Le maintien de la masse musculaire est étroitement lié à la balance entre la synthèse et la dégradation des protéines. L'exercice physique est un puissant régulateur de la balance protéique et plus particulièrement l'exercice en résistance. S'intéresser à la balance protéique après un exercice s'inscrit dans une compréhension des mécanismes cellulaires et moléculaires conduisant aux phénomènes d'hypertrophie et/ou d'atrophie musculaire. Nos travaux mettent en évidence que l'hypertrophie obtenue dans le muscle FDP après 10 semaines d'un entraînement en résistance chez le rat, est en lien avec l'activation chronique de la voie IL-6/STAT3 après chaque exercice aigu, en partie au sein du pool de cellules satellites activées. En phase proliférative, les cellules dont la voie de signalisation STAT1/STAT3 est activée, répriment l'expression des facteurs myogéniques comme MyoD et retournent ainsi à l'état quiescent, concourant à augmenter le pool de réserve. Ces mécanismes participent à la synthèse protéique par l'apport de nouveau matériel génétique au sein des fibres musculaires conduisant à une augmentation de leur surface de section ainsi qu'à leur conversion phénotypique avec l'entraînement. L'exercice en résistance favorisant la protéolyse, nos travaux ont cherché à caractériser les systèmes protéolytiques (autophagique-lysosomal, ubiquitine-protéasome) impliqués dans la balance protéique post-exercice. Les marqueurs moléculaires étudiés (activités enzymatiques du protéasome et de la cathepsine L, expression protéique et génique de LC3B, des E3 ligases…) ne permettent pas d'expliquer clairement les +30% de protéolyse obtenus une heure après des contractions excentriques sur muscle EDL isolé de rat en condition à jeun. Des perspectives d'étude des systèmes des calpaines, des caspases et/ou des métalloprotéases matricielles sont alors à envisager. / The maintain of muscle mass is closely controlled by protein synthesis and degradation balance. Physical activity and mainly resistance exercise is a powerful stimulus to positive muscle protein balance. To understand how protein balance is regulated after exercise, cellular and molecular mechanisms leading to muscular hypertrophy and/or atrophy have to be elucidated. Our works point out that FDP muscular hypertrophy after 10 weeks of resistance training in rat is partly due to the chronically activation of IL-6/STAT3 signaling pathway, occurring in the activated satellite cell pool, after each single exercise bout. Once activated and engaged in the myogenic program, cells in which STAT1/STAT3 signaling pathway is activated, could downregulate MyoD and return to a quiescent state, leading to increase satellite cell reserve's pool. These events participate to enhance protein synthesis by the incorporation of new genetic material into muscle fiber leading to increase their cross sectional area and phenotypic shift after training. As resistance exercise increases proteolysis, our works attempt to characterize the proteolysis systems (lysosomal-autophagic, ubiquitin-proteasome) involved in protein balance after exercise. The molecular markers measured ( chymotrypsin-like and cathepsin L activities, protein and gene expressions of LC3B, E3 ligases…) could not explain the +30% of proteolysis obtained one hour after resistance eccentric contractions on EDL muscle of starved rats. Further studies based on calpains, caspases and metalloproteinase activities and/or expressions should bring us valuable information.

