Structural analysis of the timber structure of Ica Cathedral, Peru

Dissertação de mestrado em Structural Analysis of Monuments and Historical Constructions

Assessment of innovative solutions for non-load bearing masonry enclosures

This paper presents some of the results of the research project “Masonry Enclosures” developed in the framework of the transnational access (TA) to LNEC’s triaxial shake table within the FP7 project SERIES.

The role of IFNγ in higher brain function: in health and under chronic stress

Tese de Doutoramento em Ciências da Saúde

Development of an in vitro model for studying aneurysms

The aims of this project was to develop an arterial aneurysm using either enzymatic or laser degradation of the arterial wall without affecting the viability of the tissue and to cultivate the arteries under pulsatile flow conditions in a vascular bioreactor with a view to investigate the progress of the disease. Characteristics of aneurysms are the degradation of smooth muscle cells, collagen and elastin. Detached smooth muscle cells and degradation of the collagen matrix and elastin fibres were observed in arteries degraded with enzymes elastase and collagenase. Only remnants of the arterial wall were detected after cultivation. This might be a suitable model for late stage aneurysms. Arteries treated with the laser system showed no charring or heat damage of the not dissected area. Collagen matrix, smooth muscle cells and elastin fibres were intact. A clear defined cut was made in a depth of 200 μm and tissue was removed. Following cultivation of these arteries a dilation of the laser-eroded area was observed. This model can mimic atherosclerotic aneurysms, when plaques weaken the tunica media of the blood vessel wall and rupture. Limitations of this study were contamination of the bioreactor system and a low number of cultivations. The aim to generate a living arterial aneurysm in vitro was not achieved. Tissue viability decreased to the level of negative controls after cultivation.

Bioequivalence of meat products treated with high pressure processing

The effects of high pressure on the composition of food products have not been evaluated extensively. Since, it is necessary to take in consideration the possible effects in basis to the changes induced in the bio molecules by the application of high pressures. The main effect on protein is the denaturation, because the covalent bonds are not affected; however hydrogen bonding, hydrophobic and intermolecular interactions are modified or destroyed. 1 High pressure can modify the activity of some enzymes. If this is done the proteolysis and lipolysis could be more or less intense and the content of free amino acids and fatty acids will be different. This could be related to the bioavailability of these compounds. Low pressures (100 MPa) have been shown to activate some enzymes (monomeric enzymes). Higher pressures induce loss of the enzyme activity. However some enzymes are very stable (ex. Lipase ~ 600 - 1000 MPa). Lipoxygenase is less stable, and there is little information about the effects on antioxidant enzymes. Other important issue is the influence of high pressure on oxidation susceptibility. This could modify the composition of lipids if the degree of the oxidation would have been higher or lower than in the traditional product. Pressure produces the damage of cell membranes favouring the contact between substrates and enzymes, exposure to oxidation of membrane fatty acids and loos of the efficiency of vitamin E. These effects can also affect to protein oxidation. In this study different compounds were analysed to establish the differences between non-treated and high-pressure treated products.

Neuropathological substrates and structural changes in late-life depression: the impact of vascular burden.

A first episode of depression after 65 years of age has long been associated with both severe macrovascular and small microvascular pathology. Among the three more frequent forms of depression in old age, post-stroke depression has been associated with an abrupt damage of cortical circuits involved in monoamine production and mood regulation. Late-onset depression (LOD) in the absence of stroke has been related to lacunes and white matter lesions that invade both the neocortex and subcortical nuclei. Recurrent late-life depression is thought to induce neuronal loss in the hippocampal formation and white matter lesions that affect limbic pathways. Despite an impressive number of magnetic resonance imaging (MRI) studies in this field, the presence of a causal relationship between structural changes in the human brain and LOD is still controversial. The present article provides a critical overview of the contribution of neuropathology in post-stroke, late-onset, and late-life recurrent depression. Recent autopsy findings challenge the role of stroke location in the occurrence of post-stroke depression by pointing to the deleterious effect of subcortical lacunes. Despite the lines of evidences supporting the association between MRI-assessed white matter changes and mood dysregulation, lacunes, periventricular and deep white matter demyelination are all unrelated to the occurrence of LOD. In the same line, neuropathological data show that early-onset depression is not associated with an acceleration of aging-related neurodegenerative changes in the human brain. However, they also provide data in favor of the neurotoxic theory of depression by showing that neuronal loss occurs in the hippocampus of chronically depressed patients. These three paradigms are discussed in the light of the complex relationships between psychosocial determinants and biological vulnerability in affective disorders.

Low doses of ionizing radiation promote tumor growth and metastasis by enhancing angiogenesis.

