Evolution of Diplodocid Sauropod dinosaurs with emphasis on specimens from Howe Ranch, Wyoming (USA)

Diplodocidae are among the best known sauropod dinosaurs. Several species were described in the late 1800s or early 1900s. Since then, numerous additional specimens were recovered in the USA, Tanzania, Portugal, as well as possibly Spain, England, and Asia. To date, the clade includes about 12 to 15 different species, some of them with questionable taxonomic status (e.g. ‘Diplodocus’ hayi or Dyslocosaurus polyonychius). However, intrageneric relationships of the multi-species, iconic genera Apatosaurus and Diplodocus are still poorly known. The way to resolve this issue is a specimen-based phylogenetic analysis, which was done for Apatosaurus, but is here performed for the first time for the entire clade of Diplodocidae. New material from different localities and stratigraphic levels on the Howe Ranch (Shell,Wyoming, USA) sheds additional light on the evolution of Diplodocidae. Three new specimens are described herein, considerably increasing our knowledge of the anatomy of the group. The new specimens (SMA 0004, SMA 0011, and SMA 0087) represent two, to possibly three new diplodocid species. They preserve material from all parts of the skeleton, including two nearly complete skulls, as well as fairly complete manus and pedes, material which is generally rare in diplodocids. Thereby, they considerably increase anatomical overlap between the sometimes fragmentary holotype specimens of the earlier described diplodocid species, allowing for significant results in a specimenbased phylogenetic analysis. Furthermore, clavicles and interclavicles are identified, the latter for the first time in dinosaurs. Their presence seems restricted to early sauropods, flagellicaudatans, and early Macronaria, and might thus be a retained plesiomorphy, with the loss of these bones being synapomorphic for Titanosauriformes and possibly Rebbachisauridae. The new material allows to test previous hypotheses of diplodocid phylogeny. In order to do so, any type specimen previously proposed to belong to Diplodocidae was included in the study, as are relatively complete referred specimens, in order to increase the degree of overlapping material. For specimens subsequently suggested to be non-diplodocid sauropods, their hypothesized sister taxa were included as outgroups. The current phylogenetic analysis thus includes 76 operational taxonomic units, 45 of which belong to Diplodocidae. The specimens were scored for 477 morphological characters, representing one of the most extensive phylogenetic analyses done within sauropod dinosaurs. The resulting cladogram recovers the classical arrangement of diplodocid relationships. Basing on a newly developed numerical approach to reduce subjectivity in the decision of specific or generic separation, species that have historically been included into well-known genera like Apatosaurus or Diplodocus, were detected to be actually generically different. Thereby, the famous genus Brontosaurus is resuscitated, and evidence further suggests that also Elosaurus parvus (previously referred to Apatosaurus) or ‘Diplodocus’ hayi represent unique genera. The study increases our knowledge about individual variation, and helps to decide how to score multi-species genera. Such a specimen-based phylogenetic analysis thus proves a valuable tool to validate historic species in sauropods, and in paleontology as a whole.

Characterization and regulation of a bacterial sugar phosphatase of the HAD superfamily, AraL, from Bacillus subtilis.

AraL from Bacillus subtilis is a member of the ubiquitous haloalkanoate dehalogenase, HAD, superfamily. The araL gene has been cloned, over-expressed in Escherichia coli and its product purified to homogeneity. The enzyme displays phosphatase activity, which is optimal at neutral pH (7.0) and 65 °C. Substrate screening and kinetic analysis showed AraL to have low specificity and catalytic activity towards several sugar phosphates, which are metabolic intermediates of the glycolytic and pentose phosphate pathways. Based on substrate specificity and gene context within the arabinose metabolic operon, a putative physiological role of AraL in detoxification of accidental accumulation of phosphorylated metabolites has been proposed. The ability of AraL to catabolise several related secondary metabolites requires regulation at the genetic level. Here, by site- directed mutagenesis, we show that AraL production is regulated by a structure in the translation initiation region of the mRNA, which most probably blocks access to the ribosome-binding site, preventing protein synthesis. Members of HAD subfamily IIA and IIB are characterised by a broad-range and overlapping specificity that anticipated the need for regulation at the genetic level. In this study we provide evidence for the existence of a genetic regulatory mechanism controlling AraL production.

