In freshwater fish, such as the rainbow trout (Oncorhynchus mykiss), higher ion concentrations in the body fluids relative to the dilute surrounding environment lead to diffusive ion loss that is countered by active ion uptake. Active ion uptake is achieved via specialised cells in the gill epithelium known as ionocytes, with the species studied to date exhibiting multiple ionocyte subtypes with specific complements of ion transport proteins. To better understand the functions and responses of each ionocyte subtype, methods are needed to isolate specific ionocyte subtypes. This thesis developed a method to use fluorescence-activated cell sorting (FACS) to isolate the peanut lectin agglutinin-positive (PNA+) ionocyte subtype of the trout gill, which is posited to be a base-secreting cell that takes up Cl- ions. A suspension of gill cells dissociated using ethylenediaminetetraacetic acid (EDTA) was labelled with biotinylated PNA that was detected using streptavidin conjugated to a fluorophore, and subjected to FACS to yield a population of PNA+ ionocytes of high viability and purity. To validate the utility of the approach, it was used in a proof-of-principle experiment to evaluate transcript abundance of cytosolic carbonic anhydrase (CAc) in PNA+ ionocytes in trout that were subjected to metabolic alkalosis. This experiment revealed that the relative transcript abundance of CAc was significantly elevated in PNA+ ionocytes of alkalotic trout relative to that of control trout (P = 0.001; N = 7), a response that is consistent with the expected role of PNA+ ionocytes in compensation for systemic alkalosis.

No / There is substantial interest in detection of human behaviour that may reveal people with deliberate malicious intent, who are engaging in deceit. Technology exists that is able to detect changes in facial patterns of movement and thermal signatures on the face. However, there is data deficiency in the research community for further study. Therefore this project aims to overcome the data deficiency in psychology study and algorithms development. A within-subjects design experiment was conducted, using immigration as a scenario for investigate participants in control and experimental conditions. A random sample of 32 volunteers were recruited, their age group is within 18 - 33. The study design required participants to answer questions on two topics, one as themselves and one as a predefined character. Data regarding visible and thermal images of facial movement and behaviour were collected. A rich FACS-coded database with high quality thermal images was established. Finally, recommendations for development and subsequent implementation of the facial analysis technique were made.

No description available.

Yes / The objective of every wind energy producer is to reduce operational costs associated to the production as a way to increase profits. One other issue that must be looked carefully is the equipment maintenance. Increase the availability of wind turbines by reducing the downtime associated to failures is a good strategy to achieve the main goal of increase profits. As a way to help in the definition of the best maintenance strategies, condition monitoring systems (CMS) have an important role to play. Informatics tools to make the condition monitoring of the wind turbines were developed and are now being installed as a way to help producers reducing the operational costs. There are a lot of developed systems to do the monitoring of a wind turbine or the whole wind park, in this paper will be made an overview of the most important systems.

Facial expressions convey non-verbal cues. Humans use facial expressions to show emotions, which play an important role in interpersonal relations and can be of use in many applications involving psychology, human-computer interaction, health care, e-commerce, and many others. Although humans recognize facial expressions in a scene with little or no effort, reliable expression recognition by machine is still a challenging problem. Automatic facial expression recognition (FER) has several related problems: face detection, face representation, extraction of the facial expression information, and classification of expressions, particularly under conditions of input data variability such as illumination and pose variation. A system that performs these operations accurately and in realtime would be a major step forward in achieving a human-like interaction between the man and machine. This document introduces novel approaches for the automatic recognition of the basic facial expressions, namely, happiness, surprise, sadness, fear, disgust, anger, and neutral using relatively low-resolution noisy sensor such as the Microsoft Kinect. Such sensors are capable of fast data collection, but the low-resolution noisy data present unique challenges when identifying subtle changes in appearance. This dissertation will present the work that has been done to address these challenges and the corresponding results. The lack of Kinect-based FER datasets motivated this work to build two Kinect-based RGBD+time FER datasets that include facial expressions of adults and children. To the best of our knowledge, they are the first FER-oriented datasets that include children. Availability of children data is important for research focused on children (e.g., psychology studies on facial expressions of children