidative stress and a decreased volume in hippocampus seem to be directly implicated in the pathophysiology of depressive episodes in MD. Furthermore, oxidative stress and inflammation are central candidates for proving a shared pathophysiology between MD and CVD. Clinical and biological pathways were found between HF and depression in patients enrolled in the HFDMP. This original program for HF management – demonstrated also in this work to be an efficient and safe way to manage HF in an outpatient setting – was shown to have a population where depression is extremely prevalent and where depression influences HF patients’ QoL and outcomes. Finally, in this work we have found an important indicator for the existence of a shared pathophysiology between depression and HF: depressive symptoms and reduced ejection fraction in HF seem to be mediated by NT-proBNP, a gold-standard indicator of the clinical status, diagnosis and severity of HF. We have found that this biological marker, extremely well established in HF, reacts differently with the existence of depressive symptoms in HF patients. This finding provides a strong hint at the common pathways between these diseases. Altogether, this work contributed to clarifications on the pathophysiology of MD as well as the biological connections between MD and CVD. Furthermore, this work has shown important indicators of common pathways between depression and HF, two highly impactful comorbid diseases. It is expected that these findings may in the future contribute to the development of better therapeutic approaches and biomarkers that will help in the diagnosis of disease progression and the evaluation of treatment response in patients with this comorbidity.

The establishment of a functional patterned vascular network is crucial for development, tissue growth and homeostasis. The mis-patterning or dysfunction of this network is associated with cancer, stroke and arteriovenous malformations. The formation of a functional vascular network requires two distinct processes – formation of primitive vascular plexuses through sprouting angiogenesis; and vascular remodelling, the reorganization of primitive plexuses into a hierarchical network of arteries, capillaries and veins. During sprouting angiogenesis endothelial cells polarise and migrate towards sources of VEGF while upon vascular remodelling, they polarise and migrate against the blood flow direction. Nevertheless, the molecular mechanisms regulating migration and polarisation during sprouting and vascular remodelling are poorly understood. Here, we investigate the mechanisms of polarisation and migration during sprouting angiogenesis by unravelling the role of Arp2/3 complex in vascular morphogenesis, using a combination of in vitro approaches with the power of mouse genetics. We show that Arp2/3 is crucial for endothelial cell migration, for the establishment of cellular protrusions and that its depletion drastically affects vascular morphogenesis during development and tumour angiogenesis. At the same time, we show that blood flow and VEGF interact to orchestrate patterns of endothelial cell polarity at the network-level. We demonstrate that this interaction defines the transition between two distinct morphogenic events, vascular sprouting and vascular remodelling. Accordingly, manipulation of VEGF gradients or blood flow in vivo compromises normal polarity patterns, resulting in delayed or premature remodelling of blood vessels. At the molecular level, we show that mechanotransduction at adherens junctions is key for VEGF-induced polarisation and negatively regulates flow-dependent collective polarisation. Specifically, we propose that vinculin recruitment is key for mechanotransduction and the establishment of a transition zone. In addition, we generated a polarity reporter mouse with endothelial cell nuclei and Golgi labelled that will enable the study the dynamics of endothelial cell polarisation downstream of VEGF and blood flow in development, homeostasis and disease. Given the physiological relevance of vascular patterning in health and disease, our approach will give insights in defining the cellular and molecular principles involved in vascular patterning.

