El concentrat de plasma assecat per polvorització (SDP) i el d’immunoglobulines (IC) són suplements dietètics que han estat àmpliament utilitzats en l’alimentació dels animals de granja i que han estat proposats com alternativa a l’ús dels antibiòtics com a promotors del creixement, especialment durant el període posterior al deslletament. Aquests suplements incrementen la taxa de creixement degut, en part, a que atenuen la resposta del sistema immunitari. Per exemple, s’ha pogut observar que la suplementació dietètica amb proteïnes plasmàtiques redueix la sobreestimulació immunitària en un model de malaltia inflamatòria intestinal. El sistema immunitari mucosal comú connecta els llocs inductors (en aquest cas la mucosa intestinal) amb els llocs efectors (com les mucoses naso- i broncoalveolar o la genito-urinària), que facilita el moviment de cèl•lules immunitàries entre teixits allunyats anatòmicament i permet que la dieta moduli la resposta associada a infeccions extraintestinals. Així doncs, ens hem plantejat si la suplementació dietètica amb proteïnes plasmàtiques podia modular la resposta immunitària en una inflamació pulmonar aguda. Primerament es va establir una pauta adequada per obtenir un model d’inflamació pulmonar robust i reproduïble. Seguidament s’han estudiat els efectes dels suplements SDP i IC sobre la resposta immunitària en aquest model d’inflamació, en ratolins acabats de deslletar. S’han analitzat cèl•lules i mediadors inflamatoris involucrats tant en la resposta innata, com en la resposta adaptativa. Finalment, també s’han estudiat els efectes dels pinsos experimentals sobre diferents elements que intervenen en la resolució de la resposta inflamatòria, com les cèl•lules reguladores i les citocines antiinflamatòries. L’administració de LPS produeix una migració de leucòcits i un alliberament massiu de citocines i quimiocines proinflamatòries, que recluten i activen monòcits i neutròfils a l’espai alveolar i al teixit pulmonar. A més estimula la immunitat adaptativa, en la que hi ha un increment considerable de limfòcits Th activats, així com una gran producció de citocines. Ambdues dietes disminueixen el reclutament de leucòcits, sobretot els neutròfils i els monòcits. Tot i que la resposta innata és més extensa, les dietes són més efectives sobre la resposta adaptativa, on redueixen les poblacions limfocitàries estimulades per l’LPS tant a pulmó com a sang. Els suplements també atenuen de forma notable l’efecte de l’LPS sobre l’expressió de citocines i quimiocines proinflamatòries a l’espai alveolar. Els efectes de l’SDP i l’IC estan mediats per canvis en l’expressió de citocines antiinflamatòries, ja que augmenten la concentració de IL-10 a pulmó i a jejú, i la del TGF-β a pulmó. Ambdues dietes redueixen el balanç entre els limfòcits Th activats i els limfòcits T reguladors, d’aquesta manera atenuen la magnitud de la resposta inflamatòria i augmenten la concentració de mediadors essencials per la reparació del teixit. Els resultats d’aquest estudi indiquen que els suplements amb proteïnes plasmàtiques poden limitar la resposta immunitària pulmonar i podrien ser útils en la prevenció i atenuació de les malalties inflamatòries extraintestinals. / The concentrated spray-dried plasma (SDP) and immunoglobulin (IC) are dietary supplements that have been widely used in the diet of farm animals and proposed as an alternative to the use of antibiotics as growth promoters, especially in the subsequent weaning period. These supplements increase the growth rate, in part because they attenuate the response of the immune system. The common mucosal immune system connects the inductive sites (in this case the intestinal mucosa) with the effector sites (as nasopharyngeal and bronchoalveolar mucosa or genito-urinary), which facilitates the movement of immune cells between anatomically distant tissues and allows the diet to modulate the associated response to intestinal infections. Consequently, the aim of the present thesis was to evaluate whether the dietary supplementation with plasma proteins could modulate the immune response in an acute lung inflammation. First, a suitable pattern was established in order to obtain a robust and reproducible inflammatory lung model. Next, the effects of SDP and IC supplementation on the immune response in this inflammation model were studied in newly weaned mice. Cells and inflammatory mediators, involved either in innate response as in adaptive response, have been analyzed. Finally, the effects of experimental feed on different elements involved in the resolution of the inflammatory response, such as regulatory cells and inflammatory cytokines, have also been assessed. Both diets decreased the recruitment of leukocytes. Although the innate response is more extensive, the diets are more effective on the adaptive response. The supplements also significantly attenuated the effect of LPS on the expression of pro-inflammatory cytokines and chemokines in the alveolar space. The effects of SDP and IC are mediated by changes in the expression of anti-inflammatory cytokines. Both diets reduced the balance between activated Th cells and regulatory T cells, in such a way that, the magnitude of the inflammatory response is attenuated and the concentration of essential mediators for tissue reparation is increased. In conclusion, the results of this study indicate that plasma protein supplementation is able to limit the pulmonary immune response and could be useful in the prevention and attenuation of the extraintestinal inflammatory diseases.

