Mutation of conserved negatively charged residues in the S2 and S3 transmembrane segments of a mammalian K+ channel selectively modulates channel gating.

Voltage-gated channel proteins sense a change in the transmembrane electric field and respond with a conformational change that allows ions to diffuse across the pore-forming structure. Site-specific mutagenesis combined with electrophysiological analysis of expressed mutants in amphibian oocytes has previously established the S4 transmembrane segment as an element of the voltage sensor. Here, we show that mutations of conserved negatively charged residues in S2 and S3 of a brain K+ channel, thought of as countercharges for the positively charged residues in S4, selectively modulate channel gating without modifying the permeation properties. Mutations of Glu235 in S2 that neutralize or reverse charge increase the probability of channel opening and the apparent gating valence. In contrast, replacements of Glu272 by Arg or Thr268 by Asp in S3 decrease the open probability and the apparent gating valence. Residue Glu225 in S2 tolerated replacement only by acidic residues, whereas Asp258 in S3 was intolerant to any attempted change. These results imply that S2 and S3 are unlikely to be involved in channel lining, yet, together with S4, may be additional components of the voltage-sensing structure.

Innovative analytical strategies for the development of sensor devices and mass spectrometry methods

Il lavoro presentato in questa tesi di Dottorato è incentrato sullo sviluppo di strategie analitiche innovative basate sulla sensoristica e su tecniche di spettrometria di massa in ambito biologico e della sicurezza alimentare. Il primo capitolo tratta lo studio di aspetti metodologici ed applicativi di procedure sensoristiche per l’identificazione e la determinazione di biomarkers associati alla malattia celiaca. In tale ambito, sono stati sviluppati due immunosensori, uno a trasduzione piezoelettrica e uno a trasduzione amperometrica, per la rivelazione di anticorpi anti-transglutaminasi tissutale associati a questa malattia. L’innovazione di questi dispositivi riguarda l’immobilizzazione dell’enzima tTG nella conformazione aperta (Open-tTG), che è stato dimostrato essere quella principalmente coinvolta nella patogenesi. Sulla base dei risultati ottenuti, entrambi i sistemi sviluppati si sono dimostrati una valida alternativa ai test di screening attualmente in uso per la diagnosi della celiachia. Rimanendo sempre nel contesto della malattia celiaca, ulteriore ricerca oggetto di questa tesi di Dottorato, ha riguardato lo sviluppo di metodi affidabili per il controllo di prodotti “gluten-free”. Il secondo capitolo tratta lo sviluppo di un metodo di spettrometria di massa e di un immunosensore competitivo per la rivelazione di prolammine in alimenti “gluten-free”. E’ stato sviluppato un metodo LC-ESI-MS/MS basato su un’analisi target con modalità di acquisizione del segnale selected reaction monitoring per l’identificazione di glutine in diversi cereali potenzialmente tossici per i celiaci. Inoltre ci si è focalizzati su un immunosensore competitivo per la rivelazione di gliadina, come metodo di screening rapido di farine. Entrambi i sistemi sono stati ottimizzati impiegando miscele di farina di riso addizionata di gliadina, avenine, ordeine e secaline nel caso del sistema LC-MS/MS e con sola gliadina nel caso del sensore. Infine i sistemi analitici sono stati validati analizzando sia materie prime (farine) che alimenti (biscotti, pasta, pane, etc.). L’approccio sviluppato in spettrometria di massa apre la strada alla possibilità di sviluppare un test di screening multiplo per la valutazione della sicurezza di prodotti dichiarati “gluten-free”, mentre ulteriori studi dovranno essere svolti per ricercare condizioni di estrazione compatibili con l’immunosaggio competitivo, per ora applicabile solo all’analisi di farine estratte con etanolo. Terzo capitolo di questa tesi riguarda lo sviluppo di nuovi metodi per la rivelazione di HPV, Chlamydia e Gonorrhoeae in fluidi biologici. Si è scelto un substrato costituito da strips di carta in quanto possono costituire una valida piattaforma di rivelazione, offrendo vantaggi grazie al basso costo, alla possibilità di generare dispositivi portatili e di poter visualizzare il risultato visivamente senza la necessità di strumentazioni. La metodologia sviluppata è molto semplice, non prevede l’uso di strumentazione complessa e si basa sull’uso della isothermal rolling-circle amplification per l’amplificazione del target. Inoltre, di fondamentale importanza, è l’utilizzo di nanoparticelle colorate che, essendo state funzionalizzate con una sequenza di DNA complementare al target amplificato derivante dalla RCA, ne permettono la rivelazione a occhio nudo mediante l’uso di filtri di carta. Queste strips sono state testate su campioni reali permettendo una discriminazione tra campioni positivi e negativi in tempi rapidi (10-15 minuti), aprendo una nuova via verso nuovi test altamente competitivi con quelli attualmente sul mercato.

