The chromosomal translocation of t(14;18)(14q32;18q21) is a characteristic aberration of follicular lymphoma and Diffuse Large B cells lymphoma. By PCR, it was proved that the rearrangement of chromosomes 14 and 18 leads to an overexpression of BCL2, an anti-apoptotic protein, which is one of the factors responsible for the maturation of the diseases. The translocation involves the promoter region of IGH gene and the transcriptional unit of BCL2 gene. Previous studies carried out in Dr Tosi’s lab showed a looping out of the BCL2 gene from its chromosome territory in 15% of the nuclei analysed. This looping out could be possibly responsible for the transcriptional activity of the gene. A further relevant finding concerns the spatial distribution of the genes involved in the translocation in the interphase nuclei. In the Pfeiffer cell line, harbouring the t(14;18) rearrangement, the translocated BCL2 gene was positioned in the cell nuclei according to a bimodal distribution. One could speculate that the distribution in the periphery and in the centre of the nuclei could divide the Pfeiffer cell line in two different subpopulations, consequently from the transcriptional activity. These preliminary data set the ground for more experimental work to test whether genes associated with the nuclear interior were transcriptionally active as opposed to the genes positioned towards the nuclear periphery, transcriptionally inactive. The work here presented focuses on this investigation using RNA-DNA FISH (Fluorescence in situ hybridization). My work enabled the detection of IGH, BCL2 and t(14;18) genes along with their transcripts inside of the nuclei of Pfeiffer cell line. Contrary to what had been hinted by previous work, my results showed multiple nuclear positions of transcriptionally active IGH/BCL2 translocation. The result will need to be further supported by software analysis in order to define its specific nuclear position and to ensure the perfect localization of the genes inside each nucleus.

Cette thèse vise à étudier le couplage entre méthodes de Galerkine discontinue (DG) et méthodes de différences finies (DF) en maillages hybrides non structuré / cartésien, en vue d'applications en aéroacoustique numérique. L'idée d'une telle approche consiste à pouvoir tirer profit localement des avantages respectifs de ces méthodes, soit, en d'autres termes, à pouvoir prendre en compte la présence de géométries complexes par une méthode DG en maillage non structuré, et les zones qui en sont suffisamment éloignées par une méthode DF en maillage cartésien, moins coûteuse. Plus précisément, il s'agit de concevoir un algorithme d'hybridation de ces deux types de schémas pour l'approximation des équations d'Euler linéarisées, puis d'évaluer avec attention le comportement numérique des solutions qui en sont issues. De par le fait qu'aucun résultat théorique ne semble actuellement atteignable dans un cas général, cette étude est principalement fondée sur une démarche d'expérimentation numérique. Par ailleurs, l'intérêt d'une telle hybridation est illustré par son application à un calcul de propagation acoustique dans un cas réaliste / This thesis aims at studying coupling techniques between Discontinuous Galerkin (DG) and finite difference (FD) schemes in a non-structured / Cartesian hybrid-mesh context,in the framework of Aeroacoustics computations. The idea behind such an approach is the possibility to locally take advantage of the qualities of each method. In other words, the goal is to be able to deal with complex geometries using a DG scheme on a non-structured mesh in their neighborhood, while solving the rest of the domain using a FD scheme on a cartesian grid, in order to alleviate the needs in computational resources. More precisely, this work aims at designing an hybridization algorithm between these two types of numerical schemes, in the framework of the approximation of the solutions of the Linearized Euler Equations. Then, the numerical behaviour of hybrid solutions is cautiously evaluated. Due to the fact that no theoretical result seems achievable at the present time, this study is mainly based on numerical experiments. What's more, the interest of such an hybridization is illustrated by its application to an acoustic propagation computation in a realistic case

