Study of the mechanism of action of anoplin, a helical antimicrobial decapeptide with ion channel-like activity, and the role of the amidated C-terminus

Anoplin, an antimicrobial, helical decapeptide from wasp venom, looses its biological activities by mere deamidation of its C-terminus. Secondary structure determination, by circular dichroism spectroscopy in amphipathic environments, and lytic activity in zwitterionic and anionic vesicles showed quite similar results for the amidated and the carboxylated forms of the peptide. The deamidation of the C-terminus introduced a negative charge at an all-positive charged peptide, causing a loss of amphipathicity, as indicated by molecular dynamics simulations in TFE/water mixtures and this subtle modification in a peptide's primary structure disturbed the interaction with bilayers and biological membranes. Although being poorly lytic, the amidated form, but not the carboxylated, presented ion channel-like activity on anionic bilayers with a well-defined conductance step; at approximately the same concentration it showed antimicrobial activity. The pores remain open at trans-negative potentials, preferentially conducting cations, and this situation is equivalent to the interaction of the peptide with bacterial membranes that also maintain a high negative potential inside. Copyright (C) 2007 European Peptide Society and John Wiley & Sons, Ltd.

Action of auxins, boron and harvest season on the rooting of kiwi fruit stem cuttings (Actinidia chinensis Pl cv Monty)

Action of auxins on the rooting of stem cuttings of kiwi (Actinidia chinensis P. cv Monty). This work studies the effects of some synthetic auxins and B in the rooting of kiwi (Actinidia chinensis Planch cv Monty) stem cuttings. The treatments used were as follows: T1 (H2O); T2 (NAA 300 ppm); T3 (IBA 300 ppm); T4 (NAA 300 ppm + B); T5 (IBA 300 ppm + B); T6 (NAA 0,5%-talc) and T7 (IBA 0,5%-talc), applied to the bases of the cuttings. These were then placed in rooting dishes with pure vermiculite in a misty nebulization chamber until collection day (120 days). The evaluation of auxin and boric acid effects on kiwi stem cuttings were made based on the following observations: 1. The percentage of rooted stem cuttings; 2. reducing sugars and total sugar analyses (in g/100 g of dry matter); and 3. tryptophan analyses (in mu g/100 mg of dry matter). The results show that summer is the best season for rooting Actinidia chinensis Planch cv Monty stem cuttings. The use of IBA talc 0,5% on the bases of the cuttings shamed positive results too.

UMP kinase from Mycobacterium tuberculosis: Mode of action and allosteric interactions, and their likely role in pyrimidine metabolism regulation

The pyrH-encoded uridine 5′-monophosphate kinase (UMPK) is involved in both de novo and salvage synthesis of DNA and RNA precursors. Here we describe Mycobacterium tuberculosis UMPK (MtUMPK) cloning and expression in Escherichia coli. N-terminal amino acid sequencing and electrospray ionization mass spectrometry analyses confirmed the identity of homogeneous MtUMPK. MtUMPK catalyzed the phosphorylation of UMP to UDP, using ATP-Mg 2+ as phosphate donor. Size exclusion chromatography showed that the protein is a homotetramer. Kinetic studies revealed that MtUMPK exhibits cooperative kinetics towards ATP and undergoes allosteric regulation. GTP and UTP are, respectively, positive and negative effectors, maintaining the balance of purine versus pyrimidine synthesis. Initial velocity studies and substrate(s) binding measured by isothermal titration calorimetry suggested that catalysis proceeds by a sequential ordered mechanism, in which ATP binds first followed by UMP binding, and release of products is random. As MtUMPK does not resemble its eukaryotic counterparts, specific inhibitors could be designed to be tested as antitubercular agents. © 2010 Elsevier Inc. All rights reserved.

