Os venenos de aranhas do gênero Loxosceles possuem uma ampla gama de atividades em diferentes células, tecidos e sistemas, e é notória a sua capacidade de causar a formação da lesão dermonecrótica. Em um trabalho prévio, relatamos uma intensa trombocitopenia induzida pelo veneno loxoscélico em coelhos. Objetivou-se no presente estudo avaliar primeiramente a interação in vitro entre as plaquetas, humanas e de coelhos, com o veneno de Loxosceles gaucho e seu principal componente, a fração esfingomielinásica. Posteriormente, investigou-se o papel da plaqueta no desenvolvimento da lesão dermonecrótica através de um modelo de trombocitopenia em coelhos. Nos estudos in vitro, os resultados de agregação em plasma rico em plaquetas evidenciou que tanto o veneno total quanto a fração esfingomielinásica induziram um aumento desta resposta nas plaquetas das duas espécies, que foi mais intensa nas plaquetas humanas. Uma vez que em plaquetas lavadas a agregação não foi constatada, confirmou-se a necessidade de componente(s) plasmático(s) para a indução desta resposta. Ainda, os dados de LDH indicaram que o veneno total, nas concentrações aqui utilizadas, não foi capaz de causar à lise plaquetária. Com relação à ativação, o veneno total, foi capaz de aumentar a expressão da LIBS1 nas plaquetas de ambas as espécies e de P-selectina na plaqueta de coelho. Sob as nossas condições experimentais, a fração não promoveu o aumento dos marcadores de ativação. Finalizando os estudos in vitro, o veneno total e a fração provocaram um aumento da adesão nas plaquetas das duas espécies, sendo que a resposta à fração esfingomielinásica mostrou-se dose-dependente. O estudo experimental, por vez, necessitou de uma etapa preliminar para a obtenção de um anticorpo anti-plaqueta, produzido em cabras, que mostrou-se eficaz em induzir um intenso quadro trombocitopênico em coelhos. Análises histopatológicas indicam que a ausência das plaquetas não diminuiu a formação de trombos intravasculares na área da derme sob a ação do veneno. O desenvolvimento da lesão dermonecrótica em coelhos injetados com o veneno loxoscélico mostrou-se mais intenso naqueles depletados de plaquetas do que nos animais com número normal de plaquetas, constatável pelo desenvolvimento de maiores áreas de equimose e das escaras necróticas. Os resultados semelhantes dos níveis plasmáticos do fator de von Willembrand, dos tempos de protrombina e de tromboplastina parcial ativada, assim como dos testes de fagocitose por polimorfonucleares isolados do sangue periférico, entre os coelhos trombocitopênicos e aqueles com contagem normal de plaquetas, evidenciam que não houve alterações expressivas nas células endoteliais, no sistema da coagulação e na capacidade fagocítica via C3b dos polimorfonucleares do sangue periférico. Os presentes resultados mostram que o veneno loxoscélico in vitro é capaz de elicitar em plaquetas humanas e de coelhos as respostas de agregação, de adesão e de ativação. Por fim, as diferenças no desenvolvimento da lesão, constatadas entre o animal trombocitopênico e o animal normal, evidenciam que a plaqueta é um fator importante para controlar a expansão e a gravidade da lesão dermonecrótica induiza em coelhos pelo veneno loxoscélico. / Venoms from Loxosceles spiders exhibit a wide range of activities on different cells, tissues and systems, being specially evident their ability to cause a dermonecrotic lesion. In a previous paper, we showed that Loxosceles gaucho spider venom induces intense thrombocytopenia in rabbits. Thus, in the present study, we firstly aimed to evaluate the in vitro interaction between human and rabbits platelets and L. gaucho venom and its major component, the sphingomyelinase fraction. Secondly, we investigated the platelet role in the dermonecrotic lesion formation using an experimental model of thrombocytopenia in rabbits. Both total venom and its sphingomyelinase fraction stimulated in vitro aggregation per se in platelet-rich plasma from both species, but a more intense response was obtained with human platelets. Taking into consideration that neither total venom nor its sphingomyelinase fraction induced aggregation of washed platelets, it can be deduced that one or more plasma components are necessary to induce this response. Moreover, LDH data indicated