Sabe-se que o sistema imune, através de um fenômeno denominado imunossenescência, gradativamente diminui a sua capacidade de resposta durante a vida. Este fato pode tornar o indivíduo mais suscetível a infecções e outras patologias. Neste contexto, seria útil procurar por fatores que alterassem esta evolução natural, principalmente os capazes de acelerar este processo. Por esta razão, nós procuramos por diferenças nos parâmetros imunológicos entre o antes e o depois da cirurgia de valva cardíaca em idosos com mais de 65 anos. Nossos resultados não apontaram, no pós-operatório, para uma diminuição da capacidade imune, uma vez que os testes cutâneos de hipersensibilidade para o PPD, tricofitina e candidina não se alteraram. Quando a resposta linfoproliferativa foi avaliada in vitro, também não apresentou diferença. Por outro lado, nós observamos um aumento na porcentagem de células T CD3 +, T CD4 + e monócitos no sangue periférico, quando comparamos os períodos. Sendo que os marcadores de ativação cellular CD25 +, CD69 + e o CD95 também se apresentaram elevados. Quanto a secreção de citocinas, nossos resultados apontam para um amento de IL-4 e IL-8. Inversamente, concentrações reduzidas de IL-2, IL-12 e IFN- foram detectadas no sobrenadante de PBMCs quando estimuladas in vitro. Em suma, nossos dados demonstram que a cirurgia de valva cardiaca é capaz de alterar vários parâmetros da resposta immune, com um aumenrto da porcentagem de células, quanto da expressão de marcadores de ativação celular e secreção de citocinas / It is known that the immune system, through a phenomenon called immunosenescence, undergoes functional changes during life which may culminate in a diminished capacity of response, turning the subject more susceptible to infections and other pathologies. In this context, it is useful to search for factors that alter this natural evolution, mainly able to delay this process. For this reason, we assessed different immunologic parameters before and after cardiac valve surgery in 65 year-old patients. Our results did not point to a postoperative immunedeficiency-like state, once that the cutaneous tests to PPD, candidin and tricophytin remained positive for most of the subjects. When the proliferative response was assessed in vitro, there were also no differences. On the other hand, we observed a post-surgical increase in the percentage of T CD3 +, T CD4 + cells and in monocytes from peripheral blood when we compare both periods. Moreover, it is important to highlight that activation markers, such as CD25, CD69 and CD95 were also presented in higher levels. According to the cytokine secretion, our results appointed to a greater secretion of IL-4 and IL-8 postoperative. Conversely, reduced concentrations of IL-2, IL-12 and IFN- were detected in supernatant of PBMCs when stimulated in vitro. In summary, our data reveal that the cardiac valve surgery with extra corporeal procedure and anesthesia is able to alter several parameters of the immune response, with an increased percentage of the major assessed cells, as well as in the expression of activation markers and cytokine secretion

A expressão de antígenos restritos a tecidos (TRAs do inglês tissue restricted antigens) no timo pelas células epiteliais medulares (mTECs de medullary thymic epithelial cells) é essencial para a tolerância central das células T. Devido à sua heterogeneidade em termos de representação de autoantígenos, esse fenômeno foi denominado como expressão gênica promíscua (PGE de promiscuous gene expression), no qual o gene Aire (de autoimmune regulator) desempenha um papel como principal regulador transcricional positivo sobre um grande conjunto de TRAs dependentes de Aire. A proteína Aire tem a capacidade de interagir com seqüências específicas de DNA desempenhando um papel como regulador direto. Neste estudo utilizamos o método dos cDNA microarrays para acessar a PGE em células mTEC CD80+ murinas cultivadas in vitro. O agrupamento hierárquico dos dados permitiu a observação de que os genes de TRAs foram diferencialmente expressos. Para testar essa hipótese, inicialmente silenciamos o gene Aire pelo método de RNA interferente (RNAi) nas células mTEC. O agrupamento hierárquico dos dados de cDNA microarray mostrou um conjunto de genes de TRAs dependentes de Aire, os quais foram reprimidos após o silenciamento deste último. Redes gênicas reconstruídas desses dados permitiram a identificação de um nó gênico (Gucy2d) estabelecendo regulação positiva sobre genes downstream nas células mTEC normais. Entretanto, sob efeito do silenciamento de Aire, Gucy2d passou a ser um repressor. Esses resultados evidenciaram que genes da PGE estão conectados em rede, que um nó gênico pode atuar como intermediário no seu controle e que Aire na rede PGE desempenha seu controle como regulador upstream. / The expression of tissue restricted antigens (TRAs) in thymus by medullary thymic epithelial cells (mTECs) is essential for the central selftolerance of T cells. Due to heterogeneity of autoantigen representation this phenomenon has been termed promiscuous gene expression (PGE), in which the autoimmune regulator (Aire) gene plays a role as main positive transcriptional regulator on a large set of Aire-dependent TRAs. Aire protein is able in binding to specific DNA sequence motifs and plays a role as a direct regulator. Here we used the cDNA microarray method to access PGE in murine CD80+ mTECs cultured in vitro. Hierarchical clustering of the data allowed observation that TRA genes were differentially expressed. To further investigate the control of PGE, we hypothesize that TRA genes establish networks contributing it selves to modulate their transcriptional levels. Aire in this case plays a role as upstream positive modulator. To test this hypothesis, initially we silenced Aire by gene knockdown (RNA interference) in mTECs. Hierarchical clustering of cDNA microarray data showed a set of Airedependent TRAs genes, which were down regulated after Aire silencing. Gene networks reconstructed from these data allowed the identification of a gene node (Gucy2d) establishing positive regulation upon downstream genes in normal mTECs. Nevertheless, under silencing of Aire, Gucy2d has become a repressor. These finding evidentiate that, genes features in PGE are connected in network; a gene node may act as intermediate in their control and that Aire in PGE network plays a role as an upstream regulator.