Tick saliva contains a lot of molecules with antihemostatic and immunosupressive effects.The goal of this thesis is to test the effects of tick salivary cystatins from I.ricinus and I.scapularis on TLR - induced maturation of bone-marrow derived dendritic cells and production of chosen cytokines. Over all, the supressive effect of tick cystatins was observed in relation to TLR-induced maturation of DC. In addition, cystatins enhanced production of IL-10 and attenuated induction of IL-12 cytokines.

Submitted by Maike Costa (maiksebas@gmail.com) on 2017-07-07T15:09:57Z No. of bitstreams: 1 arquivototal.pdf: 1854233 bytes, checksum: 535d273c615cebce265419e27f74439a (MD5) / Made available in DSpace on 2017-07-07T15:09:57Z (GMT). No. of bitstreams: 1 arquivototal.pdf: 1854233 bytes, checksum: 535d273c615cebce265419e27f74439a (MD5) Previous issue date: 2016-10-17 / Ouabain is a cardiotonic steroid initially described as a substance of plant origin. In 1991, the endogenous production of higher mammals was identified and since then their physiological actions have been studied. Work of our group have demonstrated that ouabain modulates the acute inflammatory response induced by different phlogistic agents, also being able to interfere negatively in inflammatory profile triggered by Leishmania (L.) amazonensis. Acute lung injury (ALI) is a serious inflammatory disease characterized by acute inflammation, extensive accumulation of polymorphonuclear leukocytes and accumulation of proinflammatory mediators, which culminates with diffuse alveolar damage and which may cause the patient died due to severe hypoxemia. There is no data in the literature on the effects of ouabain in ALI. Objectives: Evaluate the immunomodulatory effect of ouabain in a murine model of ALI induced by LPS. Methods: BALB / c mice were treated intraperitoneally with ouabain at a dose of 0.56 mg / kg for a period of three consecutive days, 1 hour after the last treatment the animals were challenged intranasally with 40μl of an LPS solution (2 5 mg / kg); 24 hours after challenge, the animals were euthanized, the collected biological sample and inflammatory parameters, including cell migration, protein exudates, cytokine production, TLR4 expression and histopathological changes were then evaluated. Data were analyzed by PRISMA software. Results: The treatment with ouabain decreased total leukocytes migration to the inflamed site (48,84%), this event associated with decreased neutrophil migration (70,71%) and independent of macrophage migration. The ouabain also decreased the exudate protein in the broncho-alveolar region (26,32%) and production of the cytokines TNF-α (14,80%), IL-6 (47,07%) and IL1-β (33,59%), however this reduction in the production of these mediators was not related to the expression of TLR4. Additionally, the ALI histopathology changes were also reduced by treatment with ouabain. Conclusions: The results show that ouabain has anti-inflammatory action in ALI induced by LPS. / A Ouabaína é um esteroide cardiotônico inicialmente descrito como uma substância de origem vegetal. Em 1991, a sua produção endógena por mamíferos superiores foi identificada e desde então suas ações fisiológicas vêm sendo estudadas. Trabalhos do nosso grupo demonstraram que a ouabaína modula a resposta inflamatória aguda induzida por diferentes agentes flogísticos, sendo também capaz de interferir negativamente no perfil inflamatório desencadeado pela Leishmania (L.) amazonensis. A lesão pulmonar aguda (LPA) é uma doença inflamatória caracterizada por inflamação aguda e extenso acúmulo de polimorfonucleares e de mediadores pró-inflamatórios, que culmina com dano alveolar difuso podendo levar o paciente a óbito por