Neutrophils and TNF: two key players in the immune response induced by "Leishmania major" infection

Summary Resolution of the inflammation is as important as its induction. In this thesis, we investigated the contributions of two prominent factors involved in inflammation, Tumour Necrosis Factor (TNF) and neutrophils. We studied their role in the resolution óf the inflammatory lesion induced by the infection with the protozoan parasite Leishmania major. In mice susceptible to infection with L. major, unhealing lesions are characterized by an elevated number and sustained presence of inflammatory neutrophils in the infected tissue, illustrating an acute inflammatory process. In contrast, mice from resistant strains, which resolve their lesions, can control the presence of neutrophils at the site of infection. Neutrophil persistence in the infected tissue may result from several events including an increased survival of neutrophils mediated by factors produced by the pathogen or the microenvironment. Following infection with L. major, the cellular composition of the inflammatory lesion differs significantly between susceptible and resistant mice and a higher proportion of macrophages is present in the lesions of resistant strains. In an attempt to clarify the factors involved in neutrophil persistence, we investigated the mechanisms modulating neutrophil cell death. We demonstrated that macrophages could induce neutrophil apoptosis in a process involving TNF. TNF is an essential cytokine with pro- and anti-inflammatory properties, which is expressed as a transmembrane protein that can be cleaved releasing the secreted form. Our data show the essential role of the transmembrane form of TNF (mTNF) in the induction of neutrophil apoptosis by macrophages, revealing macrophages and mTNF as important regulators of neutrophil apoptosis. TNF is critical in the resolution of the inflammatory lesion induced by L. major infection, and in L. major resistant strains its absence results in increased swelling of the lesions. We investigated the contribution of mTNF in the outcome of L. major infection. Our data demonstrate that following infection with L. major, mTNF is sufficient to support the resolution of the inflammatory lesion and optimal parasite killing. In addition, we show that the presence of mTNF is essential to induce neutrophil clearance in the infected tissue. While the persistence of neutrophils is deleterious for the host, we could demonstrate an early anti-inflammatory role of neutrophils. Altogether, this study demonstrates the importance of mTNF in the induction of neutrophil apoptosis, a process involved in the resolution of the inflammatory lesion induced by L. major infection. Résumé La résolution de l'inflammation est toute aussi importante que son initiation. Durant ce travail de thèse, nous avons étudié les contributions de deux facteurs importants impliqués dans l'inflammation, le TNF (Facteur Nécrosant des Tumeurs) et les neutrophiles, dans la résolution de la lésion inflammatoire induite par l'infection avec le parasite protozoaire Leishmania major. Chez les souris sensibles à l'infection avec L. major, des lésions importantes qui ne guérissent pas se développent ; celles-ci sont caractérisées par un nombre élevé et une présence soutenue de neutrophiles dans les tissus infectés, ce qui illustre un processus inflammatoire aigu. Au contraire, les souris résistantes à l'infection qui guérissent leurs lésions, sont capables de contrôler la présence des neutrophiles au site d'infection. La persistance des neutrophiles dans la lésion inflammatoire peut être la conséquence de plusieurs événements, dont une augmentation de la survie des neutrophiles induite par des facteurs produits par le pathogène ou le micro-environnement. Suite à l'infection avec L. major, la composition cellulaire de la lésion inflammatoire est significativement différente entre les souris sensibles et résistantes à l'infection, et une plus grande proportion de macrophages est présente dans les lésions des souris résistantes. Dans l'objectif de clarifier les facteurs impliqués dans la persistance des neutrophiles dans les tissus infectés par L. major, nous avons étudié les mécanismes de régulation de la mort des neutrophiles. Nous avons démontré que les macrophages pouvaient induire l'apoptose des neutrophiles dans un procédé impliquant le TNF. Le TNF est une cytokine aux propriétés pro- et anti-inflammatoires, exprimée sous une forme transmembranaire qui peut être clivée pour relâcher la forme sécrétée. Nos expériences illustrent le rôle essentiel de la forme transmembranaire du TNF (mTNF) dans l'induction de l'apoptose des neutrophiles par les macrophages. Lé TNF est une cytokine importante dans la résolution de la réaction inflammatoire induite par L. major, et chez les souris résistantes l'absence de TNF provoque des lésions inflammatoires plus importantes. Nous avons étudié la contribution du mTNF dans la résolution de l'infection avec L. major. Nos résultats démontrent que suite à une infection avec le parasite, la présence du mTNF est suffisante pour guérir la lésion inflammatoire et contrôler efficacement la réplication du parasite. De plus, le mTNF joue un rôle essentiel dans l'élimination des neutrophiles du tissu infecté. Alors que la persistance des neutrophiles est nocive pour l'hôte, nous avons montré que les neutrophiles avaient un rôle précoce anti-inflammatoire. En résumé, cette étude révèle l'importance du mTNF dans l'induction de l'apoptose des neutrophiles par les macrophages, un procédé impliqué dans la résolution de la lésion inflammatoire induite par l'infection avec L. major.

