Oxidative stress is responsible for deficient survival and dendritogenesis in Purkinje neurons from ataxia-telangiectasia mutated mutant mice

Atm gene-disrupted mice recapitulate the majority of characteristics observed in patients with the genetic disorder ataxia-telangiectasia (A-T). However, although they exhibit defects in neuromotor function and a distinct neurological phenotype, they do not show the progressive neurodegeneration seen in human patients, but there is evidence that ataxia-telangiectasia mutated ( Atm)-deficient animals have elevated levels of oxidized macromolecules and some neuropathology. We report here that in vitro survival of cerebellar Purkinje cells from both Atm knock-out and Atm knock-in mice was significantly reduced compared with their wild-type littermates. Although most of the Purkinje neurons from wild-type mice exhibited extensive dendritic elongation and branching under these conditions, most neurons from Atm-deficient mice had dramatically reduced dendritic branching. An antioxidant ( isoindoline nitroxide) prevented Purkinje cell death in Atm-deficient mice and enhanced dendritogenesis to wild-type levels. Furthermore, administration of the antioxidant throughout pregnancy had a small enhancing effect on Purkinje neuron survival in Atm gene-disrupted animals and protected against oxidative stress in older animals. These data provide strong evidence for a defect in the cerebellum of Atm-deficient mice and suggest that oxidative stress contributes to this phenotype.

An investigation of magnetic antennas for ground penetrating radar

For ground penetrating radar (GPR), smaller antennas would provide considerable practical advantages. Some of which are: portability; ease of use; and higher spatial sampling. A theoretical comparison of the fundamental limits of a small electric field antenna and a small magnetic field antenna shows that the minimum Q constraints are identical. Furthermore, it is shown that only the small magnetic loop antenna can be constructed to approach, arbitrarily closely, the fundamental minimum Q limit. This is achieved with the addition of a high permeability material which reduces energy stored in the magnetic fields. This is of special interest to some GPR applications. For example, applications requiring synthetic aperture data collection would benefit from the increased spatial sampling offered by electrically smaller antennas. Low frequency applications may also benefit, in terms of reduced antenna dimensions, by the use of electrically small antennas. Under these circumstances, a magnetic type antenna should be considered in preference to the typical electric field antenna. Numerical modeling data supports this assertion.

Synthesis and characterization of delta-atracotoxin-ar1a, the lethal neurotoxin from venom of the Sydney funnel-web spider (Atrax robustus)

delta-Atracotoxin-Ar1a (delta-ACTX-Ar1a) is the major polypeptide neurotoxin isolated from the venom of the male Sydney funnel-web spider, Atrax robustus. This neurotoxin targets both insect and mammalian voltage-gated sodium channels, where it competes with scorpion alpha-toxins for neurotoxin receptor site-3 to slow sodium-channel inactivation. Progress in characterizing the structure and mechanism of action of this toxin has been hampered by the limited supply of pure toxin from natural sources. In this paper, we describe the first successful chemical synthesis and oxidative refolding of the four-disulfide bond containing delta-ACTX-Ar1a. This synthesis involved solid-phase Boc chemistry using double coupling, followed by oxidative folding of purified peptide using a buffer of 2 M GdnHCl and glutathione/glutathiol in a 1:1 mixture of 2-propanol (pH 8.5). Successful oxidation and refolding was confirmed using both chemical and pharmacological characterization. Ion spray mass spectrometry was employed to confirm the molecular weight. H-1 NMR analysis showed identical chemical shifts for native and synthetic toxins, indicating that the synthetic toxin adopts the native fold. Pharmacological studies employing whole-cell patch clamp recordings from rat dorsal root ganglion neurons confirmed that synthetic delta-ACTX-Ar1a produced a slowing of the sodium current inactivation and hyperpolarizing shifts in the voltage-dependence of activation and inactivation similar to native toxin. Under current clamp conditions, we show for the first time that delta-ACTX-Ar1a produces spontaneous repetitive plateau potentials underlying the clinical symptoms seen during envenomation. This successful oxidative refolding of synthetic delta-ACTX-Ar1a paves the way for future structure-activity studies to determine the toxin pharmacophore.

