993 resultados para Lipid transport


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

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Familial renal glucosuria (FRG) is a rare co -dominantly inherited benign phenotype characterized by the presence of glucose in the urine. It is caused by mutations in the SLC5A2 gene that encodes SGLT2, a Na+ -glucose co -transporter. The purpose of our current work was twofold: to characterize the molecular and phenotype findings of an FRG cohort and, in addition, to detail the SGLT2 expression in the adult human kidney. The phenotype of FRG pedigrees was evaluated using direct sequencing for the identification of sequence variations in the SLC5A2 gene. The expression of SGLT2 in the adult human kidney was studied by immunofluorescence on kidney biopsy specimens. In the absence of renal biopsies from FRG individuals, and in order to evaluate the potential disruption of SGLT2 expression in a glucosuric nephropathy, we have selected cases of nucleoside analogues induced proximal tubular toxicity. We identified six novel SLC5A2 mutations in six FRG pedigrees and described the occurrence of hyperuricosuria associated with hypouricaemia in the two probands with the most severe phenotypes. Histopathological studies proved that SGLT2 is localized to the brush -border of the proximal tubular epithelia cell and that this normal pattern was found to be disrupted in cases of nucleoside analogues induced tubulopathy. We present six novel SLC5A2 mutations, further contributing to the allelic heterogeneity in FRG, and identified hyperuricosuria and hypouricaemia as part of the FRG phenotype. SGLT2 is localized to the brush -border of the proximal tubule in the adult human normal kidney, and aberrant expression of the co -transporter may underlie the glucosuria seen with the use of nucleoside analogues.