During early development of the nervous system, gene expression patterns are known to vary widely depending on the specific developmental trajectories of different structures. Observable changes in gene expression profiles throughout development are determined by an underlying network of precise regulatory interactions between individual genes. Elucidating the organizing principles that shape this gene regulatory network is one of the central goals of developmental biology. Whether the developmental programme is the result of a dynamic driven by a fixed architecture of regulatory interactions, or alternatively, the result of waves of regulatory reorganization is not known. Here we contrast these two alternative models by examining existing expression data derived from the developing human brain in prenatal and postnatal stages. We reveal a sharp change in gene expression profiles at birth across brain areas. This sharp division between foetal and postnatal profiles is not the result of sudden changes in level of expression of existing gene networks. Instead we demonstrate that the perinatal transition is marked by the widespread regulatory rearrangement within and across existing gene clusters, leading to the emergence of new functional groups. This rearrangement is itself organized into discrete blocks of genes, each associated with a particular set of biological functions. Our results provide evidence of an acute modular reorganization of the regulatory architecture of the brain transcriptome occurring at birth, reflecting the reassembly of new functional associations required for the normal transition from prenatal to postnatal brain development.

This thesis presents a development of Structural Health Monitoring (SHM) and Fault Diagnosis based on Artificial Immune System (AIS), a biology-inspired method motivated from the Biological Immune System (BIS). Using the antigen to model structural health or damage condition of specific characteristics and the antibody to represent an information system or a database that can identify the specific damage pattern, the AIS can detect structural damage and then take action to ensure the structural integrity. In this study the antibodies for SHM were first trained and then tested. The feature space in training includes the natural frequencies and the modal shapes extracted from the simulated structural response data including both free-vibration and seismic response data. The concepts were illustrated for a 2-DOF linear mass-spring-damper system and promising results were obtained. It has shown that the methodology can be effectively used to detect, locate, and assess damage if it occurred. Consistently good results were obtained for both feature spaces of the natural frequencies and the modal shapes extracted from both response data sets. As the only exception, some significant errors were observed in the result for the seismic response data when the second modal shape was used as the feature space. The study has shown great promises of the methodology for structural health monitoring, especially in the case when the measurement data are not sufficient. The work lays a solid foundation for future investigations on the AIS application for large-scale complex structures.

An innovative artificial immune system (AIS) is proposed herein for structural health monitoring (SHM) to ensure the structural integrity and functionality. While satisfactory results were obtained by previous AIS schemes, their performance is strongly structural-parameter-value (SPV) dependent and deviations of SPVs in testing from training due to modeling errors and measurement noises significantly deteriorates the AIS' performance. This thesis presents a less SPV-dependent AIS with a three-phase architecture, including damage-existence-detection, damage-location-determination, and damage-severity-estimation, using specially designed feature vectors (FVs) based on structural modal parameters. The maximum-relative-modal-parameter-change is used to detect the damage's existence and estimate its severity, and the pattern in normalized-modal-parameter-change is used to determinate the damage's location. Comparisons between the proposed FVs and their existing counterparts were conducted for 2/3/4-degree-of-freedom structures to illustrate the superior performance and less SPV-dependence of the proposed method, particularly in determining damage location. The proposed AIS was tested on a 4-degree-of-freedom model using 440 randomly generated damage conditions with a different SPV set per condition. A success rate of 95.23% in the determination of damage's existence and its location was obtained. The trained AIS for the 4-degree-of-freedom model was further evaluated by a four-story and two-bay by two-bay prototype structure used in the benchmark problem proposed by the IASC-ASCE Structural Health Monitoring Task Group. Results have shown great potentials of the proposed approach in its real-world applications.