alpha-conotoxins PnIA and [A10L] PnIA stabilize different states of the alpha 7-L247T nicotinic acetylcholine receptor

The effects of the native alpha-conotoxin PnIA, its synthetic derivative [ A10L] PnIA and alanine scan derivatives of [ A10L] PnIA were investigated on chick wild type alpha7 and alpha7-L247T mutant nicotinic acetylcholine receptors (nAChRs) expressed in Xenopus oocytes. PnIA and [A10L] PnIA inhibited acetylcholine (ACh)-activated currents at wtalpha7 receptors with IC50 values of 349 and 168 nM, respectively. Rates of onset of inhibition were similar for PnIA and [ A10L] PnIA; however, the rate of recovery was slower for [ A10L] PnIA, indicating that the increased potency of [ A10L] PnIA at alpha7 receptors is conveyed by its slower rate of dissociation from the receptors. All the alanine mutants of [ A10L] PnIA inhibited ACh-activated currents at wtalpha7 receptors. Insertion of an alanine residue between position 5 and 13 and at position 15 significantly reduced the ability of [ A10L] PnIA to inhibit ACh-evoked currents. PnIA inhibited the non-desensitizing ACh-activated currents at alpha7-L247T receptors with an IC50 194 nM. In contrast, [ A10L] PnIA and the alanine mutants potentiated the ACh-activated current alpha7-L247T receptors and in addition [ A10L] PnIA acted as an agonist. PnIA stabilized the receptor in a state that is non-conducting in both the wild type and mutant receptors, whereas [ A10L] PnIA stabilized a state that is non-conducting in the wild type receptor and conducting in the alpha7-L247T mutant. These data indicate that the change of a single amino acid side-chain, at position 10, is sufficient to change the toxin specificity for receptor states in the alpha7-L247T mutant.

Structure of the HERG K+ channel S5P extracellular linker - Role of an amphipathic alpha-helix in C-type inactivation

The HERG K+ channel has very unusual kinetic behavior that includes slow activation but rapid inactivation. These features are critical for normal cardiac repolarization as well as in preventing lethal ventricular arrhythmias. Mutagenesis studies have shown that the extracellular peptide linker joining the fifth transmembrane domain to the pore helix is critical for rapid inactivation of the HERG K+ channel. This peptide linker is also considerably longer in HERG K+ channels, 40 amino acids, than in most other voltage-gated K+ channels. In this study we show that a synthetic 42-residue peptide corresponding to this linker region of the HERG K+ channel does not have defined structural elements in aqueous solution; however, it displays two well defined helical regions when in the presence of SDS micelles. The helices correspond to Trp(585)-Ile(593) and Gly(604)-Tyr(611) of the channel. The Trp(585)-Ile(593) helix has distinct hydrophilic and hydrophobic surfaces. The Gly(604)-Tyr(611) helix corresponds to an N-terminal extension of the pore helix. Electrophysiological studies of HERG currents following application of exogenous S5P peptides show that the amphipathic helix in the S5P linker interacts with the pore region of the channel in a voltage-dependent manner.

Preparation and characterization of Nickel-and cobalt-doped magnetites

Trabalho apresentado no I Simpósio Mineiro de Ciências dos Materiais, Ouro Preto, Novembro de 2001.

Five generations of peace operations: from the "thin blue line" to "painting a country blue"

This article combines both international practice and analytical contributions into a systematic and synthetic presentation of the evolution of peace operations from their modern inception in 1948 to the present. It seeks to serve a didactic purpose in proposing a basic structure for Brazilian scholars' burgeoning debate on peace operations and intervention, rather than a definitive characterization of blue helmet practice. Peace operations' progression is traced through five analytical "generations," each adding a crucial factor distinguishing it from its predecessors. Each generation is placed in relation to changes in the nature of conflict and in the interpretation of the foundational principles of peace operations, and links to broader theoretical issues in International Relations are made explicit at each stage.