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

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RESUMO:Aterosclerose é uma das principais causas de morbilidade e mortalidade no mundo ocidental. É responsável, direta ou indiretamente, pela maior percentagem de gastos com a saúde na maioria dos países europeus. A “teoria lipídica” da aterosclerose, que se baseia na dislipidemia como causa primária para a doença vascular tem algumas implicações práticas importantes: permite a definição de linhas de orientação e protocolos simples e ainda estabelece alvos terapêuticos que podem ser atingidos na maior parte dos casos com a atual intervenção farmacológica. A associação da aterosclerose com o sistema imunológico (a “teoria imunológica”), forneceu por sua vez novas formas de explorar os mecanismos envolvidos e abriu novas perspetivas para um conhecimento mais completo da doença. No entanto, levanta dificuldades evidentes no que diz respeito às possibilidades terapêuticas. De todos os intervenientes no processo aterosclerótico (bioquímicos, imunológicos e anatómicos), as lipoproteínas de elevada densidade (HDL) são atualmente reconhecidas como um dos fatores mais importantes na aterogénese. Isto é baseado no reconhecimento das múltiplas propriedades anti-aterogénicas das HDL como por exemplo: a anti-oxidante, a anti-inflamatória e a antitrombótica, bem como o seu importante papel na melhoraria da função endotelial. Atualmente, é consensual que as funções anti-aterogénicas das HDL vão além do seu papel no transporte reverso do colesterol (RCT) e a importância das HDL no processo aterosclerótico baseia-se não apenas no seu papel protetor impedindo a formação da placa de ateroma, mas também na estabilização destas, prevenindo a sua ruptura e, consequentemente o evento trombótico. Como fundamentais no processo aterosclerótico estão reconhecidos dois principais conjuntos de eventos: um caracterizado por alterações no metabolismo das lipoproteínas que resultam em lipoproteínas pró-inflamatórias e pró-oxidantes que interagem com os componentes celulares da parede arterial e que conduzem à formação da placa de ateroma; o outro evento é a resposta imunológica desencadeada contra um novo conjunto de antigénios que por sua vez leva à produção de citoquinas pró-inflamatórias. Dada a complexidade da HDL e das suas múltiplas funções estas lipoproteínas tornaram-se um potencial alvo para a resposta auto-imune, e cujas consequências podem explicar algumas das associações identificados em estudos clínicos e epidemiológicos. Contudo esta interação entre o sistema imunológico e HDL nunca foi exaustivamente estudada. Portanto, pomos a hipótese de que em condições oxidativas e pró-inflamatórias, um aumento do antigénio (HDL) conduz a um consequente acréscimo na produção de anticorpos anti-HDL (aHDL) responsáveis pela alteração quantitativa e / ou qualitativa das HDL. O conceito de que estes anticorpos podem contribuir tanto para a evolução a longo prazo do processo aterosclerótico, como para o desencadeamento de eventos clínicos pode também explicar a heterogeneidade encontrada em cada doente e nos grandes estudos clínicos, no que diz respeito aos fatores de risco e outcomes clínicos. Para além disso, a confirmação desta hipótese pode permitir explicar porque é que as intervenções terapêuticas atualmente em desenvolvimento para aumentar os níveis de HDL, não conseguem mostrar a tão esperada redução do risco vascular. O objetivo geral desta tese foi identificar e caracterizar a resposta humoral contra os componentes da HDL, e avaliar possíveis mecanismos que possam contribuir para a modificação das propriedades anti-aterogénicas das HDL. Para alcançar este objetivo investigou-se: 1) A presença de anticorpos aHDL em doentes com lúpus eritematoso sistémico (SLE) e em doentes com manifestações clínicas de aterosclerose, como os doentes com doença arterial coronária (CAD), acidente vascular cerebral isquémico (IS) e diabetes tipo 2; 2) Os principais alvos antigénicos dentro do complexo das HDL e a associação entre os títulos de anticorpos aHDL e diferentes características clínicas destas doenças; 3) As modificações das funções normais associadas às HDL, em particular da função anti-oxidante e anti-inflamatória; 4) A atividade biológica dos anticorpos aHDL isolados do soro de doentes através de um conjunto de experiências in vitro de inibição da atividade da paraoxonase 1 (PON1) e da expressão de moléculas de adesão em culturas de células endoteliais. Para tal foi necessário estabelecer um método de isolamento dos anticorpos. Os anticorpos aHDL isolados do soro de doentes foram utilizados de forma a identificar as potenciais alterações dos sistemas celulares utilizados; 5) O efeito de fármacos usados no tratamento das dislipidemias, em particular o ácido nicotínico e as estatinas, na variação dos títulos de anticorpos aHDL através de ensaios clínicos randomizados, controlados com placebo e em dupla ocultação. Os métodos utilizados neste trabalho incluíram: técnicas imunológicas (como por exemplo, enzyme-linked immunoabsorbent assay - ELISA, ensaio imunoturbidimetrico e cromatografia de imuno-afinidade) técnicas bioquímicas (tais como a quantificação de atividade enzimática por espectrofotometria e por luminescência), experiências com cultura de células e citometria de fluxo. Os nossos resultados mostram que: 1) A presença de anticorpos aHDL, e mais especificamente anticorpos contra alguns do seus principais componentes como a apolipoproteína A-I (ApoA-I, principal apolipoproteína presente nas HDL) e a PON1 (o enzima que mais contribui para a propriedade anti-oxidante das HDL), quer em doentes com doenças auto-imunes, como o SLE, quer em doentes com manifestações clínicas de aterosclerose, como CAD, IS e diabetes tipo 2. Os doentes apresentaram títulos de anticorpos IgG aHDL, aApoA-I e aPON1 significativamente mais elevados do que controlos saudáveis com a mesma idade e sexo. 2) A correlação positiva estatisticamente significativa entre os títulos de aHDL e aApoA-I e aPON1 sugere que estes sejam dois dos principais alvos antigénicos dentro do complexo das HDL. Os anticorpos encontrados nestes doentes estão associados com a diminuição da atividade da PON1 e a uma redução da capacidade anti-oxidante total (TAC) do soro, um aumento dos biomarcadores de disfunção endotelial (como por exemplo dos metabolitos do óxido nítrico - NO2- e NO3-, as moléculas de adesão vascular e intracelular - VCAM-1 e ICAM-1 e os níveis de 3-nitrotirosina). Nos doentes com SLE os títulos destes estão associados a um aumento do dano cardiovascular e à atividade global da doença avaliados pelas escalas SLICC/ACR DI e BILAG score, respetivamente. Enquanto que nos doentes com diabetes tipo 2 estes anticorpos estão associados com um aumento dos níveis de glicemia em jejum (FGP) e hemoglobina glicada (HbA1c). 3) Após se ter estabelecido um método de isolamento dos anticorpos que permite isolar quantidades significativas de anticorpos do soro de doentes sem perder a sua especificidade, foi identificada a capacidade dos anticorpos isolados do soro de doentes inibirem de uma forma dependente da concentração a atividade da PON1 até um máximo de 70% no caso dos doentes com SLE e ente 7-52% no caso dos anticorpos isolados de doentes com CAD e IS. 4) O efeito anti-inflamatório das HDL na inibição da produção de VCAM-1 induzida por citoquinas (como o TNF-) foi revertido em mais de 80% pelos anticorpos aHDL isolados do soro de doentes. 