Human TRPV5 and TRPV6: key players in cadmium and zinc toxicity

TRPV5 and TRPV6 are two major calcium transport pathways in the human body maintaining calcium homeostasis. TRPV5 is mainly expressed in the distal convoluted and connecting tubule where it is the major, regulated pathway for calcium reabsorption. TRPV6 serves as an important calcium entry pathway in the duodenum and the placenta. Previously, we showed that human TRPV6 (hTRPV6) transports several heavy metals. In this study we tested whether human TRPV5 (hTRPV5) also transports cadmium and zinc, and whether hTRPV5 together with hTRPV6 are involved in cadmium and zinc toxicity. The hTRPV5 mRNA and protein were expressed in HEK293 cells transiently transfected with pTagRFP-C1-hTRPV5. The overexpression of the hTRPV5 protein at the plasma membrane was revealed by cell surface biotinylation and immunofluorescence techniques. We observed that both cadmium and zinc permeate hTRPV5 in ion imaging experiments using Fura-2 or Newport Green DCF. Our results were further confirmed using whole-cell patch clamp technique. Transient overexpression of hTRPV5 or hTRPV6 sensitized cells to cadmium and zinc. Toxicity curves of cadmium and zinc were also shifted in hTRPV6 expressing HEK293 cells clones. Our results suggest that TRPV5 and TRPV6 are crucial gates controlling cadmium and zinc levels in the human body especially under low calcium dietary conditions, when these channels are maximally upregulated.

The U7 snRNP and the hairpin binding protein: Key players in histone mRNA metabolism.

Animal replication-dependent histone mRNAs are subject to several post-transcriptional regulatory processes. Their non-polyadenylated 3' ends are formed preferentially during S phase by a unique nuclear cleavage event. This requires the base pairing between U7 snRNA and a histone spacer element 3' of the cleavage site. Cleavage occurs preferentially after adenosine, at a fixed distance from the hybrid region. A conserved RNA hairpin just upstream of the cleavage site is recognised by the hairpin binding protein (HBP) that acts as an auxiliary processing factor, stabilising the interaction of the histone pre-mRNA with the U7 snRNP. The interaction between HBP and the RNA hairpin is very stable and HBP is also found associated with histone mRNAs on polysomes. The hairpin and presumably, HBP are also required for nuclear export and translation of histone mRNA. Furthermore, histone mRNAs are selectively destabilised in the G2 phase or upon inhibition of DNA synthesis and this regulation is also associated with the hairpin. Recently, HBP-encoding cDNAs were isolated from various organisms. Human, mouse and Xenopus laevis HBPs are similar, while the Caenorhabditis elegans protein has significant homology to the others only in a central RNA binding domain.Copyright 1997 Academic Press Limited

OsteoMacs: Key players around bone biomaterials.

Osteal macrophages (OsteoMacs) are a special subtype of macrophage residing in bony tissues. Interesting findings from basic research have pointed to their vast and substantial roles in bone biology by demonstrating their key function in bone formation and remodeling. Despite these essential findings, much less information is available concerning their response to a variety of biomaterials used for bone regeneration with the majority of investigation primarily focused on their role during the foreign body reaction. With respect to biomaterials, it is well known that cells derived from the monocyte/macrophage lineage are one of the first cell types in contact with implanted biomaterials. Here they demonstrate extremely plastic phenotypes with the ability to differentiate towards classical M1 or M2 macrophages, or subsequently fuse into osteoclasts or multinucleated giant cells (MNGCs). These MNGCs have previously been characterized as foreign body giant cells and associated with biomaterial rejection, however more recently their phenotypes have been implicated with wound healing and tissue regeneration by studies demonstrating their expression of key M2 markers around biomaterials. With such contrasting hypotheses, it becomes essential to better understand their roles to improve the development of osteo-compatible and osteo-promotive biomaterials. This review article expresses the necessity to further study OsteoMacs and MNGCs to understand their function in bone biomaterial tissue integration including dental/orthopedic implants and bone grafting materials.