Riboflavin analgs from Streptomyces davawensisas antiinfectives: mode of action, mechanism of resistance of the producer and metabolization by humans

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Evaluation of ionic degradation and slot corrosion of metallic brackets by the action of different dentifrices

OBJETIVO: avaliar in vitro a degradação iônica e corrosão do fundo do slot de braquetes metálicos submetidos à escovação com dentifrícios, realizando análises da composição química por Espectroscopia de Energia Dispersiva (EDS) e qualitativa por Microscopia Eletrônica de Varredura (MEV). MÉTODOS: foram selecionados 38 braquetes divididos aleatoriamente em quatro grupos experimentais (n = 7). Dois grupos (n = 5) funcionaram como controles positivo e negativo. Aparelhos ortodônticos simulados foram confeccionados com fios de aço inoxidável 0,019" x 0,025" e anéis elastoméricos. Os grupos foram divididos de acordo com o tratamento de superfície: G1 (Máxima Proteção Anticáries®); G2 (Total 12®); G3 (Sensitive®); G4 (Branqueador®); Controle Positivo (saliva artificial) e Controle Negativo (sem tratamento). Foram realizados 28 ciclos de escovação e avaliações antes (T0) e após (T1) o experimento. RESULTADOS: o teste de Wilcoxon indicou não existir diferença nas concentrações iônicas de titânio (Ti), cromo (Cr), ferro (Fe) e níquel (Ni) entre os grupos. O grupo G2 apresentou redução significativa (p < 0,05) na concentração do íon alumínio (Al) e os grupos G3 e G4 apresentaram aumento significativo (p < 0,05) nas concentrações do íon alumínio. A análise em MEV mostrou aumento nas características indicativas de corrosão dos grupos G2, G3 e G4. CONCLUSÃO: a análise por EDS revelou que os grupos controle e G1 não sofreram alterações na composição química. O grupo G2 apresentou degradação na quantidade de íons Al, e G3 e G4 sofreram aumento na concentração de Al. A imersão em saliva artificial e o dentifrício Máxima Proteção Anticáries® não alteraram o polimento de superfície. Os dentifrícios Total 12®, Sensitive® e Branqueador® alteraram o polimento de superfície.

In vitro and in vivo acaricide action of juvenoid analogs produced from the chemical modification of Cymbopogon spp. and Corymbia citriodora essential oil on the cattle tick Rhipicephalus (Boophilus) microplus

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Evaluation of the antimutagenic activity and mode of action of the fructooligosaccharide inulin in the meristematic cells of Allium cepa culture

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Generators of supersymmetric polynomials in positive characteristic

In Kantor and Trishin (1997) [3], Kantor and Trishin described the algebra of polynomial invariants of the adjoint representation of the Lie superalgebra gl(m vertical bar n) and a related algebra A, of what they called pseudosymmetric polynomials over an algebraically closed field K of characteristic zero. The algebra A(s) was investigated earlier by Stembridge (1985) who in [9] called the elements of A(s) supersymmetric polynomials and determined generators of A(s). The case of positive characteristic p of the ground field K has been recently investigated by La Scala and Zubkov (in press) in [6]. We extend their work and give a complete description of generators of polynomial invariants of the adjoint action of the general linear supergroup GL(m vertical bar n) and generators of A(s).

Thought-Action Fusion and Neutralization Behavior

This study presents a new inventory to assess thought-action fusion (TAF). 160 college students ages 18 to 22 (M = 19.17, SD = 1.11) completed the new Modified Thought Action Scale (MTAFS). Results indicated high internal consistency in the MTAFS (Cronbach’s α = .95). A principal component analysis suggested a three factor solution of TAF-Moral (TAFM), TAFLikelihood (TAFL), and TAF-Harm avoidance-Positive (TAFHP) all with eigenvalues above 1, and factor loadings above .4. A second study examined the association between TAF, obsessivecompulsive and anxiety tendencies after the activation of TAF-like thought processes in a nonclinical sample (n=76). Subjects were randomly assigned to one of three treatment groups intended to provoke TAFL-self, TAFL-other, and TAF moral thought processes. Stepwise regression analyses revealed: 1) the Obsessive-Compulsive Inventory subscales Neutralizing and Ordering significantly predicted instructed neutralization behavior (INB) in non-clinical participants; 2) TAF-Likelihood contributed significant unique variance in INB. These findings suggest that the provocation of neutralization behavior may be mediated by specific subsets of TAF and obsessive-compulsive tendencies.