O sucesso a longo prazo dos implantes dentários depende do estabelecimento de uma condição estável. Essa estabilidade pode ser influenciada negativamente pela coexistência da doença peri-implantar, provocando a perda do tecido ósseo de suporte ao redor do implante. Em relação à etiopatogenia, duas vias distintas foram descritas: A via clássica e a via retrógrada contendo factores de origem biológica e biomecânica previamente modelados. A existência de algoritmos de risco para modelagem de doenças assume um papel importante na Medicina moderna, representando uma importante ferramenta para os clínicos no processo de diagnóstico e decisão. Atualmente não existem algoritmos de risco validados em Implantologia. Os objetivos desta investigação foram: 1. Construir um algoritmo de risco para a incidência de doença peri-implantar e estimar a fracção atribuível dos indicadores de risco; 2. Avaliar a capacidade de previsão de um algoritmo de risco para a incidência de doença peri-implantar numa amostra de validação; 3. Aferir a influência dos tempos de recall (tempo decorrido entre consultas de avaliação e manutenção) nas sub-amostras de risco da doença peri-implantar; 4. Identificar os factores associados ao prognóstico dos implantes com doença peri-implantar, construir e validar um modelo preditivo do prognóstico de implantes com doença peri-implantar. A doença peri-implantar foi definida como a presença de profundidade à sondagem ≥ 5 mm; hemorragia à sondagem e presença concomitante de perda óssea marginal / perda do nível de inserção clínico em comparação com a avaliação anterior. O primeiro estudo consistiu num estudo de caso-controlo de modo a definir uma pontuação de risco para previsão da incidência de doença peri-implantar em 1275 pacientes utilizando as variáveis: história de periodontite, o desajuste passivo ou desaperto de componentes protéticos, tipo de material utilizado na restauração, placa bacteriana, hemorragia, proximidade de outros implantes / dentes, nível ósseo, tabagismo e a interação entre a placa bacteriana e a proximidade de outros implantes / dentes. Foram estimadas a fracção atribuível (FA) para cada indicador de risco e as razões de verossimilhança positiva e negativa para a pontuação de risco. O segundo estudo consistiu num estudo de coorte retrospectivo para validação da pontuação de risco durante um seguimento de 5 anos em 353 pacientes. Foram estimados o efeito da estratificação do grupo de risco na incidência de doença periimplantar através de regressão logística e a capacidade de discriminação da pontuação de risco através da área sob a curva (AUC). O terceiro estudo compreendeu a avaliação do efeito e impacto de um recall (intervalo entre as consultas de manutenção) > 6 meses na doença peri-implantar num estudo caso-controlo aninhado na coorte do estudo 2, incluindo 170 pacientes. Foram estimados o risco relativo (RR) através de regressão logística condicional e o risco proporcional atribuível (RAP). O quarto estudo consistiu num estudo de coorte retrospectivo com 240 pacientes para derivar e validar um modelo prognóstico para implantes com doença peri-implantar, com estimativa do efeito de potenciais indicadores de prognóstico desfavorável. Foram estimados o RR para os indicadores de risco através de regressão logística e a capacidade de discriminação do modelo através da AUC em ambos os conjuntos de derivação e validação. 1. Na construção da pontuação de risco e estimativa das frações atribuíveis constatámos: Uma distribuição diferente da incidência de doença peri-implantar com 2.2%, 11.5%, 37.6% e 86.4% para pacientes de baixo risco, risco moderado, risco elevado e risco muito elevado, respectivamente. A pontuação de risco com uma capacidade de discriminação excepcional registada pela AUC com um intervalo de confiança de 95% (IC95%) de 0.963 (0.950; 0.976) (p < 0.001). As razões de verossimilhança positiva e negativa do modelo de 9.69 e 0.13, respectivamente. Uma potencial redução significativa dos casos se a exposição aos indicadores de risco modificáveis fosse suprimida, ilustrada pela redução de 5% para a falta de ajuste passivo ou desaperto de componentes protéticos; 31% para a placa bacteriana; 31% para o nível ósseo no terço médio dos implantes; 18% para a hemorragia; 26% para a restauração de metalo-cerâmica; e 15% para a interação entre placa bacteriana e proximidade de outros implantes / dentes. 2. Na validação da pontuação de risco registámos: Uma taxa de incidência da doença peri-implantar de 24.1% (n = 85 pacientes). Um RR com intervalo de confiança de 95% (IC95%) para o efeito da estratificação do grupo de risco de 5.52 (3.64; 8.35), com uma sensibilidade de 71.8%, uma especificidade de 83.2% e uma precisão de 80.5%. Uma excelente capacidade de discriminação da pontuação de risco com uma AUC (IC95%) de 0.858 (0.820; 0.896) (p < 0.001). 