Orientador: Paulo Sérgio Cerri / Banca: Lea Assed Bezerra da Silva / Banca: Mario Tanomaru Filho / Resumo: Perfurações radiculares acidentais ocorrem em cerca de 2-12% dos dentes tratados endodonticamente. Estas perfurações causam injúria aos tecidos periodontais adjacentes e, frequentemente, promovem a invasão bacteriana culminando com a inflamação do ligamento periodontal e reabsorção óssea podendo levar a perda do elemento dental. Dentre os materiais indicados para o selamento destas perfurações, o MTA tem sido largamente usado. O MTA é basicamente composto por silicatos de cálcio (80%) e óxido de bismuto (Bi2O3; 20%), usado como agente radiopacificador. Silicato tricálcico (STC) é um biomaterial utilizado associado ao óxido de zircônio (ZrO2) e o óxido de nióbio (Nb2O5) é um radiopacificador alternativo. O objetivo deste estudo foi avaliar a resposta dos tecidos periodontais promovida pelo silicato tricálcico puro (Mineral Research Processing, Meyzieu, França) associado ao ZrO2 ou ao Nb2O5 comparando-os ao MTA-Angelus, em perfuração de furca de molares de ratos. Foram utilizados 80 ratos, distribuídos em 4 grupos: GS (sham), STC+ZrO2, STC+Nb2O5 e GMTA, de acordo com o material de selamento. No grupo GS, as perfurações no assoalho da câmara pulpar não foram seladas. O acesso da cavidade foi restaurado com cimento de ionômero de vidro fotopolimerizável. Os 1os molares esquerdos, sem perfurações no assoalho da câmara pulpar, foram usados como controle (GC). Após os períodos de 7, 15, 30 e 60 as maxilas foram removidas, fixadas, descalcificadas e incluídas em parafina. Os cortes c... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Accidental radicular perforation occurs in about 2-12% of endodontically treated teeth. These perforations lead to injury of periodontal tissues and frequently promote bacterial invasion inducing to periodontal ligament inflammation and bone resorption may lead to tooth loss. Among the materials suitable for the sealing of these perforations, the MTA has been widely used. MTA is basically composed of calcium silicate (80%) and bismuth oxide (Bi2O3, 20%), a radiopacifying agent. Tricalcium silicate (TCS) is a biomaterial used associated with zirconium oxide (ZrO2) and niobium oxide (Nb2O5) is an alternative radiopacifier. Our purpose was to evaluate response of the periodontium adjacent to furcation perforations in rat molars filled with pure tricalcium silicate (Mineral Research Processing, Meyzieu, France) associated with ZrO2 or Nb2O5 and MTA-Angelus. Eighty rats were distributed into four groups: SG (sham), TCS+ZrO2, TCS+Nb2O5 and MTAG according to the sealing material. In the SG group, the perforations in the pulp chamber floors were not sealed. The access cavity was restored with light-cure glass-ionomer cement. The left upper first molars, without perforations in the pulp chamber floors, were used as controls (CG). After 7, 15, 30 and 60 days, fragments of maxilla were fixed, decalcified and embedded in paraffin. In the HE-stained sections, the volume density of inflammatory cells (VvIC), fibroblasts (VvFb) and blood vessels (VvBV) was obtained. The width of the periodo... (Complete abstract click electronic access below) / Mestre

The development of essential medicines is being slowed by a lack of efficiency in drug development as ninety per cent of drugs fail at some stage during clinical evaluation. This attrition in drug development is seen not because of a reduction in pharmaceutical research expenditure nor is it caused by a declining understanding of biology, if anything, these are both increasing. Instead, drugs are failing because we are unable to effectively predict how they will work before they are given to patients. This is due to limitations of the current methods used to evaluate a drug's toxicity and efficacy prior to its development. Quite simply, these methods do not account for the full complexity of biology in humans. Systems pharmacology models are a likely candidate for increasing the efficiency of drug discovery as they seek to comprehensively model the fundamental biology of disease mechanisms in a quantit- ative manner. They are computational models, designed and hailed as a strategy for making well-informed and cost effective decisions on drug viability and target druggability and therefore attempt to reduce this time-consuming and costly attrition. Using text mining and text classification I present a growing landscape of systems pharmacology models in literature growing from humble roots because of step-wise increases in our understanding of biology. Furthermore, I develop a case for the capability of systems pharmacology models in making predictions by constructing a model of interleukin-6 signalling for rheumatoid arthritis. This model shows that druggable target selection is not necessarily an intuitive task as it results in an emergent but unanswered hypothesis for safety concerns in a monoclonal antibody. Finally, I show that predictive classification models can also be used to explore gene expression data in a novel work flow by attempting to predict patient response classes to an influenza vaccine.

