HYAL-1 (hyaluronidase-1) appartient à la famille des hyaluronidases connues pour leur rôle dans la dégradation de l’acide hyaluronique. L’expression de HYAL-1 est élevée dans de nombreux type de cancers, notamment dans le cancer de la prostate, de la vessie, des reins et du sein où il est impliqué dans la croissance tumorale et les métastases. Récemment notre laboratoire a aussi démontré une expression élevée de HYAL-1 dans le cancer épithélial de l’ovaire (CEO) de type mucineux et à cellules claires, expression qui est inversement corrélée à celle du récepteur de l’oestrogène alpha (REα). Cependant, malgré le fait que le rôle de HYAL-1 dans le cancer soit bien établit, le mécanisme de sa régulation reste encore inconnu. Le REα est un facteur de transcription qui suite à sa liaison avec son ligand va réguler l’expression de plusieurs gènes. Le REα ainsi stimulé par l’hormone va activer la transcription de ces gènes cibles mais il est connu maintenant qu’une grande partie des gènes régulés par le REα sont en réalité réprimés par ce récepteur. Dans ce travail nous proposons d’étudier le mécanisme de la régulation du gène HYAL-1 par le REα dans le CEO à cellules claires et dans le cancer du sein. L’expression ectopique du REα dans la lignée TOV21G (RE-) de même que le traitement de la lignée MCF-7 (RE+) avec de l’oestrogène a induit une diminution du niveau d’expression de l’ARN m de HYAL-1. Ces résultats nous ont permis de confirmer que HYAL-1 est un gène cible du REα. Il est aussi connu que le REα peut exercer son action par différents mécanismes d’action, entre autres en interagissant avec une séquence d’ADN appelée élément de réponse à l’oestrogène (ERE), retrouvé sur le promoteur des gènes cibles ou bien indirectement par des interactions protéine-protéine en se liant à d’autres facteur de transcription tels que Sp1. Après avoir identifiés de telles séquences sur le promoteur proximal de HYAL-1, (1 ERE proximal à -900 pb, 3 distaux à -32350 pb, 48430, -50130 pb du site d’initiation de la transcription) en plus des 2 Sp1 connus (-60 et – 1020pb), nous avons démontrés par immunoprécipitation de la chromatine que le REα est recruté sur le promoteur de HYAL-1 au niveau de l’ERE proximal -900 pb et du distal -32350 pb de même que sur le site Sp1 -1020 pb. De plus, l’activité biologique de l’ERE -900 pb et du ii Sp1-1020pb à été confirmée par des essais de gènes rapporteurs à la luciférase. Avec son rôle connu dans la tumorigenèse, l’identification de HYAL-1 comme gène cible du REα pourrait être une avenue intéressante pour le traitement des cancers hormono-indépendants. / HYAL-1 (hyaluronidase-1) belongs to the hyaluronidase family of enzymes that degrade hyaluronic acid. HYAL-1 expression is elevated in many types of cancers including prostate, bladder, liver and breast cancer where it is involved in tumor growth and metastasis. In accordance to these observations, our group has also demonstrated high expression of HYAL-1 in clear cell and mucinous epithelial ovarian cancer (EOC) subtypes which was inversely correlated to that of estrogen receptor alpha (ERα). However, despite the fact that the role of HYAL-1 in cancer is well established, the mechanism of its regulation is still unknown. ERα is a transcriptional factor that regulates target-gene expression following ligand binding. Upon hormone stimulation, activated ERα will upregulate transcription of many target genes. However, it has been recently well documented that a large number of ERα responsive genes are in fact repressed. In this work we propose to study the mechanism by which ERα regulates HYAL-1 expression in clear cell EOC subtype as well as in breast cancer. The ectopic expression of ERα in TOV21G cell line (ERα -) and estrogen treatment of MCF-7 cells (ERα +) decreased HYAL-1 mRNA expression and allowed us to confirm that HYAL-1 is an ERα target gene. It is also known that ERα may exert its action through different mechanisms of action including interacting with a DNA sequence called estrogen response element (ERE) found in the promoter of target genes or indirectly by protein-protein interactions by binding to other transcription factor such as Sp1. Having identified such sequences in the proximal promoter of HYAL-1, (one proximal ERE -900 bp, 3 distals at -32350, -48430, -50130 bp from the start site of transcription) in addition to the two known Sp1 (-60 and -1020pb), we have demonstrated by chromatin immunoprecipitation that ERα is recruited at the HYAL-1 promoter at the ERE sites -900 pb and -32350 pb as well as at the Sp1 site -1020. Furthermore, the biological activity of the proximal ERE -900 and Sp1 -1020 sites were further confirmed by luciferase reporter gene assay. Given its known role in tumorigenesis, identification of HYAL-1 as an ERα target may provide an interesting approach for the treatment of hormono-independent cancer.