A novel steroid-like compound F90927 exerting positive-inotropic effects in cardiac muscle

Here we report a novel steroid-like compound F90363, exhibiting positive inotropy in vivo and in vitro in various cardiac muscle preparations. F90363 is a racemic mixture composed of the stereoisomers (-)-F90926 and (+)-F90927. Only F90927 exerted positive inotropy, while F90926 induced a weak negative inotropy, but only at concentrations 10(3) times higher than F90927 and most likely resulting from an unspecific interaction. The rapid time course of the action of F90927 suggested a direct interaction with a cellular target rather than a genomic alteration. We could identify the L-type Ca2+ current I(Ca(L)) as a main target of F90927, while excluding other components of cardiac Ca2+ signalling as potential contributors. In addition, several other signaling pathways known to lead to positive inotropy (e.g. alpha- and beta-adrenergic stimulation, cAMP pathways) could be excluded as targets of F90927. However, vessel contraction and stiffening of the cardiac muscle at high doses (>30 microM, 0.36 mg kg(-1), respectively) prevent the use of F90927 as a candidate for drug development. Since the compound may still find valuable applications in research, the aim of the present study was to identify the cellular target and the mechanism of inotropy of F90927.

Action Research and Learning Analytics in Higher Education

Teaching is a dynamic activity. It can be very effective, if its impact is constantly monitored and adjusted to the demands of changing social contexts and needs of learners. This implies that teachers need to be aware about teaching and learning processes. Moreover, they should constantly question their didactical methods and the learning resources, which they provide to their students. They should reflect if their actions are suitable, and they should regulate their teaching, e.g., by updating learning materials based on new knowledge about learners, or by motivating learners to engage in further learning activities. In the last years, a rising interest in ‘learning analytics’ is observable. This interest is motivated by the availability of massive amounts of educational data. Also, the continuously increasing processing power, and a strong motivation for discovering new information from these pools of educational data, is pushing further developments within the learning analytics research field. Learning analytics could be a method for reflective teaching practice that enables and guides teachers to investigate and evaluate their work in future learning scenarios. However, this potentially positive impact has not yet been sufficiently verified by learning analytics research. Another method that pursues these goals is ‘action research’. Learning analytics promises to initiate action research processes because it facilitates awareness, reflection and regulation of teaching activities analogous to action research. Therefore, this thesis joins both concepts, in order to improve the design of learning analytics tools. Central research question of this thesis are: What are the dimensions of learning analytics in relation to action research, which need to be considered when designing a learning analytics tool? How does a learning analytics dashboard impact the teachers of technology-enhanced university lectures regarding ‘awareness’, ‘reflection’ and ‘action’? Does it initiate action research? Which are central requirements for a learning analytics tool, which pursues such effects? This project followed design-based research principles, in order to answer these research questions. The main contributions are: a theoretical reference model that connects action research and learning analytics, the conceptualization and implementation of a learning analytics tool, a requirements catalogue for useful and usable learning analytics design based on evaluations, a tested procedure for impact analysis, and guidelines for the introduction of learning analytics into higher education.

Establishing optimal quantitative-polymerase chain reaction assays for routine diagnosis and tracking of minimal residual disease in JAK2-V617F-associated myeloproliferative neoplasms: a joint European LeukemiaNet/MPN&MPNr-EuroNet (COST action BM0902) study.