Electrospun silk-elastin-like fiber mats for tissue engineering applications

Protein-based polymers are present in a wide variety of organisms fulfilling structural and mechanical roles. Advances in protein engineering and recombinant DNA technology allow the design and production of recombinant protein-based polymers (rPBPs) with an absolute control of its composition. Although the application of recombinant proteins as biomaterials is still an emerging technology, the possibilities are limitless and far superior to natural or synthetic materials, as the complexity of the structural design can be fully customized. In this work, we report the electrospinning of two new genetically engineered silk-elastin-like proteins (SELPs) consisting of alternate silk- and elastin-like blocks. Electrospinning was performed with formic acid and aqueous solutions at different concentrations without addition of further agents. The size and morphology of the electrospun structures was characterized by scanning electron microscopy showing to be dependent of concentration and solvent used. Treatment with air saturated with methanol was employed to stabilize the structure and promote water insolubility through a time-dependent conversion of random coils into β-sheets (FTIR). The resultant methanol-treated electrospun mats were characterized for swelling degree (570-720%), water vapour transmission rate (1083 g/m2/day) and mechanical properties (modulus of elasticity of ~126 MPa). Furthermore, the methanol-treated SELP fiber mats showed no cytotoxicity and were able to support adhesion and proliferation of normal human skin fibroblasts. Adhesion was characterized by a filopodia-mediated mechanism. These results demonstrate that SELP fiber mats can provide promising solutions for the development of novel biomaterials suitable for tissue engineering applications.

One-sided Radial-Fundamental Matrix Estimation

For modern consumer cameras often approximate calibration data is available, making applications such as 3D reconstruction or photo registration easier as compared to the pure uncalibrated setting. In this paper we address the setting with calibrateduncalibrated image pairs: for one image intrinsic parameters are assumed to be known, whereas the second view has unknown distortion and calibration parameters. This situation arises e.g. when one would like to register archive imagery to recently taken photos. A commonly adopted strategy for determining epipolar geometry is based on feature matching and minimal solvers inside a RANSAC framework. However, only very few existing solutions apply to the calibrated-uncalibrated setting. We propose a simple and numerically stable two-step scheme to first estimate radial distortion parameters and subsequently the focal length using novel solvers. We demonstrate the performance on synthetic and real datasets.

Anotação sintética, numa perspectiva de Direito Público, ao Acórdão n.º 63/2006, em Plenário, do Tribunal Constitucional (português), de 24 de Janeiro de 2006 - qual proibição constitucional de impostos retroactivos? A tese de Oliveira Salazar.

Resumo: 1 – Sumário do Acórdão do Tribunal Constitucional n.º 63/2006, de 24 de Janeiro de 2006; 2 – Texto completo do Acórdão do Tribunal Constitucional n.º 63/2006, de 24 de Janeiro de 2006: cfr. http://www.tribunalconstitucional.pt/tc/acordaos/20060063.html , 18 de Maio de 2012; 3 – Anotação sintética; 3.1 – Introdução à anotação sintética e suas características neste caso concreto; 4 – A referência, abstracta e concreta, do princípio constitucional da «proibição de impostos retroactivos»; 5 – Alguns aspectos nucleares da eficácia temporal das normas jurídico-tributárias, brevitatis causa, das normas jurídico-fiscais; 6 – A chamada «aplicação das normas fiscais» (e/ou tributárias) no seio da «aplicação no tempo»: algumas breves notas sobre a tradição jurídico-lusitana, v.g. do ponto de vista da doutrina, a partir de 1976, designadamente até 1985; 6.1 – A tese de António de Oliveira Salazar; 7 – Conclusões. § Abstract: 1 - Summary of the Judgment of the Constitutional Court n. 63/2006 of January 24, 2006; 2 - Complete text of the Judgment of the Constitutional Court n. 63/2006 of January 24, 2006: s. http://www.tribunalconstitucional.pt/tc/acordaos/20060063.html , May 18, 2012; 3 - Synthetic Note: 3.1 - Introduction to syntheticannotation and its characteristics in this case; 4 - The reference, abstract and concrete, the constitutional principle of «prohibition of retroactive taxes»; 5 - Some aspects of nuclear the temporal validity of the legal and tax rules, brevitatis causa of legal and tax rules; 6 - the so-called «implementation of tax laws» (and/or tax) within the «Application in time»: some brief notes on the legal tradition -Lusitanian, e.g. from the standpoint of doctrine, from 1976, namely until 1985; 6.1 - The thesis of António de Oliveira Salazar; 7- Conclusions. PS: este "abstract" está tal qual como na publicação.