Analysis of the interaction of polycyclic aromatic compounds in a model organism: integration of genotoxic and histopathological effects

Due to their toxicity, especially their carcinogenic potential, polycyclic aromatic hydrocarbons (PAHs) became priority pollutants in biomonitoring programmes and environmental policy, such as the European Water Framework Directive. The model substances tested in this study, namely benzo[b]fluoranthene (B[b]F), considered potentially carcinogenic to humans and an effector carcinogenic PAH to wildlife, and phenanthrene (Phe), deemed a non-carcinogenic PAH, are common PAHs in coastal waters, owning distinct properties reflected in different, albeit overlapping, mechanisms of toxicity. Still, as for similar PAHs, their interaction effects remain largely unknown. In order to study the genotoxic effects of caused by the interaction of carcinogenic and non-carcinogenic PAHs, and their relation to histopathological alterations, juvenile sea basses, Dicentrarchus labrax, a highly ecologically- and economically-relevant marine fish, were injected with different doses (5 and 10 μg.g-1 fish ww) of the two PAHs, isolated or in mixture, and incubated for 48 h. Individuals injected with B[b]F and the PAH mixture exhibited higher clastogenic/aneugenic effects and DNA strand breakage in blood cells, determined through the erythrocytic nuclear abnormalities (ENA) and Comet assays, respectively. Also, hepatic histopathological alterations were found in all animals, especially those injected with B[b]F and the PAH mixture, relating especially to inflammation. Still, Phe also exhibited genotoxic effects in sea bass, especially in higher doses, revealing a very significant acute effect that was accordant with the Microtox test performed undergone in parallel. Overall, sea bass was sensitive to B[b]F (a higher molecular weight PAH), likely due to efficient bioactivation of the pollutant (yielding genotoxic metabolites and reactive oxygen species), when compared to Phe, the latter revealing a more significant acute effect. The results indicate no significant additive effect between the substances, under the current experimental conditions. The present study highlights the importance of understanding PAH interactions in aquatic organisms, since they are usually present in the aquatic environment in complex mixtures.

Technology transition towards electric mobility - technology assessment as a tool for policy design

Paper presented at the Colloquium Gerpisa 2013, Paris (http://gerpisa.org/node/2085), Session n°: 19 New kinds of mobility: old and new business models

Grau de dependência e grau de ajuda em idosos em serviço de apoio domiciliário: perspectivas do idoso, do ajudante familiar e do fisioterapeuta