3. Na avaliação do efeito e impacto do período de recall na incidência da doença peri-implantar registámos: Um RR (IC95%) de 1.13 (0.57; 2.21) para um recall > 6 meses. Um RAP de 5.89% que indica uma contribuição modesta do recall > 6 meses para o excesso de risco na doença peri-implantar. 4. Na derivação e validação do modelo de prognóstico para implantes com doença peri-implantar constatámos: Indicadores de prognóstico com efeito significativo no prognóstico desfavorável, conforme o RR (IC95%) de 3.13 (1.15; 8.55) para história de periodontite; 3.26 (1.37; 7.81) para doença peri-implantar grave; 3.52 (1.48; 8.37) para o comprimento do implante> 13 mm; e 3.99 (1.62; 9.82) para o desenvolvimento precoce da doença. Um modelo prognóstico com inclusão dos indicadores de prognóstico: idade, história de periodontite, doença peri-implantar grave, comprimento do implante> 13 mm e desenvolvimento precoce da doença. O modelo prognóstico obteve boa discriminação na amostra de derivação com uma AUC (IC95%) de 0.763 (0.679; 0.847) (p < 0.001); O modelo prognóstico obteve uma boa discriminação na amostra de validação com uma AUC (IC95%) de 0.709 (0.616; 0.803) (p < 0.001). Considerando os resultados, conclui-se que a gestão clinica da doença periimplantar beneficia da aplicação de algoritmos de risco para alcançar resultados previsíveis a longo prazo. Foi possível derivar e validar uma pontuação de risco para a incidência da doença peri-implantar com excelente discriminação. Um número significativo de casos poderia ser potencialmente evitado se a exposição a factores modificáveis fosse suprimida. O efeito e impacto de um recall > 6 meses foram baixos e não significativos. Foi possível derivar e validar um modelo prognóstico para implantes dentários com doença peri-implantar com boa discriminação. As presentes conclusões determinam a necessidade de intervenções na prevenção primária, secundária e terciária para maximizar a probabilidade de resultados de sucesso. Esta tese como um todo contribui para a avaliação do prognóstico em pacientes com implantes dentários e doença peri-implantar, preenchendo lacunas significativas no conhecimento contemporâneo. Sistematiza os principais factores associados à doença peri-implantar e a um prognóstico desfavorável por meio de uma abordagem epidemiológica que se pode considerar original na investigação actual. Este trabalho atesta a origem multifactorial da condição, simplificando cálculos matemáticos complexos e fornecendo ferramentas que podem ser usadas por qualquer clínico na gestão de pacientes com reabilitações implanto-suportadas. Pode servir como ponto de partida para estudos adicionais sobre o tema da doença peri-implantar e fornece uma base para intervenções preventivas e curativas. Destaca-se a importância de um acompanhamento clínico rigoroso com a inclusão de medidas diagnósticas precisas, profilaxia adequada e planeamento com envolvimento de uma equipa multidisciplinar.

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Increasing evidence supports the existence of intratumor heterogeneity (ITH) in many cancer types, with potential clinical implications for both cancer diagnosis and treatment. Expansion of genetically distinct cell populations within a tumor creates a subclonal architecture that varies dynamically throughout cancer progression. This acquired cancer trait, ITH is the substrate for Darwinian evolution to act upon, selecting the subclones that carry beneficial phenotypes. The outgrowth of such subclones impacts cancer development, drug resistance and tumor relapse. Nevertheless, despite the key role ITH plays in cancer, important questions regarding its magnitude, origin and genetic drivers across different cancer types remain largely unanswered. By facilitating the emergence of nucleotide sequence mutations, copy-number alterations (CNA), chromosomal translocations or aneuploidies, genomic instability has been regarded as the major source of ITH. However, discrepancies in the rates of genomic instability and ITH observed in previous comprehensive studies suggest that additional events congregate to increase genetic heterogeneity in tumors. It is widely accepted that genetic alterations and disruption of epigenetic regulatory mechanisms are hallmarks of cancer. Nonetheless, while the genetic contribution for cancer development is easily illustrated by mutations in tumor suppressors or oncogenes, the epigenetic involvement and its functional relevance is far more complex and has only recently become a major focus of cancer research. Besides genetic mutations, cancer cells invariably present with some degree of epigenetic alterations that contribute to the acquisition of the cancer hallmarks. Indeed, there is evidence that epigenomic reprogramming plays a seminal role in tumorigenesis by creating a progenitor-like cell state that facilitates expression of driver mutations and tumor initiation. High throughput genome sequencing (HTS) efforts have identified driver mutations in genes that regulate the epigenome, namely genome-wide chromatin and deoxyribonucleic acid (DNA) methylation. While epigenetic deregulation in cancer arises primarily as a consequence of DNA mutations, the view that altered epigenomes may also change DNA mutations rates highlights reciprocal interactions that contribute to cancer development. Accordingly, epigenomic disruption should favour the development of genetically diverse tumor cell populations, fuelling ITH. In fact, a possible relationship between genomic and epigenomic alterations during clonal evolution of tumors has been suggested in esophageal squamous cell carcinoma and glioma where high concordance was observed between the evolution of genetic and epigenetic diversification. In this PhD thesis, I conducted an exhaustive characterization of ITH across 2,807 tumor samples from 16 different carcinomas using whole-exome sequencing (WES) datasets from The Cancer Genome Atlas (TCGA). Integration of ITH scores and somatic variants detected in each tumor sample revealed that mutations in epigenetics modifier genes are the stronger determinants of ITH and display an association with increased clonal evolution across several cancer types. To investigate whether epigenomic deregulation drives the development of tumors with high levels of ITH, we focused our analysis on kidney renal clear cell carcinoma (KIRC), the cancer type with the highest frequency of mutations in epigenetic modifiers. The important role of epigenomic deregulation in the development and progression of KIRC is illustrated by the finding that patients with mutations in epigenetic modifiers have worse overall survival than those without mutations in these genes. In particular, genes that regulate genome-wide histone and DNA methylation emerged as candidate drivers of ITH. Knockout of histone methyltransferase SETD2 or DNA methyltransferase DNMT3A using the CRISPR/Cas9 system on renal carcinoma cells led to significant expansion of genetically distinct clones and culminated in highly heterogeneous cell populations which recapitulate the heterogeneity levels observed in the TCGA patients. We thus reasoned that the increased ITH observed after DNMT3A or SETD2 knockouts likely underpins variations in mitochondrial metabolism upon which natural selection can act. Indeed, changes in mitochondrial metabolism constitute an important source of variability for natural selection during cancer evolution. Upon DNMT3A or SETD2 knockout we observed that positively selected clones displayed similar mutational spectra and increased mitochondrial bioenergetic performance under stress conditions, suggesting that specific patterns of genotypic and phenotypic variation emerge upon epigenomic deregulation. Our work provides new insights on tumor development and unravels new drivers of ITH, showing an unprecedented pan-cancer portrait of the major determinants of ITH. This work validates experimentally the role of specific epigenetic modifier genes and lays a foundation for more effective cancer prognosis and treatment.

Atualmente, a maioria da população mundial vive em cidades. A qualidade de vida nas cidades é fortemente afetada pelo planeamento urbano e particularmente, pela existência de Espaços Verdes Urbanos (EVU). A inventariação destes espaços é, por isso, da maior importância. As imagens da superfície terrestre obtidas por deteção remota, através de satélites artificiais, permitem identificar a cobertura do solo através do espectro eletromagnético. Esta é, no entanto uma tarefa complexa, devido à diversidade das caraterísticas envolvidas (árvores e arbustos, suas dimensões, o tipo de folhagem, lagos e respetivas margens) e à variabilidade temporal das imagens em função das estações do ano. Algoritmos de aprendizagem automática para classificação de objetos através de imagens de satélite, como por exemplo, Support Vector Machines (SVM), árvores de decisão, redes neuronais, têm sido amplamente utilizados na análise da cobertura e uso do solo o que sugere a pertinência da sua utilização para classificar a composição dos EVU, em particular, Redes Neuronais Convolucionais (RNC) aplicadas a imagens de satélite. O objetivo desta dissertação é verificar a eficácia de métodos de aprendizagem automática para classificar a cobertura do solo em EVU, a partir de dados extraídos de imagens de satélite. Foi utilizado um conjunto de dados pré classificados obtidos no âmbito do projeto Bioveins. Este projeto é uma parceria entre várias Universidades Europeias e a BiodivERsA the network programming and funding research on biodiversity and ecosystem services across European countries and territories. A Universidade de Lisboa é uma destas Universidades e por isso os dados foram extraídos e preparados pelo Departamento de Biologia Vegetal da FCUL. O conjunto de dados engloba 220 EVU em 7 