Key players of South Korea /

"May 1993."

Visualising collaboration via email: Finding the key players

Email is an important form of asynchronous communication. Visualizing analyses of email communication patterns during a collaborative activity help us better understand the nature of collaboration, and identify the key players. By analysing the contents of email communication and adding reflective comments on its perceived importance from the participants of a collaboration new information can be gleaned not immediately obvious in its original flat form. This paper outlines a proof-of-concept prototype collaborative email visualisation schema. Data from a collaboration case study is analysed and subsequently employed to construct a display of the relative impact of both key players and the types of email used.

Evolutionary History of Arabidopsis thaliana Aminoacyl-tRNA Synthetase Dual-Targeted Proteins

Aminoacyl-transfer RNA (tRNA) synthetases (aaRS) are key players in translation and act early in protein synthesis by mediating the attachment of amino acids to their cognate tRNA molecules. In plants, protein synthesis may occur in three subcellular compartments (cytosol, mitochondria, and chloroplasts), which requires multiple versions of the protein to be correctly delivered to its proper destination. The organellar aaRS are nuclear encoded and equipped with targeting information at the N-terminal sequence, which enables them to be specifically translocated to their final location. Most of the aaRS families present organellar proteins that are dual targeted to mitochondria and chloroplasts. Here, we examine the dual targeting behavior of aaRS from an evolutionary perspective. Our results show that Arabidopsis thaliana aaRS sequences are a result of a horizontal gene transfer event from bacteria. However, there is no evident bias indicating one single ancestor (Cyanobacteria or Proteobacteria). The dual-targeted aaRS phylogenetic relationship was characterized into two different categories (paralogs and homologs) depending on the state recovered for both dual-targeted and cytosolic proteins. Taken together, our results suggest that the dual-targeted condition is a gain-of-function derived from gene duplication. Selection may have maintained the original function in at least one of the copies as the additional copies diverged.

B, NK and NKT cells contribute to suppression of immunity by dendritic cells

Among the population of antigen presenting cells, dendritic cells (DCs) are considered the sentinels of the immune system. Besides activating naı¨ ve T cells, DC can directly activate naı¨ ve and memory B cells and are also able to regulate effectors of innate immunity such as NK cells and NKT cells. Increasing evidence indicates that DCs are not only decisive for T cell priming, but are also key players to maintain self-tolerance in vivo. Previous results in our lab have shown that DCs treated with a pharmacological NFkB inhibitor (BAY11–7082) confer suppression to a previously immune response. This suppression was IL-10 dependent and results from the induction of Ag specific CD4+ regulatory T cells. To elucidate the mechanism of suppression induced by administration of Bay treated DC, we used a model of infectious tolerance transfer from DC treated mice to primed recipient mice. Our results show that both CD4 + splenic cells and non T cells from animals injected with Bay treated DC, but not from untreated DC, were capable of transferring the suppression. Moreover, sorted B cells and NK cells could transfer antigenspecific infectious tolerance after administration of Bay treated DC. In addition, this suppressive effect could not be seen either in mice depleted of NK cells nor in NKT deficient mice. These observations highlight the role of several immune cells in the maintenance of tolerance, and impact on the design of immunotherapeutic suppression of autoimmune diseases in which NKT cells are deficient or defective, such as diabetes and lupus.