Parfois posées en héritières du fantastique du dix-neuvième siècle, les écritures singulières des auteurs postcoloniaux exploitant les ressorts du surnaturel sont souvent cataloguées sous l’étiquette homogénéisante de « réalisme magique ». L'intérêt du rappro­chement des œuvres de René Depestre, Rachid Boudjedra et Ben Okri est de pouvoir interroger ces filiations à partir de leur textualisation de la figure du double. Ces littératures, adoptant une posture nouvelle face à son traitement et à son évolution, s'éloignent des écritures fantastiques occidentales. Elles s'en distinguent par l’exploitation de personnages hybrides, dont la posture face au double est conditionnée par cette hybridité-même, une conséquence inéluctable de l'Histoire. Ces auteurs postcoloniaux explorent des ethnoscapes inédits et développent une écriture résolument moderne fondée sur une hybridation générique qui contribue à revivifier le roman à l'occidentale. Leurs poétiques hybrides génèrent ainsi un genre original où le double, loin d’être une simple figure, est une clé pour aborder ces textes aux imaginaires multiples qui jettent de nouvelles bases pour une herméneutique littéraire. / The postcolonial authors who use the supernatural have sometimes been qualified as the heirs of Nineteenth Century fantastic literature, and are often categorized under the homogenizing label of “magical realism”. The relevance of the parallels between the works of René Depestre, Rachid Boudjedra and Ben Okri is apparent when interrogating these filiations through their textualisation of the figure of the Double. This writing confronts the figure's appearance and evolution with a novel posture, and has developed a varied approach different from that of Western fantastic literature. They set themselves apart by their usage of hybrid characters, whose attitude when facing the Double is conditioned by their hybridity; an inescapable consequence of history. The authors explore original ethnoscapes and have developed a thoroughly modern writing built on a generic hybridity, contributing to the revitalization of the Western novel. These hybrid poetics have generated an original genre in which the double, far from it’s basic characterization, represents a key when approaching the multi-facetted imaginations of these works, laying new groundwork for literary interpretation.

M. Tech. (Department of Biosciences, Faculty of Applied and Computer Sciences) Vaal University of Technology / Most cultivated bananas and plantains (Musa spp. sect. Eumusa), originated from two wild diploid species, Musa acuminata Colla (AA) and Musa balbisiana Colla (BB), which contributed the A and B genomes, respectively. The two genomes confer different traits to a banana plant. Intra- and interspecific hybridization between the wild diploid species, somatic mutations and selection over many thousands of years has given rise to considerable genetic variability in cultivated bananas. Bananas are classified according to its genome composition and a number of morphological traits are used to identify the genomes of a plant. Morphological classification can be misleading since the morphology of plants can be affected by environmental factors. Molecular techniques to identify the genomes of banana have many advantages. The objective of this study was to develop a SCAR (sequence characterized amplified region) marker from a previously reported A genome-specific RAPD fragment that distinguish the A genome of banana from the B genome. This fragment designated OPA17600 was cloned, sequenced and used to design longer 20-mer SCAR primers. Verification of the SCAR primers for its fidelity to the A genome was carried out on a sample of 22 homo-and heterogenomic accessions representing landraces and hybrids of different ploidy and genome combinations. Out of six primers sets that were tested one set (SC3) produced a unique 600 bp in all the A genome containing banana accessions. However, these primers also amplified an 800 bp fragment in all the BB genotypes and some accessions containing the A and B genomes. While previous reports suggested that there was considerable differentiation between the A and B genomes, recent evidence points to the contrary. The presence of the A genome fragment in the B genome genotypes and accessions may be due to recombination between the two genomes, translocations and substitutions. The study concluded that the 600-bp SCAR sequence is conserved across the A genome in Musa and can be used to identify the A genome in banana classification and Musa breeding programmes.

