Iron Overload in a Murine Model of Hereditary Hemochromatosis Is Associated with Accelerated Progression of Osteoarthritis Under Mechanical Stress

OBJECTIVE: Hereditary hemochromatosis (HH) is a disease caused by mutations in the Hfe gene characterised by systemic iron overload and associated with an increased prevalence of osteoarthritis (OA) but the role of iron overload in the development of OA is still undefined. To further understand the molecular mechanisms involved we have used a murine model of HH and studied the progression of experimental OA under mechanical stress. DESIGN: OA was surgically induced in the knee joints of 10-week-old C57BL6 (wild-type) mice and Hfe-KO mice. OA progression was assessed using histology, micro CT, gene expression and immunohistochemistry at 8 weeks after surgery. RESULTS: Hfe-KO mice showed a systemic iron overload and an increased iron accumulation in the knee synovial membrane following surgery. The histological OA score was significantly higher in the Hfe-KO mice at 8 weeks after surgery. Micro CT study of the proximal tibia revealed increased subchondral bone volume and increased trabecular thickness. Gene expression and immunohistochemical analysis showed a significant increase in the expression of matrix metallopeptidase 3 (MMP-3) in the joints of Hfe-KO mice compared with control mice at 8 weeks after surgery. CONCLUSIONS: HH was associated with an accelerated development of OA in mice. Our findings suggest that synovial iron overload has a definite role in the progression of HH-related OA

Usefulness of zebrafish model to assess glomerular and tubular alterations induced by tenofovir

Tenofovir (TFV) is one of the most used antiretroviral drugs. However, it is associated with tubular damage with mitochondria as a possible target. Tubulopathy precedes glomerular dysfunction, thus classic markers of renal function like the glomerular filtration rate (GFR) do not detect early TFV damage. Prediction and management of drug induced renal injury (DIRI) rely on the mechanisms of the drug insult and in optimal animal models to explore it. Zebrafish (Danio rerio) offers unique advantages for assessing DIRI, since the pronephros is structurally very similar to its human counterpart and is fully developed at 3.5 days postfertilization. The main aim of the present work was to evaluate the effects of TFV, as well as its pro-drug, tenofovir disoproxil fumarate (TDF), on the GFR and in mitochondria morphology in tubular cells of zebrafish larvae. Lethality curves were performed to understand the relationship between drug concentration and lethality. LC10 was selected to explore the renal function using the FITC-inulin assay and to analyze the mitochondrial toxicity by electron microscopy on larvae exposed to TDF, TFV, paracetamol and gentamicin (positive controls) or water (negative control). Lethality curves showed that gentamicin was the most lethal drug, followed by TDF, TFV and paracetamol. Gentamicin and paracetamol decreased the GFR, but no differences were found for either TDF or TFV, when compared to controls (%FITC Control = 33±8; %FITC TDF = 35±10; %FITC TFV = 30±10; %FITC Gentamicin = 46±17; %FITC Paracetamol = 83±14). Tubular mitochondria from treated larvae were notably different from non-treated larvae, showing swelling, irregular shapes, decreased mitochondria network, cristae disruption and loss of matrix granules. These results are in agreement with the effects of these drugs in humans and thus, demonstrate that zebrafish larvae can be a good model to assess the functional and structural damage associated with DIRI.

Search for the Standard Model Higgs boson produced in association with top quarks and decaying into bb in pp collisions at √s = 8 TeV with the ATLAS detector

A search for the Standard Model Higgs boson produced in association with a pair of top quarks, tt¯H, is presented. The analysis uses 20.3 fb−1 of pp collision data at s√ = 8 TeV, collected with the ATLAS detector at the Large Hadron Collider during 2012. The search is designed for the H to bb¯ decay mode and uses events containing one or two electrons or muons. In order to improve the sensitivity of the search, events are categorised according to their jet and b-tagged jet multiplicities. A neural network is used to discriminate between signal and background events, the latter being dominated by tt¯+jets production. In the single-lepton channel, variables calculated using a matrix element method are included as inputs to the neural network to improve discrimination of the irreducible tt¯+bb¯ background. No significant excess of events above the background expectation is found and an observed (expected) limit of 3.4 (2.2) times the Standard Model cross section is obtained at 95% confidence level. The ratio of the measured tt¯H signal cross section to the Standard Model expectation is found to be μ=1.5±1.1 assuming a Higgs boson mass of 125 GeV.