that total venom, at least at the concentration levels used in this study, cannot induce platelet lysis. Regarding platelet activation, total venom was able to increase the expression of LIBS1 on both human and rabbit platelet surface and P-selectin on only rabbit platelet surface. However, the sphingomyelinase fraction did not increase the expression of any marker of platelet activation, at least in the experimental conditions used in this study. Another in vitro study showed that both total venom and its sphingomyelinase fraction induce an increase in human and rabbit platelet adhesion, with the sphingomyelinase fraction exhibiting a dose-dependent response. For the study using an animal model of thrombocytopenia, it was necessary to previously produce goat antibodies against rabbit platelets, which showed to be effective to induce an intense thrombocytopenia in rabbits. Histopathological analysis of venom-induced dermic tissue lesion indicated that platelet absence does not decrease the formation of intravascular thrombi in the injured area. Nonetheless, bigger areas of equimosis and necrotic scabs could be observed in the dermonecrotic lesion of rabbits injected with loxoscelic venom and depleted of platelets. In the different groups, plasma levels of von Willembrand factor, prothrombin and activated partial thromboplastin times, and data from phagocytosis tests using polymorphonuclear cells isolated from rabbit periferical blood were similar comparing thrombocitopenic rabbits with those exhibiting normal platelet count; therefore, we can deduce that there is not any expressive alteration in endotelial cells, coagulation system and phagocytic ability of polymorphonuclear leukocytes via their complement receptor C3b. In conclusion, the loxoscelic venom is able to unchain in vitro different mechanisms leading to adhesion, activation and aggregation of both human and rabbit platelets. Furthermore, due to differences in the dermic lesion patterns between thrombocitopenic and normal animals, we can conclude that platelets must play a key role in controlling the expansion and severity of the dermonecrotic lesion induced in rabbits by L. gaucho venom.

Healthy articular joints exhibit remarkable lubrication due in large part to the complex rheological and tribological behavior of the synovial fluid (SF) that lubricates the joints. Current approaches that seek to elucidate such remarkable lubrication usually focus on the roles of high molecular weight SF components such as lubricin and hyaluronic acid but frequently overlook the role of serum albumin (SA), although it represents 90% of the protein content of SF. In this thesis, we used the Surface Forces Apparatus to investigate in detail the structural and tribological response of SA thin films when sheared between model surfaces and subjected to a large range of shearing parameters. Our data indicate that, under shear, SA films reproduce closely the shear response previously reported for SF, i.e., film thickening and formation of numerous long-lived aggregates accompanied by low friction and efficient surface protection against damage. More specifically, our detailed investigation of shear parameters reveals that (i) strong anchoring of SA to surfaces promotes the formation of large rod-like shaped aggregates that enable rolling friction and keep surfaces far apart, preventing damage, (ii) aggregation mechanism is irreversible, which makes aggregates long-lived (though mobile) in the contact, and (iii) aggregate formation only occur when SA was sheared above a ‘critical’ amplitude Ac and a critical shear velocity Vc. Collectively, our results provide experimental evidence of the role of globular proteins, such as SA, in lubrication and establish a correlation between shearing parameters, formation and stability of aggregates, low friction and wear protection. Although our findings are based on experiments involving rigid, nonporous surfaces hence can hardly be generalized to compliant and porous cartilage surfaces, they are applicable to other rigid tribosystems such as artificial joints and will certainly advance our understanding of joint implants’ lubrication in SF mediated by protein aggregation, with implications for future design of artificial joints and therapeutic interventions.