Objetivo: determinar as concentrações séricas e peritoneais das interleucinas 1ß (IL-1ß) e 6 (IL-6) e da proteína amilóide A (SAA) em pacientes com endometriose pélvica. Métodos: foram avaliadas 97 pacientes submetidas à videolaparoscopia, 57 com endometriose (A), 27 com sintomas sugestivos porém sem endometriose (B) e 13 sem sintomas e doenças (C). Foram coletados no ato cirúrgico líquido peritoneal e sangue. As concentrações dos mediadores foram determinadas em pg/ml (IL-1 e 6) e ng/ml (SAA) por método imunoenzimático e leitura óptica. Resultados: (líquido peritoneal e sangue medianas) IL-1ß: A- 11,22 and 1,83; B- 15,62 and 1,16; C- 1,92 and 0,80; IL-6: A- 6,80 and 3,70; B- 8,60 and 3,90; C- 3,40 and 2,0; SAA- A- 310,30 and 14,01; B- 306,20 and 10,39; C- 53,4 e 9,5. Conclusão: as concentrações dos mediadores de inflamação avaliados estão elevadas no líquido peritoneal e no soro das mulheres com endometriose. Estas concentrações foram semelhantes às das pacientes com sintomas sugestivos porém sem a doença. A fase do ciclo menstrual, o tipo histológico envolvido e o local de acometimento da doença não influíram de forma significante nas concentrações séricas ou peritoneais de IL-1ß, IL-6 e SAA. / Objective: to assess peritoneal and seric interleukin-1ß (IL-1ß), 6 (IL-6), and protein amyloid A (SAA) concentrations in patients with pelvic endometriosis. Methods: 97 patients were submitted to video laparoscopic surgery, 57 with endometriosis (A), 27 with suggestive symptoms but no endometriosis (B), and 13 without symptoms and diseases (C). Peritoneal fluid and blood were collected during the procedure. Mediator s concentration was determined in pg/ml (IL-1ß and IL-6) and ng/ml (SAA) through immunoenzimatic test and optic measure. Results: (peritoneal fluid and blood -medians) IL-1ß: A- 11,22 and 1,83; B- 15,62 and 1,16; C- 1,92 and 0,80; IL-6: A- 6,80 and 3,70; B- 8,60 and 3,90; C- 3,40 and 2,0; SAA- A- 310,30 and 14,01; B- 306,20 and 10,39; C- 53,4 e 9,5. Conclusion: the inflammation mediators are increased in the peritoneal fluid and blood of women with pelvic endometriosis. The concentrations are similar to those found in patients with suggestive symptoms but no endometriosis. The place of the disease, the histological type and the day of the menstrual cycle didn t alter peritoneal or seric concentration of these cytokines.

A fibrose hepática é uma patologia que acompanha outras doenças crônicas do fígado como a cirrose e o hepatocarcinoma. As células estreladas hepáticas (HSC, do inglês hepatic stellate cells) compõem uma população celular heterogênea que se caracteriza por transitar entre dois fenótipos. As células com fenótipo quiescente possuem a capacidade de armazenar vitamina A em gotas lipídicas. Os insultos ao fígado desencadeiam uma resposta inflamatória que gera estímulos parácrinos e autócrinos mediados por citocinas e espécies reativas. Neste contexto, as HSC assumem um fenótipo ativado fibrogênico e tornam-se responsáveis pela cicatrização hepática. Danos crônicos ao fígado levam a uma deposição de matriz extracelular exagerada que configura o estado patológico da fibrose. O resveratrol (RSV – 3,4’,5-tri-hidroxi-trans-estilbeno) é uma fitoalexina produzida por algumas espécies de plantas. Inúmeros efeitos benéficos à saúde são atribuídos ao RSV por causa do seu potencial antioxidante, antiinflamatório e pró-apoptótico. Estudos anteriores mostraram que tratamento da GRX, uma linhagem murina de HSC ativadas, com concentrações de RSV próximas as biodisponíveis (0,1 a 1 μM) resultou em parada do ciclo na fase S com consequente inibição de proliferação celular, um efeito associado à citotoxicidade e que pode favorecer a resolução da fibrose hepática. Neste estudo, por técnicas espectrofotométricas, foi demonstrado que tratamento da GRX por 24 horas com concentrações entre 0,1 a 50 μM de RSV promoveu um efeito pró-oxidante que causa uma citotoxicidade dependente da dose, bastante aumentada no grupo tratado com a concentração mais alta. Os efeitos citotóxicos atenuados encontrados nas células tratadas por 120 horas sugerem que a GRX pode se tornar resistente a estes efeitos. O potencial pró-oxidante do RSV foi o ponto de partida para investigar a possibilidade de que esta fitoalexina provocasse uma alteração no metabolismo mitocondrial da GRX. Para isso, os efeitos do RSV (1 a 50 μM) na função mitocondrial, na indução de morte mediada por estas organelas e na autofagia/mitofagia foram investigados por técnicas de espectrofotometria, de imunocitoquímica, de citometria de fluxo, de microscopia confocal e de microscopia eletrônica de transmissão em GRX tratadas por 24 e 120 horas. Foi demonstrado que todas as concentrações de RSV