Toluene dioxygenase-catalyzed cis-dihydroxylation of benzo[b]thiophenes and benzo[b]furans: synthesis of benzo[b]thiophene 2,3-oxide

Enzymatic cis-dihydroxylation of benzo[b]thiophene, benzo[b]furan and several methyl substituted derivatives was found to occur in both the carbocyclic and heterocyclic rings. Relative and absolute configurations and enantiopurities of the resulting dihydrodiols were determined. Hydrogenation of the alkene bond in carbocyclic cis-dihydrodiols and ring-opening epimerization/reduction reactions of heterocyclic cis/trans-dihydrodiols were also studied. The relatively stable heterocyclic dihydrodiols of benzo[b]thiophene and benzo[b]furan showed a strong preference for the trans configuration in aqueous solutions. The 2,3-dihydrodiol metabolite of benzo[b]thiophene was utilized as a precursor in the chemoenzymatic synthesis of the unstable arene oxide, benzo[b]thiophene 2,3-oxide.

Ordering the cytochrome c-initiated caspase cascade:hierarchical activation of caspases-2, -3, -6, -7, -8, and -10 in a caspase-9-dependent manner

Exit of cytochrome c from mitochondria into the cytosol has been implicated as an important step in apoptosis. In the cytosol, cytochrome c binds to the CED-4 homologue, Apaf-1, thereby triggering Apaf-1-mediated activation of caspase-9. Caspase-9 is thought to propagate the death signal by triggering other caspase activation events, the details of which remain obscure. Here, we report that six additional caspases (caspases-2, -3, -6, -7, -8, and -10) are processed in cell-free extracts in response to cytochrome c, and that three others (caspases-1, -4, and -5) failed to be activated under the same conditions. In vitro association assays confirmed that caspase-9 selectively bound to Apaf-1, whereas caspases-1, -2, -3, -6, -7, -8, and -10 did not. Depletion of caspase-9 from cell extracts abrogated cytochrome c-inducible activation of caspases-2, -3, -6, -7, -8, and -10, suggesting that caspase-9 is required for all of these downstream caspase activation events. Immunodepletion of caspases-3, -6, and -7 from cell extracts enabled us to order the sequence of caspase activation events downstream of caspase-9 and reveal the presence of a branched caspase cascade. Caspase-3 is required for the activation of four other caspases (-2, -6, -8, and -10) in this pathway and also participates in a feedback amplification loop involving caspase-9.

Comparative evaluation of 2,3-bis [2-methyloxy-4-nitro-5-sulfophenyl]-2H-tetrazolium-5-carboxanilide (XTT) and 2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2, 4-disulfophenyl)-2H-tetrazolium, monosodium salt (WST-8) rapid colorimetric assays for antimicrobial susceptibility testing of staphylococci and ESBL-producing clinical isolates

A new generation of water soluble tetrazolium salts have recently become available and in this study we compared a colorimetric assay developed using one of these salts, 2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2, 4-disulfophenyl)-2H-tetrazolium, monosodium salt (WST-8), with a previously developed 2,3-bis [2-methyloxy-4-nitro-5-sulfophenyl]-2H-tetrazolium-5-carboxanilide (XTT) colorimetric assay to determine which agent is most suitable for use as a colorimetric indicator in susceptibility testing. The MICs of 6 antibiotics were determined for 33 staphylococci using both colorimetric assays and compared with those obtained using the British Society for Antimicrobial Chemotherapy reference broth microdilution method. Absolute categorical agreement between the reference and test methods ranged from 79% (cefuroxime) to 100% (vancomycin) for both assays. No minor or major errors occurred using either assay with very major errors ranging from zero (vancomycin) to seven (cefuroxime). Analysis of the distribution of differences in the log2 dilution MIC results revealed overall agreement, within the accuracy limits of the standard test (± 1 log2 dilution), using the XTT and WST-8 assays of 98% and 88%, respectively. Further studies on 31 ESBL-producing isolates were performed using the XTT method with absolute categorical agreement ranging from 87% (nitrofurantoin) to 100% (ofloxacin and meropenem). No errors were noted for either ofloxacin or meropenem with overall agreement of 91%. The data suggests that XTT is more reliable and accurate than WST-8 for use in a rapid antimicrobial susceptibility test.

Epitaxial ferroelectric Pb(Mg1/3Nb2/3)O3-PbTiO3 thin films on La0.7Sr0.3MnO3 bottom electrode

Epitaxial (001)-oriented 0.7Pb(Mg_0.33Nb_0.67)O₃-0.3PbTiO₃ (PMN-PT) thin films were deposited by pulsed laser deposition on vicinal SrTiO₃ (001) substrates using La_0.7Sr_0.3MnO₃ as bottom electrode. Detailed microstructural investigations of these films were carried out using X-ray diffraction (XRD), atomic force microscopy (AFM) and transmission electron microscopy (TEM). Polarization-field hysteresis curves were measured at room temperature. Spontaneous polarization P s , remnant polarization P r and coercive voltage V c were found to be 25 μC/cm², 15 μC/cm² and 0.81 V, respectively. Field dependent dielectric constant measurements exhibited butterfly shaped curves, indicating the true ferroelectric nature of these films at room temperature. The dielectric constant and the dielectric loss at 100 kHz were found to be 238 and 0.14, respectively. The local piezoelectric properties of PMN-PT films were investigated by piezoelectric force microscopy and were found to exhibit a local piezoelectric coefficient of 7.8 pm/V.