In their natural environments microorganisms exist predominantly in aggregates and biofilms. The ability of bacteria to form aggregates is associated with the biosynthesis of polymers such as polysaccharides. In this study the physical mechanisms underlying bacterial aggregation by extracellular polysaccharides are investigated by utilising the bacterium Sinorhizobium meliloti. S. meliloti biosynthesises an extracellular polysaccharide called succinoglycan, which is well characterised in terms of its structure and biosynthesis. A range of previously constructed succinoglycan biosynthesis mutants were screened for altered aggregation. An S. meliloti exoS mutant (a gain of function mutation that results in a constitutively active two component regulator called ExoS) overproduces succinoglycan and has enhanced aggregation compared to the parent strain, Rm1021. The aggregates settle to the bottom of the culture vessel resulting in loss of turbidity of the cultures and phase separation. Microscopic observation showed that succinoglycan did not appear to be attached to the aggregates, which formed ordered structures of laterally aligned cells. By addition of purified succinoglycan it was found that the critical concentration of polymer required to induce aggregation and phase separation of the cultures decreased with increasing cell concentration. These observations suggest that aggregation of S. meliloti cultures in the presence of succinoglycan is driven by macromolecular crowding, otherwise known as depletion attraction. Depletion attraction can drive the ordered arrangement and aggregation of colloidal particles in the presence of polymers. Aggregation of the particles increases the volume available to the polymers, maximising their entropy and the entropy of the system. Addition of succinoglycan to stationary phase Escherichia coli cultures and polystyrene colloids also resulted in aggregation consistent with depletion attraction. Furthermore alternative polymers such as the bacterial extracellular polysaccharide xanthan produced by Xanthomonas campestris can result in aggregation of bacteria by depletion attraction. Depletion attraction may therefore be a ubiquitous force driving aggregation of crowded dispersions of bacteria and polymers. The second part of the thesis focuses on how depletion driven aggregation can lead to surface-associated biofilm formation. Imaging of the sediment formed by the exoS mutant showed that the structure formed at the base of the culture vessel leads to development of an ordered structure composed of interlinked aggregates. The role of succinoglycan in surface attachment is complex and varies with culture conditions. Depletion attraction may facilitate interaction with a surface but alternative factors may then play a role in anchoring the cells to the surface. Under certain conditions the cells may produce factors which allow binding of the cells to a surface independently of succinoglycan. This study has demonstrated for the first time that an extracellular polysaccharide produced by bacteria can result in aggregation via depletion attraction which may be an under explored mechanism by which aggregation of bacteria can occur.

Complex Event Processing (CEP) is the technical choice for high performance analytics in time-critical decision-making applications. Although current CEP systems support sequence pattern detection on continuous event streams, they do not support the computation of aggregated values over the matched sequences of a query pattern. Instead, aggregation is typically applied as a post processing step after CEP pattern detection, leading to an extremely inefficient solution for sequence aggregation. Meanwhile, the state-of-art aggregation techniques over traditional stream data are not directly applicable in the context of the sequence-semantics of CEP. In this paper, we propose an approach, called A-Seq, that successfully pushes the aggregation computation into the sequence pattern detection process. A-Seq succeeds to compute aggregation online by dynamically recording compact partial sequence aggregation without ever constructing the to-be-aggregated matched sequences. Techniques are devised to tackle all the key CEP- specific challenges for aggregation, including sliding window semantics, event purging, as well as sequence negation. For scalability, we further introduce the Chop-Connect methodology, that enables sequence aggregation sharing among queries with arbitrary substring relationships. Lastly, our cost-driven optimizer selects a shared execution plan for effectively processing a workload of CEP aggregation queries. Our experimental study using real data sets demonstrates over four orders of magnitude efficiency improvement for a wide range of tested scenarios of our proposed A-Seq approach compared to the state-of-art solutions, thus achieving high-performance CEP aggregation analytics.

