TGF-β dependent regulation of oxygen radicals during transdifferentiation of activated hepatic stellate cells to myofibroblastoid cells

Background: The activation of hepatic stellate cells (HSCs) plays a pivotal role during liver injury because the resulting myofibroblasts (MFBs) are mainly responsible for connective tissue re-assembly. MFBs represent therefore cellular targets for anti-fibrotic therapy. In this study, we employed activated HSCs, termed M1-4HSCs, whose transdifferentiation to myofibroblastoid cells (named M-HTs) depends on transforming growth factor (TGF)-β. We analyzed the oxidative stress induced by TGF-β and examined cellular defense mechanisms upon transdifferentiation of HSCs to M-HTs. Results: We found reactive oxygen species (ROS) significantly upregulated in M1-4HSCs within 72 hours of TGF-β administration. In contrast, M-HTs harbored lower intracellular ROS content than M1-4HSCs, despite of elevated NADPH oxidase activity. These observations indicated an upregulation of cellular defense mechanisms in order to protect cells from harmful consequences caused by oxidative stress. In line with this hypothesis, superoxide dismutase activation provided the resistance to augmented radical production in M-HTs, and glutathione rather than catalase was responsible for intracellular hydrogen peroxide removal. Finally, the TGF-β/NADPH oxidase mediated ROS production correlated with the upregulation of AP-1 as well as platelet-derived growth factor receptor subunits, which points to important contributions in establishing antioxidant defense. Conclusion: The data provide evidence that TGF-β induces NADPH oxidase activity which causes radical production upon the transdifferentiation of activated HSCs to M-HTs. Myofibroblastoid cells are equipped with high levels of superoxide dismutase activity as well as glutathione to counterbalance NADPH oxidase dependent oxidative stress and to avoid cellular damage.

Single acquisition electrical property mapping based on relative coil sensitivities: A proof-of-concept demonstration.

PURPOSE: All methods presented to date to map both conductivity and permittivity rely on multiple acquisitions to compute quantitatively the magnitude of radiofrequency transmit fields, B1+. In this work, we propose a method to compute both conductivity and permittivity based solely on relative receive coil sensitivities ( B1-) that can be obtained in one single measurement without the need to neither explicitly perform transmit/receive phase separation nor make assumptions regarding those phases. THEORY AND METHODS: To demonstrate the validity and the noise sensitivity of our method we used electromagnetic finite differences simulations of a 16-channel transceiver array. To experimentally validate our methodology at 7 Tesla, multi compartment phantom data was acquired using a standard 32-channel receive coil system and two-dimensional (2D) and 3D gradient echo acquisition. The reconstructed electric properties were correlated to those measured using dielectric probes. RESULTS: The method was demonstrated both in simulations and in phantom data with correlations to both the modeled and bench measurements being close to identity. The noise properties were modeled and understood. CONCLUSION: The proposed methodology allows to quantitatively determine the electrical properties of a sample using any MR contrast, with the only constraint being the need to have 4 or more receive coils and high SNR. Magn Reson Med, 2014. © 2014 Wiley Periodicals, Inc.

Correcting surface coil excitation inhomogeneities in single-shot SPEN MRI.

Given their high sensitivity and ability to limit the field of view (FOV), surface coils are often used in magnetic resonance spectroscopy (MRS) and imaging (MRI). A major downside of surface coils is their inherent radiofrequency (RF) B1 heterogeneity across the FOV, decreasing with increasing distance from the coil and giving rise to image distortions due to non-uniform spatial responses. A robust way to compensate for B1 inhomogeneities is to employ adiabatic inversion pulses, yet these are not well adapted to all imaging sequences - including to single-shot approaches like echo planar imaging (EPI). Hybrid spatiotemporal encoding (SPEN) sequences relying on frequency-swept pulses provide another ultrafast MRI alternative, that could help solve this problem thanks to their built-in heterogeneous spatial manipulations. This study explores how this intrinsic SPEN-based spatial discrimination, could be used to compensate for the B1 inhomogeneities inherent to surface coils. Experiments carried out in both phantoms and in vivo rat brains demonstrate that, by suitably modulating the amplitude of a SPEN chirp pulse that progressively excites the spins in a direction normal to the coil, it is possible to compensate for the RF transmit inhomogeneities and thus improve sensitivity and image fidelity.

Endovascular treatment of pulmonary arteriovenous malformations in hereditary haemorrhagic telangiectasia.