Tiotropium Respimat® in cystic fibrosis: Phase 3 and Pooled phase 2/3 randomized trials

Background: Tiotropium Respimat® improved lung function in a phase 2 trial in patients with cystic fibrosis (CF). We investigated its efficacy and safety in a phase 3 trial, including a pre-specified pooled analysis of the phase 2 and 3 trials. 

Methods: 12-week, randomized, double-blind, placebo-controlled trial of tiotropium Respimat® 5. μg once daily in patients with CF (N = 463). 

Results: Co-primary efficacy endpoints showed no statistical difference between tiotropium and placebo: percent-predicted forced expiratory volume in 1s (FEV₁) area under the curve from 0-4h (AUC_0-4h) (95% CI): 1.64% (0.27,3.55; p=0.092); percent-predicted trough FEV₁ (95% CI) 1.40% (0.50,3.30; p=0.15). Adverse events were similar between groups. Pooled phase 2/3 trial results showed a treatment difference in favor of tiotropium: percent-predicted FEV₁ AUC_0-4h (95% CI): 2.62% (1.34,3.90). 

Conclusion: Tiotropium was well tolerated in patients with CF; lung function improvements compared with placebo were not statistically significant in the phase 3 trial. Clinical trials: These studies are registered with clinical trial identifier numbers NCT00737100 and NCT01179347NCT00737100NCT01179347. These studies are also registered with the EudraCT number: 2008-001156-43 and 2010-019802-17.

Os alunos de risco no 2º ciclo da Escola Básica 2, 3 Ana de Castro Osório, no ano lectivo 2004/2005: um estudo de caso

O presente estudo aborda a problemática dos alunos de risco no 2° Ciclo da E.B. 2,3 Ana de Castro Osório, no ano letivo 2004/2005. Na dimensão teórica são referenciadas algumas questões de Organização Escolar (escola reflexiva, autonomia de escola, gestores intermédios e supervisão escolar, o papel do Diretor de Turma, o Projeto Curricular de Escola, algumas linhas orientadoras da política educativa portuguesa, no período em que decorre a elaboração desta investigação) e abordados alguns aspetos gerais dos pré-adolescentes, diversos conceitos de risco, a ação de diversas entidades com competência na área da infância e juventude, o papel da família e a relação desta com a escola. Foi utilizada a metodologia do “Estudo Caso”, no decurso da qual foram analisados documentos internos da escola e elaborados instrumentos de recolha de dados (questionários aplicados aos Alunos e aos seus Diretores de Turma). No tratamento das informações de natureza quantitativa, utilizou-se o programa informático Excel, enquanto nas questões abertas procedeu-se à análise de conteúdo. Em síntese, concluiu-se que: a) um terço dos alunos que frequentava o 2° Ciclo do Ensino Básico, foi sinalizado como “aluno de risco”. Cerca de 75% era do 5° Ano de Escolaridade. Os fatores de risco que mais se destacam são: assiduidade irregular, desmotivação/desinteresse/apatia, défice de atenção e concentração, indisciplina e retenções sucessivas no seu percurso escolar. A família é pouco participativa. b) a resposta curricular dá-se essencialmente dentro da sala de aula; c) a resposta organizacional assenta essencialmente nalgumas medidas previstas nos documentos legislativos. Na última parte do trabalho deixamos expressas as conclusões do estudo mais pormenorizadas, bem como algumas recomendações/sugestões que eventualmente, poderão orientar posteriores investigações. /ABSTRACT - The risk students on a 2nd Cycle of classroom of the E.B. 2, 3 Ana de Castro Osório, during academic year of 2004/2005: A Case study The present study attempts to assess risk students on a 2nd Cycle (Sth and 6th grades) classroom of the “E.B. 2, 3 Ana de Castro Osório” Public School during the academic year of 2004/2005. On a theoretical scale, are assessed some questions about the Classroom Organization (reflexive school, school autonomy, intermediate directors and school supervision, the role of the Class Tutor, the School Curricular Project, some guidelines about the Portuguese education policy during the period when this study took place), and was also assessed some general aspects of preadolescents, risk concepts, and actions taken by various organizations specialized in the areas of childhood and youth, the role of the family and the interrelation between the family and the school. It was used a “Case Study” methodology during which was assessed school documents of this particularly school, and data collecting instruments (Student-applied and Class Tutor-applied questionnaires). On dealing with quantitative data, it was used the Excel application, while for the open questions was assessed by its contents. In brief, it was concluded that: a) a third of students that attended the 2nd Cycle Basic Education schools revealed to be “risk students”. Around 75% of them were 5th grade students. Among the most important risk factors are: irregular attendance, unmotivated/aloofness /apathy, lack of attention and concentration, indiscipline and successive retentions during their education pathway. The family is scarcely participative. b) The curricular response is normally given within the classroom; c) the organizational response lies mostly on some measures included in government documents. On the last part of the paper, we focus on more detailed conclusions, as well as give recommendations/suggestions which may eventually be of guidance to future studies.