Fundação de Amparo à Pesquisa do Estado do Rio de Janeiro / As formas epimastigotas de Trypanosoma cruzi proliferam e se diferenciam no interior de diferentes compartimentos do trato digestivo dos triatomíneos. Esses ambientes antagônicos, no que diz respeito à concentração de nutrientes, pH e status redox, constituem um desafio para o protozoário por conterem moléculas e fatores capazes de deflagrar diferentes sinalizações e respostas no parasito. Por isso, testamos a influência de produtos abundantes do metabolismo do vetor e de status redox distintos, frente aos processos de proliferação e diferenciação in vivo e in vitro. Como exemplo temos o heme e a hemozoína, subprodutos da digestão da hemoglobina, e o urato, rico na urina dos insetos. O heme é uma importante molécula em todos os seres vivos. Nosso grupo mostrou seu papel na proliferação in vitro de T. cruzi e que esse sinal é governado pela enzima redox-sensível CaMKII (Lara et al., 2007; Souza et al., 2009). Esse efeito parece depender de uma sinalização redox, onde o heme e não seus análigos induz a formação de EROs, modulando a atividade da CaMKII (Nogueira et al, 2011). Apesar de gerar espécies reativas de oxigênio (EROs) em formas epimastigotas, o heme não alterou a ultraestrutura desses parasitos mostrando uma adaptação a ambientes oxidantes. Além disso, a adição de FCCP inibiu a formação de EROs mitocondrial, diminuindo a proliferação dos parasitos. Em contrapartida, a AA aumentou drasticamente a produção de EROs mitocondrial levando à morte dos epimastigotas. Estes resultados confirmam a hipótese de regulação redox do crescimento de epimastigotas. A formação de β- hematina (hemozoína) constitui uma elegante estratégia para minimizar o efeito tóxico do heme nos insetos hematófagos. Contudo, a β-hematina não influenciou a proliferação ou a metaciclogênese in vitro. Já o urato, e outros antioxidantes clássicos como o GSH e o NAC prejudicaram a proliferação in vitro de epimastigotas. Estes efeitos foram parcialmente revertidos quando os antioxidantes foram incubados juntamente com o heme. Durante a metaciclogênese in vitro, o NAC e o urato induziram um aumento significativo das formas tripomastigotas e levaram a diminuição da porcentagem de formas epimastigotas. Em contrapartida, o heme e a β-hematina apresentaram o efeito oposto, diminuindo a porcentagem de formas tripomastigotas e aumentando a de epimastigotas. No intuito de confirmar a influencia do status redox na biologia do parasito in vivo, nós quantificamos a carga parasitária nas porções anterior e posterior e no reto do triatomíneo alimentado na presença ou na ausência de NAC e urato por qPCR. O tratamento com os antioxidantes aumentou a carga parasitária em todas as partes do intestino analisadas. Posteriormente, para diferenciar as formas evolutivas responsáveis pelo incremento da carga parasitária, foram realizadas contagens diferenciais nas mesmas porções do intestino do inseto vetor. Cinco dias após a infecção foi observado aumento significativo de formas tripomastigotas e diminuição de formas epimastigotas in vivo. Em conjunto, estes dados sugerem que, assim como a concentração de nutrientes e o pH, o status redox também pode influenciar a biologia do T. cruzi no interior do inseto vetor. Neste cenário, moléculas oxidantes agiriam a favor da proliferação, e em contraste, antioxidantes parecem favorecer a metaciclogênese. / Trypanosoma cruzi epimastigotes proliferate and differentiate inside different compartments of the triatomines gut. These environments are antagonic in terms of nutrient content, pH and redox status. All these factors represent a challenge to the protozoan due to the presence of molecules and factors which are able to induce different signals to the parasite. Thus, we tested the influence of abundant metabolism products of the vector, with distinct redox status, in the proliferation and metacyclogenesis in vitro and in vivo. These molecules are heme and hemozoin, both byproducts of hemoglobin digestion, and urate, present in the urine of insects. Heme is a ubiquitous molecule present in all living organisms. Our group studied its role in T. cruzi growth in vitro, showing that this signal is governed by the redox-sensitive enzyme CaMKII (Lara et al., 2007; Souza et al., 2009). Indeed, it seems to rely on a redox signaling pathway in which heme, but not its analogs, induces ROS formation, thus modulating CaMKII activity (Nogueira et al., 2011). Although it induces ROS in epimastigotes, the heme molecule had no deleterious effect upon the parasites ultrastructure, suggesting an adaptation to oxidative environments. In addition, FCCP inhibited mitochondrial ROS formation, then decreasing the parasite proliferation. On the other hand, AA drastically increased mitochondrial ROS production leading to cell death. These results corroborate the hypothesis of redox regulation of epimastigotes proliferation. Hemozoin (β- hematin) formation is an elegant strategy to minimize the toxic effect of heme in hematophagous insects. However, β-hematin had no influence upon the proliferation or metacyclogenesis in vitro. Also, urate, GSH and NAC impaired epimastigote proliferation. These effects were partially reversed when the antioxidants were incubated along with heme. During metacyclogenesis in vitro, NAC and urate induced a significant increment of trypomastigotes and decreased the percentage of epimastigotes. Heme and β-hematin presented the opposite effect diminishing the percentage of trypomastigotes and increasing the percentage of epimastigotes. To confirm the influence