Study of interaction between Si(O,C) nanowires and the biological system

Nanomedicine is a new branch of medicine, based on the potentiality and intrinsic properties of nanomaterials. Indeed, the nanomaterials ( i.e. the materials with nano and under micron size) can be suitable to different applications in biomedicine. The nanostructures can be used by taking advantage of their properties (for example superparamagnetic nanoparticles) or functionalized to deliver the drug in a specific target, thanks the ability to cross biological barriers. The size and the shape of 1D-nanostructures (nanotubes and nanowires) have an important role on the cell fate: their morphology plays a key role on the interaction between nanostructure and the biological system. For this reason the 1D nanostructure are interesting for their ability to mime the biological system. An implantable material or device must therefore integrate with the surrounding extracellular matrix (ECM), a complex network of proteins with structural and signaling properties. Innovative techniques allow the generation of complex surface patterns that can resemble the structure of the ECM, such as 1D nanostructures. NWs based on cubic silicon carbide (3C-SiC), either bare (3C-SiC NWs) or surrounded by an amorphous shell (3C-SiC/SiO2 core/shell NWs), and silicon oxycarbide nanowires (SiOxCy NWs) can meet the chemical, mechanical and electrical requirements for tissue engineering and have a strong potential to pave the way for the development of a novel generation of implantable nano-devices. Silicon oxycarbide shows promising physical and chemical properties as elastic modulus, bending strength and hardness, chemical durability superior to conventional silicate glasses in aggressive environments and high temperature stability up to 1300 °C. Moreover, it can easily be engineered through functionalization and decoration with macro-molecules and nanoparticles. Silicon carbide has been extensively studied for applications in harsh conditions, as chemical environment, high electric field and high and low temperature, owing to its high hardness, high thermal conductivity, chemical inertness and high electron mobility. Also, its cubic polytype (3C) is highly biocompatible and hemocompatible, and some prototypes of biomedical applications and biomedical devices have been already realized starting from 3C-SiC thin films. Cubic SiC-based NWs can be used as a biomimetic biomaterial, providing a robust and novel biocompatible biological interface . We cultured in vitro A549 human lung adenocarcinoma epithelial cells and L929 murine fibroblast cells over core/shell SiC/SiO2, SiOxCy and bare 3C-SiC nanowire platforms, and analysed the cytotoxicity, by indirect and direct contact tests, the cell adhesion, and the cell proliferation. These studies showed that all the nanowires are biocompatible according to ISO 10993 standards. We evaluated the blood compatibility through the interaction of the nanowires with platelet rich plasma. The adhesion and activation of platelets on the nanowire bundles, assessed via SEM imaging and soluble P-selectin quantification, indicated that a higher platelet activation is induced by the core/shell structures compared to the bare ones. Further, platelet activation is higher with 3C-SiC/SiO2 NWs and SiOxCyNWs, which therefore appear suitable in view of possible tissue regeneration. On the contrary, bare 3C-SiC NWs show a lower platelet activation and are therefore promising in view of implantable bioelectronics devices, as cardiovascular implantable devices. The NWs properties are suitable to allow the design of a novel subretinal Micro Device (MD). This devices is based on Si NWs and PEDOT:PSS, though the well know principle of the hybrid ordered bulk heterojunction (OBHJ). The aim is to develop a device based on a well-established photovoltaic technology and to adapt this know-how to the prosthetic field. The hybrid OBHJ allows to form a radial p–n junction on a nanowire/organic structure. In addition, the nanowires increase the light absorption by means of light scattering effects: a nanowires based p-n junction increases the light absorption up to the 80%, as previously demonstrated, overcoming the Shockley-Queisser limit of 30 % of a bulk p-n junction. Another interesting employment of these NWs is to design of a SiC based epicardial-interacting patch based on teflon that include SiC nanowires. . Such contact patch can bridge the electric conduction across the cardiac infarct as nanowires can ‘sense’ the direction of the wavefront propagation on the survival cardiac tissue and transmit it to the downstream surivived regions without discontinuity. The SiC NWs are tested in terms of toxicology, biocompatibility and conductance among cardiomyocytes and myofibroblasts.