5) A angiogenesis induzida por HDL através do aumento do fator de crescimento do endotélio vascular (VEGF) foi anulada em 65% pelos anticorpos aHDL isolados do soro de doentes. 6) Os atuais agentes farmacológicos disponíveis para aumentar as concentrações de HDL-C estão associados a um aumento dos títulos de anticorpos.-------- ABSTRACTAtherosclerosis is the major cause of morbidity and mortality in the western world. It is also responsible, directly or indirectly, for the highest percentage of health costs in most European countries. Despite the use of new technologies for the diagnosis of vascular disease and regardless of the major advances in treatment, the atherosclerosis-related clinical burden is still raising. The “lipid theory” of atherogenesis, which identifies dyslipidemia as the primary cause of this vascular disease has some important practical implications: it allows the definition of simple guidelines and establishes therapeutic targets which can be generally met with current pharmacologic intervention. The association between atherosclerosis an the immune system (the immune concept) has in turn provided new ways of exploring the mechanisms involved in this condition and has opened new perspectives in the understanding of the disease. However, it raises obvious difficulties when it comes to treatment options. Of all the players (biochemical, immunological and anatomical) involved in this matter, high-density lipoproteins (HDL) are currently recognised as one of the most important factors in atherogenesis. This is based on the recognition of HDL's multiple anti-atherogenic properties: anti-oxidant, anti-inflammatory and antithrombotic, as well as its capacity to improve endothelial function. Nowadays, it is widely recognized that the anti-atherogenic functions of HDL go beyond reverse cholesterol transport (RCT), and the importance of HDL is based not just on its ability to reduce atheroma formation but also on its ability to stabilise plaques, therefore preventing their rupture and ultimately thrombosis. Two main set of events have been recognised as fundamental in atherogenesis: one, characterized by lipoprotein metabolism alterations, resulting in pro-inflammatory and pro-oxidative lipoproteins, which interact with the normal cellular elements of the arterial wall leading to atheroma formation; the other, the immune cellular response towards new sets of antigens which lead to the production of pro-inflammatory cytokines. Given to HDL complexity and multiple functions this lipoprotein has became a potential target for an auto-immune response, the consequences of which may explain some of the association identified in epidemiological and clinical studies, though the interaction between the immune system and HDL has never been thoroughly addressed. Therefore, we hypothesized that under oxidative and pro-inflammatory conditions, the increase in the antigen (HDL) would lead to a consequent increase in the production of anti-HDL (aHDL) antibodies be responsible for quantitative and/or qualitative changes of HDL. The concept that these antibodies may contribute either to the long-term evolution of atherosclerosis or to the triggering of clinical events may also explain the heterogeneity found in individual patients and in large cohorts regarding risk factors and clinical outcomes. Moreover this may be a major breakthrough in understanding why therapeutic interventions that increase HDL levels, failed to show the anticipated reduction in vascular risk. The overall aims of this thesis were to identified and characterize the humoral response towards HDL components and to evaluate the possible mechanisms that may contribute to the modifications of the anti-atherogenic properties of HDL. To achieve this objective we investigated: 1) the presence of aHDL antibodies in patients with systemic lupus erythematosus (SLE) and in patients with atherosclerosis-related clinical events, such as coronary artery disease (CAD), ischemic stroke (IS) and type 2 diabetes; 2) the association between the titres of aHDL antibodies and different clinical features of these diseases; 3) the modifications of the anti-atherogenic properties of HDL; 4) the biologic effect of aHDL antibodies isolated from serum of patients on the anti-oxidant and anti-inflammatory properties of HDL; 5) the effect of different pharmacologic treatments for dyslipidemia on the prevalence and activity of aHDL antibodies. The methodologies used in this work included immunologic-related techniques (e.g. enzyme-linked immunoabsorbent assay – ELISA, immunoturbidimetric immunoassay and immunoaffinity chromatography), biochemical techniques (enzymatic assays with quantification by spectrophotometry and luminescence methods), cell culture experiments and flow cytometry. Our results indicate that: 1) The titres of IgG aHDL, anti-apolipoprotein A-I (aApoA-I) and anti-paraoxonase 1 (aPON1) antibodies were higher in patients with SLE, CAD, IS and type 2 diabetes when compared with age and sex matched healthy controls. 2) The antibodies found in these patients were associated with decreased PON1 activity, (the enzyme responsible for most of the anti-oxidant effect of HDL), reduced total anti-oxidant capacity (TAC) of serum and increased biomarkers of endothelial dysfunction (nitric oxide metabolites, adhesion molecules, nitrotyrosine). In patients with SLE the antibody titres were associated with an increase in disease-related cardiovascular damage and activity whereas in patients with type 2 diabetes they were directly related with the fasting glucose plasma (FGP) levels and the glycosylated haemoglobin (HbA1c). 3) The antibodies isolated from serum of our patients, directly inhibited HDL-associated PON1 activity in a dose dependent way ranging from 7 to 52%. 4) The anti-inflammatory effect of HDL, measured by the percentage of inhibition of the cytokine-induced production of vascular adhesion molecules (VCAM-1), was reduced in more than 80% by aHDL antibodies isolated from our patients. 5) The HDL-induced angiogenesis by increasing vascular endothelial growth factor (VEGF) levels was abrogated in 65% by the antibodies isolated from serum of patients. 6) The current available pharmacologic agents for increasing HDL-C concentrations were associated with an increase in the titres of IgG aApoA-I antibodies. This increase was higher in the extended release niacin when compared to statins probably due to their dampening effect on oxidative stress. In conclusion, aHDL antibodies are present in different pathologic conditions. aHDL antibodies represent a family of self-reacting immunoglobulins, of which ApoA-I and PON1 might be the most relevant targets. These antibodies are biologically active, interfering with the HDL anti-oxidant and anti-inflammatory properties and, consequently, with the atherosclerotic process. The pathogenic potential of these antibodies may lead to the identification of a new biomarker for vascular disease, whilst presenting itself as a novel target for a different treatment approach which may redefine the treatment strategies and clinical trials design for HDL interventions in the future.