with autism), and also allows researchers to do deeper studies on automatic FER by analyzing possible differences between data coming from adults and children. The key contributions of this dissertation are both empirical and theoretical. The empirical contributions include the design and successful test of three FER systems that outperform existing FER systems either when tested on public datasets or in realtime. One proposed approach automatically tunes itself to the given 3D data by identifying the best distance metric that maximizes the system accuracy. Compared to traditional approaches where a fixed distance metric is employed for all classes, the presented adaptive approach had better recognition accuracy especially in non-frontal poses. Another proposed system combines high dimensional feature vectors extracted from 2D and 3D modalities via a novel fusion technique. This system achieved 80% accuracy which outperforms the state of the art on the public VT-KFER dataset by more than 13%. The third proposed system has been designed and successfully tested to recognize the six basic expressions plus neutral in realtime using only 3D data captured by the Kinect. When tested on a public FER dataset, it achieved 67% (7% higher than other 3D-based FER systems) in multi-class mode and 89% (i.e., 9% higher than the state of the art) in binary mode. When the system was tested in realtime on 20 children, it achieved over 73% on a reduced set of expressions. To the best of our knowledge, this is the first known system that has been tested on relatively large dataset of children in realtime. The theoretical contributions include 1) the development of a novel feature selection approach that ranks the features based on their class separability, and 2) the development of the Dual Kernel Discriminant Analysis (DKDA) feature fusion algorithm. This later approach addresses the problem of fusing high dimensional noisy data that are highly nonlinear distributed. / PHD

Este trabalho avalia a influência das emoções humanas expressas pela mímica da face na tomada de decisão de sistemas computacionais, com o objetivo de melhorar a experiência do usuário. Para isso, foram desenvolvidos três módulos: o primeiro trata-se de um sistema de computação assistiva - uma prancha de comunicação alternativa e ampliada em versão digital. O segundo módulo, aqui denominado Módulo Afetivo, trata-se de um sistema de computação afetiva que, por meio de Visão Computacional, capta a mímica da face do usuário e classifica seu estado emocional. Este segundo módulo foi implementado em duas etapas, as duas inspiradas no Sistema de Codificação de Ações Faciais (FACS), que identifica expressões faciais com base no sistema cognitivo humano. Na primeira etapa, o Módulo Afetivo realiza a inferência dos estados emocionais básicos: felicidade, surpresa, raiva, medo, tristeza, aversão e, ainda, o estado neutro. Segundo a maioria dos pesquisadores da área, as emoções básicas são inatas e universais, o que torna o módulo afetivo generalizável a qualquer população. Os testes realizados com o modelo proposto apresentaram resultados 10,9% acima dos resultados que usam metodologias semelhantes. Também foram realizadas análises de emoções espontâneas, e os resultados computacionais aproximam-se da taxa de acerto dos seres humanos. Na segunda etapa do desenvolvimento do Módulo Afetivo, o objetivo foi identificar expressões faciais que refletem a insatisfação ou a dificuldade de uma pessoa durante o uso de sistemas computacionais. Assim, o primeiro modelo do Módulo Afetivo foi ajustado para este fim. Por fim, foi desenvolvido um Módulo de Tomada de Decisão que recebe informações do Módulo Afetivo e faz intervenções no Sistema Computacional. Parâmetros como tamanho do ícone, arraste convertido em clique e velocidade de varredura são alterados em tempo real pelo Módulo de Tomada de Decisão no sistema computacional assistivo, de acordo com as informações geradas pelo Módulo Afetivo. Como o Módulo Afetivo não possui uma etapa de treinamento para inferência do estado emocional, foi proposto um algoritmo de face neutra para resolver o problema da inicialização com faces contendo emoções. Também foi proposto, neste trabalho, a divisão dos sinais faciais rápidos entre sinais de linha base (tique e outros ruídos na movimentação da face que não se tratam de sinais emocionais) e sinais emocionais. Os resultados dos Estudos de Caso realizados com os alunos da APAE de Presidente Prudente demonstraram que é possível melhorar a experiência do usuário, configurando um sistema computacional com informações emocionais expressas pela mímica da face. / The influence of human emotions expressed by facial mimics in decision-taking of computer systems is analyzed to improve user´s experience. Three modules were developed: the first module comprises a system of assistive computation - a digital alternative and amplified communication board. The second module, called the Affective Module, is a system of affective computation which, through a Computational Vision, retrieves the user\'s facial mimic and classifies their emotional state. The second module was implemented in two stages derived from the Facial Action Codification System (FACS) which identifies facial expressions based on the human cognitive system. In the first stage, the Affective Module infers the basic emotional