RESUMO:O envelhecimento e a preocupação com a dependência da população idosa é uma prioridade social. A permanência em casa onde se encontram todas as referências é importante em diversos aspectos, nomeadamente para uma maior independência. Quando a dependência surge, o Serviço de Apoio Domiciliário (SAD) é um dos serviços sociais a que se pode recorrer. Numa análise dos serviços de SAD nas vertentes de legislação, instituições, cuidadores e utentes, não são relacionados o grau de dependência e a atribuição da ajuda, o que parece pertinente para equacionar as necessidades das pessoas idosas em SAD. Nesse sentido, este estudo teve como objectivo verificar a percepção dos utentes de SAD do grau de dependência e do grau de ajuda na realização de taferas de auto-cuidado e mobilidade, avaliados pelo próprio idoso, pelo ajudante familiar e pelo fisioterapeuta. O presente estudo, transversal (Agosto 2013 – Abril 2014), de natureza quantitativa e qualitativa, e abordagem descritiva, observacional e correlacional, avaliou uma amostra de 51 idosos do SAD da Amadora, dos quais 80,4% eram mulheres, com média de idade de 82,4 anos (dp 6,81), principalmente viúvas, com escolaridade primária incompleta ou completa, orientada no tempo e no espaço, e com estado emocional por si considerado razoável. A investigadora, fisioterapeuta de formação, inquiriu idosos e ajudantes familiares que cuidavam dos mesmos, para recolher as suas opiniões sobre o grau de dependência e o respectivo grau de ajuda, observou o meio envolvente dos idosos, e recolheu comentários dos participantes. Os resultados obtidos apontam para um grau de dependência relacionado com a necessidade de meios e um grau de ajuda que indica ajuda de conveniência. Para a sua análise foram avaliadas tarefas de auto-cuidado e mobilidade, cuja relação foi comprovada estatisticamente. As opiniões dos intervenientes sobre a realização das tarefas e sobre o grau de dependência são parcialmente diferentes, sendo mais sobreponíveis em relação ao grau de ajuda necessária. A analise da correlação entre o grau de dependência e o grau de ajuda indica-nos que quanto maior o grau de dependência maior é o grau de ajuda. Ou seja, que a ajuda prestada nas situações de maior dependência era realmente necessária, mas também, que a ajuda prestada nas situações de menor dependência não era realmente necessária, sendo neste trabalho designada de ajuda de conveniência.------------ABSTRACT: Aging and concern about the dependence of the elderly population is a social priority. The ability to stay at home where all the references are, is important in several aspects, in particular for a greater independence. When the dependency arises, the Home Support Service (SAD in short for the portuguese "Serviço de Apoio Domiciliário") is a social service that can be appealed to. In an analysis of the SAD services in the areas of legislation, institutions, caregivers and users, the degree of dependence and aid allocation are not related, which seems relevant to equate the needs of older people in SAD. Thus, this study aimed to verify the perception of users of SAD towards the degree of dependency and the degree of help in the realization of self-care and mobility, valued by the user, the home support workers and the physiotherapist. This cross-sectional study (August 2013 - April 2014), of quantitative and qualitative nature, taking a descriptive approach, observational and correlational, assessed a sample of 51 elderly of SAD from Amadora, of which 80.4% were women, with an age average of 82.4 years (SD 6.81), mainly widows with incomplete or complete primary schooling, oriented in time and space, and emotional state by itself considered reasonable. The researcher, physiotherapist by training, inquired elderly and the home support workers who cared for them, to gather their opinions on the degree of dependency and the degree of the help provided, while observing the surrounding environment of the elderly. The results point to a degree related to the need for resources and a degree of help pointing towards a high level of convenience. For its analysis tasks of self-care and mobility were evaluated, whose relationship was confirmed statistically. The views of stakeholders on the realization of the tasks and on the degree of dependence are partly different, being more overlapping in the degree of help needed. The analysis of the correlation between the degree of dependency and the degree of help tells us that the greater the degree of dependence is, the greatest degree of help becomes. Which, that the help provided in situations of increased dependence was really needed, but also that the help provided in situations of reduced dependence was not really necessary, being designated in this work help of convenience.

Automated quantitative analysis of MCC-IMS spectra

Ion Mobility Spectrometry coupled with Multi Capillary Columns (MCC -IMS) is a fast analytical technique working at atmospheric pressure with high sensitivity and selectivity making it suitable for the analysis of complex biological matrices. MCC-IMS analysis generates its information through a 3D spectrum with peaks, corresponding to each of the substances detected, providing quantitative and qualitative information. Sometimes peaks of different substances overlap, making the quantification of substances present in the biological matrices a difficult process. In the present work we use peaks of isoprene and acetone as a model for this problem. These two volatile organic compounds (VOCs) that when detected by MCC-IMS produce two overlapping peaks. In this work it’s proposed an algorithm to identify and quantify these two peaks. This algorithm uses image processing techniques to treat the spectra and to detect the position of the peaks, and then fits the data to a custom model in order to separate the peaks. Once the peaks are separated it calculates the contribution of each peak to the data.

Evaluation of long term persistence of Diabetes Mellitus Type II treatment newly diagnosed patients in Lisbon and Tagus health administration region : a three year follow up study