promovem apoptose por meio da ativação de caspases, alteram a dinâmica/função mitocondrial e induzem o aumento de autofagia/mitofagia na GRX. No entanto, o RSV provocou biogênese de mitocôndrias nos grupos tratados com 1 e 10 μM, enquanto que o tratamento com 50 μM causou dano celular evidente na GRX, sem induzir biogênese de mitocôndrias. Desta forma, é possível que a citotoxicidade “dose-dependente” do RSV, que causa a morte celular e dano oxidativo em 24 horas de tratamento, esteja relacionada com o desequilíbrio entre a indução concomitante de apoptose mediada por dano mitocondrial, autofagia/mitofagia e biogênese de mitocôndrias. Por fim, foi investigada a liberação de TNF-α, Interleucina-6 e Interleucina-10 pela GRX tratada por 24 e 120 horas com RSV (0,1 a 50 μM), considerando o papel antiinflamatório do RSV e o papel das HSC ativadas na sinalização autócrina que contribui para a modulação fenotípica destas células. Foi demonstrado que o tratamento da GRX com RSV por 24 e 120 horas induziu a redução da liberação de Interleucina-6; enquanto que a liberação de TNF-α e Interleucina-10 foi aumentada. Estes resultados confirmam um efeito antiinflamatório do RSV que deve contribuir na prevenção da ativação ou da perpetuação do estado ativado das HSC por meio de sinalização autócrina. Ainda que a concentração do RSV seja importante para efetivamente induzir a morte das HSC ativadas, o tratamento com esta fitoalexina pode ser promissor para a resolução da fibrose hepática por diminuir a população de células ativadas e, possivelmente, prevenir a perpetuação do estado fenotípico ativado. Estudos avaliando indicadores de quiescência em células tratadas são ainda necessários para desvendar completamente os efeitos do RSV quanto às possibilidades de inibição da perpetuação ou reversão fenotípica das HSC ativadas. / Liver fibrosis is a disease that accompanies other hepatic chronic diseases such as cirrhosis and hepatocellular carcinoma. Hepatic stellate cells (HSC) are a heterogeneous cell population characterized by transiting between two phenotypes. Cells with a quiescent phenotype are able to store vitamin A into lipid droplets. Damage to the liver trigger an inflammatory response that generates paracrine and autocrine stimulation mediated by cytokines and reactive species. In this context, HSC assume an activated and fibrogenic phenotype responsive for hepatic wound-healing. Chronic insults to the liver lead to an excessive deposition of extracellular matrix that configures the pathological state of fibrosis. Resveratrol (RSV – 3,4’,5-tri-hidroxi-trans-stilbeno) is a phytoalexin produced by some species of plants. Several beneficial effects are attributed to this molecule due to its antioxidant, antiproliferative and pro-apoptotic potential. Previous studies showed that treatment with bioavailable concentrations of RSV (0.1 to 1 μM) promoted an arrest cycle at the S phase in GRX, a murine activated HSC model, leading to cell proliferation inhibition, a cytotoxic effect that contributes to the liver fibrosis resolution. In this study, it was shown by spectrophotometric techniques that GRX treatment for 24 hours at concentrations between 0.1 to 50 μM of RSV promoted a fairly clear pro-oxidant effect that causes a dose-dependent cytotoxicity that was higher in the group treated with 50 μM. The attenuated cytotoxicity found after 120 hours of GRX treatment suggest that these cells became resistant to this effect. The pro-oxidant potential of RSV was the starting point for investigating the possibility that this phytoalexin would cause a change in the GRX mitochondrial metabolism. Thus, the effects of RSV (1 to 50 μM) on altering the mitochondrial function, on inducing mitochondrial-mediated cell death, and autophagy/mitofagia were investigated in GRX treated for 24 and 120 hours by spectrophotometric techniques, immunocytochemistry, flow cytometry, confocal microscopy, and transmission electron microscopy. All the RSV concentrations promote cell apoptosis through caspases activation, alter the mitochondrial dynamics and function, and induce an increase of autophagy/mitofagia. Curiously, only 1 and 10 μM of RSV induced mitochondrial biogenesis in GRX, while the highest concentration caused an evident cell damage without inducing mitochondrial biogenesis. Thus, it is possible that the "dose-dependent" cytotoxicity of RSV, which causes cell death and oxidative damage in 24 hours of treatment, is related to an imbalance between the concomitant induction of mitochondrial-mediated apoptosis, autophagy/mitofagia, and mitochondrial biogenesis. Finally, it was investigated the release of TNF-α, Interleukin-6 and Interleukin-10 by GRX treated for 24 