Contexte : Le récepteur hormone folliculo-stimulante (FSHR) est exprimé par les cellules de l'endothélium vasculaire dans un large éventail de tumeurs humaines primaires. Notre but était d'évaluer l'intérêt de FSHR comme marqueur des vaisseaux sanguins tumoraux associés aux sarcomes, les sous-types moléculaires du cancer du sein et des métastases ainsi que comme biomarqueur prédictif de la réponse au traitement anti-angiogénique. Méthodes : Nous avons utilisé l'immunohistochimie comme technique de révélation. Ceci implique la production d'un anticorps monoclonal hautement spécifique anti-FSHR (produit chez la souris) et l'hybridation in situ pour détecter FSHR dans des échantillons de tissus provenant de patients atteints de sarcomes (308 patients), les sous-types moléculaires du cancer du sein (84 patients), et des métastases (203 patients). Pour évaluer FSHR comme marqueur prédictif du traitement anti-angiogénique du cancer du rein métastatique avec le sunitinib, nous avons utilisé la microscopie confocale à immunofluorescence. Nous avons également co-localiser FSHR avec le facteur von Willebrand, un marqueur des cellules endothéliales vasculaires (50 patients).RésultatsFSHR est exprimé dans les 11 sous-types de patients atteints de sarcomes ont analysés et dans 75% des tumeurs métastatiques examinées, ainsi que dans tous les sous-types moléculaires des cancers du sein. Dans cadre de l'étude du cancer du rein métastatique, le pourcentage de vaisseaux marqués FSHR était en moyenne cinq fois plus élevé pour les patients qui ont répondu au traitement par rapport au groupe stable et presque huit fois plus élevé que dans le groupe non-réponse (57%, 11% et 7 %, respectivement).ConclusionsNos résultats montrent que, en plus des cancers signalés précédemment, FSHR peut être considéré comme marqueur tumoral pour les sarcomes et les métastases. En outre, FSHR peut être utilisé, avec une sensibilité et une spécificité élevée, en tant que biomarqueur prédictif de la réponse au traitement par sunitinib des patients atteints de cancer du rein métastatique. / Background : Follicle Stimulating Hormone receptor (FSHR) is expressed by the vascular endothelium in a wide range of human primary tumors. Our purpose was to further evaluate FSHR as marker of tumor blood vessels associated with sarcomas, breast cancer molecular subtypes, and metastases as well as predictive biomarker of response to antiangiogenic treatment.MethodsWe used immunohistochemistry involving a highly specific mouse monoclonal anti-FSHR antibody and in situ hybridization to detect FSHR in tissue samples from patients with sarcomas (308 patients), breast cancer molecular subtypes (84 patients), and metastases (203 patients). To evaluate FSHR as predictive marker of antiangiogenic treatment of metastatic kidney cancer with sunitinib, we used immunofluorescence confocal microscopy to co-localize FSHR with von Willebrand factor, a marker of vascular endothelial cells (50 patients).ResultsFSHR is expressed in all 11 subtypes of sarcoma patients analysed, in 75% of metastatic tumors examined as well as in all different molecular subtypes of breast cancers. In metastatic kidney cancer patients the percentage of FSHR stained vessels was on average fivefold higher for the patients who responded to the treatment in comparison with the stable group and almost eightfold higher than in the non-responsive group (57%, 11%, and 7%, respectively).ConclusionsOur results suggest that, in addition to the cancers previously reported, FSHR can be considered as tumor marker for sarcomas and metastasis. Moreover, FSHR can be used, with high sensitivity and specificity, as predictive biomarker for the response to sunitinib treatment of patients with metastatic kidney cancer.

Les oligopyridine-dicarboxamides et les oligoquinoline-carboxamides sont des oligomères synthétiques capables d’adopter des conformations hélicoïdales stables et bien définies. Les premiers sont comparables à des ressorts moléculaires qui peuvent s’étirer puis s’autoassembler pour former des doubles hélices artificielles. L’étude structurale d’oligopyridines de différentes tailles par diffraction des rayons X et RMN a permis d’éclaircir les principes de l’hybridation en double hélice. Par exemple, nous constatons que la stabilité du duplex est d’autant plus grande que l’oligomère est long mais la cinétique de l’hybridation décroit avec la taille des hélices. Ces propriétés sont modulables en fonction de divers paramètres tels que le solvant ou les substituants des pyridines. Les seconds forment de simples hélices moléculaires stables dans les solvants organiques mais aussi dans l’eau. Nous avons développé leur synthèse sur support solide afin de disposer de séquences variées, à l’image des alpha-peptides. Des études par RMN suggèrent que l’introduction d’unités aminométhylpyridines au sein d’un oligoquinoline hydrosoluble apporte de la flexibilité sans perturber sa structure hélicoïdale. Cela témoigne de la stabilité de ces structures secondaires dans les solvants protiques. Par ailleurs, certains de