stages, namely, happiness, surprise, anger, fear, sadness, disgust, and the neutral state. According to most researchers, basic emotions are innate and universal. Thus, the affective module is common to any population. Tests undertaken with the suggested model provided results which were 10.9% above those that employ similar methodologies. Spontaneous emotions were also undertaken and computer results were close to human score rates. The second stage for the development of the Affective Module, facial expressions that reflect dissatisfaction or difficulties during the use of computer systems were identified. The first model of the Affective Module was adjusted to this end. A Decision-taking Module which receives information from the Affective Module and intervenes in the Computer System was developed. Parameters such as icon size, draw transformed into a click, and scanning speed are changed in real time by the Decision-taking Module in the assistive computer system, following information by the Affective Module. Since the Affective Module does not have a training stage to infer the emotional stage, a neutral face algorithm has been suggested to solve the problem of initialing with emotion-featuring faces. Current assay also suggests the distinction between quick facial signals among the base signs (a click or any other sound in the face movement which is not an emotional sign) and emotional signs. Results from Case Studies with APAE children in Presidente Prudente SP Brazil showed that user´s experience may be improved through a computer system with emotional information expressed by facial mimics.

Die molekularbiologische Untersuchung der Wechselwirkungen zwischen Pathogenen und ihren Wirtszellen ist ein wertvoller Ansatz zur Erschließung bakterieller Pathogenitätsmechanismen und hilft, unser ständig wachsendes Wissen über fundamentale Prozesse in eukaryotischen Zellen zu erweitern. Das Gram-negative Bakterium Salmonella Typhimurium ist ein gängiger Modellorganismus, um den intrazellulären Lebensstil bakterieller Pathogene und deren Einfluss auf Wirtszellprozesse zu erforschen. In der vorliegenden Arbeit wurde ein FACS-basierter Hochdurchsatz RNA Interferenz Screen etabliert und durchgeführt, um Wirtszellfaktoren zu entschlüsseln, welche in die intrazelluläre Replikation von Salmonella Typhimurium in humanen Epithelzellen involviert sind. Ein Salmonellen-Stamm, der zwei fluoreszierende Reporterproteine exprimiert, wurde konstruiert, um die bakterielle Replikation und die metabolische Aktivität in infizierten Wirtszellen zu detektieren. Der Einsatz einer humanen Kinase-Bibliothek lieferte 48 potentielle Kandidatengene, von denen 15 in einer anschließenden Validierung als relevante Faktoren identifiziert werden konnten. Die Mitogen-aktivierte Protein Kinase MKK7, deren Depletion eine verminderte bakterielle Replikation zur Folge hatte, wurde für eine weitergehende funktionelle Charakterisierung ausgewählt. Es zeigte sich, dass reduzierte MKK7 Proteinmengen eine Verringerung des Proteins zytosolische Phospholipase A2 (cPLA2) durch eine transkriptionelle Regulation zur Folge hatten. Die Bedeutung von cPLA2 für die bakterielle Infektion wurde durch die Salmonellen-induzierte, dauerhafte Phosphorylierung des Faktors deutlich und konnte durch Replikationsvergleiche in cPLA2-depletierten und nicht-depletierten Zellen bestätigt werden. Mikroskopische Ergebnisse deuteten darauf hin, dass die Phospholipase für den fehlerfreien Aufbau von Salmonellen-induzierten Filamenten notwendig ist, welche unerlässlich für die Salmonellenreplikation in Epithelzellen sind. / The study of pathogen-host cell interactions on the molecular level is a valuable tool to reveal bacterial pathogenicity mechanisms and, moreover, contributes to our increasing knowledge of fundamental cellular processes of eukaryotic cells. The Gram negative bacterium Salmonella Typhimurium is a well established model organism to investigate the intracellular lifestyle of bacterial pathogens and their modulation of host cell processes. In this work, a FACS-based high-throughput RNA interference screen was established and performed to elucidate host cell factors involved in the intracellular replication of Salmonella Typhimurium. A Salmonella strain expressing two fluorescent reporter constructs was generated which allowed for monitoring the bacterial replication and metabolic activity within infected cells. A human kinome-wide siRNA library was screened and 48 candidates were chosen for further validation. Among these, 15 host cell genes were identified to influence Salmonella intracellular replication. The mitogen activated protein kinase MKK7, whose depletion caused a decrease in bacterial replication, was selected for a more profound functional characterization. It could be demonstrated that the knock down of MKK7 caused a decrease in phospholipase A2 (cPLA2) protein levels due to a transcriptional regulation. A role for cPLA2 during the bacterial intracellular lifestyle was implicated by the finding that Salmonella induced a permanent phosphorylation of the phospholipase. The necessity of cPLA2 was confirmed with replication assays in cPLA2 depleted cells using siRNA and shRNA mediated knock down strategies. Microscopic experiments indicated that the phospholipase A2 is involved in the accurate generation of Salmonella-induced filaments, structures that were reported to be indispensable for replication.