and 120 hours with RSV (0.1 to 50 μM), considering the anti-inflammatory role of RSV and the autocrine signalling role of HSC that contributes to the perpetuation of its activated phenotype. It was demonstrated that GRX treatment with RSV for 24 and 120 hours reduced the release of Interleukin-6 in the culture medium; whereas the release of TNF-α and Interleukin-10 was increased. These results confirm the anti-inflammatory properties of RSV and may contribute to the prevention of HSC activation through autocrine signalling. Although RSV concentration is important to effectively induce activated HSC death, cells treatment with this phytoalexin may be promising for liver fibrosis resolution through decreasing the population of activated cells or through preventing the perpetuation of activated state of HSC. Future studies evaluating the quiescence indicators of GRX under RSV treatment are still needed to fully unravel the effects of this phytoalexin on inhibiting the perpetuation of activated HSC or reversing its activated phenotype.

INTRODUÇÃO: O transplante de intestino (TI) estabeleceu-se como tratamento para pacientes com falência intestinal e complicações da nutrição parenteral. Entretanto, sepse continua sendo a principal causa de mortalidade. A lesão causada pela isquemia seguida de reperfusão (LIR) é apontada como um dos fatores de ruptura da barreira mucosa intestinal, com consequente translocação bacteriana e sepse, seja precocemente, por lesão epitelial direta, seja mais tardiamente pela sua associação com o desenvolvimento da rejeição celular aguda. Criou-se um modelo de TI em porcos jovens com a finalidade de estudar a LIR e seus efeitos no epitélio intestinal. MÉTODOS: Para a padronização do modelo, foram realizados 25 procedimentos, tendo sido testados os tamanhos dos animais, as soluções de preservação, o tipo de drenagem venosa, o tipo de reconstrução intestinal e o tempo de duração do experimento. Na pesquisa propriamente dita, 20 porcos jovens foram submetidos a TI ortotópico. Dois grupos foram determinados conforme o tempo de isquemia fria a que foi submetido o intestino: grupo 1 (n=12) 90 minutos (min) e grupo 2 (n=8) 180 min. O procedimento foi realizado sob técnica asséptica e as anastomoses vasculares realizadas entre a aorta do doador e a aorta infra-renal do receptor e a veia porta do doador e a veia cava inferior do receptor. O trânsito intestinal foi reconstruído através de anastomoses entre o jejuno proximal do doador e do receptor e o íleo terminal do doador e do receptor. A solução de preservação utilizada foi Euro Collins. Não foi administrada medicação imunossupressora, exceto pela metilprednisolona (20mg/kg) no momento da reperfusão. Fragmentos de intestino foram obtidos: 1 no momento da laparotomia do doador, o fragmento basal, considerado controle, 2 30 min após a reperfusão e 3 3 dias após o transplante. Os fragmentos assim obtidos foram submetidos a: 1 análise histológica com coloração de hematoxilina-eosina (HE), 2 análise imunoistoquímica para a detecção de infiltração da mucosa por neutrófilos (marcados pelos grânulos ricos em mieloperoxidase MPO), 3 análise histoquímica para quantificação de células epiteliais em apoptose pelo método TUNEL, 4 análise da expressão dos genes da endotelina-1 (ET-1) e da interleucina-6 (IL-6), do gene antiapoptótico Bcl-XL e do gene pró-apoptótico Bak. A análise estatística foi realizada utilizando-se o teste de Mann-Whitney na comparação entre os grupos e, na análise da evolução temporal da LIR em cada grupo, o teste de Friedman seguido do teste post hoc de Dunn quando detectada diferença estatisticamente significante. RESULTADOS: Não foram encontradas diferenças entre os grupos quanto às alterações histológicas estudadas. O grau de infiltração da mucosa por neutrófilos elevou-se significantemente nos dois grupos 30 min após perfusão, tendo persistido elevado 3 dias após o TI somente no grupo 2. O número de células epiteliais em apoptose detectadas pelo método TUNEL sofreu incremento significante apenas no grupo 1, 3 dias após o procedimento. As duas citocinas estudadas, IL-6 e ET-1 mostraram elevação significante 30 minutos após a reperfusão, tendo retornado aos níveis basais 3 dias após a cirurgia em ambos os grupos. Detectou-se redução significante da expressão do Bcl-XL somente no grupo 1, 3 dias após o TI. CONCLUSÕES: As citocinas estudadas estão envolvidas no processo de LIR nas fases iniciais do TI. Ocorre diminuição da expressão de gene anti-apoptótico e aumento do número de células em processo de morte celular de maneira mais intensa no grupo submetido a menor tempo de isquemia / INTRODUCTION: Intestinal transplantation (ITx) has become an accepted mode of