of the redox status in the parasite biology in vivo, we quantified the parasite loads in the anterior and posterior midguts and in the rectum of the triatomine vector fed with or without NAC and urate by qPCR. The treatment with the antioxidants increased the parasite loads in all midgut sections analyzed. Afterwards, in order to distinguish the evolutive forms responsible for the increment of parasite loads, we performed differential counting of the midgut sections. Five days post infection we observed an increment of trypomastigotes and a decrease of epimastigote forms in vivo. Taken together, these data suggests that, as well as nutrient content and pH, the redox status may also influence T. cruzi biology in the vector. In this scenario, oxidants act to turn on proliferation and in contrast, antioxidants seem to switch the cycle towards metacyclogenesis.

L'acidose ruminale est l'une des préoccupations majeures des exploitations laitières actuelles. Les levures vivantes (LV) ont été largement étudiées et utilisées chez les vaches laitières pour stabiliser la fermentation ruminale. Récemment, la mesure du potentiel redox ruminal (Eh, en mV) a été considérée comme un outil intéressant pour indiquer le trouble de la fermentation ruminale. L'effet positif de LV sur Eh ruminal a été rapporté, mais il reste variable selon les conditions expérimentales. Les objectifs de ce travail étaient de fournir une meilleure compréhension du mode d'action de LV et de définir la condition optimale de l'utilisation de LV chez les vaches laitières. La première partie de ce travail a consisté en une analyse quantitative des résultats de 22 expériences avec des vaches laitières canulées. La deuxième partie de ce travail a consisté à vérifier certains des résultats de l'analyse quantitative par une expérience chez des vaches en lactation. En utilisant l'analyse quantitative de données existantes provenant d'expériences antérieures, nous avons clarifié la relation entre le Eh ruminal et d'autres paramètres ruminaux principaux tels que le pH et le profil VFA, et suggéré que les variations de Eh pourraient être liées au transfert d'électrons dans les réactions dans le rumen. En outre, la réponse du Eh après la supplémentation en LV était également liée à celle du profil AGV ruminal, suggérant que l'effet de LV sur le profil VFA était atteint par l'augmentation du pouvoir réducteur, reflétant un meilleur transfert d'électrons dans le rumen. L'analyse a en outre démontré que la régulation du Eh ruminal par LV serait particulièrement efficace lorsque le risque de troubles digestifs est élevé. Puisque l'influence des caractéristiques de la ration sur le Eh ruminal a été quantifiée, l'effet de LV dans un régime donné pourrait être estimé indirectement. En outre, l'analyse quantitative a également révélé que la réponse de Eh suite à la supplémentation en LV était associée à la quantité de sucres solubles ingérée. L'expérience in vivo chez des vaches en début de lactation a confirmé un effet plus important de LV sur Eh ruminal avec une ration riche en sucres solubles, et a démontré que la supplémentation en LV avait un impact sur la richesse des bactéries, et que les métabolites ont également été influencés par la supplémentation en LV, probablement associée à la diminution du Eh ruminal. / Ruminal acidosis is one of the major concerns of current dairy farms. Live yeasts (LY) have been extensively studied and used in dairy cows for stabilization of rumen fermentation. Recently, measurement of ruminal redox potential (Eh, in mV) has been considered as an interesting tool to indicate ruminal fermentation disorder. The positive effect of LY on ruminal Eh has been reported, but it remains variable according to the experimental conditions. The aims of this work was to provide better understanding of mode of actions of LY, and to define the optimal condition of LY utilization in dairy cows. The first part of this work consisted to quantitative analysis of existing results from 22 experiments with cannulated dairy cattle. The second part of this work consisted to verify some of the results from quantitative analysis by an in vivo experiment in lactating cows. By using quantitative analysis of existing data from previously conducted experiments, we clarified the relationship between ruminal redox and other main ruminal parameters such as pH and VFA profile, and suggested that Eh variations might be related to the transfer of electrons in the reactions producing VFAs in the rumen. Moreover, response of ruminal Eh following live yeast supplementation was also related to that of ruminal VFA profile, which suggested that the effect of LY on VFA profile was achieved via the increase of reducing power, possibly reflected improved electron transfer and use in the rumen. The analysis further demonstrated that the regulation of ruminal Eh by LY would be particularly effective when risk of digestive disorder is high. Since the influence of dietary characteristics on ruminal Eh was quantified, the effect of LY in a given diet could be indirectly estimated. In addition, quantitative analysis also associated the response of ruminal Eh following LY supplementation to the intake of soluble sugars. The in vivo experiment in early-lactating cows confirmed greater effect of LY on ruminal Eh in diet rich in soluble sugars, and further demonstrated that i) LY supplementation tended to impact the richness of ruminal bacteria, and ii) some unidentified metabolites were also influenced by LY supplementation, probably associated to the decrease of ruminal Eh.