Anotação sintética, numa perspectiva de Direito Público-penal e Direito Privado-civil, ao Acórdão do Supremo Tribunal de Justiça Português, de 19 de Abril de 2012-- A poluição sonora e o direito humano ao descanso e à saúde, sem descurar a necessidade duma adequada política tributária.

Resumo: 1 – Sumário do Acórdão do Supremo Tribunal de Justiça, de 19 de Abril de 2012; 2 – Texto completo do Acórdão do Supremo Tribunal de Justiça, de 19 de Abril de 2012: cfr. http://www.dgsi.pt/jstj.nsf/954f0ce6ad9dd8b980256b5f003fa814/fc664c231f3e73cf802579ea003d91d2?OpenDocument&Highlight=0,polui%C3%A7%C3%A3o , 2 de Junho de 2012; 3 – Anotação sintética; 3.1 – Introdução à anotação sintética e suas características neste caso concreto; 4 – Algumas referências constitucionais centrais em relação a Direitos humanos e, nomeadamente, a um Direito humano a um meio-ambiente sadio, saudável em todas as suas vertentes e sentidos – o exemplo central do artigo 9.º da CRP; 4.1 – Algumas referências constitucionais centrais em relação a Direitos humanos e, nomeadamente, a um Direito humano a um meio-ambiente sadio, saudável em todas as suas vertentes e sentidos – o exemplo central do artigo 66.º da CRP e o Regime Geral do Ruído; 5 – O direito humano ao descanso e à saúde, rectius o direito ao ambiente sadio vs o direito ao lazer e/ou exploração económica de indústrias de diversão, rectius o direito à liberdade de iniciativa económica privada; 6 – A violação do direito humano, de personalidade, ao descanso e à saúde, rectius o direito a um ambiente sadio, numa perspectiva de Direito privado e Direito civil; 7 – A criminalização da poluição, designadamente a criminalização da poluição sonora – uma perspectiva de Direito público e Direito penal; 8 - A necessidade duma adequada política tributária que compatibilize desenvolvimento sustentado com a protecção dum meio ambiente sadio e com qualidade de vida; 9 – Conclusões. § Abstract: 1 - Summary of the Judgment of the Supreme Court of April 19, 2012, 2 - Complete text of the Judgment of the Supreme Court of April 19, 2012: cf. http://www.dgsi.pt/jstj.nsf/954f0ce6ad9dd8b980256b5f003fa814/fc664c231f3e73cf802579ea003d91d2?OpenDocument&Highlight=0,polui%C3%A7%C3%A3o , June 2, 2012, 3 - Synthetic Note: 3.1 - Introduction to synthetic annotation and its characteristics in this case 4 - Some references constitutional power over human rights and in particular to a human right to a healthy environment, healthy in all its forms and meanings - the central example of Article 9. of CRP; 4.1 - Some references constitutional power over human rights and in particular to a human right to a healthy environment, healthy in all its forms and meanings - the central example of Article 66. No of CRP and the General Noise; 5 - the human right to rest and health, rectius the right to healthy environment vs. the right to leisure and / or economic exploitation of industries fun, rectius the right to freedom of private economic initiative; 6 - the violation of human personality, to rest and health, rectius the right to a healthy environment, a perspective of private law and civil law; 7 - criminalization of pollution, including the criminalization of noise - a perspective of public law and criminal law; 8 - the need for appropriate tax policy that reconciles sustainable development with the protection of a healthy environment and quality of life; 9 - Conclusions.