treatment of intestinal failure patients who develop parenteral nutrition-related complications. Overall outcomes have dramatically improved but sepsis remains the leading cause of mortality. Ischemia-reperfusion injury (IRI) has been related to the development of sepsis due either to direct mucosal damage or to increased risk of acute cellular rejection. An experimental ITx model has been idealized in order to better characterize IRI-associated mucosal damage. METHOD: 25 procedures involving 75 outbred pigs were necessary to standardize the procedure. Weight of the animals, venous drainage, intestinal transit reconstruction as well as the time period the animals should be maintained alive were evaluated. Orthotopic ITx was performed in 20 hybrid pigs. Two groups were assigned according to cold ischemia time (CI): group 1 (n=12) 90 minutes (min), group 2 (n=8) 180 min. The procedure was performed under aseptic technique and portal drainage was adopted as standard. Intestinal transit reconstruction involved the performance of termino-terminal anastomosis between donor and recipient jejunum and donor and recipient terminal ileum. Euro-Collins was used as preservation solution. 20mg/kg of metilprednisolone was administered at reperfusion and no other immunosuppressive drug has been employed. Specimens were collected from the donor at laparotomy, and from the receptor, 30 min, and 3 days after reperfusion. Mucosal damage was assigned by histological evaluation with hematoxylin-eosin dye. Neutrophilic infiltration was quantified using myeloperoxidase (MPO) immunohistochemical assay and epithelial cell apoptosis was also assigned by means of TUNEL assay. Molecular biology involved the quantification of the expression of the IL-6, ET-1, Bak, and Bcl-XL genes. RESULTS: No statistical difference was detected between the groups as far as plain histological evaluation is regarded. Neutrophilic infiltration increased in a similar fashion in both groups, but lasted longer in group 2. Apoptosis detected by TUNEL showed significant increase in group 1, 3 days after surgery. Anti-apoptotic gene Bcl-XL had its expression decreased in group 1, in 3 days as well. Endothelin-1 and IL-6 genes expression increased 30 min after the procedure and had already returned to baseline 3 d after surgery. CONCLUSION: IL-6 and ET-1 are involved precociously in the development of intestinal IRI. Neutrophilic infiltration lasted longer in the group submitted to longer CI. Although there were no significant differences between the groups, significant increase in the number of apoptotic epithelial cells 3 days after reperfusion could be detected in animals

Les maladies cardiovasculaires représentent un important problème de santé publique à travers le monde. Parmi ces pathologies, l’infarctus du myocarde (IM) a fait l’objet de nombreux essais visant à diminuer sa sévérité. Néanmoins peu ont remporté leur pari. Cet échec peut avoir plusieurs composantes parmi lesquelles la méconnaissance de la complexité des mécanismes moléculaires impliqués. Notre compréhension de l’IM a cependant été nettement améliorée grâce à des méthodes utilisées pendant les dernières décennies et qui consistaient à étudier séparément un nombre limité d’acteurs moléculaires impliqués dans un mécanisme simple et linéaire. Cependant, l’échec des essais cliniques basés sur ces approches réductionnistes ont montré leurs limites. L’émergence de la biologie des systèmes, ces dernières années, a stimulé les recherches visant à mieux intégrer et comprendre la nature complexe et stochastique des réseaux moléculaires et leur dynamique dans la progression des pathologies. L’objectif général de cette thèse a consisté en le développement et l’amélioration de méthode d’analyses et de combinaison des données spatio-temporelles issus d’expériences réalisées sur un modèle d’infarctus du myocarde chez la souris. L’objectif scientifique visait à caractériser les principaux signaux dynamiques au cours de la séquence d’ischémie-reperfusion. A cet effet, nous avons tout d’abord développé une chaîne de méthode utilisant la clarification d’organe et la microscopie de fluorescence permettant de quantifier, en 3 dimensions, la zone du myocarde soumise au choc oxydant lors de la reperfusion. Dans une seconde partie, nous avons développé une nouvelle chaîne analytique pour caractériser la dynamique d’expression des transcrits. Appliquée aux animaux contrôles (soumis à la chirurgie et l’anesthésie), nous mettrons, grâce à cette chaîne de méthode, le rôle majeur de la voie de l’interleukine 6 dans le développement de la réponse immunitaire et nous concluons ainsi sur la nécessité de réaliser une analyse dynamique du modèle