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Dissertação para obtenção do Grau de Mestre em Engenharia Civil – Perfil de Estruturas

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

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INTRODUCTION: HIV-infected children and adolescents treated with highly active antiretroviral therapy (HAART) regimens that include a protease inhibitor (PI) can show significant improvements in clinical outcomes, nutritional status and quality of life. The study aimed to report nutritional and metabolic alterations for pediatric patients continuously exposed to HAART and for healthy controls for up to 1 year. METHODS: Clinical, anthropometric, lipid profile and food intake data were collected prospectively over approximately 12-months for each patient. RESULTS: Fifty-one individuals were studied, of these, 16 were healthy. After 12 months follow-up, HIV-positive individuals remained below the healthy control group parameters. No change was observed concerning food intake. Triglyceride serum levels were higher in patients using protease inhibitor at the onset of the study [PI groups: 114 (43 - 336), and 136 (63 - 271) versus control group: 54.5 (20 - 162); p = 0.003], but after twelve months follow-up, only the group using protease inhibitor for up to two months presented higher values [140 (73 - 273) versus 67.5 (33 - 117); p = 0.004]. HDL-cholesterol was lower in HIV-positive individuals [HIV-positive groups: 36 (27 - 58) and 36 (23 - 43); control 49.5 (34 - 69); p = 0.004]. CONCLUSIONS: HIV-infected children and adolescents treated with highly active antiretroviral therapy showed compromised nutritional parameters compared to a paired healthy control group. Individuals using protease inhibitor presented worse triglyceride serum levels compared to their healthy counterparts.