A cavidade peritoneal de camundongos abriga uma variedade de células do sistema imune. Inicialmente, devido as limitações metodológicas, acreditava-se que, aproximadamente, 90% das células peritoneais era representada por macrófagos. Em seguida, graças aos extensos estudos com células peritoneais, observou-se que, além dos macrófagos, o peritônio abrigava muitos linfócitos B, principalmente do subtipo B-1. Utilizando metodologias contemporâneas de FACS - citometria de fluxo, este trabalho mostra que, aproximadamente, 30% das células peritoneais são macrófagos, 55% são linfócitos B-1, dos quais 40% pertencem ao subtipo B-1a e 15% ao subtipo B-1b. Os 15%-20% restantes representam outros subtipos celulares, como linfócitos T, linfócitos B-2, linfócitos NK, eosinófilos, além da presença de outras populações de células que não foram possíveis de ser identificadas com os marcadores de superfície utilizados neste trabalho. Em contraste com a literatura, nossos estudos mostraram que os macrófagos da cavidade peritoneal de camundongos representam uma população heterogênea. Por meio da co-expressão de CD11b e F4/80, foi possível descrever dois subtipos funcionais de macrófagos, denominados aqui de PM-1 (CD11bhigh, F4/80high) e PM-2 (F4/8010W, CD11bint). As células PM-2 encontram-se em menor abundância (-10% do total de macrófagos) no peritônio não estimulado e, diferentemente de células PM-1, espraiam-se de forma alongada (bipolar) quando colocadas em cultura. Curiosamente, PM-2 são os únicos macrófagos peritoneais que expressam MHC-II e, ainda, 30% dessa população expressa CD11c, o marcador típico de células dendríticas. Já as células PM-1 possuem morfologia arredondada, estão em abundância no peritônio e produzem altas doses de NO após estímulo com LPS. Ambas as células são capazes de fagocitar bactérias in vivo, no entanto, as células PM-2 parecem mais eficientes, sendo capazes de internalizar quantidades maiores de bactérias. Após infecção in vivo, o número absoluto e a porcentagem de PM-2 aumenta muito no peritônio, chegando a representar metade dos fagócitos da cavidade peritoneal. Os macrófagos PM-1 e PM-2 parecem não representar monócitos já que não compartilham do fenótipo de monócitos presentes no sangue periférico. Deve-se ressaltar que, além das células PM-1 e PM-2, outras subpopulações do peritônio também expressam CD11b e F4/80. Com base na literatura atual, acredita-se que todas as células B-1 do peritônio expressam CD11b, uma molécula que é co-expressa com CD18 para formar o receptor de complemento e a molécula de adesão Mac-1. Entretanto, os estudos, apresentados neste trabalho, mostram que metade de cada subtipo de B-1 (B-1a e B-1b) não expressa CD11b. As células B-1 CD11b+ são maiores, mais granulosas e expressam quantidades maiores de IgM de superfície. As células CD11b+ possuem uma curiosa tendência de se juntar e formar duplas (doublets) fortemente ligadas entre si que estão presentes em abundância no peritônio de camundongos adultos. Além de ligarem entre si, as células B-1 podem formar duplas com macrófagos PM-1 e PM-2. Durante a ontogenia, as células CD11b- surgem primeiro no peritônio e são as progenitoras das células B-1 CD11b+. Após estímulo inflamatório (LPS í. v.) os linfócitos B-1 CD11b+ migram do peritônio para o baço, onde proliferam e transformam-se em células secretoras de anticorpos (plasmócitos). No peritônio, as células B-1 não são capazes de se diferenciar em plasmócitos (células CD138high). Os resultados apresentados aqui mostraram que, diferentemente dos macrófagos PM-2, os PM-1 são os responsáveis por inibir a formação de células secretoras de anticorpos derivadas de B-1. Em suma, nossos dados sugerem que, mediante estímulo inflamatório, as células B-1 maduras (CD11b+) migram do peritônio para o baço, afastandose de macrófagos PM-1. Já no baço, as células B-1 encontram um micro-ambiente favorável para se proliferarem e diferenciarem-se em plasmócitos, secretando a maioria dos anticorpos naturais vistos logo nas primeiras horas pós-estímulo. / Mouse peritoneal cavity (PerC) represents the source for a variety of cellular subsets of the immune system. In early studies, marred by Iimited methodological tools, it was thought that macrophages comprise roughly 90% of total PerC cells. Shortly thereafter, it was recognized that, beside macrophages, the mouse peritoneal cavity shelters large amount of B-lymphocytes, specially the B-1 subset. In essence, by applying contemporary technology, studies presented here show that, roughly 30% of PerC cells comprise macrophages, 55% comprise B-1 cells, which 40% represents the B-1 a subset and roughly 15% are B-1b cells. The remaining 15%-20% of PerC cells comprise a variety of known immune cells, including B-2, T, NK and eosinophils. In addition, there were some other cellular subsets that could not be identified in these studies, probably due to limited cell surface molecules analyzed. Surprisingly, the data presented here show that peritoneal macrophages represent a highly heterogeneous population. Based on the co-expression of both CD11b and F4/80, we have identified two functionally distinct subsets of peritoneal macrophages, named here as PM-1 (CD11bhigh, F480high) and PM-2 (CD11bint, F4/80low). PM-2 are less abundant (-10% within total macrophages) in the PerC of unstimulated mouse, however, unlike PM-1, these cells are able to spread in spindle (bipolar) morphology upon sorting and in vitro culture. Curiously, PM-2 is the only PerC macrophage that expresses surface MHC-II. In addition, one third of PM-2 population expresses CD11c, a universal marker for dendritic cells. In turn, PM-1 cells are round shaped cells, represent the majority of PerC macrophages (-90%) and secrete high amounts of NO upon LPS stimulation. In vivo phagocytosis assay revealed that both cells could internalize bacteria, however, PM-2 cells showed to be more efficient, in that it was able to phagocyte higher amounts of bacteria when compared to PM-1. After i.p. in vivo stimulation, the absolute number and percentage of PM-2 cells increase drastically in the peritoneum, reaching almost half of the total PerC macrophages. Importantly, PM-1 and PM-2 macrophages seem not to represent PerC monocytes since it does not share any of the phenotype expressed by blood monocytes. Interestingly, in addition to PM-1 and PM-2, some other cellular subsets in the PerC, such as eosinophils, are able to express CD11b and F4/80. PerC B-1 cells have long been known to express CD11b, which is co-expressed with CD18 to form the Mac-1/CR3 complement receptor and adhesion molecule. However, although all PerC B-1 cells are commonly believed to express CD11b, we show here that nearly half of the cells in each of the PerC B-1 subsets (B-1a and B-1b) do not express this surface receptor. The CD11b+ cells in each B-1 subset are larger, more granular and express higher levels of surface IgM than the CD11b- B-1 cells. Surprisingly, the CD11b+ B-1 subset has the curious tendency to initiate the formation of tightly associated doublets that are present at high frequency in adult PerC. In addition to binding to each other, B-1 cells can also form doublets with PM-1 and PM-2. During ontogeny, CD11b- B-1 cells arise earlier in the mouse PerC and are the progenitors of CD11b+ B-1 cells. Upon LPS stimulation, PerC CD11b+ B-1 cells migrate to the spleen where they proliferate and differentiate into antibody-secreting cells (plasma cells). Within the PerC, B-1 cells are not able to differentiate into plasma cells (CD138high cells). The data shown in here reveal that, unlike PM-2, PM-1 macrophages are the cellular subset responsible for inhibiting the formation of B-1-derived antibody-secreting cells. Altogether, our data suggest that, upon inflammatory stimuli, mature CD11b+ B-1 cells migrate from the PerC to the spleen, avoiding the inhibitory effect of PM-1 cells. In the spleen, B-1 cells find an appropriate environment to proliferate and terminally differentiate into antibody-secreting cells, thus, secreting the majority of immunoglobulin seen in few hours after in vivo stimulation.