PRINCIPLES: To assess the efficiency and complication rates of vaso-occlusion of pulmonary arteriovenous malformations (PAVMs) in Rendu-Osler-Weber disease (hereditary haemorrhagic telangectasia; HHT). METHODS: Seventy-two patients were investigated in our institution for HHT between March 2000 and November 2011. Sixteen presented PAVMs (22.2%), and 11 (68.8%) were treated with vaso-occlusion for a total of 18 procedures. Procedures included coils, plugs and combined approaches. Immediate success and recurrence rate, complication were recorded, as well as persistent and new PAVMs during clinical and computed tomography (CT) follow-up. RESULTS: Eighteen procedures were performed and a total of 37 PAVMs were treated, 19 with coils, 16 with plugs and 2 with combined treatment. Mean CT follow-up time was 41 months (1‒164). No major complication was observed. One distal translocation was treated during the same intervention. Two PAVMs persisted after treatment (5.7%), both treated by means of plug embolisation. One new PAVM was observed during follow-up CT. PAVMs with an afferent artery of less than 3mm or asymptomatic PAVMs were not treated. CONCLUSION: Recent studies have demonstrated that vaso-occlusion has become the gold standard treatment for PAVM. This study is in accordance with previous results and shows a minimal complication rate and little recurrence, whether by coils, plugs, or combined treatments.

High-Resolution Magnetic Resonance Imaging Can Reliably Detect Orbital Tumor Recurrence after Enucleation in Children with Retinoblastoma.

PURPOSE: Orbital tumor recurrence is a rare but serious complication in children with retinoblastoma, leading to a high risk of metastasis and death. Therefore, we assume that these recurrences have to be detected and treated as early as possible. Preliminary studies used magnetic resonance imaging (MRI) to evaluate postsurgical findings in the orbit. In this study, we assessed the diagnostic accuracy of high-resolution MRI to detect orbital tumor recurrence in children with retinoblastoma in a large study cohort. DESIGN: Consecutive retrospective study (2007-2013) assessing MRI findings after enucleation. PARTICIPANTS: A total of 103 MRI examinations of 55 orbits (50 children, 27 male/23 female, mean age 16.3±12.4 months) with a median time of 8 months (range, 0-93) after enucleation for retinoblastoma. METHODS: High-resolution MRI using orbital surface coils was performed on 1.5 Tesla MRI systems to assess abnormal orbital findings. MAIN OUTCOME MEASURES: Five European experts in retinoblastoma imaging evaluated the MRI examinations regarding the presence of abnormal orbital gadolinium enhancement and judged them as "definitive tumor," "suspicious of tumor," "postsurgical condition/scar formation," or "without pathologic findings." The findings were correlated to histopathology (if available), MRI, and clinical follow-up. RESULTS: Abnormal orbital enhancement was a common finding after enucleation (100% in the first 3 months after enucleation, 64.3% >3 years after enucleation). All histopathologically confirmed tumor recurrences (3 of 55 orbits, 5.5%) were correctly judged as "definitive tumor" in MRI. Two orbits from 2 children rated as "suspicious of tumor" received intravenous chemotherapy without histopathologic confirmation; further follow-up (67 and 47 months) revealed no sign of tumor recurrence. In 90.2%, no tumor was suspected on MRI, which was clinically confirmed during follow-up (median follow-up after enucleation, 45 months; range, 8-126). CONCLUSIONS: High-resolution MRI with orbital surface coils may reliably distinguish between common postsurgical contrast enhancement and orbital tumor recurrence, and therefore may be a useful tool to evaluate orbital tumor recurrence after enucleation in children with retinoblastoma. We recommend high-resolution MRI as a potential screening tool for the orbit in children with retinoblastoma to exclude tumor recurrence, especially in high-risk patients within the critical first 2 years after enucleation.