RESUMO: Raional: A persistência à terapêutica é o tempo em qualquer antidiabético oral, desde o seu início até à descontinuação de todas as medicações ou até ao fim do período do estudo. Os objetivos deste estudo foi a análise da persistência à terapêutica no segundo e terceiro anos após início do tratamento em doentes adultos diagnosticados na região de Lisboa e Vale do Tejo e determinar o efeito de determinadas variáveis na persistência a longo prazo. Métodos: Um estudo retrospetivo não interventivo foi desenhado com base nos dados a obter do SIARS (prescrições e aquisições na farmácia) e Pordata. A persistência foi quantificada como a percentagem de doentes que continuam a adquirir pelo menos um antidiabético oral ao segundo e terceiro anos após a compra da primeira receita. A associação entre a persistência e o segundo e terceiro anos com cada uma das co-variáveis foi aferido pelo teste qui-quadrado e os odd ratios foram calculados com recurso a um modelo de regressão logística. Resultados: A persistência à terapêutica obtida foi de 80% e 62% para o segundo e terceiro anos após início da terapêutica. Odd ratios para primeiro e segundo ano: número de grupos farmacoterapêuticos diferentes (OR = 2.167, 1.807 – 2.598, p = 0.000 / OR = 1.863, 1.621 – 2.142, p = 0.000); idade (OR = 0.914, 0.772 – 1.081, p = 0.294 / OR = 0.875, 0.764 – 1.002, p = 0.054); sexo (OR = 1.163, 0.983 – 1.377, p = 0.079); número de diferentes prescritores (OR = 3.594, 3.030 – 4.262, p = 0.000 / OR = 2.167, 1.886 – 2.491, p = 0.000); instituição de prescrição (OR = 0.725, 0.698 – 0.753, p = 0.000 / OR = 0.683, 0.650 – 0.717, p = 0.000); grupo farmacoterapêutico (OR = 1.056, 1.043 – 1.069, p = 0.000 / OR = 1.077, 1.060 – 1.095, p = 0.000); relação com o médico (OR = 0.834, 0.816 – 0.852, p = 0.000 / OR = 0.799, 0.777 – 0.821, p = 0.000) e custo médio mensal por grupo farmacoterapêutico (OR = 0.954, 0.942 – 0.968, p = 0.000 / OR = 0.930, 0.914 – 0.947, p = 0.000). Conclusões: O valor da persistência à terapêutica no segundo ano é ligeiramente acima do que é mencionado na literatura e não existem dados para comparar os resultados do terceiro ano. Relativamente ao efeito das co-variáveis no segundo e terceiro anos após o início do tratamento, os resultados são sobreponíveis, sendo que o sexo não está associado à persistência ao terceiro ano.----------------------------------ABSTRACT: Background: Therapy persistence is the time on any antidiabetic medication, from initiation of therapy to discontinuation of all medications or the end of the study period. The aim of the study was to analyse the therapy persistence in the second and third years after treatment initiation in newly diagnosed adult patients in the Lisbon and Tagus Valley region and to determine the effect of several co-variables in the long term persistence. Methods: A retrospective non-interventional study based on SIARS data (drug prescriptions and acquisitions) and Pordata was designed. Persistence was quantified as the percentage of patients that continued to purchase at least one type of antidiabetic at year 2 and 3 after the date of first prescription acquisition. Association between persistence at second and third years with each of the other co-variables were verified by using the Chi-Square test and odds ratio were calculated using a regression logistic model. Results: Therapy persistence obtained was 80% and 62% for the second and third years after treatment initiation. Odd ratios for second and third years: number of different pharmacotherapeutic groups (OR = 2.167, 1.807 – 2.598, p = 0.000 / OR = 1.863, 1.621 – 2.142, p = 0.000); age (OR = 0.914, 0.772 – 1.081, p = 0.294 / OR = 0.875, 0.764 – 1.002, p = 0.054); gender (OR = 1.163, 0.983 – 1.377, p = 0.079); number of different prescribers (OR = 3.594, 3.030 – 4.262, p = 0.000 / OR = 2.167, 1.886 – 2.491, p = 0.000); institution of prescription (OR = 0.725, 0.698 – 0.753, p = 0.000 / OR = 0.683, 0.650 – 0.717, p = 0.000); pharmacotherapeutic group (OR = 1.056, 1.043 – 1.069, p = 0.000 / OR = 1.077, 1.060 – 1.095, p = 0.000); relationship with the physician (OR = 0.834, 0.816 – 0.852, p = 0.000 / OR = 0.799, 0.777 – 0.821, p = 0.000) and average cost per month and per pharmacotherapeutic group (OR = 0.954, 0.942 – 0.968, p = 0.000 / OR = 0.930, 0.914 – 0.947, p = 0.000). Conclusions: Second year therapy persistence value is slightly above of what is referenced in literature and no data was found to compare the third year value. Regarding the effect of the co-variables analysed at second and third years after treatment initiation, the results were overlapping with gender being not associated with persistence at the third year.