hipoxemia severa. Não há dados na literatura sobre os efeitos da ouabaína na LPA. Objetivos: Avaliar o efeito imunomodulador de ouabaína em modelo murino de LPA induzida por LPS. Métodos: Camundongos BALB/c machos foram tratados via intraperitoneal com ouabaína na dose de 0,56 mg/Kg por um período de três dias consecutivos, 1h após o último tratamento os animais foram desafiados via intranasal com LPS (2,5 mg/Kg); 24h após o desafio, os animais foram eutanásiados, as amostras biológicas coletadas e os parâmetros inflamatórios, incluindo migração celular, exsudato proteico, produção de citocinas, expressão de TLR4 e alterações histopatológicas foram então avaliados. Os dados foram analisados pelo software PRISMA. Resultados: O tratamento com a ouabaína diminuiu a migração de leucócitos totais para o sítio inflamado (48,84%), evento este, associado a diminuição da migração de neutrófilos (70,7%) e independente da migração de macrófagos. Ouabaína também diminuiu o exsudato proteico na região bronco-alveolar (26,32%) e a produção das citocinas TNF-α (14,80%), IL-6 (47,07%) e IL1-β (33,59%), entretanto essa redução na produção desses mediadores não mostrou relação com a expressão do TLR4. Adicionalmente, as alterações histopatológicas características da LPA também foram reduzidas pelo tratamento com ouabaína. Conclusões: Os resultados obtidos demonstram que ouabaína possui ação anti-inflamatória na LPA induzida por LPS.

Submitted by Automa??o e Estat?stica (sst@bczm.ufrn.br) on 2016-03-22T19:38:42Z No. of bitstreams: 1 FranciscoCanindeDeSousaJunior_TESE.pdf: 1665223 bytes, checksum: a3c9c1fa573d5e42243d0808e537d798 (MD5) / Approved for entry into archive by Arlan Eloi Leite Silva (eloihistoriador@yahoo.com.br) on 2016-03-28T19:44:15Z (GMT) No. of bitstreams: 1 FranciscoCanindeDeSousaJunior_TESE.pdf: 1665223 bytes, checksum: a3c9c1fa573d5e42243d0808e537d798 (MD5) / Made available in DSpace on 2016-03-28T19:44:15Z (GMT). No. of bitstreams: 1 FranciscoCanindeDeSousaJunior_TESE.pdf: 1665223 bytes, checksum: a3c9c1fa573d5e42243d0808e537d798 (MD5) Previous issue date: 2015-02-27 / O crescente interesse e aplica??es dos produtos biotecnol?gicos v?m aumentando o desenvolvimento de novos processos de recupera??o e purifica??o de prote?nas. A adsor??o em leito expandido (ALE) tem se destacado como uma t?cnica promissora para essa finalidade, pois combina em uma ?nica opera??o as etapas de clarifica??o, concentra??o e purifica??o da prote?na alvo, reduzindo assim tempo e custos de opera??o. Neste contexto, o objetivo desta tese foi avaliar a recupera??o e purifica??o do ant?geno 503 de Leishmania i. chagasi expresso em E. coli M15 e a remo??o de endotoxina por ALE. Na primeira etapa do trabalho foram realizados ensaios em taques agitados sob a forma de dois planejamentos experimentais, para definir as condi??es ?timas de adsor??o e elui??o do ant?geno na resina Streamline chelating. Nos ensaios de adsor??o usando o leito na forma expandida empregou-se uma coluna de 2,6 cm de di?metro por 30,0 cm de altura, acoplada a uma bomba perist?ltica. Na segunda etapa do trabalho, avaliou-se a remo??o de endotoxina durante o processo de recupera??o do ant?geno, empregando o tensoativo n?oi?nico triton X-114 na etapa de lavagem da ALE. Na terceira etapa, buscou-se elaborar um modelo matem?tico capaz de prever as curvas de ruptura do ant?geno 503 em coluna na forma expandida. Os resultados do planejamento experimental para adsor??o do ant?geno 503 mostraram o pH 8,0 e a concentra??o de NaCl de 2,4 M como melhores condi??es de adsor??o. No segundo planejamento, o ?nico fator significativo para elui??o foi a concentra??o de imidazol, definida em 600 mM. A isoterma de adsor??o do ant?geno 503 mostrou