Virranahto ja jänniterasitukset taajuusmuuttajakäytöissä

Nykyiset IGB-transistorit voivat tuottaa jopa 30 ns nousureunoja, jolloin jänniterasitus jakautuu hyvin epätasaisesti käämien ja kierrosten kesken. Tämä ja eristeissä esiintyvät ilmataskut asettavat käämikierrosten välisten eristeiden jännitekestävyyden koetukselle, ja aiheuttavat riskin osittaispurkaukselle. Urassa hajavuon epätasaisen jakauman aiheuttama virranahto on yleensä ollut esillä suurissa koneissa, jotka toimivat 50 Hz:llä. Nykyään taajuusmuuttajien soveltaminen on mahdollistanut nimellistaajuudeltaan satojen hertsien suurnopeuskoneiden käämitysten valmistamisen pyörölangasta, jolloin virranahto tulee merkittäväksi, koska kierroksia on muutama urassa. Työssä mitattiin jännitteen jakautumista 3.5 MW:n tuulivoimageneraattorin segmentissä käyttäen käämitysten syöttöön erilaisia jännitteen nousuaikoja, ja pyrittiin arvioimaan onko kyseinen kone vaarassa kokea osittaispurkauksia elinikänsä aikana. Työssä on myös käsitelty virranahtoa käyttäen finiittielementtimenetelmää simuloimaan ja vertaamaan erilaisia tilanteita käämitystavoissa pyörölankakoneessa. Tämä on myös osittain sovellettavissa muotokuparilla käämittyihin koneisiin. Käämivyyhdin vyyhdenpään alueella tehdyn johdinnipun 180 asteen kierron vaikutusta simuloitiin ja sen aiheuttamaa parannusta varmennettiin käytännön mittauksilla.

Efeito da temperatura no desempenho catalítico de óxidos de ferro contendo cobre e alumínio

Aluminum and copper doped hematite was evaluated in the high temperature shift (HTS) reaction at several temperatures in order to find catalysts that can work in different operational conditions. It was found that the catalysts work in kinetic regime in the range of 300-400 ºC. Both copper and aluminum increases the activity and selectivity. Aluminum acts as textural promoter whereas copper acts as structural one. The most promising catalyst is that with both copper and aluminum which showed higher activity and selectivity than a commercial sample. This catalyst has the advantages of being non toxic and can work at low temperatures.

Integração das técnicas de triagem virtual e triagem biológica automatizada em alta escala: oportunidades e desafios em P&D de fármacos

High-throughput screening (HTS) and virtual screening (VS) are useful methods employed in drug discovery, allowing the identification of promising hits for lead optimization. The efficiency of these approaches depends on a number of factors, such as the organization of high quality databases of compounds and the parameterization of essential components of the screen process. This brief review presents the basic principles of the HTS and VS methods, as well as a perspective of the utility and integration of these drug design approaches, highlighting current opportunities and future challenges in medicinal chemistry.

Hidróxidos duplos lamelares à base de escória de alto forno

Hydrotalcite like compounds (HT) were prepared by co-precipitation (Mg/Al = 3.5), using an acid solution with blast furnace slag and MgCl2.6H2O and aqueous solutions of NaOH. The following synthesis variables were investigated: temperature (30 and 45 ºC) and pH (9 and 12). Depending of the temperature two systems were observed: Mg-Al-CO3 (T = 30 ºC); Mg-Al-Cl-CO3 (T = 45 ºC). An increase in the pH of synthesis and Mg2+ concentration produced HTs well-crystallized and with greater values of all cell parameters. The study showed the potentiality of BFS in the synthesis of well-crystallized LDHs without the presence of other crystalline phases.

Magnetic sputtering of complex oxides

HTSC materials are relevant in modern microelectronics, because of their transformation from the normal state to the superconducting. That is why the idea of producing HTSC in industrial amounts is actual nowadays. To decrease cost of their production it is important to use magnetron sputtering systems which give the best results for essential parameters. Modeling is the simplest and the fastest way to determine optimum sputtering condition. This thesis concentrates on determination the phases of the whole sputtering process and to find out basic factors of each phase using the modeling. It was find out, that the main factors which influence on the mode of occurrence of the initial stages are the current density of the magnetron discharge and the pressure of sputtering gas. With the modeling also velocity dependences were obtained for YBCO and SmFeAsO. These were compared and difference between them was examined. To support represented model comparison was made with experimental results. This showed that the model gives good results, very similar to the experimental ones. The results of this work were published in annual conference of the finnish physical society.