Geographical distribution of Trypanosoma cruzi in triatomine vectors in the State of Mato Grosso do Sul, Brazil

Introduction This work presents the initial findings of a molecular epidemiological investigation of Trypanosoma cruzi in triatomine insects in State of Mato Grosso do Sul. Methods A total of 511 triatomines from different regions of the state were examined. Deoxyribonucleic acid (DNA) was extracted from the intestinal contents of the insects using phenol-chloroform-isoamyl alcohol (25:24:1). Polymerase chain reaction (PCR) using primers 121/122 targeting DNA kinetoplast (kDNA) was then performed to identify T. cruzi, and positive samples were subjected to PCR using the primer pair TcSC5D-F/R followed by restriction fragment length polymorphism (RFLP) with the restriction enzymes SphI and HpaI (1 U/reaction), cloning and sequencing. Results One hundred samples were positive for T. cruzi, and three discrete typing units (DTUs) were identified (TcI, TcII, and TcBat). Triatoma sordida had the highest T. cruzi occurrence (83.3%), and DTUs were found in three samples: 58.3% of the samples were TcI, 33.3% were TcII and 8.3% were TcBat. There was a clear geographical distribution of the DTUs throughout the state, with TcI, TcII and TcBat located in the center, TcI located in the east, and TcII located in the west. Conclusions This study showed the occurrence of overlapping DTUs in State of Mato Grosso do Sul. The distributions of the DTUs were different, with TcI, TcII and TcBat in the center of the state, TcI predominantly in the east, and TcII in the west. Further studies may reveal a more defined mosaic distribution of DTUs in MS.

Congenital hyperthyroidism: autopsy report

We report the autopsy of a stillborn fetus with congenital hyperthyroidism born to a mother with untreated Graves' disease, whose cause of death was congestive heart failure. The major findings concerned the skull, thyroid, heart, and placenta. The cranial sutures were closed, with overlapping skull bones. The thyroid was increased in volume and had intense blood congestion. Histological examination showed hyperactive follicles. The heart was enlarged and softened, with dilated cavities and hemorrhagic suffusions in the epicardium. The placenta had infarctions that involved at least 20% of its surface, and the vessels of the umbilical cord were fully exposed due to a decrease in Wharton 's jelly. Hyperthyroidism was confirmed by the maternal clinical data, the fetal findings of exophthalmia, craniosynostosis, and goiter with signs of follicular hyperactivity. Craniosynostosis is caused by the anabolic action of thyroid hormones in bone formation during the initial stages of development. The delayed initiation of treatment in the present case contributed to the severity of fetal hyperthyroidism and consequent fetal death.

A wave of change: Labor market and social security impacts of the EU refugee inflow

The recent massive inflow of refugees to the European Union (EU) raises a number of unanswered questions on the economic impact of this phenomenon. To examine these questions, we constructed an overlapping-generations model that describes the evolution of the skill premium and of the welfare benefit level in relevant European countries, in the aftermath of an inflow of asylum-seekers. In our simulation, relative wages of skilled workers increase between 8% and 11% in the period of the inflow; their subsequent time path is dependent on the initial skill premium. The entry of migrants creates a fiscal surplus of about 8%, which can finance higher welfare benefits in the subsequent periods. These effects are weaker in a scenario where refugees do not fully integrate into the labor market.

Sexing the human skull through the mastoid process

OBJECTIVE: The purpose of this study was to evaluate the significance for sex determination of the measurement of the area formed by the xerographic projection of 3 craniometric points related to the mastoid process: the porion, asterion, and mastoidale points. METHOD: Sixty skulls, 30 male and 30 female, were analyzed. A xerographic copy of each side of the skull was obtained. On each xerographic copy, the craniometric points were marked to demarcate a triangle. The area (mm²) of the demarcated triangle for each side of the skull (right (D) and left (E) sides) was determined, and the total value of these measures (T) was calculated. RESULTS: Concerning the right area of the male and female skulls, 60% of the values overlapped; for the left area, 51.67% overlapped, and for the total area, 36.67% overlapped. The analysis of the differences between the sexes in the areas studied was significant for the 3 areas. Regarding the total area, which is the preferred measurement because of the asymmetry between the sides of the skull, the value of the mean was 1505.32 mm² for male skulls, which was greater than the maximum value obtained in the female skulls. The value of the mean for female skulls was 1221.24 mm², less than the minimum value obtained for the male skulls. CONCLUSIONS: This study demonstrates a significant result in the 3 studied areas, (D), (E), and (T). The total area values show less overlapping of values between the sexes, and therefore can be used for sexing human skulls. For the population studied, values of the total area that were greater than or equal to 1447.40 mm² belonged to male crania (95% confidence). Values for this area that were less than or equal to 1260.36 mm² belonged to female crania (95% confidence).

