Generation and characterization of function-blocking anti-ectodysplasin A (EDA) monoclonal antibodies that induce ectodermal dysplasia.

Development of ectodermal appendages, such as hair, teeth, sweat glands, sebaceous glands, and mammary glands, requires the action of the TNF family ligand ectodysplasin A (EDA). Mutations of the X-linked EDA gene cause reduction or absence of many ectodermal appendages and have been identified as a cause of ectodermal dysplasia in humans, mice, dogs, and cattle. We have generated blocking antibodies, raised in Eda-deficient mice, against the conserved, receptor-binding domain of EDA. These antibodies recognize epitopes overlapping the receptor-binding site and prevent EDA from binding and activating EDAR at close to stoichiometric ratios in in vitro binding and activity assays. The antibodies block EDA1 and EDA2 of both mammalian and avian origin and, in vivo, suppress the ability of recombinant Fc-EDA1 to rescue ectodermal dysplasia in Eda-deficient Tabby mice. Moreover, administration of EDA blocking antibodies to pregnant wild type mice induced in developing wild type fetuses a marked and permanent ectodermal dysplasia. These function-blocking anti-EDA antibodies with wide cross-species reactivity will enable study of the developmental and postdevelopmental roles of EDA in a variety of organisms and open the route to therapeutic intervention in conditions in which EDA may be implicated.

Avaliação do website educacional em Primeiros Socorros

Avaliar a estrutura, a qualidade da informação e a navegabilidade do website em Primeiros Socorros. A avaliação foi realizada por profissionais de informática, da saúde e por estudantes, utilizando-se instrumentos específicos e validados. O método Kappa foi aplicado para avaliar a concordância das respostas e o coeficiente α de Cronbach , para avaliar a confiabilidade do instrumento. Nas respostas obtidas pelos profissionais, observou-se que não houve concordância das respostas dos profissionais de informática (0.047), indicando que a estrutura do website deve ser revisada. Na avaliação dos profissionais da saúde (-0.062), verificou-se que, apesar de não haver concordância, a qualidade da informação é adequada em razão dos escores positivos assinalados. Na avaliação da confiabilidade do instrumento de navegabilidade obteve-se α=0,974. Apesar de melhorias na estrutura do website serem indicadas, a qualidade da informação é boa e seu uso colaborou para o aprendizado dos estudantes.

Desenvolvimento de Aplicações Web e Multimédia: Desenvolvimento de uma Website

Este relatório descreve o estágio realizado pela autora, no âmbito da licenciatura em Tecnologias de Informação e Comunicação (TIC), da Universidade Pública de Cabo Verde (Uni-CV), realizado na empresa Tera C&D, que decorreu entre 2 Março a 31 Julho de 2009. O referido é o fruto de um projecto, que consistiu no desenvolvimento de um Website dinâmico, com o suporte de uma Base de Dados. Durante o período de estágio, a autora integrada numa equipa composta por quatro elementos, que actuam na área de Gestão de Sistemas Informáticos, Redes de Computadores, Desenvolvimento Web e Multimédia e Programação. Este trabalho foi desenvolvido sob o desígnio de trabalho colaborativo, em que a autora teve o apoio de várias pessoas. No decorrer deste período do estágio, em que foi desenvolvido o projecto, foram executadas um conjunto de actividades desde a análise dos documentos até a criação do site efectivamente. Paralelamente a essas actividades, decorreu uma formação, cujo grande objectivo foi orientar os formandos na execução das actividades e adquirir conhecimentos sobre Novas Tecnologias de Desenvolvimento Web. Considerando os objectivos referidos, o relatório ficará composto por uma parte introdutória, na qual se fará o enquadramento e também serão especificadas, as motivações associadas à escolha do tema. Seguidamente far-se-á a caracterização da empresa acolhedora, focando os aspectos tais como: a actividade principal da empresa, os objectivos os princípios e valores que conduzema mesma. Após fazer a caracterização da empresa acolhedora, far-se-á uma descrição sucinta das actividades realizadas no estágio. O desenvolvimento continuará com o relato de sobre a preparação do ambiente de desenvolvimento, para demonstrar detalhadamente a preparação deste ambiente, antes do inicio do projecto.Tecnologias e Ferramentas utilizadas será o ponto seguinte, no qual vão ser abordadas as diversas tecnologias e ferramentas utilizadas no desenvolvimento do referido projecto. O desenvolvimento segue-se com o tópico “Processo de Desenvolvimento Website”. Neste ponto será descrito pormenorizadamente as diferentes fases do processo de desenvolvimento de um Website, descrevendo com detalhes, o que significa cada fase e as actividades desenvolvidas em cada uma delas. Após descrever o Processo de Desenvolvimento, falar-se-á do projecto desenvolvido propriamente dito, expondo imagens que ilustram cada fase ou actividade realizada, caso for possível. Na parte conclusiva, falar-se-á da importância do desenvolvimento deste trabalho para a autora, do sentimento, das habilidades e competências adquiridas ou aperfeiçoadas.