Infusing the use of seasonal climate forecasting into crop management practice in North East Australia using discussion support software

Seasonal climate forecasting offers potential for improving management of crop production risks in the cropping systems of NE Australia. But how is this capability best connected to management practice? Over the past decade, we have pursued participative systems approaches involving simulation-aided discussion with advisers and decision-makers. This has led to the development of discussion support software as a key vehicle for facilitating infusion of forecasting capability into practice. In this paper, we set out the basis of our approach, its implementation and preliminary evaluation. We outline the development of the discussion support software Whopper Cropper, which was designed for, and in close consultation with, public and private advisers. Whopper Cropper consists of a database of simulation output and a graphical user interface to generate analyses of risks associated with crop management options. The charts produced provide conversation pieces for advisers to use with their farmer clients in relation to the significant decisions they face. An example application, detail of the software development process and an initial survey of user needs are presented. We suggest that discussion support software is about moving beyond traditional notions of supply-driven decision support systems. Discussion support software is largely demand-driven and can compliment participatory action research programs by providing cost-effective general delivery of simulation-aided discussions about relevant management actions. The critical role of farm management advisers and dialogue among key players is highlighted. We argue that the discussion support concept, as exemplified by the software tool Whopper Cropper and the group processes surrounding it, provides an effective means to infuse innovations, like seasonal climate forecasting, into farming practice. Crown Copyright (C) 2002 Published by Elsevier Science Ltd. All rights reserved.

Psicologia das motivações ajurídicas do sentenciar : a emergência do saber em detrimento do poder

RESUMO:A pretensão deste trabalho reside na indagação sobre a eventual existência de um acervo de motivações ajurídicas que fundamentem as decisões judicias, erigindo como actor principal o Aplicador da Lei. Para tanto – recorreu-se à utilização de um instrumento construído pela autora, em 2005 – a Grelha de Observação das Motivações Ajurídicas do Sentenciar – destinado a captar aspectos considerados principais na comunicação dos protagonistas do processo de criminalização secundária, quer ao nível da comunicação verbal (C.V.) quer no concerne à comunicação não verbal (C.N.V.), verificando, depois, em presença das decisões, as razões justificativas evidenciadas. E que constituíram o suporte ajurídico de cada sentença. Assim, realizou-se um trabalho de observação, tendo a Grelha de Observação sido aplicada por licenciados em Psicologia, estudantes do 2º ciclo de Psicologia Forense e da Exclusão Social (Universidade Lusófona de Humanidades e Tecnologias), e alunos da Faculdade de Direito (Universidade Nova de Lisboa), que operaram em três tribunais criminais de Lisboa A amostra é constituída por 25 julgamentos, com um total de 64 personagens judiciais. Os resultados, ainda que meramente indicativos, demonstram uma tendência para a validade e consistência interna da grelha, indiciando também a verificação de motivações ajurídicas nas decisões judiciais. ABSTRACT: The intention of this work resides in the investigation of the eventual existence of an amount of non judicial motivations which support judicial decisions, where the law applicator arises as the main actor. For such - the use of an instrument built by the author in 2005 was resorted to - the Observation grid of the non judicial motivations of the sentencing - destined to capture aspects considered primary in the communication of the key players of the secondary criminalization process, in both the verbal communication level and in what concerns non verbal communication, later verifying, in the presence of the decisions, the evidenced justifiable reasons, which established the non judicial support of each sentence. Thus, an observational work was performed, having been applied the observation grid by Psychology graduates, students of the masters in Forensic Psychology and of Social Exclusion (Lusófona University of Humanities and Technology), as well as students of the Faculty of Law (New University of Lisbon), which operated in three criminal courts in Lisbon. The sample consists of 25 trials, with a total of 64 judicial characters. The results, even if only indicative, show a trend towards internal consistency and validity of the grid, also indicating the verification of non judicial motivations in judicial decisions.