Radiotherapy is a widely used treatment option in cancer. However, recent evidence suggests that doses of ionizing radiation (IR) delivered inside the tumor target volume, during fractionated radiotherapy, can promote tumor invasion and metastasis. Furthermore, the tissues that surround the tumor area are also exposed to low doses of IR that are lower than those delivered inside the tumor mass, because external radiotherapy is delivered to the tumor through multiple radiation beams, in order to prevent damage of organs at risk. The biological effects of these low doses of IR on the healthy tissue surrounding the tumor area, and in particular on the vasculature remain largely to be determined. We found that doses of IR lower or equal to 0.8 Gy enhance endothelial cell migration without impinging on cell proliferation or survival. Moreover, we show that low-dose IR induces a rapid phosphorylation of several endothelial cell proteins, including the Vascular Endothelial Growth Factor (VEGF) Receptor-2 and induces VEGF production in hypoxia mimicking conditions. By activating the VEGF Receptor-2, low-dose IR enhances endothelial cell migration and prevents endothelial cell death promoted by an anti-angiogenic drug, bevacizumab. In addition, we observed that low-dose IR accelerates embryonic angiogenic sprouting during zebrafish development and promotes adult angiogenesis during zebrafish fin regeneration and in the murine Matrigel assay. Using murine experimental models of leukemia and orthotopic breast cancer, we show that low-dose IR promotes tumor growth and metastasis and that these effects were prevented by the administration of a VEGF receptor-tyrosine kinase inhibitor immediately before IR exposure. These findings demonstrate a new mechanism to the understanding of the potential pro-metastatic effect of IR and may provide a new rationale basis to the improvement of current radiotherapy protocols.

Intra-articular osteoid osteoma of the knee: clinical and therapeutical particularities

The intra-articular osteoid osteoma (10-13% of the cases) is often difficult to identify. They present frequent atypical clinical signs and radiological images that eventually lead to inadequate treatment. For example, it has been observed that this pathology leads to inappropriate arthroscopies (up to 40%). Meniscal tear and then osteochondritis were initially suspected on a patient with an intra-articular osteoid osteoma at the tibia level. For the treatment, any damage of the cartilage has to be avoided. Thermoablation with radiofrequency is the treatment of choice

Magnetic resonance imaging determinants of intraindividual variability in the elderly: combined analysis of grey and white matter.

Elderly individuals display a rapid age-related increase in intraindividual variability (IIV) of their performances. This phenomenon could reflect subtle changes in frontal lobe integrity. However, structural studies in this field are still missing. To address this issue, we computed an IIV index for a simple reaction time (RT) task and performed magnetic resonance imaging (MRI) including voxel based morphometry (VBM) and the tract based spatial statistics (TBSS) analysis of diffusion tensor imaging (DTI) in 61 adults aged from 22 to 88 years. The age-related IIV increase was associated with decreased fractional anisotropy (FA) as well as increased radial (RD) and mean (MD) diffusion in the main white matter (WM) fiber tracts. In contrast, axial diffusion (AD) and grey matter (GM) densities did not show any significant correlation with IIV. In multivariate models, only FA has an age-independent effect on IIV. These results revealed that WM but not GM changes partly mediated the age-related increase of IIV. They also revealed that the association between WM and IIV could not be only attributed to the damage of frontal lobe circuits but concerned the majority of interhemispheric and intrahemispheric corticocortical connections.

Derris (Lonchocarpus) urucu (Leguminosae) Extract Modifies the Peritrophic Matrix Structure of Aedes aegypti (Diptera:Culicidae)

Aqueous suspension of ethanol extracts of Derris (Lonchocarpus) urucu (Leguminosae), collected in the state of Amazonas, Brazil, were tested for larvicidal activity against the mosquito Aedes aegypti (Diptera:Culicidae). The aim of this study was to observe the alterations of peritrophic matrix in Ae. aegypti larvae treated with an aqueous suspension of D. urucu extract. Different concentrations of D. urucu root extract were tested against fourth instar larvae. One hundred percent mortality was observed at 150 µg/ml (LC50 17.6 µg/ml) 24 h following treatment. In response to D. urucu feeding, larvae excreted a large amount of amorphous feces, while control larvae did not produce feces during the assay period. Ultrastructural studies showed that larvae fed with 150 µg/ml of D. urucu extract for 4 h have an imperfect peritrophic matrix and extensive damage of the midgut epithelium. Data indicate a protective role for the peritrophic matrix. The structural modification of the peritrophic matrix is intrinsically associated with larval mortality.

In vitro measurement of enzymatic markers as a tool to detect mouse cardiomyocytes injury

Primary cultures of cardiomyocytes represent a useful model for analyzing cardiac cell biology as well as pathogenesis of several cardiovascular disorders. Our aim was to standardize protocols for determining the damage of cardiac cells cultured in vitro by measuring the creatine kinase and its cardiac isotype and lactate dehydrogenase activities in the supernatants of mice cardiomyocytes submitted to different protocols of cell lysis. Our data showed that due to its higher specificity, the cardiac isotype creatine kinase was the most sensitive as compared to the others studied enzymatic markers, and can be used to monitor and evaluate cardiac damage in in vitro assays.

Regulation of inflammasome activity.