Peptide modification or blocking of CD8, resulting in weak TCR signaling, can activate CTL for Fas- but not perforin-dependent cytotoxicity or cytokine production.

This study describes a form of partial agonism for a CD8+ CTL clone, S15, in which perforin-dependent killing and IFN-gamma production were lost but Fas (APO1 or CD95)-dependent cytotoxicity preserved. Cloned S15 CTL are H-2Kd restricted and specific for a photoreactive derivative of the Plasmodium berghei circumsporozoite peptide PbCS 252-260 (SYIPSAEKI). The presence of a photoactivatable group in the epitope permitted assessment of TCR-ligand binding by TCR photoaffinity labeling. Selective activation of Fas-dependent killing was observed for a peptide-derivative variant containing a modified photoreactive group. A similar functional response was obtained after binding of the wild-type peptide derivative upon blocking of CD8 participation in TCR-ligand binding. The epitope modification or blocking of CD8 resulted in an > or = 8-fold decrease in TCR-ligand binding. In both cases, phosphorylation of zeta-chain and ZAP-70, as well as calcium mobilization were reduced close to background levels, indicating that activation of Fas-dependent cytotoxicity required weaker TCR signaling than activation of perforin-dependent killing or IFN-gamma production. Consistent with this, we observed that depletion of the protein tyrosine kinase p56(lck) by preincubation of S15 CTL with herbimycin A severely impaired perforin- but not Fas-dependent cytotoxicity. Together with the observation that S15 CTL constitutively express Fas ligand, these results indicate that TCR signaling too weak to elicit perforin-dependent cytotoxicity or cytokine production can induce Fas-dependent cytotoxicity, possibly by translocation of preformed Fas ligand to the cell surface.

Organização e Publicação dos Termos do Website da ANACOM sob uma Perspetiva Linked Open Data

O propósito da Web Semântica é conseguir uma Web de dados totalmente ligada, isto é, numa perspetiva Linked Open Data. A Web Semântica deve garantir (estabelecendo padrões tecnológicos, vocabulários, linguagens lógicas, etc.) que os conteúdos publicados na Websejam inteligíveis quer por agentes humanos, quer por agentes máquina. Esta dissertação tem como objetivo responder à um problema delimitado, propondo uma solução no quadro da Web Semântica e suas tecnologias. Partindo-se de uma lista de termos em linguagem natural utilizados no Website da ANACOM (Autoridade Nacional de Comunicações), propomos uma organização de acordo com metodologias de construção de ontologias e vocabulários. Inspirámo-nos em duas metodologias, o Ontology Development 101 e o Process and Methodology for Core Vocabularies. O vocabulário controlado resultante, tem como base tecnológica o modelo de organização de conhecimento, recomendado pelo W3C (World Wide Web Consortium), o SKOS (Simple Knowledge Organization System). Trata-sede uma tecnologia standard da W3C desde 2009, utilizada na criação de tesauros,esquemas de classificação, taxonomias, glossários e outros tipos de vocabulários controlados. Como resultado da nossa intervenção, conseguimos organizar e codificar em SKOS, cerca de cinco centenas de termos identificados no Website da ANACOM. Para além da proposta do vocabulário controlado, passámos em revista às tecnologias e teorias que sustentam a temática da Web Semântica.