A cavidade peritoneal de camundongos abriga uma variedade de células do sistema imune. Inicialmente, devido as limitações metodológicas, acreditava-se que, aproximadamente, 90% das células peritoneais era representada por macrófagos. Em seguida, graças aos extensos estudos com células peritoneais, observou-se que, além dos macrófagos, o peritônio abrigava muitos linfócitos B, principalmente do subtipo B-1. Utilizando metodologias contemporâneas de FACS - citometria de fluxo, este trabalho mostra que, aproximadamente, 30% das células peritoneais são macrófagos, 55% são linfócitos B-1, dos quais 40% pertencem ao subtipo B-1a e 15% ao subtipo B-1b. Os 15%-20% restantes representam outros subtipos celulares, como linfócitos T, linfócitos B-2, linfócitos NK, eosinófilos, além da presença de outras populações de células que não foram possíveis de ser identificadas com os marcadores de superfície utilizados neste trabalho. Em contraste com a literatura, nossos estudos mostraram que os macrófagos da cavidade peritoneal de camundongos representam uma população heterogênea. Por meio da co-expressão de CD11b e F4/80, foi possível descrever dois subtipos funcionais de macrófagos, denominados aqui de PM-1 (CD11bhigh, F4/80high) e PM-2 (F4/8010W, CD11bint). As células PM-2 encontram-se em menor abundância (-10% do total de macrófagos) no peritônio não estimulado e, diferentemente de células PM-1, espraiam-se de forma alongada (bipolar) quando colocadas em cultura. Curiosamente, PM-2 são os únicos macrófagos peritoneais que expressam MHC-II e, ainda, 30% dessa população expressa CD11c, o marcador típico de células dendríticas. Já as células PM-1 possuem morfologia arredondada, estão em abundância no peritônio e produzem altas doses de NO após estímulo com LPS. Ambas as células são capazes de fagocitar bactérias in vivo, no entanto, as células PM-2 parecem mais eficientes, sendo capazes de internalizar quantidades maiores de bactérias. Após infecção in vivo, o número absoluto e a porcentagem de PM-2 aumenta muito no peritônio, chegando a representar metade dos fagócitos da cavidade peritoneal. Os macrófagos PM-1 e PM-2 parecem não representar monócitos já que não compartilham do fenótipo de monócitos presentes no sangue periférico. Deve-se ressaltar que, além das células PM-1 e PM-2, outras subpopulações do peritônio também expressam CD11b e F4/80. Com base na literatura atual, acredita-se que todas as células B-1 do peritônio expressam CD11b, uma molécula que é co-expressa com CD18 para formar o receptor de complemento e a molécula de adesão Mac-1. Entretanto, os estudos, apresentados neste trabalho, mostram que metade de cada subtipo de B-1 (B-1a e B-1b) não expressa CD11b. As células B-1 CD11b+ são maiores, mais granulosas e expressam quantidades maiores de IgM de superfície. As células CD11b+ possuem uma curiosa tendência de se juntar e formar duplas (doublets) fortemente ligadas entre si que estão presentes em abundância no peritônio de camundongos adultos. Além de ligarem entre si, as células B-1 podem formar duplas com macrófagos PM-1 e PM-2. Durante a ontogenia, as células CD11b- surgem primeiro no peritônio e são as progenitoras das células B-1 CD11b+. Após estímulo inflamatório (LPS í. v.) os linfócitos B-1 CD11b+ migram do peritônio para o baço, onde proliferam e transformam-se em células secretoras de anticorpos (plasmócitos). No peritônio, as células B-1 não são capazes de se diferenciar em plasmócitos (células CD138high). Os resultados apresentados aqui mostraram que, diferentemente dos macrófagos PM-2, os PM-1 são os responsáveis por inibir a formação de células secretoras de anticorpos derivadas de B-1. Em suma, nossos dados sugerem que, mediante estímulo inflamatório, as células B-1 maduras (CD11b+) migram do peritônio para o baço, afastandose de macrófagos PM-1. Já no baço, as células B-1 encontram um micro-ambiente favorável para se proliferarem e diferenciarem-se em plasmócitos, secretando a maioria dos anticorpos naturais vistos logo nas primeiras horas pós-estímulo. / Mouse peritoneal cavity (PerC) represents the source for a variety of cellular subsets of the immune system. In early studies, marred by Iimited methodological tools, it was thought that macrophages comprise roughly 90% of total PerC cells. Shortly thereafter, it was recognized that, beside macrophages, the mouse peritoneal cavity shelters large amount of B-lymphocytes, specially the B-1 subset. In essence, by applying contemporary technology, studies presented here show that, roughly 30% of PerC cells comprise macrophages, 55% comprise B-1 cells, which 40% represents the B-1 a subset and roughly 15% are B-1b cells. The remaining 15%-20% of PerC cells comprise a variety of known immune cells, including B-2, T, NK and eosinophils. In addition, there were some other cellular subsets that could not be identified in these studies, probably due to limited cell surface molecules analyzed. Surprisingly, the data presented here show that peritoneal macrophages represent a highly heterogeneous population. Based on the co-expression of both CD11b and F4/80, we have identified two functionally distinct subsets of peritoneal macrophages, named here as PM-1 (CD11bhigh, F480high) and PM-2 (CD11bint, F4/80low). PM-2 are less abundant (-10% within total macrophages) in the PerC of unstimulated mouse, however, unlike PM-1, these cells are able to spread in spindle (bipolar) morphology upon sorting and in vitro culture. Curiously, PM-2 is the only PerC macrophage that expresses surface MHC-II. In addition, one third of PM-2 population expresses CD11c, a universal marker for dendritic cells. In turn, PM-1 cells are round shaped cells, represent the majority of PerC macrophages (-90%) and secrete high amounts of NO upon LPS stimulation. In vivo phagocytosis assay revealed that both cells could internalize bacteria, however, PM-2 cells showed to be more efficient, in that it was able to phagocyte higher amounts of bacteria when compared to PM-1. After i.p. in vivo stimulation, the absolute number and percentage of PM-2 cells increase drastically in the peritoneum, reaching almost half of the total PerC macrophages. Importantly, PM-1 and PM-2 macrophages seem not to represent PerC monocytes since it does not share any of the phenotype expressed by blood monocytes. Interestingly, in addition to PM-1 and PM-2, some other cellular subsets in the PerC, such as eosinophils, are able to express CD11b and F4/80. PerC B-1 cells have long been known to express CD11b, which is co-expressed with CD18 to form the Mac-1/CR3 complement receptor and adhesion molecule. However, although all PerC B-1 cells are commonly believed to express CD11b, we show here that nearly half of the cells in each of the PerC B-1 subsets (B-1a and B-1b) do not express this surface receptor. The CD11b+ cells in each B-1 subset are larger, more granular and express higher levels of surface IgM than the CD11b- B-1 cells. Surprisingly, the CD11b+ B-1 subset has the curious tendency to initiate the formation of tightly associated doublets that are present at high frequency in adult PerC. In addition to binding to each other, B-1 cells can also form doublets with PM-1 and PM-2. During ontogeny, CD11b- B-1 cells arise earlier in the mouse PerC and are the progenitors of CD11b+ B-1 cells. Upon LPS stimulation, PerC CD11b+ B-1 cells migrate to the spleen where they proliferate and differentiate into antibody-secreting cells (plasma cells). Within the PerC, B-1 cells are not able to differentiate into plasma cells (CD138high cells). The data shown in here reveal that, unlike PM-2, PM-1 macrophages are the cellular subset responsible for inhibiting the formation of B-1-derived antibody-secreting cells. Altogether, our data suggest that, upon inflammatory stimuli, mature CD11b+ B-1 cells migrate from the PerC to the spleen, avoiding the inhibitory effect of PM-1 cells. In the spleen, B-1 cells find an appropriate environment to proliferate and terminally differentiate into antibody-secreting cells, thus, secreting the majority of immunoglobulin seen in few hours after in vivo stimulation.