bom ajuste ao modelo de Langmuir (R=0,98) e os valores de qmax (capacidade m?xima de adsor??o) e Kd (constante de equil?brio) estimados foram de 1,95 mg/g e 0,34 mg/mL, respectivamente. Atrav?s dos testes de purifica??o diretamente do homogeneizado n?o clarificado obteve-se uma recupera??o de 59,2% da prote?na de interesse e um fator de purifica??o de 6,0. A adi??o do tensoativo n?o-i?nico Triton X-114 ? etapa de lavagem da ALE proporcionou altos valores (>99%) de remo??o do LPS inicialmente presente nas amostras para todas as condi??es estudadas. O modelo matem?tico obtido para descrever a curva de ruptura do ant?geno 503 na resina Streamline Chelanting em leito expandido apresentou bom ajuste aos dados experimentais, tanto para etapa de estimativa de par?metros quanto para de valida??o. O modelo validado foi utilizado na otimiza??o das efici?ncias, obtendo-se os valores m?ximos de efici?ncia do processo e efici?ncia da coluna de 89,2% e 75,9%, respectivamente. Portanto, ALE mostrou ser uma alternativa eficiente na recupera??o da prote?na-alvo e remo??o de endotoxina a partir de um extrato de E. coli n?o clarificado em apenas uma etapa. / The growing interest and applications of biotechnology products have increased the development of new processes for recovery and purification of proteins. The expanded bed adsorption (EBA) has emerged as a promising technique for this purpose. It combines into one operation the steps of clarification, concentration and purification of the target molecule. Hence, the method reduces the time and the cost of operation. In this context, this thesis aim was to evaluate the recovery and purification of 503 antigen of Leishmania i. chagasi expressed in E. coli M15 and endotoxin removal by EBA. In the first step of this study, batch experiments were carried out using two experimental designs to define the optimal adsorption and elution conditions of 503 antigen onto Streamline chelating resin. For adsorption assays, using expanded bed, it was used a column of 2.6 cm in diameter by 30.0 cm in height coupled to a peristaltic pump. In the second step of study, the removal of endotoxin during antigen recovery process was evaluated employing the non-ionic surfactant Triton X-114 in the washing step ALE. In the third step, we sought developing a mathematical model able to predict the 503 antigen breakthrough curves in expanded mode. The experimental design results to adsorption showed the pH 8.0 and the NaCl concentration of 2.4 M as the optimum adsorption condition. In the second design, the only significant factor for elution was the concentration of imidazole, which was taken at 600 mM. The adsorption isotherm of the 503 antigen showed a good fit to the Langmuir model (R = 0.98) and values for qmax (maximum adsorption capacity) and Kd (equilibrium constant) estimated were 1.95 mg/g and 0.34 mg/mL, respectively. Purification tests directly from unclarified feedstock showed a recovery of 59.2% of the target protein and a purification factor of 6.0. The addition of the non-ionic surfactant Triton X-114 to the washing step of EBA led to high levels (> 99%) of LPS removal initially present in the samples for all conditions tested. The mathematical model obtained to describe the 503 antigen breakthrough curves in Streamline Chelanting resin in expanded mode showed a good fit for both parameter estimation and validation steps. The validated model was used to optimize the efficiencies, achieving maximum values of the process and of the column efficiencies of 89.2% and 75.9%, respectively. Therefore, EBA is an efficient alternative for the recovery of the target protein and removal of endotoxin from an E. coli unclarified feedstock in just one step.