Highly efficient DNA incorporation of intratumourally injected [125I]iododeoxyuridine under thymidine synthesis blocking in human glioblastoma xenografts.

Intratumoural (i.t.) injection of radio-iododeoxyuridine (IdUrd), a thymidine (dThd) analogue, is envisaged for targeted Auger electron- or beta-radiation therapy of glioblastoma. Here, biodistribution of [(125)I]IdUrd was evaluated 5 hr after i.t. injection in subcutaneous human glioblastoma xenografts LN229 after different intravenous (i.v.) pretreatments with fluorodeoxyuridine (FdUrd). FdUrd is known to block de novo dThd synthesis, thus favouring DNA incorporation of radio-IdUrd. Results showed that pretreatment with 2 mg/kg FdUrd i.v. in 2 fractions 0.5 hr and 1 hr before injection of radio-IdUrd resulted in a mean tumour uptake of 19.8% of injected dose (% ID), representing 65.3% ID/g for tumours of approx. 0.35 g. Tumour uptake of radio-IdUrd in non-pretreated mice was only 4.1% ID. Very low uptake was observed in normal nondividing and dividing tissues with a maximum concentration of 2.9% ID/g measured in spleen. Pretreatment with a higher dose of FdUrd of 10 mg/kg prolonged the increased tumour uptake of radio-IdUrd up to 5 hr. A competition experiment was performed in FdUrd pretreated mice using i.t. co-injection of excess dThd that resulted in very low tumour retention of [(125)I]IdUrd. DNA isolation experiments showed that in the mean >95% of tumour (125)I activity was incorporated in DNA. In conclusion, these results show that close to 20% ID of radio-IdUrd injected i.t. was incorporated in tumour DNA after i.v. pretreatment with clinically relevant doses of FdUrd and that this approach may be further exploited for diffusion and therapy studies with Auger electron- and/or beta-radiation-emitting radio-IdUrd.

Note on the blocking flow shop problem

We present some results attained with different algorithms for the Fm|block|Cmax problem using as experimental data the well-known Taillard instances.

Allergologie: 1. Anti-TNFalpha: effets indésirables et implications pratiques [Management of adverse events induced by TNFalpha blocking agents].

TNFalpha blocking agents are effective and essential tools in the management of many inflammatory conditions including rheumatoid arthritis, spondylarthropathies and chronic inflammatory bowel disease. With time, some known side-effects have gained in importance and others have appeared. This article focuses on the potential risks of infection and autoimmunity induced by TNFalpha blocking agents and on the strategy to prevent and treat such adverse events.

Blocking Junctional Adhesion Molecule C Enhances Dendritic Cell Migration and Boosts the Immune Responses against Leishmania major.

The recruitment of dendritic cells to sites of infections and their migration to lymph nodes is fundamental for antigen processing and presentation to T cells. In the present study, we showed that antibody blockade of junctional adhesion molecule C (JAM-C) on endothelial cells removed JAM-C away from junctions and increased vascular permeability after L. major infection. This has multiple consequences on the output of the immune response. In resistant C57BL/6 and susceptible BALB/c mice, we found higher numbers of innate immune cells migrating from blood to the site of infection. The subsequent migration of dendritic cells (DCs) from the skin to the draining lymph node was also improved, thereby boosting the induction of the adaptive immune response. In C57BL/6 mice, JAM-C blockade after L. major injection led to an enhanced IFN-γ dominated T helper 1 (Th1) response with reduced skin lesions and parasite burden. Conversely, anti JAM-C treatment increased the IL-4-driven T helper 2 (Th2) response in BALB/c mice with disease exacerbation. Overall, our results show that JAM-C blockade can finely-tune the innate cell migration and accelerate the consequent immune response to L. major without changing the type of the T helper cell response.