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Zero valent iron nanoparticles (nZVI) are considered very promising for the remediation of contaminated soils and groundwaters. However, an important issue related to their limited mobility remains unsolved. Direct current can be used to enhance the nanoparticles transport, based on the same principles of electrokinetic remediation. In this work, a generalized physicochemical model was developed and solved numerically to describe the nZVI transport through porous media under electric field, and with different electrolytes (with different ionic strengths). The model consists of the Nernst–Planck coupled system of equations, which accounts for the mass balance of ionic species in a fluid medium, when both the diffusion and electromigration of the ions are considered. The diffusion and electrophoretic transport of the negatively charged nZVI particles were also considered in the system. The contribution of electroosmotic flow to the overall mass transport was included in the model for all cases. The nZVI effective mobility values in the porous medium are very low (10−7–10−4 cm2 V−1 s−1), due to the counterbalance between the positive electroosmotic flow and the electrophoretic transport of the negatively charged nanoparticles. The higher the nZVI concentration is in the matrix, the higher the aggregation; therefore, low concentration of nZVI suspensions must be used for successful field application.

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Introduction Although the initiation of highly active antiretroviral therapy (HAART) is accompanied by an attenuation of viral load, metabolic disorders characterized by hyperglycemia, dyslipidemia, and lipodystrophy are often observed in patients under this treatment. Certain foods, such as oat bran, soy protein, and flaxseed, have been shown to improve a patient's lipid profile despite possible increases in uricemia. Thus, a bioactive compound was formulated using these foods to help patients with HIV/AIDS control metabolic disorders resulting from HAART. Methods An uncontrolled before and after study was performed. The total cholesterol, HDL-cholesterol, LDL-cholesterol, triglycerides, and uric acid before and after 3 months of consuming the formulation were compared in patients. The compound was formulated such that 40g (the recommended daily intake) contained approximately 10g of flaxseed, 20g of oat bran, and 10g of textured soy protein. Results The study population consisted of 139 patients, 31 of whom were included in the final analysis. There were no significant variations between the laboratory results obtained before and after consumption of the compound. Conclusions The regular consumption of the formulation together with individualized dietary guidance did not reduce lipid levels and did not contribute to an increase in uricemia in the study group. However, new studies with higher doses of the foods that compose the formulation should be encouraged to investigate whether these foods can positively influence the lipid profiles of these patients.

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This thesis is a study of how heat is transported in non-steady-state conditions from a superconducting Rutherford cable to a bath of superfluid helium (He II). The same type of superconducting cable is used in the dipole magnets of the Large Hadron Collider (LHC). The dipole magnets of the LHC are immersed in a bath of He II at 1.9 K. At this temperature helium has an extremely high thermal conductivity. During operation, heat needs to be efficiently extracted from the dipole magnets to keep their superconducting state. The thermal stability of the magnets is crucial for the operation of the LHC, therefore it is necessary to understand how heat is transported from the superconducting cables to the He II bath. In He II the heat transfer can be described by the Landau regime or by the Gorter-Mellink regime, depending on the heat flux. In this thesis both measurements and numerical simulation have been performed to study the heat transfer in the two regimes. A temperature increase of 8 2 mK of the superconducting cables was successfully measured experimentally. A new numerical model that covers the two heat transfer regimes has been developed. The numerical model has been validated by comparison with existing experimental data. A comparison is made between the measurements and the numerical results obtained with the developed model.