cidades europeias (Lisboa, Almada, Zurique, Paris, Antuérpia, Poznan e Tartu). Os dados iniciais estão estruturados em três grupos. O primeiro grupo inclui as coordenadas geográficas do centróide do espaço de 10 metros por 10 metros que representa cada instância. O segundo grupo inclui 27 caraterísticas obtidas das imagens de satélite, nomeadamente as frequências do espectro eletromagnético visível e invisível ao olho humano, e um conjunto de índices de vegetação, solo, água, densidade urbana e outras variáveis biofísicas. Finalmente o terceiro grupo, corresponde à classificação dos componentes dos espaços, efetuada por observação no terreno. Esta classificação foi efetuada atribuindo uma percentagem a cada uma de 11 categorias: Allotment gardens, Artificial surfaces, Ornamental bed, Barelands, Grassland, Water surfaces, Riparian vegetation, Shrubs, Broad deciduous, Broad ever e Coniferous. Foram efetuadas três abordagens de aprendizagem automática. Numa primeira abordagem ao problema, aplicámos um método ensemble, floresta aleatória, criando um classificador binário para cada categoria. Para esta abordagem foram utilizados como unidade de entrada, os centróides dos espaços, sem qualquer transformação estrutural, mas o conjunto de dados foi transformado em 11 réplicas, uma para cada categoria, em que a cada instância, foi atribuída uma classe com o valor 0 ou 1, correspondendo à ausência ou presença da respetiva categoria. Na segunda abordagem, utilizámos um único classificador multiclasse, constituído por uma RNC. Os dados foram agrupados de forma a constituirem imagens para input da rede. A cada imagem foi atribuída uma classe através de um número inteiro correspondendo a uma das categorias, aquela com maior predominância nessa imagem, considerando todos os pontos que a formaram. A última abordagem baseou-se também numa RNC mas utilizando tantos classificadores quantas as categorias existentes. À semelhança do que foi feito para o método ensemble, cada imagem foi classificada com valores de 0 ou 1, correspondendo à ausência ou presença da categoria respetiva na imagem. A classificação binária pelo método ensemble não produziu resultados muito satisfatórios, mas ambas as abordagens com RNC evidenciaram serem bastante eficazes para a classificação dos EVU. A abordagem multiclasse apresenta vantagens pela simplicidade de implementação pois utiliza um único classificador. Os dados utilizados para aprendizagem apresentam um elevado desequilíbrio de classes, por isso a obtenção de mais dados e mais variados, poderá representar um desafio para futuro trabalho sobre o tema, assim como uma abordagem multilabel que pode permitir um resultado mais detalhado para cada espaço.

Ngodo e marrabenta são palavras que, Para um público não familiarizado com a história e as práticas culturais moçambicanas da região sul do país, soam bastante misteriosas. Não é possível explica- -las em apenas um parágrafo. Seus significados cambiantes ao longo do século XX, assim como a complexidade das relações sociais, políticas e culturais que rodeiam o ngodo e a marrabenta, serão explorados ao longo do texto. Porém, cabe apresentar, de antemão e minimamente, o que ambas significam. A mbiÌa (no plural, timbila) é uma espécie de xilofone, denominada corriqueiramente pelos Portugueses ao longo do século XX como marimba, muito comum no grupo étnico chopi, que ocupa, principalmente, a região de ZavaIa. Possui diferentes tamanhos e afinações. A mbila foi usada em orquestras, compostas também por tambores e bailarinos, financiadas pelos chefes chopi, e funcionavam, desde pelo menos meados da primeira metade do século XIX, como um importante demarcador de pertencimento cultural empregado nas apresentações do ngodo (no plural, migodo). O ngodo é "um conjunto de canções e instrumentos organizados em uma composição".3 A apresentação de um ngodo, com seus diferentes movimentos, compostos por dança, canções (chamadas de mzeno) e instrumentos, era previamente elaborada Por um compositor, que a coordenava e comandava.

En 1955, Fernand Deligny publie « La caméra outil pédagogique », dont le statut se situe entre le manifeste et la proposition d’expérimentation. Quoique éloigné de la théorie au sens strict, le texte est composé d’une série de thèses implicites et présente ce qu’on pourrait appeler la première synthèse de l’auteur sur le cinéma et l’image. Il constitue en outre un document historique, révélant une prise de position originale par rapport aux polémiques de l’après-guerre sur le rôle sociopolitique du cinéma. Je voudrais ici m’attarder sur les thèses sous-jacentes à ce texte. Le geste de Deligny – celui de mettre une caméra entre les mains de jeunes délinquants – est certes original et radical, mais mon hypothèse est qu’il s’inscrit dans une tradition matérialiste et un courant soviéto-marxiste souterrain qui se sont transmis de manière discontinue et indirecte.