Reliable detection of JAK2-V617F is critical for accurate diagnosis of myeloproliferative neoplasms (MPNs); in addition, sensitive mutation-specific assays can be applied to monitor disease response. However, there has been no consistent approach to JAK2-V617F detection, with assays varying markedly in performance, affecting clinical utility. Therefore, we established a network of 12 laboratories from seven countries to systematically evaluate nine different DNA-based quantitative PCR (qPCR) assays, including those in widespread clinical use. Seven quality control rounds involving over 21,500 qPCR reactions were undertaken using centrally distributed cell line dilutions and plasmid controls. The two best-performing assays were tested on normal blood samples (n=100) to evaluate assay specificity, followed by analysis of serial samples from 28 patients transplanted for JAK2-V617F-positive disease. The most sensitive assay, which performed consistently across a range of qPCR platforms, predicted outcome following transplant, with the mutant allele detected a median of 22 weeks (range 6-85 weeks) before relapse. Four of seven patients achieved molecular remission following donor lymphocyte infusion, indicative of a graft vs MPN effect. This study has established a robust, reliable assay for sensitive JAK2-V617F detection, suitable for assessing response in clinical trials, predicting outcome and guiding management of patients undergoing allogeneic transplant.

Studies on the role of BCR in Philadelphia chromosome-positive leukemias

It is well established that the chimeric Bcr-Abl oncoprotein resulting from fusing 3$\sp\prime$ ABL sequences on chromosome 9 to 5$\sp\prime$ BCR sequences on chromosome 22 is the primary cause of Philadelphia chromosome-positive (Ph$\sp1$) leukemias. Although it is clear that the cis-Bcr sequence present within Bcr-Abl is able to activate the tyrosine kinase activity and F-actin binding capacity of Bcr-Abl which is critical for the transforming ability of BCR-ABL, the biological role of normal BCR gene product (P160 BCR) remains largely unknown. The previous finding by our lab that P160 BCR forms stable complexes with Bcr-Abl oncoprotein in Ph$\sp1$-positive leukemic cells implicated P160 BCR in the pathogenesis of Ph$\sp1$-positive leukemias. Here, we demonstrated that P160 BCR physically interacts with P210 BCR-ABL and become tyrosine phosphorylated when co-expressed with P210 BCR-ABL in COS1 cells while no tyrosine phosphorylation of P160 BCR can be detected when it is expressed alone. The results suggest that P160 BCR is a target for the Bcr-Abl tyrosine kinase. Although we were unable to detect stable physical interaction between P160 BCR and P145 c-ABL (Ib) in COS1 cells overexpressing both proteins, P160 BCR was phosphorylated on tyrosine residues when co-expressed with activated tyrosine kinase of P145 c-ABL (Ib). In addition, studies of tyrosine phosphorylation of BCR deletion mutants and 2-dimensional tryptic mapping of in vitro phosphorylated wild type and mutant (tyrosine to phenylalanine) Bcr-Abl indicated that tyrosine 177, 283 and 360 of Bcr represent some of the phosphorylation sites. Even though the significance of tyrosine phosphorylation of residues 283 and 360 of Bcr has not been determined, tyrosine phosphorylation of residue 177 within Bcr-Abl has been reported to be critical for its interaction with Grb2 molecule and subsequent activation of Ras signaling pathway. Here, we further demonstrated that tyrosine 177 phosphorylated P160 BCR is also able to bind to Grb2 molecule suggesting the role of P160 BCR in the Ras signaling pathway.^ Surprisingly, using 3$\sp\prime$ BCR antisense oligonucleotide to reduce the expression of P160 BCR without interfering with the expression of BCR-ABL resulted in increased growth or survival of B15 cells and M3.16 cells expressing either P185 BCR-ABL or P210 BCR-ABL respectively. The results provided strong arguments that P160 BCR may function as a negative regulator for cell growth.^ Considering all these results, we hypothesize that P160 BCR negatively regulate cell growth and tyrosine phosphorylation of P160 BCR turns off its growth suppressor function and turns on its growth stimulatory function. We further speculate that Bcr-Abl oncoprotein in leukemia cells stably interacts with and constitutively phosphorylates portions of P160 BCR converting it into a growth stimulatory state. In normal cells, the growth suppressor effects of P160 BCR could only be transiently and conditionally switched to growth stimulatory action by a strictly regulated cellular tyrosine kinase such as c-ABL. The model will be further discussed in the text. ^