expérimental pour caractériser sa réponse à l’échelle moléculaire et éviter toute surinterprétation de la réponse à l’IM / Cardiovascular diseases represent a major health burden worldwide. According to the World Health Organization, 17 million people are dying each year by heart diseases which account to 31% of total deaths globally. Among these diseases is myocardial infarction (MI). Several efforts have been made to decrease the associated mortality rates but unfortunately, only few has succeeded to date. This failure is contributed to several factors, among them is the misunderstanding of the mechanism responsible for the progression of the disease.Our understanding of the MI pathology has been greatly improved by the approaches that have been widely used in the previous decades, relying mainly on studying molecules/pathways separately. However, this knowledge was not enough to make a difference clinically. Therefore, deciphering the interconnections between molecules has become an urge for better understanding of the diseases’ progression. In this regard, the work in this doctoral thesis involves different aspects of the MI pathology. The general aim of this work is to improve the dynamic analytical approach using systems biology tools, where new mechanistic information is decoded. Firstly, in a 3D heart model, we propose a chain of methods using clarified mouse heart and fluorescence microscopy to molecularly characterize the area at risk in the myocardium of IR and cardioprotected mice based on its redox state. In addition, we aim to develop a new analytical approach using dynamical large-scale transcriptomic data for characterizing the dynamic transcripts expression. Applying this approach on a control mouse model (mice subjected to anesthesia and surgical interventions), we show that Il-6 is a major mediator of the activated inflammatory reaction. In conclusion, this analytical approach highlights the necessity of the sapatio-temporal analysis to characterize the molecular events occurring in response to MI

Cancer-associated cachexia is a condition defined by a sustained net-negative energy imbalance. Although the different types of adipose tissue – white, beige, and brown – have been implicated in contributing to cancer-associated cachexia, the mechanisms of these maladaptive changes and their impact on whole-body energy expenditure have not been fully elucidated. Using breast cancer as our model, we demonstrate white adipose tissue browning in murine and human breast cancer; furthermore, we demonstrate that this effect is extremely localized and takes place early in tumor progression. We utilized in vitro cell culture techniques and demonstrate that cancer secreted factors and cross-talk with white adipocytes decrease expression of classic white adipose tissue-related genes. We also demonstrate in murine and human culture models that cancer secreted factors reduce white adipocyte lipid droplet size, and cross-talk between cancer cells and adipocytes results in an increase in lipolysis-related gene expression. Interestingly, our results strongly suggest that in mice, neither cancer secreted factors nor cross talk with adipocytes can induce white adipose tissue browning, indicate that this process likely occurs independently of direct cancer interactions with local white adipocytes. We demonstrate that interleukin 6, a cytokine with previous implications in white adipose tissue browning, induces interleukin 6-mediated signaling; however, that signaling alone is not enough to directly induce white adipose tissue browning. We present preliminary data suggesting that immune cell population shifts within the white adipose tissue of mice with breast cancer tumors may be source of white adipose tissue browning. We show that the Virginia Commonwealth University Health System has an identifiable population of patients with cancer with what we hypothesize as maladaptive thermogenic adipose tissue activity, and discuss ongoing experiments aimed at understanding the implications of these changes on whole body energy expenditure in human patients. Lastly, in a case of autoimmune diabetes mellitus in the setting of an extra-adrenal paraganglioma, we demonstrate that the interaction between cancer and whole-body metabolism is multifaceted. Together, these experiments demonstrate that adipose tissue plasticity occurs in breast cancer (and other cancers), and that different drivers for individual changes exist within the tumor microenvironment. We predict that further exploration of the exact mechanisms and translational implications will provide useful information to lead to new therapeutic treatments for patients with cancer-associated cachexia.