Effects of blocking farm minkś feed access with open water

Selostus: Tarhatun minkin syömään pääsyn estäminen vesialtaalla

Blocking the B7-H4 pathway with novel recombinant antibodies enhances T cell-mediated antitumor responses.

B7-H4 inhibits T-cell activation and is widely expressed by solid neoplasms. We have recently demonstrated that the expression of B7-H4 on the surface of malignant cells in vivo is inducible, and that novel anti-B7-H4 recombinant antibodies can reverse the inhibition of tumor-specific T cells. Thus, antibodies targeting the B7-H4 pathways may extend the survival of cancer patients by restoring T cell-mediated antitumor responses.

Blocking hypoxia-induced autophagy in tumors restores cytotoxic T-cell activity and promotes regression.

The relationship between hypoxic stress, autophagy, and specific cell-mediated cytotoxicity remains unknown. This study shows that hypoxia-induced resistance of lung tumor to cytolytic T lymphocyte (CTL)-mediated lysis is associated with autophagy induction in target cells. In turn, this correlates with STAT3 phosphorylation on tyrosine 705 residue (pSTAT3) and HIF-1α accumulation. Inhibition of autophagy by siRNA targeting of either beclin1 or Atg5 resulted in impairment of pSTAT3 and restoration of hypoxic tumor cell susceptibility to CTL-mediated lysis. Furthermore, inhibition of pSTAT3 in hypoxic Atg5 or beclin1-targeted tumor cells was found to be associated with the inhibition Src kinase (pSrc). Autophagy-induced pSTAT3 and pSrc regulation seemed to involve the ubiquitin proteasome system and p62/SQSTM1. In vivo experiments using B16-F10 melanoma tumor cells indicated that depletion of beclin1 resulted in an inhibition of B16-F10 tumor growth and increased tumor apoptosis. Moreover, in vivo inhibition of autophagy by hydroxychloroquine in B16-F10 tumor-bearing mice and mice vaccinated with tyrosinase-related protein-2 peptide dramatically increased tumor growth inhibition. Collectively, this study establishes a novel functional link between hypoxia-induced autophagy and the regulation of antigen-specific T-cell lysis and points to a major role of autophagy in the control of in vivo tumor growth.

Combination of lentivector immunization and low-dose chemotherapy or PD-1/PD-L1 blocking primes self-reactive T cells and induces anti-tumor immunity.

In the last two decades, anti-cancer vaccines have yielded disappointing clinical results despite the fact that high numbers of self/tumor-specific T cells can be elicited in immunized patients. Understanding the reasons behind this lack of efficacy is critical in order to design better treatment regimes. Recombinant lentivectors (rLVs) have been successfully used to induce antigen-specific T cells to foreign or mutated tumor antigens. Here, we show that rLV expressing a murine nonmutated self/tumor antigen efficiently primes large numbers of self/tumor-specific CD8(+) T cells. In spite of the large number of tumor-specific T cells, however, no anti-tumor activity could be measured in a therapeutic setting, in mice vaccinated with rLV. Accumulating evidence shows that, in the presence of malignancies, inhibition of T-cell activity may predominate overstimulation. Analysis of tumor-infiltrating lymphocytes revealed that specific anti-tumor CD8(+) T cells fail to produce cytokines and express high levels of inhibitory receptors such as programmed death (PD)-1. Association of active immunization with chemotherapy or antibodies that block inhibitory pathways often leads to better anti-tumor effects. We show here that combining rLV vaccination with either cyclophosphamide or PD-1 and PD-L1 blocking antibodies enhances rLV vaccination efficacy and improves anti-tumor immunity.