Monoclonal antibodies (mAbs) represent one of the fastest growing classes of therapeutic proteins. This success is due to a number of attractive properties such as high binding affinity, specificity, low immunogenicity and high aqueous solubility. Despite this, mAbs can suffer from undesirable physical instabilities, especially reversible self-association (RSA), which can lead to aggregation and phase separation. One aspect of formulation is therefore to find solution conditions which minimise mAb aggregation propensity during storage at high concentrations. Hence, the buffer, excipient and pH must be carefully considered to obtain the optimal formulation. Currently, if a platform formulation process is non-ideal for a particular candidate mAb, then an alternative strategy is to utilise high-throughput screening to measure various physical parameters indicative of physical stability. Arginine (in the form of hydrochloride salt Arg·HCl) is often used in formulations exhibiting high RSA and a propensity for aggregation. The interaction of Arg with the protein surface is complex and dependent on both the salt form and concentration. Here the focus was on the glutamate salt of arginine (Arg·Glu), to quantify its effect on mAb conformational and colloidal stability under different pH conditions. Arg·Glu was able to decrease the propensity of the mAbs to aggregate, particularly at pH values closer to their pI.The work also included the use of in vitro cell culture models to examine cell viability in the presence of the various arginine salts over a range of osmolalities. Whilst Arg·Glu is composed of two naturally occurring amino acids and both of which are considered non-toxic individually, the effect of the increased concentrations of their combination, on cells has not been explored previously. In vitro cell lines were chosen to represent the subcutaneous tissue, the effect of Arg·Glu on cell viability was compared against NaCl, Arg·HCl and sodium glutamate (NaGlu). The work concluded there was no additional toxicity associated with the presence of Arg·Glu in the cell culture models studied, therefore Arg·Glu has the potential as an excipient as it reduces aggregation and is nontoxic. Another aspect of the work was to assess the use of solution NMR spectroscopy as an orthogonal technique in mAb formulation characterisation. 1H NMR spectroscopy was used to measure a number of experimental parameters for high concentration mAb solution. The work proposed that 1H NMR spectroscopy can serve as a valuable orthogonal method for mAb characterization and formulation.

Protein aggregation is a frequently cited problem during the development of liquid protein formulations, which is especially problematic since each protein exhibits different aggregation behaviour. Aggregation can be controlled by judicious choice of solution conditions, such as salt and buffer type and concentration, pH, and small molecule additives. However, finding conditions is still a trial and error process. In order to improve formulation development, a fundamental understanding of how excipients impact upon protein aggregation would significantly contribute to the development of stable protein therapeutics. The underlying mechanisms that control effects of excipients on protein behaviour are poorly understood. This dissertation is directed at understanding how excipients alter the conformational and colloidal stability of proteins and the link to aggregation. This knowledge can be used for finding novel ways of either predicting or preventing/inhibiting protein aggregation. Experiments using static and dynamic light scattering, intrinsic fluorescence, turbidity and electrophoretic light scattering were conducted to study the effect of solution conditions such as pH, salt type and concentration on protein aggregation behaviour for three model systems: lysozyme, insulin and a monoclonal antibody. Emphasis is placed on understanding the effects of solution additives on protein-protein interactions and the link to aggregation. This understanding has allowed the rational development of stable formulations with novel additives, such as arginine containing dipeptides and polycations.