The role of the immune system in periodontal disease has been well established. Individuals who smoke are more prone to developing periodontitis because of the excess plaque buildup and the immune system's attack of the bacteria on the gingiva. This study aims to examine the role of immunotherapy in the reversal of periodontal disease in individuals who smoke. Further research will not only assist with the reversal of periodontitis, but may also improve other debilitating co-morbid diseases including autoimmune diseases such as human immunodeficiency virus. To examine this question, a variety of research studies from various sources were examined. There are numerous factors, which contribute to the initiation and progression of periodontal disease. Use of nicotine products enhances the accumulation of plaque formation. Accumulation of large amounts of plaque and calculus assist in the progression of periodontal disease. Disease of the gingiva may lead numerous other debilitating diseases such as respiratory infections, Alzheimer's disease and unfavorable pregnancy outcomes. Two components of the immune system play a major role in gingival inflammation observed in smokers; these include cytokine production and inflammation. The concentration of pro-inflammatory cytokines released from the macrophages of smokers is significantly higher than that observed in non-smokers. Pro-inflammatory cytokines, such as TNF-alpha, are responsible for inflammation of the gingiva. The use of pre-existing immunotherapy treatments may be beneficial and assist in the reversal of periodontal disease. Further researcher on immune therapy, which has been shown to be successful in other disease treatments will not only be beneficial for the oral cavity, but for overall systemic health. Both immune and bacterial components should be taken into consideration when developing therapies for periodontal disease. Pro-inflammatory cytokines such as TNF-alpha are dramatically increased in smokers. Researchers should model therapies after existing therapies that have been successful for other diseases such as immunotherapy studies with lamvidine (Hepatitis B), lambrolixumab (melanoma) and infliximad (oral plaque). The mTOR signaling pathway controls the release of pro-inflammatory and anti-inflammatory cytokines. Deletion of the raptor protein causes more inflammation in the colon due to an increased release of pro-inflammatory cytokines. Therapies which directly act upon this signaling pathway should be further researched upon to reverse the effects of periodontal disease. Chromogranin A release showed a significant increase in smokers because of the body's ability to want to automatically attach the harsh environment created by tobacco. Modulating the immune response induced by Chromagranin A or possibly through the use of anti-TNF-alpha therapies which have shown success in patients with rheumatoid arthritis also is open to future research. The shift from gram-negative to gram-positive bacteria was noted in periodontal disease patients. The less diverse bacteria lead to the loss of the ability of ligaments to attach to one another in the oral cavity. The development and testing of bacteria specific antibiotics such as Fusobacterium, Prevotella, and Selenomonas may prove to be very beneficial. There is much optimism among the dental professionals that immunotherapy may lead to the development of future therapies for periodontitis in both smokers and non-smokers. However we must also encourage patients to stop smoking through awareness and education.