A ausência de XPC, uma proteína canonicamente envolvida em reparo de DNA por excisão de nucleotídeos, está associada a vários fenótipos característicos de disfunção mitocondrial como o desequilíbrio entre os complexos da cadeia transportadora de elétrons (CTE), redução no consumo de oxigênio, maior produção de peróxido de hidrogênio, e maior sensibilidade a agentes que causam estresse mitocondrial. Contudo, uma descrição mecanística da relação entre deficiência de XPC e disfunção mitocondrial ainda não está bem estabelecida. Aqui mostramos que a deficiência de XPC está associada ao aumento na expressão do supressor de tumor p53. Essa alteração é acompanhada pelo aumento da expressão de diversas proteínas que participam em importantes funções mitocondriais. A inibição de p53 reverte a superexpressão de algumas dessas proteínas. O tratamento com o inibidor do Complexo III da CTE antimicina A induz aumento da expressão de p53 de forma mais acentuada na linhagem Xpc-/-, enquanto o tratamento com o antioxidante N-acetilcisteína diminue a produção basal de H2O2, expressão de p53 e sensibilidade aumentada ao tratamento com antimicina A. Em conjunto, nossos resultados suportam a hipótese de que o aumento da produção de H2O2 em células Xpc-/- tem um papel causal na regulação da expressão de p53 e na disfunção mitocondrial / Although XPC has been initially implicated in the nucleotide excision DNA repair pathway, its deficiency is associated with mitochondrial dysfunction, including unbalanced electron transport chain (ETC) activity, lower oxygen consumption, increased hydrogen peroxide production, and greater sensitivity to mitochondrial stress. However, a mechanistic understanding of the role of XPC in regulating mitochondrial function is still not well established. Here we show that XPC deficiency is associated with increased expression of the tumor suppressor p53, which is accompanied by increased expression of several proteins that participate in important mitochondrial functions. Inhibition of p53 reverses the overexpression of some of these proteins. In addition, treatment with the ETC inhibitor antimycin A induces p53 expression more robustly in the Xpc-/- cells, while treatment with the antioxidant N-acetylcysteine decreases basal H2O2 production, p53 expression and sensitivity to antimycin A treatment. Together, our results support a model in which increased H2O2 production in Xpc-/- causes upregulation of p53 expression and mitochondrial dysfunction

O objetivo do presente trabalho foi comparar os efeitos dos distintos ambientes endócrinos peri-ovulatórios na proliferação celular, no metabolismo das poliaminas e no ambiente redox do útero bovino durante o diestro inicial. 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). Trinta 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 D7 um subgrupo dos animais foi abatido e amostras de endométrio e lavado uterino foram coletadas para as análises laboratoriais. Os animais do grupo FG-CLG apresentaram no D0 um folículo 1,2 vezes maior e estradiol 2,3 vezes maior em relação ao FP-CLP. Consequentemente, as vacas do FG-CLG desenvolveram CL maiores (1,6 vezes) e mais pesados (1,4 vezes) o que proporcionou no D7 concentrações de P4 1,5 vezes maior em relação ao FP-CLP. Primeiramente, uma análise do transcriptoma endometrial foi realizada e indicou que processos celulares como proliferação, composição de matrix extracelular, processos metabólicos e processos de oxirredução foram afetados pelo modelo experimental. Para avaliar a condição proliferativa e/ou apoptótica endometrial análises de imunomarcação foram realizadas (Ki-67, marcador de proliferação e caspase 3 ativada, marcador de apoptose). O número de células positivas para Ki-67 no epitélio luminal foi 4,1 vezes maior no FP-CLP, porém não houve diferença na marcação da caspase 3 ativada. No epitélio glandular tanto o ki-67 (1,4 vezes) como a caspase 3 ativada (2,6) estavam aumentados no grupo FG-CLG. Em relação à síntese das poliaminas não houve diferença entre os grupos nas concentrações das poliaminas e abundância dos transcritos e proteína das enzimas de síntese. Por outro lado, foi detectada uma forte associação entre a expressão do gene SAT1 com o gene ODC1 (r2: 0,73; P<0,01) e também entre os genes AZIN1 e AMD1 (r2: 0,61; P<0,01). Quanto ao ambiente redox não houve diferença entre os tratamentos no lavado uterino mas no endométrio, no grupo FP-CLP, houve redução da atividade das enzimas Catalase (0,5 vs 0,79 U/mg proteína; P<0,01), glutationa peroxidase (2,0 vs 2,43 nmol NADPH/min/mg proteína; P<0,01), aumento da atividade da Superóxido dismutase (44,77 vs 37,76 U; P=0,04) e da peroxidação lipídica (28,5 vs 17,43 nmol/MDA/mg proteína; P<0,001), mas sem alterar as quantidades da