Wireless sensor networks used to have a main role as a monitoring tool for environmental purposes and animal tracking. This spectrum of applications, however, has dramatically grown in the past few years. Such evolution means that what used to be application-specific networks are now multi application environments, often with federation capabilities. This shift results in a challenging environment for data privacy, mainly caused by the broadening of the spectrum of data access points and involved entities. This thesis first evaluates existing privacy preserving data aggregation techniques to determine how suitable they are for providing data privacy in this more elaborate environment. Such evaluation led to the design of the set difference attack, which explores the fact that they all rely purely on data aggregation to achieve privacy, which is shown through simulation not to be suitable to the task. It also indicates that some form of uncertainty is required in order to mitigate the attack. Another relevant finding is that the attack can also be effective against standalone networks, by exploring the node availability factor. Uncertainty is achieved via the use of differential privacy, which offers a strong and formal privacy guarantee through data perturbation. In order to make it suitable to work in a wireless sensor network environment, which mainly deals with time-series data, two new approaches to address it have been proposed. These have a contrasting effect when it comes to utility and privacy levels, offering a flexible balance between privacy and data utility for sensed entities and data analysts/consumers. Lastly, this thesis proposes a framework to assist in the design of privacy preserving data aggregation protocols to suit application needs while at the same time complying with desired privacy requirements. The framework's evaluation compares and contrasts several scenarios to demonstrate the level of flexibility and effectiveness that the designed protocols can provide. Overall, this thesis demonstrates that data perturbation can be made significantly practical through the proposed framework. Although some problems remain, with further improvements to data correlation methods and better use of some intrinsic characteristics of such networks, the use of data perturbation may become a practical and efficient privacy preserving mechanism for wireless sensor networks.

Os venenos de aranhas do gênero Loxosceles possuem uma ampla gama de atividades em diferentes células, tecidos e sistemas, e é notória a sua capacidade de causar a formação da lesão dermonecrótica. Em um trabalho prévio, relatamos uma intensa trombocitopenia induzida pelo veneno loxoscélico em coelhos. Objetivou-se no presente estudo avaliar primeiramente a interação in vitro entre as plaquetas, humanas e de coelhos, com o veneno de Loxosceles gaucho e seu principal componente, a fração esfingomielinásica. Posteriormente, investigou-se o papel da plaqueta no desenvolvimento da lesão dermonecrótica através de um modelo de trombocitopenia em coelhos. Nos estudos in vitro, os resultados de agregação em plasma rico em plaquetas evidenciou que tanto o veneno total quanto a fração esfingomielinásica induziram um aumento desta resposta nas plaquetas das duas espécies, que foi mais intensa nas plaquetas humanas. Uma vez que em plaquetas lavadas a agregação não foi constatada, confirmou-se a necessidade de componente(s) plasmático(s) para a indução desta resposta. Ainda, os dados de LDH indicaram que o veneno total, nas concentrações aqui utilizadas, não foi capaz de causar à lise plaquetária. Com relação à ativação, o veneno total, foi capaz de aumentar a expressão da LIBS1 nas plaquetas de ambas as espécies e de P-selectina na plaqueta de coelho. Sob as nossas condições experimentais, a fração não promoveu o aumento dos marcadores de ativação. Finalizando os estudos in vitro, o veneno total e a fração provocaram um aumento da adesão nas plaquetas das duas espécies, sendo que a resposta à fração esfingomielinásica mostrou-se dose-dependente. O estudo experimental, por vez, necessitou de uma etapa preliminar para a obtenção de um anticorpo anti-plaqueta, produzido em cabras, que mostrou-se eficaz em induzir um intenso quadro trombocitopênico em coelhos. Análises histopatológicas indicam que a ausência das plaquetas não diminuiu a formação de trombos intravasculares na área da derme sob a ação do veneno. O