Le Brain-derived Neurotrophic Factor (BDNF) est impliqué dans les processus cognitifs impliquant l'hippocampe et les structures corticales. Cette étude avait pour but d’analyser les effets postopératoires sur l’expression du BDNF dans ces structures cérébrales.Ainsi, les effets d’une anesthésie de courte durée au propofol, d’une chirurgie mineure et du lipopolysaccharide (LPS) sur l'altération de l’expression du BDNF ont été analysée dans l'hippocampe et le cortex de deux modèles de rongeurs nocturnes et diurnes, correspondant respectivement des jeunes rats Sprague Dawley mâles et des Arvicanthis ansorgei.Dans un premier temps, la rythmicité nycthémérale de l’expression du BDNF a été analysée.Les quantités de BDNF présentes dans le cortex et l'hippocampe ont été déterminées par une technique ELISA. Il s’est avéré que, dans l'hippocampe et le cortex de rat et d’A. ansorgei, le BDNF suit une rythmicité sur 24 heures. La quantité de BDNF atteint un maximum à ZT5(i.e., 5 heures après le début de l’exposition à la lumière). Dans un deuxième temps, les effets sur l’expression du BDNF ont été analysés après administration de propofol et/ou LPS, ainsi que lors d’une chirurgie légère. Parallèlement,l’impact de ces traitements sur la mémoire a été testé à l’aide d’un test d'évitement passif.Nos résultats indiquent que la quantité de BDNF est régulée positivement dans l'hippocampe et le cortex de rats lorsque les animaux ont subit une anesthésie de courte durée au propofol en présence ou absence de LPS. A l’opposé, une chirurgie mineure (sous anesthésie propofol)n’a aucun effet sur les quantités de BDNF.En conclusion, ces études mettent en évidence des effets majeurs d’une anesthésie sur l’expression du BDNF, ainsi que les effets protecteurs du propofol sur la neuroinflammation induite par le LPS. / Brain-derived neurotrophic factor (BDNF) is involved in cognition and hippocampus and cortical structures are important in cognition. The present study was designed to analyse the post-operative effects on BDNF. For this purpose, we examined the effects of short duration propofol anaesthesia, LPS and minor surgery on the BDNF protein alteration in the hippocampus and cortex.Young male Sprague Dawley rats were used in all experiments except Arvicanthis ansorgei which were used to study the circadian rhythmicity of BDNF protein as model for diurnal rodents. The quantity of BDNF protein present in the cortex and hippocampal supernatants was determined with an ELISA technique. Memory was tested by fear conditioning using the classic fear conditioning preparation (passive avoidance apparatus).The major finding on BDNF protein in control conditions is that the BDNF protein followed a circadian rhythmicity during the 24 hours of day in the rat’s hippocampus and cortex. The concentration of BDNF protein has been observed to reach a maximum at ZT5 (5 hours afterthe light on period) whether the animals were nocturnal or diurnal. In addition, we found that BDNF protein amount is up regulated in the hippocampus and cortex of rats when they were submitted to short duration propofol anaesthesia, as well as LPS and no effect when minor surgery under propofol anaesthesia was performed.In conclusion, these studies illustrate the dramatic effects of post-operative conditions and neuroinflammation induced by LPS on cognition and the potential mechanism involved. This study also suggests the protective effects of the short duration propofol anaesthesia against neuroinflammation induced by LPS.

A terapia endodôntica visa o selamento biológico do complexo sistema apical, contribuindo para isso, as substâncias usadas no tratamento e a resposta imune do paciente. A LILT tem mostrado atividade antiinflamatória, favorecendo o processo reparacional. Sendo assim, este trabalho objetivou analisar o efeito da LILT na atividade secretória de macrófagos, previamente ativados por IFN-? e LPS de E.coli, e estimulados por substâncias liberadas de três tipos de cimentos endodônticos, um a base de óxido de zinco e eugenol, outro a base de hidróxido de cálcio e um terceiro resinoso. A citotoxicidade dessas substâncias foi avaliada usando a técnica de análise do MTT. Macrófagos ativados foram estimulados por essas substâncias ou não (controle) e então, irradiados ou não (controle) e a secreção de proteínas próinflamatórias (interleucina-1 b, fator de necrose tumoral-a e metaloproteinase da matriz-1) foram analisadas pelo teste ELISA. As irradiações foram realizadas com um laser GaAlAs (780 nm, 70 mW, ponta da fibra de 4 mm2, 1.67 seg, 3 J/cm2). Foram usadas duas aplicações de irradiação com intervalo de 6 h. Os dados obtidos foram tratados por Análise de Variância, quando de distribuição normal, ou teste de Friedman, quando de distribuição não normal, com nível de significância de 5 % (p = 0,05). A viabilidade dos controles e células tratados pelos cimentos endodônticos foi similar. Produção de IL-1 b e TNF-a foram observadas. Houve alta produção de MMP-1. Entretanto, sem diferenças estatísticas entre os grupos experimentais. Os grupos irradiados apresentaram resultados similares aos não irradiados. Substâncias