The door of the tomb is open: Participatory action research to assess the impact of strengthened community capacity on maternal health care in rural Mali

This participatory action-research project addressed the hypothesis that strengthened community and women's capacity for self-development will lead to action to address maternal health problems and the prevention of maternal morbidity and mortality in Mali. Research objectives were: (1) to undertake a comparative cross-sectional study of the association of community capacity with improved maternal health in rural areas of Sanando, Mali, where capacity building interventions have taken place in some villages but not in others. (2) to describe women's maternal health status, access to and use of maternal health services given their residence in program or comparison communities.^ The participatory action research project was an integrated qualitative and quantitative study using participatory rural appraisal exercises, semi-structured group interviews and a cross-sectional survey.^ Factors related to community capacity for self-development were identified: community harmony; an understanding of the benefits of self-development; dynamic leadership; and a structure to implement collective activities.^ A distinct difference between the program and comparison villages was the commitment to train and support traditional birth attendants (TBAs). The TBAs in the program villages work in the context of the wider, integrated self-development program and, 10 years after their initial training, the TBAs continue to practice.^ Many women experience labor and childbirth alone or are attended by an untrained relative in both program and comparison villages. Nevertheless a significant change is apparent, with more women in program villages than in comparison villages being assisted by the TBAs. The delivery practices of the TBAs reveal the positive impact of their training in the "three cleans" (clean hands of the assistant, clean delivery surface and clean cord-cutting). The findings of this study indicate a significant level of unmet need for child spacing methods in all villages.^ The training and support of TBAs in the program villages yielded significant improvements in their delivery practices, and resulting outcomes for women and infants. However, potential exists for further community action. Capacities for self-development have not yet been directed toward an action plan encompassing other Safe Motherhood interventions, including access to family planning services and emergency obstetric care services. ^

Estrogen action during early mouse mammary gland development

Estrogens have been implicated in the normal and neoplastic development of the mammary gland. Although estradiol is essential for early mammary differentiation, its role in postnatal ductal morphogenesis is poorly defined. We have found that neonatal estradiol exposure promotes precocious ductal outgrowth and terminal end bud formation in 21 day-old female mice. In contrast to this precocious phenotype, day 21 estradiol-treated epithelium, transplanted into control host fatpads, grows more slowly than control epithelium. Western and immunohistochemical (IHC) analyses indicate that neonatally-estrogenized glands have significantly less total ER than controls at days 7 and 21, and significantly more stromal ER at day 35. Estrogen receptor α (ER) is present in the gland when treatment is initiated at day 1. We propose that the premature activation of ER by neonatal estradiol exposure, during this critical perinatal period, is a key factor in the alteration of mammary growth and ER expression. ^ To address the role of ER function in mammary morphogenesis, we have developed an in vitro system to study the effect of estradiol exposure in vivo. Keratin and ER-positive mammary epithelial cell lines from 7, 21 and 35 day-old oil or estradiol treated mice have been established. Cell lines derived from estradiol-treated mice grow significantly slower than cells from control glands. Although the level of ER expressed by each cell line is correlated to its rate of growth, epithelial growth in vitro is estradiol-independent and antiestrogen-insensitive. Estradiol-induced transcription from an ERE-reporter in transiently-transfected cell lines confirms the functionality of the ER detected by western and IHC. However, there are no differences in estradiol-stimulated transcription between cell lines. ^ In conclusion, neonatal estradiol treatment alters the pattern of ER expression in mammary epithelial and stromal cells in vivo, and the growth of mammary epithelial cells in vivo and in vitro. When grown outside of the estrogenized host, exposed epithelium grows more slowly than the control. Therefore, an extra-epithelial factor is necessary for enhanced epithelial growth. Our model, which couples an in vivo-in vitro approach, can be used in the future to identify factors involved in the period of early mammary outgrowth and carcinogen susceptibility. ^