desenvolvimento da lesão dermonecrótica em coelhos injetados com o veneno loxoscélico mostrou-se mais intenso naqueles depletados de plaquetas do que nos animais com número normal de plaquetas, constatável pelo desenvolvimento de maiores áreas de equimose e das escaras necróticas. Os resultados semelhantes dos níveis plasmáticos do fator de von Willembrand, dos tempos de protrombina e de tromboplastina parcial ativada, assim como dos testes de fagocitose por polimorfonucleares isolados do sangue periférico, entre os coelhos trombocitopênicos e aqueles com contagem normal de plaquetas, evidenciam que não houve alterações expressivas nas células endoteliais, no sistema da coagulação e na capacidade fagocítica via C3b dos polimorfonucleares do sangue periférico. Os presentes resultados mostram que o veneno loxoscélico in vitro é capaz de elicitar em plaquetas humanas e de coelhos as respostas de agregação, de adesão e de ativação. Por fim, as diferenças no desenvolvimento da lesão, constatadas entre o animal trombocitopênico e o animal normal, evidenciam que a plaqueta é um fator importante para controlar a expansão e a gravidade da lesão dermonecrótica induiza em coelhos pelo veneno loxoscélico. / Venoms from Loxosceles spiders exhibit a wide range of activities on different cells, tissues and systems, being specially evident their ability to cause a dermonecrotic lesion. In a previous paper, we showed that Loxosceles gaucho spider venom induces intense thrombocytopenia in rabbits. Thus, in the present study, we firstly aimed to evaluate the in vitro interaction between human and rabbits platelets and L. gaucho venom and its major component, the sphingomyelinase fraction. Secondly, we investigated the platelet role in the dermonecrotic lesion formation using an experimental model of thrombocytopenia in rabbits. Both total venom and its sphingomyelinase fraction stimulated in vitro aggregation per se in platelet-rich plasma from both species, but a more intense response was obtained with human platelets. Taking into consideration that neither total venom nor its sphingomyelinase fraction induced aggregation of washed platelets, it can be deduced that one or more plasma components are necessary to induce this response. Moreover, LDH data indicated that total venom, at least at the concentration levels used in this study, cannot induce platelet lysis. Regarding platelet activation, total venom was able to increase the expression of LIBS1 on both human and rabbit platelet surface and P-selectin on only rabbit platelet surface. However, the sphingomyelinase fraction did not increase the expression of any marker of platelet activation, at least in the experimental conditions used in this study. Another in vitro study showed that both total venom and its sphingomyelinase fraction induce an increase in human and rabbit platelet adhesion, with the sphingomyelinase fraction exhibiting a dose-dependent response. For the study using an animal model of thrombocytopenia, it was necessary to previously produce goat antibodies against rabbit platelets, which showed to be effective to induce an intense thrombocytopenia in rabbits. Histopathological analysis of venom-induced dermic tissue lesion indicated that platelet absence does not decrease the formation of intravascular thrombi in the injured area. Nonetheless, bigger areas of equimosis and necrotic scabs could be observed in the dermonecrotic lesion of rabbits injected with loxoscelic venom and depleted of platelets. In the different groups, plasma levels of von Willembrand factor, prothrombin and activated partial thromboplastin times, and data from phagocytosis tests using polymorphonuclear cells isolated from rabbit periferical blood were similar comparing thrombocitopenic rabbits with those exhibiting normal platelet count; therefore, we can deduce that there is not any expressive alteration in endotelial cells, coagulation system and phagocytic ability of polymorphonuclear leukocytes via their complement receptor C3b. In conclusion, the loxoscelic venom is able to unchain in vitro different mechanisms leading to adhesion, activation and aggregation of both human and rabbit platelets. Furthermore, due to differences in the dermic lesion patterns between thrombocitopenic and normal animals, we can conclude that platelets must play a key role in controlling the expansion and severity of the dermonecrotic lesion induced in rabbits by L. gaucho venom.