liberadas pelos cimentos endodônticos testados não se mostraram citotóxicas nas condições deste experimento. Essas substâncias, bem como a LILT, no parâmetro utilizado, não causam alteração na atividade de secreção de MMP-1, IL-1 b e TNF-a por macrófagos ativados. / The endodontic therapy seeks the dental root canal biological sealing, depending on substances used in this process and patient’s defense immune factors. LILT has shown an anti-inflammatory activity, improving the periapical repair process. This in vitro study aimed to analyze the effect of LILT at the secretory activity of macrophages previously activated by interferon-gamma and lypopolisaccharide from E.coli, and stimulated by substances leached from three endodontic sealers (zinc oxide-eugenol based, resinous and calcium hydroxide-based). Cytotoxicity of these substances was assessed by the MTT test. Activated macrophages were stimulated by the substances or not (control) and then, irradiated or not (control) and the secretion of pro-inflammatory proteins (interleukin-1 b, tumor necrosis factor-a and matrix metalloproteinase-1) was analyzed by ELISA test. The LILT was performed using a GaAlAs laser (780 nm, 70 mW, focal spot of 4.0 mm2, 1.67 sec, 3 J/cm2). Two irradiations with 6 h-intervals were done. The data was compared by either ANOVA test or Friedman’s test. The cell viabilities of controls and cells treated by the sealers were similar. Production of IL -1 b and TNF-a were observed. There was a high production of MMP-1. However, statistical differences were not observed amongst the groups. The irradiated groups presented results similar to those of non irradiated groups. Substances leached from the endodontic sealers are non cytotoxic at these experiments conditions . These substances, as well as the LILT, at the parameter used, were not able to change the secretion of MMP-1, IL-1 b e TNF-a by activated macrophages.

Pesquisa descritivo-explicativa e exploratória sobre a relação entre Turismo e desenvolvimento regional que apresenta o modelo de análise APL Tur, elaborado com base em elementos de referenciais teóricos de economia, geografia, sociologia, administração, comunicação, antropologia e Turismo. O objetivo é mostrar se é possível realizar o desenvolvimento regional por meio do Turismo. Uma região é um contexto territorial delimitado por critérios geográficos, econômicos e políticos. Um dos modelos da análise e gestão do desenvolvimento regional é o Arranjo Produtivo Local APL, uma categoria especial de cluster. Em razão das limitações do APL, elabora-se o modelo particular de análise APL Tur Arranjo Produtivo Local de Turismo. O modelo APL Tur estrutura-se nas dimensões econômica, cultural e organizacional. Os elementos do desenvolvimento regional na dimensão econômica são: delimitação espacial; disparidades intra-regionais; externalidades; sustentabilidade ambiental; e inclusão social. Os elementos do Turismo circunscritos na dimensão econômica são: oferta e demanda; desempenho; priorização; exportação; circuito produtivo; interatividade extra-regional; e acessibilidade. Na dimensão cultural, destacam-se: aspectos históricos; acervos e incentivos; estética; produtos e atrativos; animação; e motivação e satisfação da comunidade. Na dimensão organizacional, têm-se os elementos: poder e capital social; gestão sistêmica; divulgação e imagem; mercadologia e comercialização; planejamento; empreendedorismo e inovação; e conhecimento. Realizou-se o teste do modelo APL Tur, aplicando-o à Região das Hortênsias (Rio Grande do Sul, Brasil), formada pelos municípios de Gramado, Canela, Nova Petrópolis e São Francisco de Paula. Com base no quadro de indicadores dos elementos das dimensões do modelo APL Tur, verifica-se que o Turismo contribui para o desenvolvimento regional, pois proporciona o ingresso, produção e distribuição de riquezas. Para que uma região se desenvolva economicamente por meio do Turismo, é preciso, entretanto, uma gestão adequada das dimensões e elementos do APL Tur, visando à realização dos seus indicadores. O modelo APL Tur é um instrumento adequado para diagnosticar e para identificar oportunidades de melhoria da atividade turística como polarizadora ou como alternativa do desenvolvimento regional. / This is a descriptive-explanatory and exploratory research about the relationship between Tourism and regional development that presents the LPS Tour analysis model and that was worked out taking its basis elements of theoretical references from economics, geography, sociology, administration, communication, anthropology and Tourism. The objective is to show if it is possible to accomplish regional development through Tourism. A region is a territorial context delimitated by geographical, economic, and political criteria. It is taken into consideration that the general model of Local Productive System of Tourism LPS, a especific cathegory of cluster, has limitations to the analysis and management of Tourism. For this reason, the LPS Tour Local Productive System of Tourism - particular model of analysis - is developed. The LPS model is structured in the