A Meta-Analytic Review of Mindfulness-Based Interventions on Symptom Reduction: A Critique and Guide for Future Investigation.

Over the past decade, mindfulness practices have been used with increasing frequency as therapeutic components within cognitive behavioral treatment regimens. As is standard practice, prescriptive uses of mindfulness intervention are incorporated to improve end-state functioning by ameliorating problematic symptoms and conditions. Common change-targets include the control of cognitive and emotional content for purposes of enhancing psychological self-regulation and physical well-being. The term mindfulness applies to a heterogeneous range of practices, methods, and techniques. While there is no singular agreed upon definition for mindfulness, as a process concept, the term connotes an immediate, non-thetic access to events, wherein each occasioning event is experienced in toto within the broader contextual event-field, and distinct from intervening conceptual themes being noticed. Training in mindfulness practices may be conducted using individual, group, or small class formats. The current paper provides a meta-analytic review of 44 treatment outcome studies (extracted 1982 through 2006), which examines the clinical utility of mindfulness as the primary therapeutic approach. Results indicated that average effect sizes for mindfulness based interventions fell within the medium range for construct category variables examined (d = .56). These findings suggest that mindfulness training is a cost-effective treatment for a wide array of contemporary psychological problems and diagnoses, in addition to fostering positive psychology attributes such as quality and satisfaction with life. A critique of the research and recommendations for future research, including a need to examine the role of mindfulness as a tool for cultivating increased psychological acceptance and life satisfaction, is presented.

An un-mounted horse care group for children and youth with an Autism Spectrum Disorder: Recommendations for program development and treatment intervention

Equine Assisted Activities and Therapies (EAAT) including Therapeutic Horseback Riding (THR) and un-mounted equine assisted activities are interventions aimed at improving the daily functioning and success of individuals with disabilities, including those with an autism spectrum disorder (ASD). While THR is frequently utilized as a treatment intervention for children with ASD, there are many limitations (individual's weight, horse health, weather, physical limitations, health conditions, etc.) that prevent this population from participating in mounted programs. Un-mounted equine assisted activities are often utilized as an alternative, but they are not informed by empirical research or a standardized treatment model. This paper provides a comprehensive review of the literature for EAAT including un-mounted programs, examination of organizational guidelines as they apply to un-mounted programs, and consultation with program directors regarding current practices in the field, and finally it establishes recommendations for the development of a standard curriculum that would strengthen un-mounted horse care group programs serving children with ASD.

Educating a New Cadre of Experts Specializing in Digital Collections and Digital Curation: Experiential Learning in Digital Library Curriculum

Integration of experiential learning into the library and information science (LIS) courses has been a theme in LIS education, but the topic deserves renewed attention with an increasing demand for professionals in the digital library field and in light of the new initiative announced by the Library of Congress (LC) and the Institution of Museum and Library Services (IMLS) for national residency program in digital curation. The balance between theory and practice in digital library curricula, the challenges of incorporating practical projects into LIS coursework, and the current practice of teaching with hands on activities represent the primary areas of this panel discussion.

Computational approaches to subjectivity and sentiment analysis: Present and envisaged methods and applications

Recent years have witnessed a surge of interest in computational methods for affect, ranging from opinion mining, to subjectivity detection, to sentiment and emotion analysis. This article presents a brief overview of the latest trends in the field and describes the manner in which the articles contained in the special issue contribute to the advancement of the area. Finally, we comment on the current challenges and envisaged developments of the subjectivity and sentiment analysis fields, as well as their application to other Natural Language Processing tasks and related domains.

Magnetars: Properties, Origin and Evolution

Magnetars are neutron stars in which a strong magnetic field is the main energy source. About two dozens of magnetars, plus several candidates, are currently known in our Galaxy and in the Magellanic Clouds. They appear as highly variable X-ray sources and, in some cases, also as radio and/or optical pulsars. Their spin periods (2–12 s) and spin-down rates (∼10−13–10−10 s s−1) indicate external dipole fields of ∼1013−15 G, and there is evidence that even stronger magnetic fields are present inside the star and in non-dipolar magnetospheric components. Here we review the observed properties of the persistent emission from magnetars, discuss the main models proposed to explain the origin of their magnetic field and present recent developments in the study of their evolution and connection with other classes of neutron stars.