Les complexes de rhénium jouent un rôle important dans le domaine de la médecine nucléaire. Si pendant longtemps le rhénium a été utilisé comme modèle structural du technétium-99m, les caractéristiques physiques prometteuses des isotopes 186Re et 188Re, font des complexes de 186/188Re des candidats prometteurs en tant que radiopharmaceutiques thérapeutiques. De même, les propriétés intéressantes de photoluminescence des complexes de rhénium non-radioactifs en font d'excellents outils comme catalyseurs, matériaux luminescents et capteurs d'imagerie. Dans ce travail, notre objectif était (i) de développer, en utilisant une stratégie de chimie Click, des ligands multidentes pour la stabilisation de différents cœurs rhéniés de type [Re(CO)3]+ et [ReO]3+ (M = Re ou 188Re) ainsi que pour les coeurs analogues à base de 99mTc dans certains exemples, (ii) d'évaluer le potentiel des complexes de rhénium (technétium) en tant que sondes d'imagerie (Re naturel ou 99mTc) ou agents thérapeutiques (188Re). Pour ce faire, deux systèmes de chélatants spécifiques du rhénium (technétium) ont été utilisés: un système tripodal semi-rigide dans le deuxième chapitre et un fragment pyta au troisième chapitre, ces deux chélatants ayant été développés précédemment dans notre groupe. Ainsi, sur la base d'un ligand click tridente de type N2O, deux études différentes ont été réalisées dans le chapitre II. Dans la première étude, deux voies de synthèse ont permis de préparer une famille de ligands potentiellement tétradentes de type N3O, conçus pour coordonner les cœurs rhéniés aux états d'oxydation + I et + V. Les études de coordination envers ces différents noyaux de rhénium, ont également été étudiées. Les premiers résultats de radiomarquage combinés aux récents travaux rapportés par l'équipe de Dugave[xii] sur des systèmes similaires indiquent que ce ligand pourrait être un chélatant prometteur pour le 99mTc...voire le 188Re. A court terme, l'extension du radiomarquage au coeur [188ReVO]3+ devra être effectuée et la stabilité in vitro du complexe radioactif testée dans des conditions physiologiques classiques, dans le plasma humain et par des expériences d'échange de cystéine. La deuxième étude portait sur le développement de nouveaux radiopharmaceutiques du 99mTc sélectifs contre l'hypoxie. Notre ligand tripodal de départ a été décoré avec un groupe nitro (soit un groupe nitrobenzyle ou une entité métronidazole (Mtz)). Des positions différentes ont été considérées mais seulement deux ligands contenant du métronidazole (Mtz) et un ligand contenant un groupe nitro ainsi que les complexes correspondants de tricarbonylrhénium(I) ont été obtenus et caractérisés, notamment par électrochimie. Les potentiels de réduction du groupe NO2 dans les complexes [Re(CO)3Cl(L2)] et [Re(CO)3(L6)] sont similaires à ceux reportés dans la littérature pour des complexes de rhénium(I). Le chapitre III est axé sur l'étude de l'effet AIE (émission induite par l'agrégation) dans les complexes tricarbonylrhénium(I), l'association de cet effet avec les propriétés intrinsèques des complexes de Re(I) pouvant conduire à des composés très attrayants. Pour ce faire, nous avons combiné un fluorophore organique (motif benzoxazole ou PBO) qui présente une excellente stabilité et des propriétés optiques intéressantes, avec un complexe tricarbonylrhénium(I) basé sur une unité pyta (soit le 2-pyridyl-1,2,3-triazole ou le 2-pyridyl-1,2,4-triazole). Quatre composés ont été étudiés. Les structures RX ont révélé des différences structurales spectaculaires entre les deux premiers complexes, ReL8 et ReL9. Cette étude étant une nouvelle orientation dans notre groupe, ce travail sera un excellent point de départ pour d'autres recherches. Divers colorants organiques et/ou des modifications structurales de la fraction organique seront bientôt pris en compte pour développer des sondes luminescentes de rhénium(I) hautement émissives. / Rhenium complexes play a significant role in nuclear medicine. Rhenium has been widely used as a surrogate of technetium for a long time, and the promising physical features of 186Re and 188Re, make 186/188Re-complexes promising candidates as therapeutic radiopharmaceuticals.Similarly, the interesting photoactive and photoluminescence properties of non-radioactive Re-complexes make them excellent catalysts, luminescent materials and imaging sensors.Thus, in this work, our goal was to (i) develop, using a click chemistry strategy, multidentate ligands for the stabilization of different rhenium cores [Re(CO)3]+ and [ReO]3+ (M = Re or 188Re) as