Summary Interleukin-1beta (IL-1beta) is a potent inflammatory cytokine, which is implicated in acute and chronic inflammatory disorders. The activity of IL-1beta is regulated by the proteolytic cleavage of its inactive precursor resulting in the mature, bioactive form of the cytokine. Cleavage of the IL-1beta precursor is performed by the cysteine protease caspase-1, which is activated within protein complexes termed 'inflammasomes'. To date, four distinct inflammasomes have been described, based on different core receptors capable of initiating complex formation. Both the host and invading pathogens need to control IL-1beta production and this can be achieved by regulating inflammasome activity. However, we have, as yet, little understanding of the mechanisms of this regulation. In particular the negative feedbacks, which are critical for the host to limit collateral damage of the inflammatory response, remain largely unexplored. Recent exciting findings in this field have given us an insight into the potential of this research area in terms of opening up new therapeutic avenues for inflammatory disorders.

The role of platelet and plasma markers of antioxidant status and oxidative stress in thrombocytopenia among patients with vivax malaria

Malaria remains an important health problem in tropical countries like Brazil. Thrombocytopenia is the most common hematological disturbance seen in malarial infection. Oxidative stress (OS) has been implicated as a possible mediator of thrombocytopenia in patients with malaria. This study aimed to investigate the role of OS in the thrombocytopenia of Plasmodium vivax malaria through the measurement of oxidant and antioxidant biochemical markers in plasma and in isolated platelets. Eighty-six patients with P. vivax malaria were enrolled. Blood samples were analyzed for total antioxidant and oxidant status, albumin, total protein, uric acid, zinc, magnesium, bilirubin, total thiols, glutathione peroxidase (GPx), malondialdehyde (MDA), antibodies against mildly oxidized low-density lipoproteins (LDL-/nLDL ratio) and nitrite/nitrate levels in blood plasma and GPx and MDA in isolated platelets. Plasma MDA levels were higher in thrombocytopenic (TCP) (median 3.47; range 1.55-12.90 µmol/L) compared with the non-thrombocytopenic (NTCP) patients (median 2.57; range 1.95-8.60 µmol/L). Moreover, the LDL-/nLDL autoantibody ratio was lower in TCP (median 3.0; range 1.5-14.8) than in NTCP patients (median 4.0; range 1.9-35.5). Finally, GPx and MDA were higher in the platelets of TPC patients. These results suggest that oxidative damage of platelets might be important in the pathogenesis of thrombocytopenia found in P. vivax malaria as indicated by alterations of GPx and MDA.

X-ray structures of Sap1 and Sap5: structural comparison of the secreted aspartic proteinases from Candida albicans.

Proteolytic activity is an important virulence factor for Candida albicans (C. albicans). It is attributed to the family of the secreted aspartic proteinases (Saps) from C. albicans with a minimum of 10 members. Saps show controlled expression and regulation for the individual stages of the infection process. Distinct isoenzymes can be responsible for adherence and tissue damage of local infections, while others cause systemic diseases. Earlier, only the structures of Sap2 and Sap3 were known. In our research, we have now succeeded in solving the X-ray crystal structures of the apoenzyme of Sap1 and Sap5 in complex with pepstatin A at 2.05 and 2.5 A resolution, respectively. With the structure of Sap1, we have completed the set of structures of isoenzyme subgroup Sap1-3. Of subgroup Sap4-6, the structure of the enzyme Sap5 is the first structure that has been described up to now. This facilitates comparison of structural details as well as inhibitor binding modes among the different subgroup members. Structural analysis reveals a highly conserved overall secondary structure of Sap1-3 and Sap5. However, Sap5 clearly differs from Sap1-3 by its electrostatic overall charge as well as through structural conformation of its entrance to the active site cleft. Design of inhibitors specific for Sap5 should concentrate on the S4 and S3 pockets, which significantly differ from Sap1-3 in size and electrostatic charge. Both Sap1 and Sap5 seem to play a major part in superficial Candida infections. Determination of the isoenzymes' structures can contribute to the development of new Sap-specific inhibitors for the treatment of superficial infections with a structure-based drug design program.

Regulation of inflammasome activity.

Summary Interleukin-1beta (IL-1beta) is a potent inflammatory cytokine, which is implicated in acute and chronic inflammatory disorders. The activity of IL-1beta is regulated by the proteolytic cleavage of its inactive precursor resulting in the mature, bioactive form of the cytokine. Cleavage of the IL-1beta precursor is performed by the cysteine protease caspase-1, which is activated within protein complexes termed 'inflammasomes'. To date, four distinct inflammasomes have been described, based on different core receptors capable of initiating complex formation. Both the host and invading pathogens need to control IL-1beta production and this can be achieved by regulating inflammasome activity. However, we have, as yet, little understanding of the mechanisms of this regulation. In particular the negative feedbacks, which are critical for the host to limit collateral damage of the inflammatory response, remain largely unexplored. Recent exciting findings in this field have given us an insight into the potential of this research area in terms of opening up new therapeutic avenues for inflammatory disorders.