ces peptidomimes s’avèrent capables de reconnaitre et stabiliser des motifs structuraux particuliers de l’ADN : les G-quadruplex. Etant donné que ces architectures se forment à des endroits clés du génome impliqués dans des cancers, ces hélices moléculaires font figure de potentiels agents antitumoraux d’un nouveau genre. / Oligopyridine-dicarboxamides and oligoquinoline-carboxamides are synthetic oligomers able to fold into stable and well defined helical conformations. The first ones are comparable to molecular springs which can extend then associate to form artificial double helices. A structural study of oligopyridines of various sizes by X-ray diffraction and NMR provided a better understanding of the hybridization process. For example, we noticed that the stability of the duplex is all the higher as the oligomer is long but the kinetics of hybridization decrease with length. These properties depend on diverse parameters such as the solvent or the substituants of pyridine rings. The second family forms stable single helices in organic solvents and also in water. We adapted their synthesis on solid support to promote accessibility to a variety of sequences, just like for alphas-peptides. NMR studies suggested that the introduction of aminomethylpyridine units within a hydrophilic oligoquinoline strand brings some flexibility without disrupting its helical structure, showing the high stability of these secondary structures in protic solvents. Besides, some of these peptidomimetics turn out to be capable of recognizing and stabilizing a particular DNA motif: G-quadruplex structure. Given that these architectures form in critical places of the genome involved in cancers, these molecular helices may represent a new class of potential antitumoral agents.

Les foldamères de type oligopyridine biscarboxamide constituent une famille d’oligomères synthétiques pouvant adopter une structure hélicoïdale et s’hybrider pour former des hélices doubles. Ce travail a eu pour objectif de synthétiser une nouvelle génération de foldamères π-fonctionnels porteurs de groupements photoactifs ou électroactifs, d’étudier les facteurs gouvernant l’équilibre entre hélice simple et hélice double, d’analyser l’impact de cet équilibre sur les propriétés optiques, et enfin de mettre en place un nouveau type de stimulus permettant de contrôler cet équilibre. Deux foldamères photoactifs dotés d’unités Disperse Red, ont été synthétisés. Leurs structures cristallographiques confirment la formation de structures hélicoïdales. Un choix précis du solvant permet d’orienter sélectivement l’équilibre vers la formation d’une hélice simple ou double.Le contrôle de l’équilibre d’hybridation par dilution permet de moduler l’activité en Génération de Seconde Harmonique du foldamère. En outre, la cavité générée par l'hélice permet la reconnaissance de divers anions. Enfin, les premiers efforts fournis pour induire une hélicité donnée à ces foldamères par voie supramoléculaire sont décrits. Par ailleurs, un foldamère électroactif fonctionnalisé par deux unités tétrathiafulvalène (TTF) a été synthétisé selon une méthodologie originale. La présence des unités TTF permet un contrôle redox inédit de la structuration du foldamère, par dimérisation de cations radicaux. Le concept a été élargi via l’immobilisation d’un foldamère sur surface d’or (SAMs). Enfin, une capsule électroactive capable de complexer l’acide tartrique a également été synthétisée et caractérisée. / Oligopyridine biscarboxamide-based foldamers constitute a family of synthetic oligomers that can fold into helical structures and hybridize to form double helices. This work aims at synthesizing a new generation of π-functionalized foldamers featuring photoactive and electroactive moieties, in order: to study the factors governing the equilibrium between simple and double helices, to analyze the impact of this equilibrium on the optical and recognition properties, and to set up a new type of stimulus to control this equilibrium. Two photoactive foldamers of different lengths and bearing two Disperse Red units were synthesized. Their crystallographic structures confirm the formation of helical structures. A precise choice of the solvent allows to drive the equilibrium towards the single or the double helix selectively.The cavity generated within the helix presents a good affinity for anions. The control over the hybridization equilibrium allows modulating the Second Harmonic Generation activity. Eventually, our first efforts to control the helicity of these foldamers through supramolecular chiral induction are described. On the other hand, an electroactive foldamer featuring two tetrathiafulvalene (TTF) units was synthesized according to an original methodology. The presence of TTF units allows an unprecedented redox control of the structure of foldamer, by dimerization of radical cations. The concept has been extended by immobilizing a foldamère on a gold surface (SAMs). Finally, an electroactive capsule capable of complexing tartaric acid has also been synthesized and characterized.