economic, cultural, and organizational dimensions. The regional development elements in the economic dimension are: spatial delimitation; intra-regional dissimilarities; externalities; environmental sustainability; and social inclusion. The Tourism elements circumscribed in the economic dimension are: supply and demand; performance; prioritization; exportation; productive circuit; extra-regional interactivity; and accessibility. In the cultural dimension, the elements that stand out are: historical aspects; collections and incentives; esthetics; products and attractions; animation; and motivation and satisfaction of the community. In the organizational dimension, we have the following elements: power and stock capital; systemic management; divulgation and image; marketing and commercialization; planning; entrepreneurship and innovation; and knowledge. The test of the LPS Tour model was performed applying it to the Região das Hortênsias (Rio Grande do Sul, Brasil) (The Hydrangeas Region), that is formed by the municipal districts of Gramado, Canela, Nova Petrópolis, and São Francisco de Paula. Based on the panel of indicators of the dimensions elements of the LPS Tour model, it is possible to verify that Tourism contributes to the regional development because it accomplishes the ingression, production and distribution of richness. However, for a region to develop economically through Tourism, an adequate management of its dimensions and elements is necessary, aiming at the accomplishment of the LPS Tour indicators. The LPS Tour model is an adequate instrument to diagnose and to identify opportunities of improvement of the tourist activity, as a polarizer or as an alternative to regional development.

An exceptionally important stress response of Pseudomonas putida strains to toxic chemicals is the induction of efflux pumps that extrude solvents, as well as other toxicants, into the surrounding medium. However, the bacterial tolerance mechanisms are still not fully understood, thus in this thesis metabolomic approaches were used to detect and identify metabolites involved in P. putida DOT-T1E tolerance to abiotic stresses, in particular focussing on the role of efflux pumps. To elucidate any metabolome alterations several strains of P. putida, including the wild type DOT-T1E, and the efflux pump knockouts DOT-T1E-PS28 and DOT-T1E-18, were challenged with different levels of propranolol. Fourier-transform infrared (FT-IR) spectroscopy, which provided a rapid, high-throughput metabolic fingerprint of P. putida strains, was used to investigate any phenotypic changes resulting from exposure to propranolol. FT-IR data illustrated phenotypic changes associated with the presence of propranolol within the cell that could be assigned to the bacterial protein components. To complement this phenotypic fingerprinting approach metabolic profiling on the same samples was performed using gas chromatography mass spectrometry (GC-MS) to identify metabolites of interest during growth of bacteria following this toxic perturbation with propranolol. GC-MS revealed significant changes in ornithine levels which can be directly linked to bacterial tolerance mechanisms, and alterations in the levels of several other metabolites which were also modified in response to propranolol exposure. Moreover, the effect of the organic solvent toluene was also investigated using the same approach. Examination of FT-IR data indicated that protein and fatty acids were the most affected components of P. putida strains due to the presence of toluene within the cell. Moreover, application of GC-MS allowed for the identification and quantification of several metabolites which were differentially produced or consumed in the presence of toluene. To investigate the role of efflux pumps in P. putida DOT-T1E, several analytical techniques were employed including Raman spectroscopy, gas and liquid chromatography to identify and quantify the level of propranolol or toluene in P. putida cells. These analyses showed that propranolol and toluene accumulated in the mutant P. putida DOT-T1E-18 (lacking the TtgABC pump) at higher levels in comparison with the levels found in the wild-type DOT-T1E and the mutant DOT-T1E-PS28 (lacking the TtgGHI pump), indicating the key role of efflux pumps in solvent tolerance. Furthermore, the effect of Mg2+ and Ca2+ on the stabilisation of the toluene tolerance of P. putida DOT-T1E strains was examined in order to elucidate whether divalent cations interact with efflux pumps or other resistant mechanisms to improve solvent tolerance. FT-IR analysis suggested that the influence of divalent cations on the stabilisation of the toluene tolerance could be due to the contribution of metal ions towards other tolerance mechanisms such as lipopolysaccharide (LPS) instead of enhancing the activity of efflux pumps. In conclusion, this thesis presents evidence that phenotypic fingerprinting and metabolic profiling approaches in combination with chemometric methods can generate valuable information on phenotypic responses occurring within microbial cultures subjected to abiotic stress.