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Ocean acidification, recognized as a major threat to marine ecosystems, has developed into one of the fastest growing fields of research in marine sciences. Several studies on fish larval stages point to abnormal behaviours, malformations and increased mortality rates as a result of exposure to increased levels of CO2. However, other studies fail to recognize any consequence, suggesting species-specific sensitivity to increased levels of CO2, highlighting the need of further research. In this study we investigated the effects of exposure to elevated pCO2 on behaviour, development, oxidative stress and energy metabolism of sand smelt larvae, Atherina presbyter. Larvae were caught at Arrábida Marine Park (Portugal) and exposed to different pCO2 levels (control: ~600μatm, pH=8.03; medium: ~1000μatm, pH=7.85; high: ~1800μatm, pH=7.64) up to 15days, after which critical swimming speed (Ucrit), morphometric traits and biochemical biomarkers were determined. Measured biomarkers were related with: 1) oxidative stress - superoxide dismutase and catalase enzyme activities, levels of lipid peroxidation and DNA damage, and levels of superoxide anion production; 2) energy metabolism - total carbohydrate levels, electron transport system activity, lactate dehydrogenase and isocitrate dehydrogenase enzyme activities. Swimming speed was not affected by treatment, but exposure to increasing levels of pCO2 leads to higher energetic costs and morphometric changes, with larger larvae in high pCO2 treatment and smaller larvae in medium pCO2 treatment. The efficient antioxidant response capacity and increase in energetic metabolism only registered at the medium pCO2 treatment may indicate that at higher pCO2 levels the capacity of larvae to restore their internal balance can be impaired. Our findings illustrate the need of using multiple approaches to explore the consequences of future pCO2 levels on organisms.

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Combinatorial Optimization Problems occur in a wide variety of contexts and generally are NP-hard problems. At a corporate level solving this problems is of great importance since they contribute to the optimization of operational costs. In this thesis we propose to solve the Public Transport Bus Assignment problem considering an heterogeneous fleet and line exchanges, a variant of the Multi-Depot Vehicle Scheduling Problem in which additional constraints are enforced to model a real life scenario. The number of constraints involved and the large number of variables makes impracticable solving to optimality using complete search techniques. Therefore, we explore metaheuristics, that sacrifice optimality to produce solutions in feasible time. More concretely, we focus on the development of algorithms based on a sophisticated metaheuristic, Ant-Colony Optimization (ACO), which is based on a stochastic learning mechanism. For complex problems with a considerable number of constraints, sophisticated metaheuristics may fail to produce quality solutions in a reasonable amount of time. Thus, we developed parallel shared-memory (SM) synchronous ACO algorithms, however, synchronism originates the straggler problem. Therefore, we proposed three SM asynchronous algorithms that break the original algorithm semantics and differ on the degree of concurrency allowed while manipulating the learned information. Our results show that our sequential ACO algorithms produced better solutions than a Restarts metaheuristic, the ACO algorithms were able to learn and better solutions were achieved by increasing the amount of cooperation (number of search agents). Regarding parallel algorithms, our asynchronous ACO algorithms outperformed synchronous ones in terms of speedup and solution quality, achieving speedups of 17.6x. The cooperation scheme imposed by asynchronism also achieved a better learning rate than the original one.

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Abstract INTRODUCTION : The human T-lymphotropic virus-1 (HTLV-1) is associated with chronic inflammatory diseases such as HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP), a chronic inflammatory disease. Disturbances in lipid metabolism are involved in inflammatory and demyelinating diseases. METHODS : Plasma levels of triglycerides, total cholesterol, and fractions of HTLV-1-infected individuals of both sexes with different clinical progressions were determined. RESULTS : Elevated levels of triglyceride and very low-density lipoproteins (VLDL) were exclusively detected in HTLV-1-infected women from asymptomatic and HAM/TSP groups compared with uninfected individuals (p = 0.02). CONCLUSIONS : Elevated triglyceride and VLDL levels in HTLV-1-infected women may be related to the predominance of HAM/TSP in women.