Forest structure of artificial islands in the Tucuruí dam reservoir in northern Brazil: a test core-area model

Construction of hydroelectric dams in tropical regions has been contributing significantly to forest fragmentation. Alterations at edges of forest fragments impact plant communities that suffer increases in tree damage and dead, and decreases in seedling recruitment. This study aimed to test the core-area model in a fragmented landscape caused by construction of a hydroelectric power plant in the Brazilian Amazon. We studied variations in forest structure between the margin and interiors of 17 islands of 8-100 hectares in the Tucuruí dam reservoir, in two plots (30 and >100m from the margin) per island. Mean tree density, basal area, seedling density and forest cover did not significantly differ between marginal and interior island plots. Also, no significant differences were found in liana density, dead tree or damage for margin and interior plots. The peculiar topographic conditions associated with the matrix habitat and shapes of the island seem to extend edge effects to the islands' centers independently of the island size, giving the interior similar physical microclimatic conditions as at the edges. We propose a protocol for assessing the ecological impacts of edge effects in fragments of natural habitat surrounded by induced (artificial) edges. The protocol involves three steps: (1) identification of focal taxa of particular conservation or management interest, (2) measurement of an "edge function" that describes the response of these taxa to induced edges, and (3) use of a "Core-Area Model" to extrapolate edge function parameters to existing or novel situations.

Development of a model for occupational risk acceptance criteria definition in industrial settings

Doctoral Dissertation for PhD degree in Industrial and Systems Engineering

Encapsulation and controlled release of bioactive compounds in lactoferrin-glycomacropeptide nanohydrogels : curcumin and caffeine as model compounds

Curcumin and caffeine (used as lipophilic and hydrophilic model compounds, respectively) were successfully encapsulated in lactoferrin-glycomacropeptide (Lf-GMP) nanohydrogels by thermal gelation showing high encapsulation efficiencies (>90 %). FTIR spectroscopy confirmed the encapsulation of bioactive compounds in Lf-GMP nanohydrogels and revealed that according to the encapsulated compound different interactions occur with the nanohydrogel matrix. The successful encapsulation of bioactive compounds in Lf-GMP nanohydrogels was also confirmed by fluorescence measurements and confocal laser scanning microscopy. TEM images showed that loaded nanohydrogels maintain their spherical shape with sizes of 112 and 126 nm for curcumin and caffeine encapsulated in Lf-GMP nanohydrogels, respectively; in both cases a polydispersity of 0.2 was obtained. The release mechanisms of bioactive compounds through Lf-GMP nanohydrogels were evaluated at pH 2 and pH 7, by fitting the Linear Superimposition Model to the experimental data. The bioactive compounds release was found to be pH-dependent: at pH 2, relaxation is the governing phenomenon for curcumin and caffeine compounds and at pH 7 Ficks diffusion is the main mechanism of caffeine release while curcumin was not released through Lf-GMP nanohydrogels.

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.

A Multisectorial model of prices: the SAM approach

Social Accounting Matrices (SAM) are normally used to analyse the income generation process. They are also useful, however, for analysing the cost transmission and price formation mechanisms. For price contributions, Roland-Holst and Sancho (1995) used the SAM structure to analyse the price and cost linkages through a representation of the interdependence between activities, households and factors. This paper is a further analysis of the cost transmission mechanisms, in which I add the capital account to the endogenous components of the Roland-Holst and Sancho approach. By doing this I reflect the responses of prices to the exogenous shocks in savings and investment. I also present an additive decomposition of the global price effects into categories of interdependence that isolates the impact on price levels of shocks in the capital account. I use a 1994 Social Accounting Matrix to make an empirical application of the Catalan economy. Keywords: social accounting matrix, cost linkages, price transmission, capital account. JEL Classification: C63, C69, D59.