Cytogenetics of spiders of the infraorder Mygalomorphae is largely unknown. My thesis is focused on the karyotype evolution of spiders of families Hexathelidae Dipluridae, which are basal groups of the superfamily Avicularioidea. In this thesis the evolution of diploid chromosome number, chromosome morfology, and also sex chromosomes are studied. The karyotypes of the mygalomorphs are quite interesting for their diversity. Many members of the family Hexathelidae exhibit extreme systems of a chromosomal sex determination which are composed by many gonosomes X. The meiotic division of males is analyzed. Some species exhibit modifications of this division such as achiazmatic meiosis. Selected markers have been detected by fluorescence in situ hybridization. Keywords: evolution, fluorescent in situ hybridization, karyotype, nucleolus organizer region, sex chromosomes, spider

INTRODUÇÃO: Recentemente, a classificação molecular do câncer gástrico (CG) emergiu como opção promissora para definir subgrupos prognósticos, distinguir o comportamento biológico, além de permitir a identificação de potenciais alvos terapêuticos para drogas específicas. Por meio de técnicas moleculares, o CG foi dividido em 4 subtipos: instabilidade de microssatélites (MSI), vírus Epstein-Barr (EBV)-positivo, genomicamente estável (GS) e instabilidade cromossômica (CIN). Os custos associados à complexidade técnica das metodologias moleculares são ainda um obstáculo para sua implantação na prática de rotina. OBJETIVOS: Determinar os grupos da classificação molecular por meio da expressão proteica de marcadores associados a cada subtipo. Comparar as diferentes classificações moleculares existentes e propor um novo modelo. MÉTODOS: Foram avaliados, retrospectivamente, 287 pacientes com CG submetidos à gastrectomia-D2 curativa, por meio da construção de tissue microarray. A expressão das proteínas de reparo do DNA, E-caderina e p53 foram avaliadas por imuno-histoquímica (IH), determinando os grupos MSI, GS e CIN, respectivamente. O EBV foi detectado por hibridização in situ (ISH). RESULTADOS: Após avaliação histopatológica, 179 (62,4%) pacientes foram classificados como CIN, 58 (20,2%) MSI, 30 (10,5%) EBV e 20 (7%) como GS. Os subtipos associaram-se com características distintas, tais como: gênero masculino (EBV, p=0.101); idade avançada (MSI, p=0,017), menor relação neutrófilo-linfócito (CIN, p=0,029), gastrectomia total (EBV, p < 0,001), localização distal (MSI, p=0,004), Laurén difuso (GS, p < 0,001), e estádio avançado (GS, p=0,014). O subtipo MSI apresentou melhor sobrevida livre de doença (SLD) (82,8%), seguido pelo EBV e CIN (ambos com 70%) e GS (50%) (p=0.005). A sobrevida global (SG) foi maior nos tumores MSI (75,9%), seguido pelo EBV (73,3%), CIN (65.4%) e GS (45%, mediana de 25 meses) (p=0.007). Em análise multivariada, gastrectomia total, pT, pN e os subtipos tumorais foram fatores significativos associados à SLD (MSI p=0,012; EBV p=0,037; CIN p=0,018; GS referência). Do mesmo modo, o tipo de cirurgia, pT, e os subtipos tumorais foram fatores independentes associados a SG (MSI p=0,010; EBV p=0,006; CIN p=0,025; GS referência). Com base no risco de recidiva dos pacientes do estudo, nova classificação, que inclui 5 subtipos, foi proposta. Evidenciou-se que a classificação por risco e cluster tiveram melhor acurácia para identificar recidivas e óbitos respectivamente. CONCLUSÃO: A análise IH/ISH foi capaz de determinar subtipos de CG com características clinicopatológicas e prognósticos distintos, reproduzindo os subtipos obtidos pela classificação molecular / BACKGROUND: Recently, the molecular classification of gastric cancer (CG) emerged as a promising option to define prognostic subgroups, to distinguish biological behavior, and to identify potential therapeutic targets for specific drugs. Through molecular techniques, GC was divided into 4 subtypes: microsatellite instability (MSI), Epstein-Barr virus (EBV) positive, genomically stable (GS) and chromosomal instability (CIN). The costs associated with the technical complexity of the molecular methodologies are still an obstacle to its implementation in routine practice. OBJECTIVES: To determine molecular classification groups by means of protein expression of markers associated with each subtype. To compare the different existing molecular classifications and propose a new model. METHODS: We retrospectively evaluated 287 CG patients submitted to curative D2-gastrectomy through the construction of tissue microarray. Expression of the DNA repair proteins, E-cadherin and p53 were evaluated by immunohistochemistry (IH), determining the MSI, GS and CIN subtypes, respectively. EBV was detected by in situ hybridization (ISH). RESULTS: After the histopathological evaluation, 179 (62.4%) patients were classified as CIN, 58 (20.2%) MSI, 30 (10.5%) EBV and 20 GS (7%) as GS. The subtypes presented associations with distinct characteristics, such as: male gender (EBV, p=0.101); advanced age (MSI, p=0.017), Laurén diffuse type (GS, p < 0.001), lower neutrophil-lymphocyte ratio (CIN, p=0.029), total gastrectomy (EBV, p < 0.001), distal location (MSI, p=0.004), and advanced stage (GS, p=0.014). The MSI subtype presented better disease-free survival (DFS) (82.8%), followed by the EBV and CIN subtypes (both with 70%) and GS (50%) (p=0.005). Overall survival (OS) was higher in MSI tumors (75.9%), followed by EBV (73.3%), CIN (65.4%) and GS (45%, median of 25 months) (p=0.007). In multivariate analysis, total gastrectomy, tumor invasion, lymph node metastasis, and tumor subtypes were significant factors associated with SLD (MSI p=0.012, EBV p=0.037, CIN p=0.018, GS reference). Likewise, type of surgery, pT, and tumor subtypes were independent factors associated with OS (p=0.010, p=0.010, EBV p=0.006, CIN p=0.025, GS reference). Based on the risk of recurrence, a new classification including 5 subtypes was proposed. It was evidenced that the risk classification and cluster had better accuracy to identify recurrences and deaths respectively. CONCLUSION: The IH / ISH analysis was able to determine CG groups with clinicopathological characteristics and distinct prognoses, reproducing the subtypes obtained by the molecular classification

A esquistossomose é uma doença causada por trematódeos do gênero Schistosoma. Dentre os genes identificados no transcriptoma do parasita, membros da família gênica SmVAL (Schistosoma mansoni Venom Allergen-Like) foram apontados como candidatos vacinais. SmVALs foram identificadas em secreções de cercárias e esquistossômulos cultivados in vitro, os transcritos SmVAL4 e 24 foram localizados nas glândulas acetabulares de germ ball e a proteína nativa SmVAL4 foi identificada em extrato de cercárias, indicando funções durante a penetração da pele. Já os transcritos SmVAL13 e 14 foram localizados na glândula esofágica anterior de vermes adultos, sugerindo papéis no processo de alimentação sanguínea. A imunização com as proteínas rSmVAL4, 6, 7, 13, 14 e 18 coadministradas não protegeu camundongos contra o desafio experimental, porém, observou-se uma diminuição do número de fêmeas e do número de ovos no grupo imunizado. A investigação de funções para as proteínas secretadas mostrou que a rSmVAL18 interage com plasminogênio in vitro favorecendo a invasão do hospedeiro. / Schistosomiasis is a disease caused by trematodes of the genus Schistosoma. Among the genes identified in the parasite transcriptome, members of SmVAL (Schistosoma mansoni Venom Allergen-Like) gene family were proposed as vaccine candidates. SmVALs were identified in cercariae and schistosomule secretions in vitro, the SmVAL4 and 24 transcripts were located to the germ ball acetabular glands and SmVAL4 native protein was identified in cercariae extract, indicating functions in skin penetration. On the other hand, SmVAL13 and 14 transcripts were located to the anterior esophageal gland of adult worms, suggesting roles in the blood feeding processes. Immunization with rSmVAL4, 6, 7, 13, 14 and 18 proteins co-administered did not protected mice against experimental challenge, however, there was a decrease in the number of females and the number of eggs in the immunized group. The investigation of functions for secreted proteins showed that rSmVAL18 interacts with plasminogen in vitro thus favoring the host invasion.