espécies reativas. De acordo com os resultados obtidos, pode-se concluir que o ambiente endócrino peri-ovulatório regula a proliferação endometrial mas sem alterar a via de síntese das poliaminas e modula também o ambiente redox uterino / This work aimed to compare the effects of the different periovulatory endocrine milieu on the several molecular pathways: as cellular proliferation, polyamine metabolism and redox environment of the bovine uterus during the early diestrus. For this, we controlled pharmacologically the follicular growth to induce the ovulation of larger follicle (Large Follicle-Large Corpus lutem group LF-LCL) or smaller follicle (Small follicle-Small Corpus luteum group SF-SCL). Thirty multiparous Nelore cows were synchronized. Fifteen cows were assigned for each group. On the LF-LCL, the cows received a dose of prostaglandin F2&#945; (PGF), a progesterone device and an estradiol benzoate on D10. On the time of progesterone device was withdraw (between D1.75 e D2.5) all the animals received a dose of PGF. The ovulation was induced with buserelin acetate (D0). The unique difference between the groups was that animals of SF-SCL did not received a dose of PGF on D10 and the progesterone device withdraw was between D1.25 e o D1.5. On D7 a subset of animals was slaughtered and the endometrial and uterine washings samples were collected for laboratorial analyses. On D0 the animals of LF-LCL had a 1.2 times larger follicles and 2.3 times higher levels of estradiol than animals of SF-SCL. Firstly, an endometrial transcriptome analyses was realized and suggested that several cellular processes as proliferation, metabolic processes and oxi-reduction processes were affected by experimental model. To evaluate the proliferative and/or apoptotic condition, endometrial immunohistochemistry analyses were realized (ki-67, proliferation marker and caspase 3 activated, apoptose marker). The number of Ki-67 positive cells in the luminal epithelium was 4.1 times higher on SF-SCL. However, there was no difference in caspase 3 activated analyses. On the glandular epithelium both ki-67 (1.4) and caspase 3 activated (2.6) were increased on LF-LCL. About polyamine synthesis pathway there was no difference in polyamines levels and gene and protein expression of synthesis enzymes between the groups. In other hand, it was detected a strong association between SAT1 gene expression and ODC1 gene expression (r2: 0.73; P<0.01) and also between AZIN1 and AMD1 gene expression (r2: 0.61; P<0.01). On the redox environment analysis there was no difference between the groups on the uterine washings but the SF-SCL group had lower endometrial catalase (0.5 vs. 0.79 U/mg protein, P<0.001) and glutathione peroxidase (GPx; 2.0 vs. 2.43 nmol NADPH/min/mg protein, P=0.04) activity, as well as higher lipid peroxidation (28.5 vs. 17.43 nmol MDA/mg of protein, P < 0.001) and superoxide dismutase (SOD) activity (44.77 vs. 37.76 U; P=0.04). There were no differences in the endometrial reactive species (RS) between the groups. In conclusion, the periovulatory endocrine milieu regulates the endometrial proliferation but without alterations in polyamine metabolism and also modulates the uterine redox environment in beef cattle

Nesta pesquisa são apresentados os resultados de uma investigação que teve como objetivo acessar o conhecimento docente de futuros professores de química, no contexto do Programa Institucional de Bolsa de Iniciação à Docência (Pibid). Esta pesquisa figura como um estudo de caso. Os dados se baseiam nas narrativas registradas por meio de gravações audiovisuais de três participantes da pesquisa durante as reuniões do subprojeto Pibid-química, regências e entrevistas semiestruturadas. Além destes, foram analisados relatórios, planos de aula, questionários dos pibidianos e um instrumento de percepção dos alunos da escola conveniada sobre o conhecimento docente dos licenciandos. Investigaram-se ainda os possíveis impactos do Pibid na formação da coordenadora e supervisora que orientaram os pibidianos. Buscou-se, por meio dos diversos documentos, indícios do conhecimento docente dos licenciandos bolsistas Pibid. A análise dos dados qualitativos teve como fundamentação teórica a análise de conteúdo, sendo as categorias provenientes do modelo do conhecimento base em conjunto com o modelo pentagonal. A análise do instrumento de percepção permeou métodos estatísticos. Para analisar as marcas deixadas pelo Pibid na formação da coordenadora e supervisora, adotaram-se as categorias do modelo conhecimento docente e outras emergentes. O pibidiano Antônio apresentou um bom conhecimento de conteúdo, de seus alunos e de suas limitações em relação ao conteúdo redox. Por outro lado, em algumas passagens das aulas, teve dificuldades