Numerous studies have focused on identifying novel probiotic-based treatment options for immune homeostasis maintenance and favorable modulation of the gut microbiota which acts as a key regulator of the gut-brain axis. Recently, probiotics interventions are gaining interest as effective approaches to treat neuropsychiatric disorders through the gut-brain axis. However, there is limiting knowledge about probiotics' effects during puberty on the developing brain and immune responses. Probiotic intake could offer a strategy to counteract the immune, microbial and behavioral disturbances induced by inflammatory LPS. Thus, we hypothesized that the intake of a novel probiotic bacterium Rouxiella badensis subsp. acadiensis would modulate the immune response and that pubertal administration could mitigate LPS- induced inflammation and prevent enduring behavioral changes later in life. We investigated the interaction of the probiotic with the intestinal mucosa and its ability of modulating the gut mucosal immunity (Article 1). Next, we examined the ability of pubertal treatment with R. badensis subsp. acadiensis to alleviate LPS-induced anxiety-like and depression-like behaviors in adult male and female mice and to affect the expression of 5HT1A receptors in specific brain areas of adult mice (Article 2). We finally studied the ability of R. badensis subsp. acadiensis treatment during puberty to mitigate the effects of LPS on the immune system and on the gut microbiome composition (Article 3). These studies have demonstrated the ability of R. badensis subsp. acadiensis to survive the gastrointestinal conditions, interact with the gut epithelium and modulate the intestinal homeostasis. Pubertal use of the bacterium was associated with sex-specific effects on the acute immune response, microbiome structure, enduring neurobehavioral outcomes and the expression of 5HT1A receptors in specific brain areas, later in life. This dissertation emphasizes on the importance of puberty as a window of opportunities during which probiotic use can alleviate the long-term neural, behavioral, immunological and microbiome alterations induced by stress.

<p> Although historically treated as separate systems, there is considerable interaction between the immune system and brain. It has become increasingly clear that immunebrain communication is important to both health and disease. An immunogenic challenge given during the first postnatal week in rodents impacts the developing central nervous system (CNS) leading to long-term behavioural and molecular alterations reflective of enhanced stress-reactivity. Anxiety and depression are stress-related pathologies with a proposed neurodevelopmental origin suggesting that perturbation to neonatal immunebrain signalling may contribute to psychopathology. The current body of work examined the long-term impact of an early immune challenge on behavioural and molecular phenotypes associated with anxiety and depression. Mice were administered lipopolysaccharide (LPS) on postnatal days three and five. The emergence of anxietyrelated behaviour was characterized along the developmental trajectory of LPS-mice concurrent with changes to serotonergic neurocircuitry. Adult depressive-related behaviour was assessed in the forced swim test (FST) along with hippocampal neurogenesis as revealed by immunoreactivity for bromodeoxyuridine (BrdU) and doublecortin (DCX). The results demonstrated a sex-specific alteration in both the temporal emergence and phenotypic variant of anxiety-related behaviours displayed by LPS-mice. This was accompanied by changes to CNS serotonergic-related gene expression that coincided with a critical developmental time window essential to the establishment of emotionality. Adult LPS-mice exhibited hyperactivity during the FST that was accompanied by increased doublecortin immunoreactivity in the dorsal and ventral hippocampus, reflecting enhanced immature neuronal differentiation. The current results demonstrate that an early immune challenge impacts the developing CNS leading to enhanced emotional-reactivity. Altered serotonergic neurocircuitry and adult hippocampal neurogenesis may underlie behavioural abnormalities. The current body of work demonstrates a preeminent role for early-life immune disturbance in psychopathology and advances understanding of how immune-brain signalling impacts the developing CNS and confers risk for later disease. </p> / Thesis / Doctor of Philosophy (PhD)