Acute lymphoblastic leukemia (ALL) is the most frequent cancer found in children and results from the clonal expansion of transformed lymphoid precursors. Approximately 15% of pediatric ALL patients present with a T-cell phenotype (T-ALL). Despite the recent improvements in the treatment of T-ALL, there are still a high number of relapses and the intensive chemotherapeutic regiments used are associated with long-term severe complications. In order to develop new therapeutic strategies that can further increase efficacy while reducing side effects, one needs to better understand the pathobiology of T-ALL. In particular, it is necessary to understand how microenvironmental and cell-autonomous mechanisms influence the initiation and the progression of leukemia. The present thesis has the preocupation of exploring the mechanisms by which both an extracellular cue (IL-4) and a cell-intrinsic transcription factor (TAL1) may partake in leukemia development and maintenance. Interleukin-4 (IL-4) is a γ-common chain cytokine produced within the bone marrow microenvironment that is known to promote the in vitro proliferation of T-ALL cells. In Chapter 2, we present evidence that IL-4 induces primary T-ALL cell cycle progression from G0/G1 into S and G2/M, by up-regulating cyclin D2, E and A and down-regulating the cyclin-dependent kinase inhibitor p27kip1. Transfection of T-ALL cells with the VP22-p27kip1 fusion protein, which is able to translocate into the cytoplasm and nucleus of target cells, abrogates IL-4-mediated proliferation. This indicates that p27kip1 downregulation is mandatory for cell cycle progression of T-ALL cells stimulated with IL-4. Furthermore, IL-4 stimulates mTOR activation, as determined by increased phosphorylation of its downstream targets p70S6K, S6 and 4E-BP1. Inhibition of mTOR signaling with rapamycin prevents IL-4-induced T-ALL cell growth, cell cycle progression and proliferation. Our results identify mTOR as a critical regulator of IL-4-mediated effects in T-ALL cells and support the rationale for using mTOR pharmacological inhibitors in T-ALL therapy (Cardoso et al. Leukemia 2009). The basic helix-loop-helix transcription factor TAL1 is aberrantly expressed in up to 65% of T-ALL patients. LMO2, a Lim-only domain protein, is often co-expressed ectopically with TAL1 in this malignancy. These genes appear to have leukemogenic potential, since both TAL1 and LMO2 transgenic mice develop leukemias of T-cell phenotype. However, it is still unclear whether TAL1 is effectively leukemogenic in humans, or whether merely participates as a secondary event in the transformation Abstract xiii process in T-ALL. To address this question, we transduced hematopoietic progenitors with TAL1 and/or LMO2 and co-cultured them with OP9-Dll1 stromal cells, which have the capacity to induce T-cell differentiation in vitro. We found that TAL1 and LMO2 genes deregulate human T-cell differentiation in stromal cell co-cultures. Interestingly, the coordinated expression of both TAL1 and LMO2 led to a relative increase in CD3+CD4+CD8+ T-cell precursors with increased cell size. This observation is particularly interesting given that TAL1-expressing patients normally display a similar phenotype. These preliminary results show that TAL1 and LMO2 can disrupt normal human T-cell development, therefore likely predisposing thymocytes to malignant transformation (Chapter 3). In our effort to characterize the mechanisms by which TAL1 might promote T-cell leukemogenesis, we developed a TAL1 inducible system, by fusing TAL1 with the hormone binding domain (HBD) of the estrogen receptor (ER), which we expressed in a TAL1-negative T-cell line. Upon 4-Hydroxi-Tamoxifen (4OHT) treatment, ER-TAL1 fusion protein is able to translocate into the nucleus and consequently trigger its transcriptional program. Gene expression profiling of 4OHT-treated HPB-ALL cells stably transduced with the ER-TAL1 fusion revealed a total of 26 genes up- or down-regulated by TAL1 activation, in at least two independent experiments. We selected seven of those genes on the basis of their function/potential interest in cancer and confirmed the differential expression of three (CASZ1, DMGDH and OR5M3) by qRT-PCR. Accordingly, transfection of another TAL1-negative T-ALL cell line, P12, with TAL1, also led to increased expression of the validated TAL1 target genes. The possible involvement of CASZ1 in TAL1-mediated anti-apoptotic and proliferative effects in T-ALL cells was subsequently investigated. Knock-down of TAL1 with siRNA in the TAL1-positive T-ALL cell line Jurkat decreased the expression of CASZ1, correlating with loss of cell viability. Moreover, CASZ1 knockdown in Jurkat cells led to functional effects similar to those of TAL1 knockdown, namely a decrease in survival