Are pulsars born with a hidden magnetic field?

The observation of several neutron stars in the centre of supernova remnants and with significantly lower values of the dipolar magnetic field than the average radio-pulsar population has motivated a lively debate about their formation and origin, with controversial interpretations. A possible explanation requires the slow rotation of the protoneutron star at birth, which is unable to amplify its magnetic field to typical pulsar levels. An alternative possibility, the hidden magnetic field scenario, considers the accretion of the fallback of the supernova debris on to the neutron star as responsible for the submergence (or screening) of the field and its apparently low value. In this paper, we study under which conditions the magnetic field of a neutron star can be buried into the crust due to an accreting, conducting fluid. For this purpose, we consider a spherically symmetric calculation in general relativity to estimate the balance between the incoming accretion flow and the magnetosphere. Our study analyses several models with different specific entropy, composition, and neutron star masses. The main conclusion of our work is that typical magnetic fields of a few times 1012 G can be buried by accreting only 10−3–10−2 M⊙, a relatively modest amount of mass. In view of this result, the central compact object scenario should not be considered unusual, and we predict that anomalously weak magnetic fields should be common in very young (< few kyr) neutron stars.

The differential evolution of hot- and cold-dust in galaxies in the last 7.5 GYR

Tese de mestrado em Física, apresentada à Universidade de Lisboa, através da Faculdade de Ciências, 2016

The resource wealth burden - oil and gas sectors in the former USSR. OSW Study 12/2003

The former USSR area plays a great role in the international oil and gas market. Russia is a real gas giant, with the richest deposits of this material in the world. Russia is also the main exporter of natural gas to many European countries. Keeping a strong position in this market remains a priority for the Russian Federation's economic policy. Europe is a very attractive region because its demand for gas is expected to grow steadily, while its own gas production keeps decreasing. In the long term, the Far East will be an important market for Russian exports, too. According to estimates, demand there will grow even faster than in Europe. Caspian gas producers, for the time being, can not really compete with Russia in this field, and this status quo will most probably be preserved in the nearest future.

Controlled ‘Europeanisation’? The KMW–Nexter merger and the Germany’s new strategy for the arms industry. OSW COMMENTARY NUMBER 180/26.08.2015

On 29 July a deal was signed in Paris concerning a merger between Krauss-Maffei Wegmann (KMW), Germany’s largest manufacturer of tanks, infantry fighting vehicles and artillery systems, and its French counterpart Nexter. The new holding formed as a result of the merger will be Europe’s largest producer of arms systems for land forces, comparable to the Airbus Group in the aerospace industry. While work on finalising the merger was underway, the German government was developing a new strategy for Germany’s arms industry, which was published on 9 June 2015. The strategy’s provisions show that German politicians, despite holding negative opinions on previous mergers between German arms companies and foreign businesses, have concluded that consolidation at the European level is nonetheless the only way to go. However, the strategy also states that the German government should exercise more influence than previously on the terms and conditions of any such consolidation. To this end, it identified key national technologies which will be supported and protected through various instruments, including also the conclusion of intergovernmental agreements on strategic defence co-operation. Such agreements may regulate questions such as the ownership structures of the new companies, the locations for developing technologies and for manufacturing products, subcontractors and exports of jointly developed arms and military equipment. In relation to the KMW–Nexter merger, such a deal between France and Germany is expected to be signed this autumn.

The next step after Japan? (Virtual reality, training and crisis management). Transatlantic Security Paper No. 2, June 2012

The recent crisis in Japan, which combined tsunami and technological events, shows that any crisis, especially those in developed and developing countries, is from here out a hybrid crisis, mixing natural factors and human/technological (NATECH). Faced with such dramatic events, which exceed any means available for emergency rescue service, it is necessary a) to remain prudent and b) to prepare. One of the means for preparing is unquestionably training. However, here, undoubtedly there are important constraints: How to train, for example, while reproducing vividly and realistically, an event? How to exceed the admittedly useful, although very limited, level of the table-top exercise? How also to avoid the unnecessary mobilization of dozens, even hundreds, of field and operation staffers to take part in an exercise which could lead to a disappointing outcome? A major crisis, a major exercise, in effect. The solution of virtual reality has emerged, in Europe and in the United States. It is also sometimes called “serious game”.