Terapia celular com células mesenquimais por via intracoronária: contributos do estudo invasivo da microcirculação coronária

RESUMO: Introdução - A utilização de células e das suas propriedades para o tratamento das doenças cardiovasculares, é uma promessa para o futuro e talvez a única forma de ultrapassar algumas das insuficiências das terapêuticas atuais. A via de entrega das células mais utilizada na investigação tem sido a intracoronária, ganhando a microcirculação especial relevância, por ser onde ocorre a primeira interação com o tecido nativo. As células estaminais mesenquimais (CEM) têm propriedades que as tornam particularmente aptas para a Terapia Celular, mas as suas dimensões, superiores ao diâmetro dos capilares, tem motivado controvérsia quanto à sua entrega intracoronária. A cardiologia de intervenção tem atualmente técnicas que permitem a avaliação em tempo real e in vivo do estado da microcirculação coronária. A determinação do índice da resistência da microcirculação (IRM) fornece informação sobre a circulação dos pequenos vasos, de forma independente da circulação coronária e do estado hemodinâmico, mas a aplicabilidade clínica deste conhecimento encontra-se ainda por definir. Objectivos Esclarecer o potencial do IRM no estudo dos efeitos do transplante de CEM por via intracoronária. População e Métodos . Estudo pré-clínico com modelo animal (suíno) desenvolvido em 3 fases. Na Primeira Fase foram utilizados 8 animais saudáveis para estudar e validar a técnica de determinação de estudo da microcirculação. Efetuou-se a determinação do IRM com duas doses diferentes de papaverina para a indução da resposta hiperémica máxima (5 e 10 mg) e após a disfunção da microcirculação com injeção intracoronária de microesferas de embozene com 40 μm de diâmetro. Na Segunda Fase foram utilizados 18 animais saudáveis, randomizados em grupo controlo e grupo recetor de 30 x 106 CEM por via intracoronária. Foram avaliados de forma cega o IRM, a pressão aórtica, o fluxo coronário epicárdico e a ocorrência de alterações electrocardiográficas. Na Terceira Fase foram utilizados 18 animais, com enfarte agudo do miocárdio provocado (EAM), randomizados em grupo controlo, grupo recetor de CEM expandidas de forma convencional e grupo recetor de CEM expandidas com metodologia inovadora e de menores dimensões. Foi realizada uma exploração da dose/efeito com infusão faseada de 10 x 106, 15 x 106 e 20 x 106 CEM, com determinação do IRM, da pressão aórtica, do fluxo coronário epicárdico e da ocorrência de alterações eletrocardiográficas. Quatro semanas após a entrega das células foi novamente avaliado o IRM e foi efetuado o estudo anatomopatológico dos animais na procura de evidência de neoangiogénese e de regeneração miocárdica, ou de um efeito positivo da resposta reparadora após o enfarte. Resultados Nas 3 fases todos os animais mantiveram estabilidade hemodinâmica e eletrocardiográfica, com exceção da elevação de ST de V1-V3 verificada após a injeção das microesferas. Na Primeira Fase as duas doses de papaverina induziram uma resposta hiperémica eficaz, sem tradução com significado na determinação do IRM (variação da pressão distal de - 11,4 ± 5 e de - 10,6± 5 mmHg com as doses de 5 e 10 mg respetivamente (p=0,5). Com a injeção das microesferas o IRM teve uma elevação média de 310 ± 190 %, para um valor médio de 41,3 ± 16 U (p = 0,001). Na Segunda Fase não houve diferenças significativas dos parâmetros hemodinâmicos, do fluxo epicárdico e da avaliação eletrocardiográfica entre os dois grupos. O IRM de base foi semelhante e após a infusão intracoronária observou-se uma elevação expressiva do IRM nos animais que receberam células em comparação com o grupo controlo (8,8 U ± 1 vs. 14,2 U ± 1,8, P=0,02) e quanto ao seu valor de base (aumento de 112%, p=0,008). Na terceira Fase não houve novamente diferenças significativas dos parâmetros hemodinâmicos, do fluxo epicárdico e da avaliação eletrocardiográfica entre os três grupos. Houve uma elevação do IRM nos animais que receberam células a partir da 2ª dose (72% nas células convencionai e 108% nas células inovadoras) e que se manteve com a 3ª dose (100% nas células convencionais e 88% nas inovadoras) com significado estatístico em comparação com o grupo controlo (p=0,034 com a 2ªdose e p=0,024 com a 3ª dose). Quatro semanas após a entrega das CEM observou-se a descida do IRM nos dois grupos que receberam células, para valores sobreponíveis aos do grupo controlo e aos valores pós-EAM. Na avaliação anatomopatológica e histológica dos corações explantados não houve diferenças entre os três grupos. Conclusões O IRM permite distinguir alterações da microcirculação coronária motivadas pela entrega intracoronária de CEM, na ausência de alterações de outros parâmetros clínicos da circulação coronária utilizados em tempo real. As alterações do IRM são progressivas e passíveis de avaliar o efeito/dose, embora não tenha sido possível determinar diferenças com os dois tipos de CEM. No nosso modelo a injeção intracoronária não se associou a evidência de efeito benéfico na reparação ou regeneração miocárdica após o EAM.