Em bovinos, o desenvolvimento embrionário pré implantacional depende das funções do endométrio bovino que tem suas funções mediadas por uma complexa interação da ação e dos efeitos dos hormônios esteroides ovarianos E2 e P4. Estes hormônios regulam a expressão gênica e controlam o ambiente uterino modulando, entre outros, a expressão de microRNAs e a rede de citocinas relacionadas ao sistema imune. Os objetivos do presente trabalho foram abordados em dois capítulos, sendo (I) comparar os efeitos dos distintos ambientes endócrinos peri-ovulatórios sobre a expressão de microRNAs (II) e na modulação do sistema IL1/TLR no endométrio bovino nos dias 4 e 7 após a indução da ovulação. Para isso, controlou-se farmacologicamente o crescimento do folículo objetivando induzir a ovulação de folículos de maior diâmetro (grupo folículo grande-CL grande, FG-CLG) ou de menor diâmetro (grupo folículo pequeno-CL Pequeno, FP-CLP). Vinte e duas vacas multíparas nelore, foram pré-sincronizadas, metade destes animais foram destinados para o grupo FG-CLG e receberam uma dose de prostaglandina F2&#945; (PGF) e um dispositivo de progesterona, juntamente com benzoato de estradiol no D10. No momento da retirada dos dispositivos de progesterona (entre D1,75 e D2,5) todos os animas receberam uma dose de PGF. A ovulação foi induzida com acetato de buserelina (D0). O que diferiu entre os tratamentos foi que os animais do grupo FP-CLP não receberam uma dose de PGF no D10 e o momento da retirada dos dispositivos foi entre D1,25 e o D1,5. No capítulo I, o a expressão de microRNAs foi determinada por qPCR nos dias 4 e 7. Dos 90 microRNAs testados, 21 apresentaram se up-regulated e dois down-regulated no grupo FG-CLG (P<0.1) no D4. No D7, quatro microRNAs foram diferentemente expressos, sendo um up-regulated e três down-regulated no grupo FG-CLG (P<0.1) no D7. Para os microRNAs diferentemente expressos determinou-se mRNA-alvos preditos. Uma análise de ontologia demonstrou que os mRNAs-alvos apresentaram enriquecimento funcional na via dos receptores de hormônios esteroides, entre outras. No capítulo II, o sistema IL1/TLR foi avaliado quanto a abundância de transcriptos envolvidos neste sistema, do microRNA bta-mir-155 e das proteínas IL1&#946; e IL1R1. A abundância relativa de mRNA apresentou diferença (P<0.1) na abundância dos mRNAs de IL1R1, TAB1 e FOXP3, das proteínas IL1&#946; e IL1R1, sendo essas moléculas up-regulated no grupo FG-CLG. O microRNA bta-mir-155 foi down-regulated no grupo FG-CLG (P<0.1). Diante disto, pode-se concluir que o ambiente endócrino peri-ovulatório determina o perfil de expressão de microRNAs e modula o sistema IL1/TLR no endométrio bovino / In cattle, the pre implantation embryo development depends on the functions of the bovine endometrium that has its functions mediated by a complex interaction of action and the effects of ovarian steroid hormones E2 and P4. These hormones regulate gene expression and control the modulating uterine environment among others, the expression of microRNAs and the network of cytokines related to the immune system. The objectives of this study were discussed in two chapters, (I) to compare the effects of different peri-ovulatory endocrine environment on the expression of microRNAs (II) and modulation of the IL-1 system / TLR in bovine endometrium on days 4 and 7 after induction of ovulation. For this, it was controlled pharmacologically follicle growth aiming to induce ovulation of follicles larger diameter (great grand-CL follicle group, FG-CLG) or smaller in diameter (small-CL Small follicle group, FP-PLC). Twenty two nelore multiparous cows were pre-sync, half of these animals were used for the FG-NCG group and received a dose of F2á prostaglandin (PGF) and progesterone device along with oestradiol benzoate in D-10. Upon withdrawal of progesterone devices (between 1.75 and D-D-2,5) all animas received a dose of PGF. Ovulation was induced with buserelin acetate (D0). What differed between treatments was that animals FP-CLP group did not receive a dose of PGF in the D-10 and the time of removal of the devices was between D-1,25 and D-1.5. In Chapter I, the expression of microRNAs was determined by qPCR on 4 and 7. Of the 90 microRNAs tested, 21 showed was up-regulated and down-regulated in two FG-CLG group (P <0.1) in the D4. In D7 four microRNAs were differently expressed, one up-regulated and down-regulated in three FG-CLG group (P <0.1) at D7. For differently expressed microRNAs was determined predicted mRNA-target. An ontology analysis showed that the mRNA-targets had functional enrichment in via the steroid hormone receptors, among others. In Chapter II, the IL-1 / TLR system was evaluated as the abundance of transcripts involved in this system, the bta-mir-155 microRNA and IL1&#946; and IL1R1 proteins. The relative abundance of mRNA was different (P <0.1) in the abundance of mRNAs IL1R1, TAB1 and FOXP3, the IL1&#946; and IL1R1 proteins, and these up-regulated molecules in the FG-CLG group. The bta-mir-155 microRNA was down-regulated in the FG-CLG group (P <0.1). Given this, we can conclude that the peri-ovulatory endocrine milieu determines the profile of microRNA expression and modulates the IL1 / TLR system in bovine endometrium