Luminescence-based assay methods in drug discovery

The drug discovery process is facing new challenges in the evaluation process of the lead compounds as the number of new compounds synthesized is increasing. The potentiality of test compounds is most frequently assayed through the binding of the test compound to the target molecule or receptor, or measuring functional secondary effects caused by the test compound in the target model cells, tissues or organism. Modern homogeneous high-throughput-screening (HTS) assays for purified estrogen receptors (ER) utilize various luminescence based detection methods. Fluorescence polarization (FP) is a standard method for ER ligand binding assay. It was used to demonstrate the performance of two-photon excitation of fluorescence (TPFE) vs. the conventional one-photon excitation method. As result, the TPFE method showed improved dynamics and was found to be comparable with the conventional method. It also held potential for efficient miniaturization. Other luminescence based ER assays utilize energy transfer from a long-lifetime luminescent label e.g. lanthanide chelates (Eu, Tb) to a prompt luminescent label, the signal being read in a time-resolved mode. As an alternative to this method, a new single-label (Eu) time-resolved detection method was developed, based on the quenching of the label by a soluble quencher molecule when displaced from the receptor to the solution phase by an unlabeled competing ligand. The new method was paralleled with the standard FP method. It was shown to yield comparable results with the FP method and found to hold a significantly higher signal-tobackground ratio than FP. Cell-based functional assays for determining the extent of cell surface adhesion molecule (CAM) expression combined with microscopy analysis of the target molecules would provide improved information content, compared to an expression level assay alone. In this work, immune response was simulated by exposing endothelial cells to cytokine stimulation and the resulting increase in the level of adhesion molecule expression was analyzed on fixed cells by means of immunocytochemistry utilizing specific long-lifetime luminophore labeled antibodies against chosen adhesion molecules. Results showed that the method was capable of use in amulti-parametric assay for protein expression levels of several CAMs simultaneously, combined with analysis of the cellular localization of the chosen adhesion molecules through time-resolved luminescence microscopy inspection.

The spindle assembly checkpoint as a drug target - Novel small-molecule inhibitors of Aurora kinases

Cell division (mitosis) is a fundamental process in the life cycle of a cell. Equal distribution of chromosomes between the daughter cells is essential for the viability and well-being of an organism: loss of fidelity of cell division is a contributing factor in human cancer and also gives rise to miscarriages and genetic birth defects. For maintaining the proper chromosome number, a cell must carefully monitor cell division in order to detect and correct mistakes before they are translated into chromosomal imbalance. For this purpose an evolutionarily conserved mechanism termed the spindle assembly checkpoint (SAC) has evolved. The SAC comprises a complex network of proteins that relay and amplify mitosis-regulating signals created by assemblages called kinetochores (KTs). Importantly, minor defects in SAC signaling can cause loss or gain of individual chromosomes (aneuploidy) which promotes tumorigenesis while complete failure of SAC results in cell death. The latter event has raised interest in discovery of low molecular weight (LMW) compounds targeting the SAC that could be developed into new anti-cancer therapeutics. In this study, we performed a cell-based, phenotypic high-throughput screen (HTS) to identify novel LMW compounds that inhibit SAC function and result in loss of cancer cell viability. Altogether, we screened 65 000 compounds and identified eight that forced the cells prematurely out of mitosis. The flavonoids fisetin and eupatorin, as well as the synthetic compounds termed SACi2 and SACi4, were characterized in more detail utilizing versatile cell-based and biochemical assays. To identify the molecular targets of these SAC-suppressing compounds, we investigated the conditions in which SAC activity became abrogated. Eupatorin, SACi2 and SACi4 preferentially abolished the tensionsensitive arm of the SAC, whereas fisetin lowered also the SAC activity evoked by lack of attachments between microtubules (MTs) and KTs. Consistent with the abrogation of SAC in response to low tension, our data indicate that all four compounds inhibited the activity of Aurora B kinase. This essential mitotic protein is required for correction of erratic MT-KT attachments, normal SAC signaling and execution of cytokinesis. Furthermore, eupatorin, SACi2 and SACi4 also inhibited Aurora A kinase that controls the centrosome maturation and separation and formation of the mitotic spindle apparatus. In line with the established profound mitotic roles of Aurora kinases, these small compounds perturbed SAC function, caused spindle abnormalities, such as multi- and monopolarity and fragmentation of centrosomes, and resulted in polyploidy due to defects in cytokinesis. Moreover, the compounds dramatically reduced viability of cancer cells. Taken together, using a cell-based HTS we were able to identify new LMW compounds targeting the SAC. We demonstrated for the first time a novel function for flavonoids as cellular inhibitors of Aurora kinases. Collectively, our data support the concept that loss of mitotic fidelity due to a non-functional SAC can reduce the viability of cancer cells, a phenomenon that may possess therapeutic value and fuel development of new anti-cancer drugs.