Chronic murine myocarditis due to Trypanosoma cruzi: an ultrastructural study and immunochemical characterization of cardiac interstitial matrix

In an attempt to define the mouse-model for chronic Chagas' disease, a serological, histopathological and ultrastructural study as well as immunotyping of myocardium collagenic matrix were performed on Swiss mice, chronically infected with Trypanosoma cruzi strains: 21 SF and mambaí (Type II); PMN and Bolivia (Type III), spontaneously surviving after 154 to 468 days of infection. Haemagglutination and indirect immunofluorescence tests showed high titres of specific antibodies. The ultrastructural study disclosed the cellular constitution of the inflammatory infiltrate showing the predominance of monocytes, macrophages with intense phagocytic activity, fibroblasts, myofibroblasts and abundant collagen matrix suggesting the association of the inflammatory process with fibrogenesis in chronic chagasic cardiomyopathy. Artertolar and blood capillary alterations together with dissociation of cardiac cells from the capillary wall by edema and inflammation were related to ultrastructural lesions of myocardial cells. Rupture of parasitized cardiac myocells contribute to intensify the inflammatory process in focal areas. Collagen immunotyping showed the predominance of Types III and IV collagen. Collagen degradation and phagocytosis were present suggesting a reversibility of the fibrous process. The mouse model seems to be valuable in the study of the pathogenetic mechanisms in Chagas cardiomyopathy, providing that T. cruzi strains of low virulence and high pathogenecity are used.

Cell-matrix interactions in Schistosomal portal fibrosis: a dynamic event

In recent years, one of the most significant progress in the understanding of liver diseases was the demonstration that liver fibrosis is a dynamic process resulting from a balance between synthesis and degradation of several matrix components, collagen in particular. Thus, fibrosis has been found to be a very early event during liver diseases, be it of toxic, viral or parasitic origin, and to be spontaneously reversible, either partially or totally. In liver fibrosis cell matrix interactions are dependent on the existence of the many factors (sometimes acting in combination) which produce the same events at the cellular and molecular levels. These events are: (i) the recruitment of fiber-producing cells, (ii) their proliferation, (iii) the secretion of matrix constituents of the extracellular matrix, and (iv) the remodeling and degradation of the newly formed matrix. All these events represent, at least in principle, a target for a therapeutic intervention aimed at influencing the experimentally induced hepatic fibrosis. In this context, hepatosplenic schistosomiasis is of particular interest, being an immune cell-mediated granulomatous disease and a model of liver fibrosis allowing extensive studies in human and animals as well as providing original in vitro models.

Estimation of the Spatial Weights Matrix under Structural Constraints

While estimates of models with spatial interaction are very sensitive to the choice of spatial weights, considerable uncertainty surrounds de nition of spatial weights in most studies with cross-section dependence. We show that, in the spatial error model the spatial weights matrix is only partially identi ed, and is fully identifi ed under the structural constraint of symmetry. For the spatial error model, we propose a new methodology for estimation of spatial weights under the assumption of symmetric spatial weights, with extensions to other important spatial models. The methodology is applied to regional housing markets in the UK, providing an estimated spatial weights matrix that generates several new hypotheses about the economic and socio-cultural drivers of spatial di¤usion in housing demand.

Adaptive Continuous time Markov Chain Approximation Model to General Jump-Diusions

We propose a non-equidistant Q rate matrix formula and an adaptive numerical algorithm for a continuous time Markov chain to approximate jump-diffusions with affine or non-affine functional specifications. Our approach also accommodates state-dependent jump intensity and jump distribution, a flexibility that is very hard to achieve with other numerical methods. The Kolmogorov-Smirnov test shows that the proposed Markov chain transition density converges to the one given by the likelihood expansion formula as in Ait-Sahalia (2008). We provide numerical examples for European stock option pricing in Black and Scholes (1973), Merton (1976) and Kou (2002).

Human GnRH deficiency: a unique disease model to unravel the ontogeny of GnRH neurons.