em organizar a classe e o tempo de sua aula ao conteúdo. Antônio fez uso, em suas sequências didáticas, de diferentes instrumentos avaliativos. As suas estratégias de ensino se baseavam principalmente em práticas experimentais, na leitura, entre outros. A pibidiana Melissa demonstrou não dominar o conteúdo de reações redox, não ter conhecimento de currículo e de estratégias para desenvolver o conceito que ensinava. Grande parte dos conhecimentos docentes que emergiram dos documentos analisados da pibidiana foi reflexo das orientações que recebia das tutoras e também da experiência oportunizada pelo Pibid de como acessar as dificuldades dos alunos com o conteúdo específico, experimentar a gestão de classe e a organização de aulas. As avaliações adotadas pela pibidiana tinham por objetivo coletar dados para o subprojeto. Durante a sua regência não demonstrou uma preocupação com a aprendizagem dos alunos, mas em executar o currículo pretendido e aplicar as avaliações. O pibidiano Mateus demonstrou também limitações com o conteúdo e em selecionar os conceitos necessários para a aprendizagem dos discentes. Apresentou ainda dificuldades com a gestão do tempo e da classe. Por outro lado, passou a conhecer as barreiras conceituais dos discentes com o conteúdo redox, a partir de sua vivência no Pibid. Diversas das decisões sobre o fazer no ensino de ciências eleitas por Mateus foram orientações da supervisora. Pelo mapeamento infere-se que os conhecimentos para a docência acessados pelos pibidianos foram reflexo do Pibid personificado na figura das tutoras e pela experiência em sala. Evidenciou-se ainda que o Pibid vem contribuindo para a formação continuada da coordenadora, por possibilitar que a professora universitária conheça, por exemplo, a realidade da escola da educação básica; e da supervisora, por conceber-se como co-formadora dos licenciandos. Conclui-se que o conhecimento docente acessado por Mateus e Melissa foi influenciado pelas orientações da coordenadora. Acredita-se que o contato com o futuro ambiente profissional e a tutoria pelas docentes experientes oportunizaram aos pibidianos acessar os conhecimentos inerentes ao desenvolvimento do conhecimento base e do PCK. / This study presents the results of a research that aimed to access the teacher knowledge of future chemistry teachers, in the context of the Institutional Program for Scholarships for Initiation in Teaching (Pibid). This is a case study. Data were collected from audiovisual recordings of narratives from three participants during the Pibid-chemistry meetings, from regencies and from semi-structured interviews. In addition, Pibid-participants\' reports, lesson plans and questionnaires were analyzed, and an instrument that assessed the students\' perception about the teacher knowledge of the undergraduate students regarding teaching practices. The possible impacts of Pibid on the training of the coordinator and supervisor who guided the Pibid participants were also analyzed. Several documents were analyzed for evidence of the teacher knowledge of the Pibid students. The theoretical background for the analysis of qualitative data was based on content analysis, with categories from the base knowledge and the pentagonal model. The analysis of the students\' perception instrument was based on statistical methods. In order to analyze the marks left by Pibid in the training of the coordinator and supervisor, the study adopted the categories of teaching knowledge´model and other emerging ones. Pibid participant Antônio presented consistent content knowledge, students knowledge and students difficulties knowledge in relation to the redox content. On the other hand, in some occasions during classes, he had difficulties organizing the class and the content into the class time. Antônio used, in his didactic sequences, different evaluation instruments. His teaching strategies were based mainly on experimental practices, reading, among others. Pibid-participant Melissa demonstrated not master the redox reactions content, she did not have any knowledge about curricula and so could not develop strategies to work with the subject she taught. Much of the teaching knowledge that emerged from the documents analyzed from Melissa reflects the guidelines that she received from the coordinator and supervisor and from the experience offered by Pibid: how to access students\' difficulties with a specific content, how to deal with class management and organizing the classrooms. The assessment instruments adopted by Melissa were aimed at collecting data for the subproject. During her regency, she did not show concern for student learning, but in executing the intended curriculum and applying the assessments. Another Pibid participant, Mateus also