Este trabalho avalia a influência das emoções humanas expressas pela mímica da face na tomada de decisão de sistemas computacionais, com o objetivo de melhorar a experiência do usuário. Para isso, foram desenvolvidos três módulos: o primeiro trata-se de um sistema de computação assistiva - uma prancha de comunicação alternativa e ampliada em versão digital. O segundo módulo, aqui denominado Módulo Afetivo, trata-se de um sistema de computação afetiva que, por meio de Visão Computacional, capta a mímica da face do usuário e classifica seu estado emocional. Este segundo módulo foi implementado em duas etapas, as duas inspiradas no Sistema de Codificação de Ações Faciais (FACS), que identifica expressões faciais com base no sistema cognitivo humano. Na primeira etapa, o Módulo Afetivo realiza a inferência dos estados emocionais básicos: felicidade, surpresa, raiva, medo, tristeza, aversão e, ainda, o estado neutro. Segundo a maioria dos pesquisadores da área, as emoções básicas são inatas e universais, o que torna o módulo afetivo generalizável a qualquer população. Os testes realizados com o modelo proposto apresentaram resultados 10,9% acima dos resultados que usam metodologias semelhantes. Também foram realizadas análises de emoções espontâneas, e os resultados computacionais aproximam-se da taxa de acerto dos seres humanos. Na segunda etapa do desenvolvimento do Módulo Afetivo, o objetivo foi identificar expressões faciais que refletem a insatisfação ou a dificuldade de uma pessoa durante o uso de sistemas computacionais. Assim, o primeiro modelo do Módulo Afetivo foi ajustado para este fim. Por fim, foi desenvolvido um Módulo de Tomada de Decisão que recebe informações do Módulo Afetivo e faz intervenções no Sistema Computacional. Parâmetros como tamanho do ícone, arraste convertido em clique e velocidade de varredura são alterados em tempo real pelo Módulo de Tomada de Decisão no sistema computacional assistivo, de acordo com as informações geradas pelo Módulo Afetivo. Como o Módulo Afetivo não possui uma etapa de treinamento para inferência do estado emocional, foi proposto um algoritmo de face neutra para resolver o problema da inicialização com faces contendo emoções. Também foi proposto, neste trabalho, a divisão dos sinais faciais rápidos entre sinais de linha base (tique e outros ruídos na movimentação da face que não se tratam de sinais emocionais) e sinais emocionais. Os resultados dos Estudos de Caso realizados com os alunos da APAE de Presidente Prudente demonstraram que é possível melhorar a experiência do usuário, configurando um sistema computacional com informações emocionais expressas pela mímica da face. / The influence of human emotions expressed by facial mimics in decision-taking of computer systems is analyzed to improve user´s experience. Three modules were developed: the first module comprises a system of assistive computation - a digital alternative and amplified communication board. The second module, called the Affective Module, is a system of affective computation which, through a Computational Vision, retrieves the user\'s facial mimic and classifies their emotional state. The second module was implemented in two stages derived from the Facial Action Codification System (FACS) which identifies facial expressions based on the human cognitive system. In the first stage, the Affective Module infers the basic emotional stages, namely, happiness, surprise, anger, fear, sadness, disgust, and the neutral state. According to most researchers, basic emotions are innate and universal. Thus, the affective module is common to any population. Tests undertaken with the suggested model provided results which were 10.9% above those that employ similar methodologies. Spontaneous emotions were also undertaken and computer results were close to human score rates. The second stage for the development of the Affective Module, facial expressions that reflect dissatisfaction or difficulties during the use of computer systems were identified. The first model of the Affective Module was adjusted to this end. A Decision-taking Module which receives information from the Affective Module and intervenes in the Computer System was developed. Parameters such as icon size, draw transformed into a click, and scanning speed are changed in real time by the Decision-taking Module in the assistive computer system, following information by the Affective Module. Since the Affective Module does not have a training stage to infer the emotional stage, a neutral face algorithm has been suggested to solve the problem of initialing with emotion-featuring faces. Current assay also suggests the distinction between quick facial signals among the base signs (a click or any other sound in the face movement which is not an emotional sign) and emotional signs. Results from Case Studies with APAE children in Presidente Prudente SP Brazil showed that user´s experience may be improved through a computer system with emotional information expressed by facial mimics.