O trabalho em equipas docentes e o desenvolvimento da Educação Física

RESUMO: O sucesso das respostas aos novos desafios, exigências e expectativas que, nos dias de hoje, recaem sobre a escola, parece depender da concepção das estratégias organizacionais que globalmente a escola define, tendo em conta as características do contexto e das populações que serve. Depende por isso do modo como os actores chave se organizam e trabalham. Os modos de entender o trabalho docente em equipa prenunciam formas de entender a acção educativa e de conduzir a aprendizagem dos alunos. Tomando como ponto de partida as nossas experiências profissionais, este trabalho constitui uma reflexão sobre as possibilidades de desenvolvimento da Educação Física, baseada na escola e sustentada na organização do trabalho dos seus professores. Elaborando uma síntese dos principais problemas do seu desenvolvimento discutem-se as possibilidades e as vantagens da organização do trabalho dos professores em equipas, num contexto organizacional marcado por avanços e retrocessos nos processos de descentralização e atribuição de autonomia às escolas. Enquadra-se a organização do trabalho docente no âmbito das culturas de escola. Revisita-se o conceito de trabalho em equipa, distinguindo-o da simples participação nas estruturas da escola, realçando a sua importância na construção e gestão de conhecimento profissional através do estabelecimento de parcerias sinérgicas, onde o desempenho de papéis de liderança tem uma enorme importância. ABSTRACT: Successful responses to the new challenges, demands and expectations that currently are ascribed to schools seem to depend on the design of organizational strategies that broadly define the school, taking into account the characteristics of its context and the population it serves. Therefore, their success depends on how the key players are organized and work. The ways of perceiving the teaching team work forecast the ways of understanding the educational activity and how to lead learning. Starting from our own professional experience, this study is a reflection on the school based development of physical education, sustained by the organization of their teachers’ work. On organizing a summary of the major problems found in the development of this subject, we discuss the feasibilities and the gains got from the organization of teachers' teamwork, in an organizational context marked by promises and setbacks in the processes of decentralization and schools autonomy. The organization of teaching is to be seen as part of the school culture. We review the concept of teamwork as distinct from mere participation in the structures of the school, and its importance in the construction and management of professional knowledge will be highlighted by the establishment of synergistic partnerships, in which the performance of leadership roles is of the greatest importance. RÉSUMÉ: Le succès des réponses aux nouveaux défis, les exigences et les attentes que dans ces jours, s’accomplissent sur l'école, semble dépendre de la conception des stratégies d'organisation qui définit de façon générale l'école, en tenant compte des caractéristiques du contexte et des populations qu'elle dessert. Tout cela dépend donc de la façon dont les acteurs principaux sont organisés et travaillent. Les façons de comprendre l'équipe pédagogique démontrent des formes de comprendre l'activité éducative et de conduire à l'apprentissage des élèves. En prenant comme point de départ nos expériences professionnelles, ce travail est une réflexion sur le potentiel de développement de l'Éducation Physique, dépendante de l'école et soutenue dans l'organisation du travail de leurs enseignants. En préparant une synthèse des principaux problèmes de leur développement, on discute les possibilités et les avantages de l'organisation du travail des enseignants dans les équipes, dans un contexte organisationnel marqué par des avancées et des reculs dans le processus de décentralisation et l'accomplissement de l'autonomie aux écoles. On encadre l'organisation du travail des enseignants dans le champ des cultures de l’école. On revoit le concept du travail en équipe, en le distinguant de la simple participation dans les structures de l'école, en soulignant son importance dans la construction et gestion des connaissances professionnelles à travers l'établissement de partenariats synergiques, où les rôles de leadership joue un rôle d'une grande importance.

Análise de risco em organizações: caso de estudo

Dissertação Final de Mestrado para obtenção do grau de Mestre em Engenharia Mecânica no perfil de Manutenção e Produção

A explicitação do raciocínio na resolução de problemas no 2º ciclo do ensino básico: contributos para o desenvolvimento das competências orais e escritas

Dissertação apresentada à Escola Superior de Educação de Lisboa para a obtenção de grau de Mestre em Didática da Língua Portuguesa no 1.º e 2.º Ciclos do Ensino Básico

O som da Sé Patriarcal de Lisboa no final do século XX: os cânticos do Pe. Manuel Luís e os acompanhamentos de Antoine Sibertin-Blanc

A novidade do cerimonial em língua vernácula, resultante do Concílio Vaticano II, levou a uma vasta actividade composicional em todos os países católicos. Em Portugal, e muito especialmente na região de Lisboa, um dos principais protagonistas desta acção foi o Pe. Manuel Luís (1924-81). Apesar de os seus cânticos (em particular, os Salmos Responsoriais) serem actualmente utilizados de norte a sul de Portugal, a actividade do Pe. Manuel Luís esteve centrada em Lisboa, nomeadamente, no contexto das celebrações realizadas na Sé Patriarcal, onde foi inevitável o contacto assíduo com Antoine Sibertin-Blanc (1930-2012), organista daquele templo desde 1965. As harmonizações de Sibertin-Blanc viriam a tornar-se, para os frequentadores da Catedral lisboeta, indissociáveis das melodias do Pe. Manuel Luís.