well as the analogous 99mTc-cores in some examples, (ii) assess the potential of the rhenium(technetium) complexes as imaging (natRe or 99mTc) or therapeutic (188Re) agents. To do so, two rhenium(technetium) specific-chelating systems were used: a semi-rigid tripodal system in the second chapter and a pyta moiety in the third chapter, these two chelators being developed previously in our group. Thus, based on a N2O tridentate click ligand, two different studies were carried out in chapter II. In the first one, two synthetic pathways to a range of potentially N3O tetradentate ligands, designed to coordinate rhenium cores as well as their coordination behaviors towards different rhenium cores (oxidation states +I and +V), were investigated. The first radiolabeling results combined with the recent work reported by Dugave and co-workers indicated that this ligand could be a promising 99mTc-chelator for nuclear imaging applications. As perspectives to this work, the extension of the radiolabelling work using the [188ReVO]3+ core should be performed, and the in vitro stability should be tested under physiological conditions in human plasma and by cysteine exchange experiments. The second study was focused on the development of novel hypoxia-selective 99mTc radiopharmaceuticals. Our semi-rigid tripodal click framework was decorated with an appended nitro group (either a nitrobenzyl group or a metronidazole (Mtz) unit). Different positions were considered and at least only two metronidazole (Mtz)-containing ligands and one nitro group-containing ligand as well as their corresponding tricarbonyl rhenium(I) complexes were obtained and characterized, in particular by electrochemistry. The reduction potentials of NO2 group in complexes [Re(CO)3Cl(L2)] and [Re(CO)3(L6)] were similar to those of reported hypoxic imaging agents, prompting us to further investigate other properties of these complexes. Chapter III was focused on the study of AIE (aggregation-induced emission) effect in tricarbonyl Re(I) complexes, the association of this effect with the intrinsic properties of Re(I) complexes being expected to lead to very attractive compounds. To do that, we combined an organic fluorophore (PBO) which exhibits excellent stability and optical properties, with a tricarbonylrhenium(I) complex based on a pyta unit (either a 2-pyridyl-1,2,3-triazole or a 2-pyridyl-1,2,4-triazole ligands). Four compounds were studied. The X-Ray structures revealed spectacular discrepancies between the two first triazole-based complexes ReL8 and ReL9. Moreover, this study being a novel orientation in our group, this work is a great starting point for further investigations. Various organic dyes and/or structural modifications of the organic moiety will soon be considered in order to develop highly emissive rhenium(I) luminescent probes.

Microarrays have been widely used to study differential gene expression at the genomic level. They can also provide genome-wide co-expression information. Robust approaches are needed for integration and validation of independently-collected datasets which may contribute to a common hypothesis. Previously, attempts at meta-analysis have contributed to solutions to this problem. As an alternative, for microarray data from multiple highly similar biological experimental designs, a more direct combined approach is possible. In this thesis, a novel approach is described for microarray combined analysis, including gene-level unification into a virtual platform followed by normalization and a method for ranking candidate genes based on co-expression information - called Coex-Rank. We applied this approach to our Sppar (a PPARγ mutant) dataset, which illustrated an improvement in statistical power and a complementary advantage of the Coex-Rank method from a biological perspective. We also performed analysis to other PPARγ-related microarray datasets. From the perspective of gene sets, we observed that up-regulated genes from mice treated with the PPARγ ligand rosiglitazone were significantly down-regulated in mice with a global knock-in dominant-negative mutation of PPARγ. Integrated with publicly available PPRE (PPAR Response Element) datasets, we found that the genes which were most up-regulated by rosiglitazone treatment and which were also down-regulated by the global knock-in mutation of PPARγ were robustly enriched in PPREs near transcription start sites. In addition, we identified several potential PPARγ targets in the aorta and mesenteric artery for further experimental validation, such as Rhobtb1 and Rgs5.