NOVA ANOTAÇÃO SINTÉTICA, NUMA PERSPECTIVA DE DIREITO PÚBLICO, AINDA AO ACÓRDÃO 212/95, DO TRIBUNAL CONSTITUCIONAL (PORTUGUÊS), DE 20.04.1995 - O DEVER FUNDAMENTAL DE EXISTIR NA LEI EM VIGOR UMA RESPONSABILIDADE CRIMINAL DAS "PESSOAS COLECTIVAS" (OU "PESSOAS JURÍDICAS") E A CONSTITUIÇÃO DA REPÚBLICA PORTUGUESA, V.G. O CASO DO ART. 7º DO REGIME GERAL DAS INFRACÇÕES TRIBUTÁRIAS E DO ART. 3.º DO REGIME DAS INFRACÇÕES ANTIECONÓMICAS E CONTRA A SAÚDE PÚBLICA

Resumo: 1 – Sumário do Acórdão do Tribunal Constitucional n.º 212/1995, de 20 de Abril; 2 – Parte principal do Acórdão do Tribunal Constitucional n.º 212/1995, de 20 de Abril: cfr. http://www.tribunalconstitucional.pt/tc/acordaos/19950212.html , 18 de Maio de 2012; 3 – Anotação sintética; 3.1 – Introdução à anotação sintética e suas características neste caso concreto; 4 – O RIAECSP (Regime das Infracções Anti-Económicas e Contra a Saúde Pública) e a responsabilidade criminal das pessoas colectivas e equiparadas, prevista no seu art. 3.º; 5 – Societas delinquere non potest?; 6 – Breves traços históricos do brocardo societas delinquere non potest; 7 - Alguns dos marcos relevantes na Doutrina penal portuguesa recente acerca da responsabilidade penal das organizações, entes colectivos, pessoas colectivas, pessoas jurídicas; 8 - Os art.os 12.º/2 e 2.º da CRP e a Responsabilidade Criminal dos Entes Colectivos, pessoas jurídicas; 9 - O art. 29.º/5 da CRP - ou o princípio non bis in idem - e a responsabilidade criminal das organizações, dos entes colectivos, das pessoas colectivas; 10 – Conclusões. § Summary: 1 - Summary Judgment of the Constitutional Court No. 212/1995 of 20 April; 2 - The main part of the Constitutional Court Ruling No. 212/1995 of 20 April: cfr. http://www.tribunalconstitucional.pt/tc/acordaos/19950212.html, May 18, 2012; 3 - short annotation; 3.1 - Introduction to synthetic note and its features in this particular case; 4 - The RIAECSP (Status of Anti-Economic Offences and Against Public Health) and the criminal liability of companies and similar persons, provided for in his art. 3; 5 - delinquere Societas non potest ?; 6 - Brief historical traces of societas aphorism delinquere non potest; 7 - Some of the important milestones in recent Portuguese criminal Doctrine about the criminal liability of organizations, public entities, legal persons, legal entities; 8 - art.os 12/2 and 2 of CRP and Criminal Responsibility of loved Collective, legal entities; 9 - Art. 29/5 CRP - or the principle of non bis in idem - and the criminal liability of organizations, collective entities, of legal persons; 10 - Conclusions.

Dense motion field estimation from myocardial boundary displacements

Minimally invasive cardiovascular interventions guided by multiple imaging modalities are rapidly gaining clinical acceptance for the treatment of several cardiovascular diseases. These images are typically fused with richly detailed pre-operative scans through registration techniques, enhancing the intra-operative clinical data and easing the image-guided procedures. Nonetheless, rigid models have been used to align the different modalities, not taking into account the anatomical variations of the cardiac muscle throughout the cardiac cycle. In the current study, we present a novel strategy to compensate the beat-to-beat physiological adaptation of the myocardium. Hereto, we intend to prove that a complete myocardial motion field can be quickly recovered from the displacement field at the myocardial boundaries, therefore being an efficient strategy to locally deform the cardiac muscle. We address this hypothesis by comparing three different strategies to recover a dense myocardial motion field from a sparse one, namely, a diffusion-based approach, thin-plate splines, and multiquadric radial basis functions. Two experimental setups were used to validate the proposed strategy. First, an in silico validation was carried out on synthetic motion fields obtained from two realistic simulated ultrasound sequences. Then, 45 mid-ventricular 2D sequences of cine magnetic resonance imaging were processed to further evaluate the different approaches. The results showed that accurate boundary tracking combined with dense myocardial recovery via interpolation/ diffusion is a potentially viable solution to speed up dense myocardial motion field estimation and, consequently, to deform/compensate the myocardial wall throughout the cardiac cycle. Copyright © 2015 John Wiley & Sons, Ltd.