Le criblage virtuel est largement employé pour la recherche de nouveaux médicaments.La sélection de structures pour les méthodes de criblage virtuel basées sur la structure reste problématique. Nous avons montré que les propriétés physico-chimiques du site de liaison, critères simples et peu coûteux en temps de calcul, pouvaient être utilisées pour guider celle-ci.L’évaluation des méthodes de criblage virtuel, critique pour vérifier leur fiabilité, repose sur la qualité de banques d’évaluation. Nous avons construit la NRLiSt BDB, n’incluant que des données vérifiées manuellement et prenant en compte le profil pharmacologique des ligands. Une étude à l’aide du logiciel Surflex-Dock montre qu’elle devrait devenir la base de données de référence, pour l’évaluation des méthodes de criblage virtuel et pour rechercher de nouveaux ligands des récepteurs nucléaires. L’application d’un protocole hiérarchique de criblage in silico/in vitro, a permis d’identifier de nouveaux composés inhibiteurs de l’IL-6, potentiellement utilisables dans le traitement de la polyarthrite rhumatoïde. Les résultats in vitro devront être confirmés par des tests in vivo. / Virtual screening is widely used in drug discovery processes.Structure selection in structure-based virtual screening methods is still problematic. We showed that simple and “low cost” binding site physico-chemical properties could be used to guide structure selection.The evaluation of virtual screening methods, necessary to ensure their reliability, relies on benchmarking databases quality. We created the NRLiSt BDB, gathering only manually curated data and taking into account ligands pharmacological profiles. A study using Surflex-Dock showed that the NRLiSt BDB should become the reference, both for the evaluation of virtual screening methods and for the identification of new ligands of the nuclear receptors.The use of a in silico/invitro hierarchical approach screening allowed to identify new IL-6 inhibitors, that could be used in rheumatoid arthritis treatment. In vitro results should be confirmed in vivo.

Background: Obesity-related diseases are arising as a major problem among children. inflammation has recently been identified to play an important role in the relationship between obesity.- as well as stunting-related diseases. Objectives: The aim of this study was to assess the association between serum tumour necrosis factor-alpha (TNF-α), interleukin-6 (IL-6) and C-reactive protein (CRP) concentrations and a variety of cardiometabolic and anthropometric indices of children in a township outside Potchefstroom, South Africa. Methods: Blood samples of 115 girls and 78 boys (mean age 15.6 ± 1.35) in the Physical Activity in the Young (PLAY) study were cross-sectionally analysed. Trained fieldworkers collected the demographic, Tanner growth stage and habitual physical activity information. Physiologists measured the children’s blood pressure. Anthropometric measurements were taken by. trained post-graduate students with level 1 or 2 qualifications in anthropometrics. A standard test battery was administered by trained postgraduate students in Human Movement Science to assess muscular strength. flexibility and endurance of the children. Blood samples were collected, centrifuged and stored frozen until further analyses. Results: Stunted girls had a significantly higher serum TNF-α concentration than the non-stunted girls (p=0.03). The factor analyses showed that the inflammatory. status clustered with the height for age-z-scores (HAZ) scores and the waist-hip-ratio (WHR). The HAZ-score of the over-fat boys (- 1.46) was significantly smaller than the lean boys (- 1.14, p=0.0 1). whereas the over-fat girls had a trend for a smaller HAZ-score (-1.07) than the lean girls (-0.89). No significant differences were found between the over-fat and the lean children-s inflammatory status. TNF-α and CRP levels tended to be higher in the over-fat children than in lean children. The girls' scrum IL-6 and CRP concentrations correlated significantly with their body mass index (BMI) and WHR (p<0.05 )and their TNF-α and IL-6 concentrations correlated significantly with their WHR (p<0.01 and p<0.05, respectively). Conclusion: In comparison to the non-stunted girls, stunted girls had a statistically significantly higher TNF-α concentration. Unusual fat distribution that is found in over-fat and stunted children may be associated with low-grade inflammation in children. More research is needed on these associations with markers of inflammation in a long-term longitudinal study. / Thesis (M.Sc. (Nutrition))--North-West University, Potchefstroom Campus, 2007.