seemed to have limited content knowledge and about selecting the necessary concepts for students learning. He also presented difficulties with time and class management. On the other hand, he came to understand the students\' conceptual difficulties with the redox content from his experience in Pibid. Several of the decisions that must be made in science education chosen by Mateus were guidelines given by his supervisor. From the mapping, it is inferred that the teaching knowledge accessed by the participants was a reflection of Pibid participation, personified in the figure of the tutor and by the experience in the classroom. It was also evidenced that Pibid contributes to the continued professional developing of the coordinator, by enabling university professors to know, for instance, the reality of basic education schools; and of supervisors, for conceiving themselves as co-teachers of the teaching students. The conclusion drawn is that the teaching knowledge accessed by Mateus and Melissa were influenced mainly by the coordinator\'s guidelines. It is believed that contact with their future work environment and tutoring by experienced teachers were fundamental for Pibid participants to access the knowledge inherent to the development of basic knowledge and the PCK.

Ce travail de thèse s’est intéressé au développement de complexes de métaux non-nobles portant des ligands non-innocents et leurs applications en catalyse comme alternatives efficaces aux complexes de métaux nobles dans un contexte de fortes préoccupations économiques et environnementales. Ainsi, nous avons synthétisé et caractérisé des complexes de fer et de nickel portant des ligands non-innocents, à l’aide de différentes techniques spectroscopiques. Ces ligands pourraient moduler la réactivité du métal et étendre ainsi le champ d’applications de ces métaux de transition. Dans le but de développer de nouvelles méthodes de synthèse en catalyse au fer, des complexes de ce dernier avec des ligands bis(imino)pyridines ont été évalués pour une réaction tandem d’activation/arylation d’arènes non activés. Des études mécanistiques préliminaires, basées sur des données spectroscopiques (RMN, IR in situ, RPE) et théoriques (DFT), ont permis de suggérer un mécanisme différent de ceux connus pour le fer dans la littérature et n’est pas compatible avec un mécanisme de substitution aromatique radicalaire (HAS). Nous nous sommes également intéressés à un autre domaine majeur en catalyse : les réactions de couplage croisés catalysées par des complexes de nickel portant des ligands redox non-innocents. Les défis actuels de la catalyse au nickel sont la réalisation de couplage croisés d’halogénures d’alkyle non activés et les mécanismes impliqués diffèrent généralement de ceux mis en jeu dans les réactions de couplage croisé pallado-catalysées. Les réactions de couplage croisé catalysées par des complexes de nickel portant des ligands redox non-innocents ont été étudiées afin de découvrir de nouvelles réactivités et d’avoir une meilleure compréhension des cycles catalytiques mis en jeu. / This PhD work has focused on the development of complexes of non-noble metals bearing non-innocent ligands and their catalytic applications as efficient alternatives to noble metal complexes, in the light of increasing concerns regarding cost and sustainability-related issues of noble metals. Towards this goal, we have developed and characterized complexes of non-noble metals (Fe, Ni) with non-innocent ligands using multiple spectroscopic techniques. This work was aimed at broadening the field of useful catalytic applications of these particular complexes. For our dedicated program in iron catalysis, a new method for tandem C–H activation/arylation of unactivated arenes catalyzed by iron complexes bearing redox-active bis(imino)pyridine ligands was developed. Preliminary mechanistic insights were gained based on combined spectroscopic data (NMR, in situ IR, EPR), reactivity studies as well as DFT calculations. The results obtained are clearly in favor of a mechanism distinct to that previously reported for iron-based catalytic systems and are not compatible with homolytic aromatic substitution (HAS). We have also focused on another challenging field in catalysis: cross-coupling reactions catalyzed by nickel as base metal, bearing redox non-innocent ligands. Several challenges in cross-coupling reactions remain among which coupling of non-activated alkyl halides. Mechanisms involving nickel catalysts often differ from those involved in palladium catalyzed cross-coupling reactions. The behavior of nickel complexes bearing redox non-innocent ligands was studied in order to unveil new reactivites and gain a better understanding of the catalytic cycles at stake.