---------------------------- ABSTRACT: ABSTRACT Introduction The use of cells for the treatment of cardiovascular disease is a promise for the future and perhaps the only option to overcome some of the shortcomings of current therapies. The strategy for the delivery of cells most often used in current research has been the intracoronary route and due to this microcirculation gains special relevance, mainly because it is the first interaction site of transplanted cells with the native tissue. Mesenchymal stem cells (MSC) have properties that make them suitable for Cell Therapy, but its dimensions, larger than the diameter of capillaries, have prompted controversy about the safety of intracoronary delivery. The interventional cardiology currently has techniques that allow for real-time and in vivo assessment of coronary microcirculation state. The determination of the index of microcirculatory resistance index (IMR) provides information about small vessels, independently of the coronary circulation and hemodynamic status, but the clinical applicability of this knowledge is yet to be defined. Objectives To clarify the potential use of IMR in the study of the effects of MSC through intracoronary transplantation. Population and Methods Preclinical study with swine model developed in three phases. In Phase One 8 healthy animals were used to study and validate the IMR assessment in our animal model. IMR was assessed with two different doses of papaverine for inducing the maximal hyperaemic response (5 and 10 mg) and microcirculation dysfunction was achieved after intracoronary injection with embozene microspheres with 40 μm in diameter. In Phase Two we randomized 18 healthy animals divided between the control group and the one receiving 30 x 106 MSC through an intracoronary infusion. There we blindly evaluated IMR, the aortic pressure, the epicardial coronary flow and the occurrence of ECG changes. In Phase Three we used 18 animals with a provoked acute myocardial infarction (AMI), randomized into a control group, a MSC expanded conventionally receiver group and a MSC expanded with an innovative methodology receiver group. There was a stepwise infusion with doses of 10 x 106, 15 x 106 and 20 x 106 MSC with determination of IMR, the aortic pressure, the epicardial coronary flow and occurrence of electrocardiographic abnormalities. Four weeks after cell delivery we again measured the IMR and proceeded with the pathological study of animals in the search for evidence of neoangiogenesis and myocardial regeneration, or a positive effect in the reparative response following the infarction. Results All animals remained hemodynamically stable and with no electrocardiographic abnormalities, except for the ST elevation in V1-V3 observed after injection of the microspheres. In Phase One the two doses of papaverine achieved an hyperemic and effective response without significant differences in IMR (variation of the distal pressure -11.4 ± 5 and -10.6 ± 5 mmHg with the doses of 5 and 10 mg respectively (p = 0.5). With the injection of the microspheres the IMR had an average increase of 310 ± 190% for an average value of 41.3 ± 16 U (p = 0.001). In the second phase there were no significant differences in hemodynamic parameters, epicardial flow and electrocardiographic assessment between the two groups. The baseline IMR was similar and after intracoronary infusion there was a significant increase in animals receiving cells compared with the control group (8.8 ± U 1 vs. 14.2 ± 1.8, p = 0.02) and with their baseline (112% increase, p = 0.008). In the third phase again there were no significant differences in hemodynamic parameters, the epicardial flow and electrocardiographic evaluation between the three groups. There was a significant increase in IMR in animals that received cells from the 2nd dose (72% in conventional cells and 108% in the innovative cells) that remained with the 3rd dose (100% in conventional cells and 88% in the innovative) with statistical significance compared with the control group (p = 0.034 with 2nd dose, p = 0.024 with 3rd dose). Four weeks after delivery of the MSC we observed the fall of the IMR in the two groups that received cells with values overlapping those of the control group. In pathological and histological evaluation of removed hearts there were no differences among the three groups. Conclusions The IMR allows for the differentiation of changes in coronary microcirculation motivated by intracoronary delivery of MSC in the absence of modification in other clinical parameters. IMR changes are progressive and enable the evaluation of the effect / dose, though it has not been possible to determine differences in the two types of MSC. In our model, intracoronary injection of MSC was not associated with evidence of repair or myocardial regeneration after AMI.