Evolutionary survival of a species is largely a function of its reproductive fitness. In mammals, a sparsely populated and widely dispersed network of hypothalamic neurons, the gonadotropin-releasing hormone (GnRH) neurons, serve as the pilot light of reproduction via coordinated secretion of GnRH. Since it first description, human GnRH deficiency has been recognized both clinically and genetically as a heterogeneous disease. A spectrum of different reproductive phenotypes comprised of congenital GnRH deficiency with anosmia (Kallmann syndrome), congenital GnRH deficiency with normal olfaction (normosmic idiopathic hypogonadotropic hypogonadism), and adult-onset hypogonadotropic hypogonadism has been described. In the last two decades, several genes and pathways which govern GnRH ontogeny have been discovered by studying humans with GnRH deficiency. More importantly, detailed study of these patients has highlighted the emerging theme of oligogenicity and genotypic synergism, and also expanded the phenotypic diversity with the documentation of reversal of GnRH deficiency later in adulthood in some patients. The underlying genetic defect has also helped understand the associated nonreproductive phenotypes seen in some of these patients. These insights now provide practicing clinicians with targeted genetic diagnostic strategies and also impact on clinical management.

First Report of a Newborn Rat Ventilation Model for Bronchopulmonary Dysplasia Permitting Evaluation of Long-Term Outcome

Background: Bronchopulmonary dysplasia (BPD) remains the leading cause of chronic pulmonary morbidity among preterm neonates. However, the exact pathophysiology is still unknown. Here we present the first results from a new model inteAbstracts, 25th International Workshop on Surfactant Replacement 400 Neonatology 2010;97:395-400 grating the most common risk factors for BPD (lung immaturity, inflammation, mechanical ventilation (MV), oxygen), which allows long-term outcome evaluation due to a non-traumatic intubation procedure. Objectives: To test the feasibility of a new rat model by investigating effects of MV, inflammation and oxygen applied to immature lungs after a ventilation-free interval. Methods: On day 4, 5, or 6 newborn rats were given an intraperitoneal injection of lipopolysaccharides to induce a systemic inflammation. 24 h later they were anesthetized, endotracheally intubated and ventilated for 8 h with 60% oxygen. After weaning of anesthesia and MV the newborn rats were extubated and returned to their mothers. Two days later they were killed and outcome measurements were performed (histology, quantitative RT-PCR) and compared to animals investigated directly after MV. Results: Directly after MV, histological signs of ventilator-induced lung injury were found. After 48 h, the first signs of early BPD were seen with delayed alveolar formation. Expression of inflammatory genes was only transiently increased. After 48 h genes involved in alveolarization, such as matrix metalloproteinase-9 and tropoelastin, showed a significant change of their expression. Conclusion: For the first time we can evaluate in a newborn rat model the effects of MV after a ventilation-free interval. This allows discrimination between immediate response genes and delayed changes of expression of more structural genes involved in alveolarization.

Calomys callosus: an alternative model to study fibrosis in Schistosomiasis mansoni: the pathology of the acute phase

Twenty Calomys callosus, Rengger, 1830 (Rodentia-Cricetidae) were studied in the early stage of the acute schistosomal mansoni infection (42nd day). The same number of Swiss Webster mice were used as a comparative standard. Liver and intestinal sections, fixed in formalin-Millonig and embedded in paraffin, were stained with hematoxilin and eosin, PAS-Alcian Blue, pH = 1.0 and 2.5, Lennert's Giemsa, Picrosirius plus polarization microscopy, Periodic acid methanamine silver, Gomori's silver reticulin and resorcin-fuchsin. Immunohistological study (indirect immunofluorescence and peroxidase labeled extravidin-biotin methods) was done with antibodies specific to pro-collagen III, fibronectin, elastin, condroitin-sulfate, tenascin, alpha smooth muscle actin, vimentin and desmin. The hepatic granulomas were small, reaching only 27 of the volume of the hepatic Swiss Webster granuloma. They were composed mainly by large immature macrophages, often filled by schistosomal pigment, characterizing an exsudative-macrophage granuloma type. The granulomas were situated in the parenchyma and in the portal space. They were often intravascular, poor of extracellular matrix components, except fibronectin and presented, sometimes alpha smooth muscle actin and vimentin positive cells. The C. callosus intestinal granulomas were similar to Swiss Webster, showing predominance of macrophages. Therefore, the C. callosus acquire very well the Schistosoma mansoni infection, without developing strong hepatic acute granulomatous reaction, suggesting lack of histopathological signs of hypersensitivity.