La synthèse et la caractérisation de la structure électronique de nanosystèmes contenant des complexes redox en forme d'étoile ou dendritiques ont été conduites en vue d'applications dans le domaine des matériaux et de la catalyse. La première partie est consacrée à la description de ces nanomatériaux, la seconde partie à la synthèse et aux propriétés de nanoparticules métalliques pour des applications biomédicales et catalytiques, et la troisième partie à la mise en œuvre et optimisation des propriétés catalytique de nanoréacteurs moléculaires et nanoparticulaires. Ces nanoréacteurs catalytiques se sont montrés particulièrement efficaces pour des réactions de métathèse oléfiniques et pour des réactions de couplage carbone-carbone de type Miyaura-Suzuki. Les réactions catalytiques ont pu être conduites dans l'eau avec des quantités extrêmement faibles de catalyseur. Dans ce dernier cas, le caractère « homéopathique » des catalyseurs a même pu être démontré. / Synthesis and characterization of the electronic structures of star-shape and dendritic and nanoparticle-centered nanosystems containing redox-active groups have been carried out for applications in materials chemistry and catalysis. The first part of the manuscript describes molecular nanomaterials, the second part nanoparticles for catalytic and biomedical applications, and the third part catalytic processes optimized in nanoreactors of molecular or nanoparticle types. The catalytic nanoreactors have been shown to be extremely efficient for olefin metathesis reactions and for Miyaura-Suzuki reactions. Catalytic reactions could be performed in water using extremely low amounts of nanoreactor-stabilized catalysts. In the latter example, catalysis is even « homeopathic ».

A modelagem matemática por meio de equações diferenciais é uma importante ferramenta para prever o comportamento de baterias redox de vanádio, pois ela pode contribuir para o aperfeiçoamento do produto e melhor entendimento dos princípios da sua operação. Os estudos de modelagem podem ser aliados à análise assintótica no intuito de promover reduções ou simplificações que tornem os modelos menos complexos, isso é feito a partir da observação da importância que cada termo exerce sobre as equações. Tais simplificações são úteis neste contexto, visto que os modelos geralmente abordam uma célula apenas - a menor unidade operacional da bateria - enquanto aplicações reais exigem o uso de dezenas ou centenas delas implicando em uma maximização do uso de recursos computacionais. Neste trabalho, foram investigadas múltiplas formas de reduções assintóticas que empregadas na construção dos modelos puderam acelerar o tempo de processamento em até 2,46 vezes ou reduzir os requisitos de memória principal em até 11,39%. As simulações computacionais foram executadas pelo software COMSOL Multiphysics v. 4.4, e também por scripts desenvolvidos em ambiente de programação MATLAB. A validação dos resultados foi feita comparando-os a dados experimentais presentes na literatura. Tal abordagem permitiu também validar as rotinas implementadas para a simulação dos modelos comparando suas soluções com aquelas providas pelo COMSOL. / Mathematical modelling using differential equations is an important tool to predict the behavior of vanadium redox batteries, since it may contribute to improve the device performance and lead to a better understanding of the principles of its operation. Modelling can be complemented by asymptotic analysis as a mean to promote reductions or simplifications that make models less complex. Such simplifications are useful in this context, whereas these models usually addresses one cell only the smallest operating unit while real applications demand tens or hundreds cells implying on larger computational requirements. In this research, several options for asymptotic reductions were investigated and, applied to different models, were able to speed up the processing time in 2.46× or reduce the memory requirements up to 11.39%. The computational simulations were executed by COMSOL Multiphysics v.4.4, also by in-house code developed in MATLAB. The validation of results was done by comparing it to experimental results available in literature. Additionally, correlating the results provided by COMSOL with the ones arising from the implemented sub-routines allowed to validate the developed algorithm.