and proliferation. Overall, these studies allowed the identification of three novel TAL1 downstream targets, likely with functional relevance for TAL1-mediated leukemogenic potential (Chapter 4). TAL1 binds to repressive chromatin complexes, namely involving HDAC1, and incubation with HDAC inhibitors (HDACis) promotes apoptosis of leukemia cells derived from TAL1 transgenic mice. In Chapter 5, we evaluated the impact of HDACis on TAL1 from a somewhat different perspective, namely by analyzing their impact on Abstract xiv TAL1 expression rather than transcriptional activity. We found that incubation of T-ALL cells with HDACis strikingly down-regulates TAL1 protein expression. This is due to decreased TAL1 gene transcription in cells with an intact TAL1 locus, and to impaired TAL1 mRNA translation in cells that harbor the TAL1d deletion. Importantly, HDACi-induced apoptosis of T-ALL cells is significantly reversed by TAL1 forced over-expression. Our results indicate that the HDACi-mediated apoptotic program in T-ALL cells is partially dependent on the down-regulation of TAL1 expression, and suggest that integration of HDACis into T-ALL treatment protocol may be of potential therapeutic benefit (Cardoso et al, Leukemia 2011, advance online publication). Taken together, the results described in this thesis highlight the importance that microenviromental factors, such as IL-4, might have in the progression of T-ALL (for instance, by activating mTOR and promoting cell cycle progression), and hint on the importance that cell-autonomous factors, such as TAL1 and LMO2, may have in predisposing T-cells for malignant transformation and promoting survival of T-ALL cells. Importantly, our results further demonstrate that both extracellular cues and intracellular molecular lesions can constitute targets for therapeutic intervention in T-ALL.

A 66-year-old man presented with a 10-month history ofmultiple pruritic flesh-colored translucent papules on both eye-brows. Two of the lesions were located on the left eyebrowand four grouped on the right eyebrow (Fig. 1a,b). He had noother symptoms or signs. Clinical examination showed noother skin lesions nor evidence of lymphadenopathy. A punchbiopsy specimen revealed a dense nodular infiltrate ofmonomorphic small to medium-sized B lymphocytes involvingthe superficial and deep dermis (Fig. 2a–c). Immunohistochem-ical studies disclosed immunoreactivity of the lymphoid cellsfor CD5, CD20, and CD23 (Fig. 2d). The complete blood cellcount was normal, but the flow cytometry of the peripheralblood, the myelogram, and the osteomedullary biopsy wereconsistent with B-cell chronic lymphocytic leukemia (B-CLL)diagnosis. Fluorescence in situ hybridization (FISH) analysisperformed on peripheral blood and bone marrow disclosed nochromosomal abnormalities. Imaging studies showed no signifi-cant alterations. The patient was treated with local irradiationof lesions (total cumulative dose of 12 Gray), with completeclinical resolution. There were no complications or recurrenceat 12 months follow-up.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has put severe strain on global economies, healthcare systems, and public health. The coronavirus disease 2019 (COVID-19) seems to carry significantly higher mortality compared to the viral respiratory infections we were accustomed to.

While public health campaigns against SARS-CoV-2 pandemic are underway to promote frequent hand washing as one of the ancillary strategies to reduce transmission of the newly identified virus, we are seeing a rise in the frequency of hand dermatitis, many of which are predominantly irritative in nature. While irritative contact dermatitis due to the use of Personal Protection Equipment (PPE) and frequent hand sanitizing has been under the mediatic spotlight we must not forget the significant impact social isolation measures may have on our allergic contact dermatitis patients, who may find themselves deprived of the diagnostic work-up or clinical follow-up necessary for their improvement, during this crisis.

This report analyzes the case of a 63-year-old Caucasian male, Fitzpatrick III phototype, who was admitted to the Intensive Care Unit (ICU) due to a severe coronavirus disease 2019 (COVID-19) pneumonia. The patient’s medical history was remarkable for valvular heart disease, the presence of a biological aortic valve, and primary arterial hypertension, but he had no prior history of any dermatological complaints. The patient was chronically medicated with furosemide 40mg bid, apixaban 5mg bid, and amlodipine 5mg od. At admission, laboratory studies revealed mild lymphopenia (900/µL). Lopinavir/ritonavir and hydroxychloroquine were administered for 14 days. He was ventilated at admission for 21 days until his respiratory function improved.