Homogeneous assay technologies in drug screening: Quenching Resonance Energy Transfer (QRET) technique

Information gained from the human genome project and improvements in compound synthesizing have increased the number of both therapeutic targets and potential lead compounds. This has evolved a need for better screening techniques to have a capacity to screen number of compound libraries against increasing amount of targets. Radioactivity based assays have been traditionally used in drug screening but the fluorescence based assays have become more popular in high throughput screening (HTS) as they avoid safety and waste problems confronted with radioactivity. In comparison to conventional fluorescence more sensitive detection is obtained with time-resolved luminescence which has increased the popularity of time-resolved fluorescence resonance energy transfer (TR-FRET) based assays. To simplify the current TR-FRET based assay concept the luminometric homogeneous single-label utilizing assay technique, Quenching Resonance Energy Transfer (QRET), was developed. The technique utilizes soluble quencher to quench non-specifically the signal of unbound fraction of lanthanide labeled ligand. One labeling procedure and fewer manipulation steps in the assay concept are saving resources. The QRET technique is suitable for both biochemical and cell-based assays as indicated in four studies:1) ligand screening study of β2 -adrenergic receptor (cell-based), 2) activation study of Gs-/Gi-protein coupled receptors by measuring intracellular concentration of cyclic adenosine monophosphate (cell-based), 3) activation study of G-protein coupled receptors by observing the binding of guanosine-5’-triphosphate (cell membranes), and 4) activation study of small GTP binding protein Ras (biochemical). Signal-to-background ratios were between 2.4 to 10 and coefficient of variation varied from 0.5 to 17% indicating their suitability to HTS use.

Design of a 1 kW 150 rpm permanent-magnet synchronous generator for stand-alone wind-power applications

A program for calculating low-speed low-power synchronous machine is presented. A permanent-magnet synchronous generator for 1 kW 150 rpm is designed. Optimization of magnet’s and coil’s dimensions was made.

Wind Turbine Direct-Drive Permanent-Magnet Generator with Direct Liquid Cooling for Mass Reduction

Today’s electrical machine technology allows increasing the wind turbine output power by an order of magnitude from the technology that existed only ten years ago. However, it is sometimes argued that high-power direct-drive wind turbine generators will prove to be of limited practical importance because of their relatively large size and weight. The limited space for the generator in a wind turbine application together with the growing use of wind energy pose a challenge for the design engineers who are trying to increase torque without making the generator larger. When it comes to high torque density, the limiting factor in every electrical machine is heat, and if the electrical machine parts exceed their maximum allowable continuous operating temperature, even for a short time, they can suffer permanent damage. Therefore, highly efficient thermal design or cooling methods is needed. One of the promising solutions to enhance heat transfer performances of high-power, low-speed electrical machines is the direct cooling of the windings. This doctoral dissertation proposes a rotor-surface-magnet synchronous generator with a fractional slot nonoverlapping stator winding made of hollow conductors, through which liquid coolant can be passed directly during the application of current in order to increase the convective heat transfer capabilities and reduce the generator mass. This doctoral dissertation focuses on the electromagnetic design of a liquid-cooled direct-drive permanent-magnet synchronous generator (LC DD-PMSG) for a directdrive wind turbine application. The analytical calculation of the magnetic field distribution is carried out with the ambition of fast and accurate predicting of the main dimensions of the machine and especially the thickness of the permanent magnets; the generator electromagnetic parameters as well as the design optimization. The focus is on the generator design with a fractional slot non-overlapping winding placed into open stator slots. This is an a priori selection to guarantee easy manufacturing of the LC winding. A thermal analysis of the LC DD-PMSG based on a lumped parameter thermal model takes place with the ambition of evaluating the generator thermal performance. The thermal model was adapted to take into account the uneven copper loss distribution resulting from the skin effect as well as the effect of temperature on the copper winding resistance and the thermophysical properties of the coolant. The developed lumpedparameter thermal model and the analytical calculation of the magnetic field distribution can both be integrated with the presented algorithm to optimize an LC DD-PMSG design. Based on an instrumented small prototype with liquid-cooled tooth-coils, the following targets have been achieved: experimental determination of the performance of the direct liquid cooling of the stator winding and validating the temperatures predicted by an analytical thermal model; proving the feasibility of manufacturing the liquid-cooled tooth-coil winding; moreover, demonstration of the objectives of the project to potential customers.