Les protéines destinées à la sécrétion ou adressées à la membrane transitent par le réticulum endoplasmique (RE) où elles acquièrent leur conformation native et subissent des modifications post-traductionnelles comme la formation de ponts disulfures. Dans ce compartiment, la formation de ponts disulfures repose sur l’activité de l’oxydase Ero1 et de la Protein Disulfure Isomerase (PDI). Ero1 catalyse la formation de ponts disulfures et les transmet à la PDI qui à son tour oxyde les substrats. L’isomérisation ou la réduction terminale des ponts disulfures non-natifs repose sur un système de réduction dans le RE encore non élucidé. Des études suggèrent l’importance du glutathion réduit (GSH) dans ce système de réduction. Le GSH est un tripeptide redox exclusivement synthétisé dans le cytosol. Notre étude s’attache à (i) décrire les flux de glutathion entre RE et cytosol et (ii) identifier les acteurs de ce transport (iii) comprendre l’impact d’une modification de l’homéostasie redox du glutathion sur la physiologie du RE.Nous avons établi un système permettant d’étudier les flux de glutathion entre cytosol et RE. Afin de démasquer ces flux intracellulaires, nous avons utilisé une souche de S. cerevisiae surexprimant le transporteur plasmatique du glutathion, HGT1. Ce système permet de modifier rapidement et drastiquement la concentration cytosolique de glutathion. Les flux intracellulaires engendrés sont ensuite suivis grâce à des sondes redox spécifiques du glutathion adressées dans le RE ou le cytoplasme.(i) Nos résultats suggèrent que le GSH et le GSSG sont importés dans le RE depuis le cytosol. Le GSH est transporté selon un gradient de concentration via un système de transport de diffusion facilité. Ces flux sont également observés lors de stress stimulant la synthèse de GSH (stress thermique, arsenite…).(ii) Le transport de GSH dans le lumen est assuré par le translocon Sec61, et une régulation de cet import par la chaperone luminale Kar2 est observée.(iii) une réduction rapide de l’état redox du glutathion dans le RE conduit à une mort cellulaire programmée non apoptotique, également observée lors d’autre stress RE (traitement tunicamycine). / The endoplasmic reticulum (ER) is the first intracellular compartment of the protein secretion pathway. Protein maturation in this compartment involves protein folding and post-traductionnal modification including formation of disulfide bonds. The formation of disulfide bonds is operated by a highly conserved redox relay made of the thiol oxidase Ero1 and the protein disulfide isomerase (PDI). Ero1p catalyzes disulfide bond formation and relays them by thiol-disulfide exchange to PDI, which in turn oxidizes substrates. Isomerization and terminal reduction of non-native disulfide bonds both rely on a reduction system that remains to be formally identified. Studies however suggest the importance of reduced glutathione in this reducing system. GSH is small redox tripeptide exclusively synthesized in the cytosol. In this study we (i) describe the main parameters of glutathione traffic across the ER membrane (ii) identify the main actors involved in the transport and (iii) analyze the physiological impact of a modification of the ER glutathione redox state.We established a system to monitor the fluxes of glutathione from the cytosol to the ER in S. cerevisiae. To artificially increase fluxes of glutathione, we used a cell over-expressing the GSH plasma membrane transporter HGT1, which when grown in presence of glutathione import high levels of this compound. Consequently, we monitored the intracellular relocation of imported GSH by following GSH fluxes using two specific redox probes. Our data indicate that:(i) GSH is transported into the ER by facilitated diffusion along a concentration gradient. GSSG can also be imported into the ER. Similarly, stress conditions that stimulate GSH synthesis, such as heat shoc, arsenite treatment, also triggered a GSH import in the ER.(ii) GSH import in the ER is achieved by the translocon Sec61, and is regulated by the lumenal chaperone Kar2.(iii) A rapid reduction of glutathione ER redox state leads to the activation of a non-apoptotic programmed cell death pathway, usually observed during high ER stress.

The thesis deals with the treatment of the input of the wine with the subsequent applications to the manufacture of aromatized wine-based drinks. A key element is the theoretical knowledge of the chemical composition of wine with his practical patent processing up to the real form of the final products, which can be placed on the market. Part of the thesis is a legislative preview over the issue and sensory and analytical evaluation in graphical and tabular form.