Our murine model of mild traumatic brain injury (mTBI) has shown improved survival after Pseudomonas aeruginosa (Psd) challenge as compared to controls (tail trauma or sham injury). Previous work suggests an mTBI-specific involvement of the neuro-immune axis which augments the innate immune response, increasing survival. Additional factors for the enhanced mTBI survival were explored via microarray analysis of lungs harvested 48 hours post-trauma, the point prior to Psd challenge in our model. At 48 hours post-trauma, mTBI lungs have a number of upregulated ATP synthesis and mitochondrial gene sets. Increased available energy could prime the mTBI lungs, allowing an earlier and more robust response to Psd infection, possibly contributing to the increased mTBI survival. This is supported by increased neutrophil recruitment in the bronchoalveolar lavage of mTBI mice four hours after Psd instillation. Downregulated gene sets related to cellular connections suggest that neutrophils recruited to the lung have an easier extravasation pathway into the air space of mTBI lungs compared to control. Based on genetic and neutrophil recruitment data, it is possible that mTBI creates an energetically prepared and easily accessible lung better tailored for recruiting and allowing entry of neutrophils in response to an infection compared to control.

Parasitas e agentes infecciosos devem representar uma importante pressão seletiva atuante sobre a evolução de aspectos da história de vida dos hospedeiros. Os desafios impostos aos hospedeiros pelos patógenos resultam em uma séria de respostas fisiológicas e comportamentais, respostas estas pouco conhecidas para a maioria das espécies de hospedeiros ectotermos, como anfíbios anuros. O objetivo deste estudo foi quantificar os efeitos da ativação do sistema imune via injeção de LPS (lipopolissacarídeo), em duas diferentes concentrações, sobre o comportamento alimentar, taxa metabólica de repouso, atividade locomotora espontânea e comportamento anti-predatório de machos adultos da perereca das bromélias: Scinax gr. perpusillus . O tratamento com LPS provocou redução na taxa de alimentos ingeridos, confirmando o efeito anorexigênico deste componente, e aumento na atividade locomotora espontânea, porém não apresentou efeito sobre a taxa metabólica de repouso ou sobre o comportamento anti-predatório das pererecas / Parasites and infectious agents should represent important selective pressure acting on the evolution of aspects of life history of the hosts. The challenges posed by pathogens to hosts result in a series of physiological and behavioral responses, which are not well known to most ecthoterm hosts species, such as anuran amphibians. The aim of this study was to quantify the effects of activation of the immune system via injection of LPS (lipopolysaccharide) in two different concentrations, on feeding behavior, resting metabolic rates, spontaneous locomotor activity, and antipredatory behavior of adult males of the bromeliad tree-frog: Scinax gr. perpusillus . Treatment with LPS caused a reduction in the rate of food intake confirming the anorexigenic effect of this component and an increase in spontaneous locomotor activity, but had no effect on the resting metabolic rate or in the the antipredator behavior of frogs

Despite the convergence of rapid technological advances in genomics and the maturing field of ecoimmunology, our understanding of the genes that regulate immunity in wild populations is still nascent. Previous work to assess immune function has relied upon relatively crude measures of immunocompetence. However, with next-generation RNA-sequencing, it is now possible to create a profile of gene expression in response to an immune challenge. In this study, captive zebra finch (Taeniopygia guttata; adult males) were challenged with bacterial lipopolysaccharide (2 mg/Kg BW; dissolved in 0.9% saline) or vehicle (0.9% saline) to stimulate the immune system. Two hours after injection, birds were euthanized and hypothalami, spleen, and red blood cells (RBCs) were collected. Taking advantage of the fully sequenced genome of zebra finch, total RNA was isolated, sequenced, and partially annotated in these tissue/cells. The data show 628 significantly upregulated transcripts in the hypothalamus, as well as 439 and 121 in the spleen and RBCs, respectively, relative to controls. Also, 134 transcripts in the hypothalamus, 517 in the spleen, and 61 in the RBCs were significantly downregulated. More specifically, a number of immunity-related transcripts (e.g., IL-1β, RSAD2, SOCS3) were upregulated among tissues/cells. Additionally, transcripts involved in metabolic processes (APOD, LRAT, RBP4) were downregulated, suggesting a potential trade-off in expression of genes that regulate immunity and metabolism. Unlike mammals, birds have nucleated RBCs, and these results suggest a novel transcriptomic response of RBCs to immune challenge. Lastly, molecular biomarkers could be developed to rapidly screen bird populations by simple blood sampling in the field.