Release of insulin from PLGA-alginate dressing stimulates regenerative healing of burn wounds in rats

Burn wound healing involves a complex set of overlapping processes in an environment conducive to ischemia, inflammation, and infection costing $7.5 billion/year in the US alone, in addition to the morbidity and mortality that occur when the burns are extensive. We previously showed that insulin, when topically applied to skin excision wounds, accelerates re-epithelialization, and stimulates angiogenesis. More recently, we developed an alginate sponge dressing (ASD) containing insulin encapsulated in PLGA microparticles that provides a sustained release of bioactive insulin for >20days in a moist and protective environment. We hypothesized that insulin-containing ASD accelerates burn healing and stimulates a more regenerative, less scarring, healing. Using a heat-induced burn injury in rats, we show that burns treated with dressings containing 0.04mg insulin/cm2, every three days for 9 days, have faster closure, faster rate of disintegration of dead tissue, and decreased oxidative stress.In addition, in insulin-treated wounds the pattern of neutrophil inflammatory response suggests faster clearing of the burn dead tissue. We also observe faster resolution of the pro-inflammatory macrophages. We also found that insulin stimulates collagen deposition and maturation with the fibers organized more like a basket weave (normal skin) than aligned and crosslinked (scar tissue). In summary , application of ASD-containing insulin-loaded PLGA particles on burns every three days stimulates faster and more regenerative healing. These results suggest insulin as a potential therapeutic agent in burn healing and, because of its long history of safe use in humans, insulin could become one of the treatments of choice when repair and regeneration are critical for proper tissue function.

Human and murine clonal CD8+ T cell expansions arise during tuberculosis because of TCR selection

The immune system can recognize virtually any antigen, yet T cell responses against several pathogens, including Mycobacterium tuberculosis, are restricted to a limited number of immunodominant epitopes. The host factors that affect immunodominance are incompletely understood. Whether immunodominant epitopes elicit protective CD8+ T cell responses or instead act as decoys to subvert immunity and allow pathogens to establish chronic infection is unknown. Here we show that anatomically distinct human granulomas contain clonally expanded CD8+ T cells with overlapping T cell receptor (TCR) repertoires. Similarly, the murine CD8+ T cell response against M. tuberculosis is dominated by TB10.44-11-specific T cells with extreme TCRß bias. Using a retro genic model of TB10.44-11-specific CD8+ Tcells, we show that TCR dominance can arise because of competition between clonotypes driven by differences in affinity. Finally, we demonstrate that TB10.4-specific CD8+ T cells mediate protection against tuberculosis, which requires interferon-? production and TAP1-dependent antigen presentation in vivo. Our study of how immunodominance, biased TCR repertoires, and protection are inter-related, provides a new way to measure the quality of T cell immunity, which if applied to vaccine evaluation, could enhance our understanding of how to elicit protective T cell immunity.

“Fantasia Lusitana”, de João Canijo: O Portugal ficcional vs. o país real. O Estado Novo e a ‘portugalidade’. A construção da identidade

ISBN: 978-989-96858-3-3