La conception de la théorie de l’interprétation est constante. Elle repose sur une distinction entre les textes contractuels clairs et les textes contractuels ambigus. L’interprétation des textes ambigus relève du pouvoir souverain des juges du fond tandis que l’interprétation des textes clairs est considérée comme une dénaturation, cas d’ouverture à cassation. Toutefois, cette conception classique de la théorie de l’interprétation semble contestable. Les textes législatifs relatifs à cette théorie ne justifient pas le clivage entre les textes clairs et les textes ambigus, que nous proposons d’écarter au profit de l’admission de l’absoluité du pouvoir d’interprétation des juges du fond. Par ailleurs, l’opération d’interprétation des contrats est régie par les directives déclaratives, les directives protectrices de la partie faible et les directives complétives. Elle est marquée par l’emprise de l’équité sur l’interprétation complétive. L’ensemble des directives d’interprétation conduit à la mise en oeuvre de « l’équité subjective originelle », ou subsidiairement de « l’équité complétive », « subjective » par référence à la volonté contractuelle sinon « objective » en se fondant sur la conception raisonnable de la société. L’interprétation s’avère un processus au service de l’équité contractuelle. Elle contribue à la préservation de l’équité subjective contractuelle et conduit à défaut à l’élaboration d’une équité objective contractuelle. Différentes conceptions juridiques constituent des applications avérées de cette lecture de la théorie de l’interprétation, à savoir l’interdiction de détournement de la finalité d’un droit, l’abandon de l’acte ostensible simulé pour l’application de l’acte secret sincère et l’adjonction d’obligations à certains contrats. En parallèle, de nouvelles applications sont proposées. La théorie de l’interprétation peut constituer le complément de la notion de cause, le dénouement de la problématique de la révision pour imprévision et une technique de révision des contrats nés déséquilibrés. / The concept of the theory of interpretation has been always based on the distinction between the clear contractual texts and the ambiguous contractual texts. The ambiguous texts are interpreted freely by the judges on merits whereas the interpretation of the clear texts is considered as a denaturation and a cause to claim cassation. However, this classic concept of the theory of interpretation seams disputable. As the difference in the interpretation of the clear and ambiguous texts is not justified by legal texts, we suggest to drop out this distinction and to give the judges the absolute authority to interpret all texts. Besides, the interpretation of contracts is governed by three rules: the declaration of the real common intention of the contracting parties rather than paying attention to the literal meaning of the terms, the protection of the weak party and the completion of the intention of the parties. The interpretation theory leads to the application of the equity which prevails over the completion rules. The judges will initially ascertain the “original subjective equity”. If not applicable, they will then establish the “subjective completive equity” depending on the contractual will, or the “objective completive equity” based on the reasonable conception of the society. The interpretation theory favors the contractual equity. It contributes to preserve the subjective contractual equity and creates an objective contractual equity. Several legal concepts constitute a genuine application of our understanding of the theory of interpretation, such as the interdiction of using a contractual right contrary to the reason created for, the abandonment of the fake apparent act in order to apply the real unrevealed one, and the addition of obligations to some contracts. In parallel, new applications are suggested. The theory of interpretation may complete the notion of cause, handle the problem of the unforeseen circumstances and permit to revise the imbalanced contracts.

O carcinoma de célula renal (RCC) subtipo célula clara é o câncer mais letal e prevalente do sistema urinário. O diagnóstico deste tipo de câncer frequentemente é tardio em conseqüência da falta de sintomas perceptíveis aos pacientes. Um dos objetivos deste trabalho é a identificação de novos marcadores moleculares para diagnóstico precoce, o que ajudaria a diminuir a mortalidade em função de complicações resultantes do avanço da doença. Outro objetivo é a identificação de um conjunto de marcadores moleculares de prognóstico, de modo à prever com acurácia a evolução clínica da doença e, por conseqüência, o tempo de sobrevida do paciente. As modificações transcricionais associadas à carcinogênese e à progressão do câncer de rim ainda não foram completamente elucidadas. Além dos oncogenes e genes supressores de tumor, RNAs não-codificadores (ncRNAs) recentemente foram apontados como importantes reguladores da expressão gênica em humanos, e podem ter um papel importante na transformação maligna do câncer de rim. Para analisar a expressão gênica de ncRNAs e de genes codificadores para proteína foram utilizados dois microarranjos desenvolvidos por nosso grupo, enriquecidos em sondas para ncRNAs. Uma das plataformas possui 4 mil sondas de cDNA, das quais 822 sondas são para ncRNAs mapeando em regiões intrônicas. Outra possui 44 mil elementos e combina sondas de oligonucleotídeos (60-mer) intrônicas e exônicas de um mesmo locus genômico. Análises estatísticas foram feitas com a ferramenta Significance Analysis of Microarrays (q &#8804; 0,05) combinadas ou com a técnica de \"patient leave-one-out\" (genes com presença em 8 100% dos subconjuntos), ou alternativamente com o teste discriminante de Golub (p &#8804; 0,01 ou p < 0,05). Com a plataforma de 4 mil sondas foram estudadas 30 amostras de tecido renal de 18 pacientes com RCC subtipo célula clara. Um conjunto de 36 ncRNAs foi identificado como diferencialmente expresso entre amostras tumorais e não-tumorais. Uma assinatura adicional de 265 genes codificadores de proteínas foi identificada, indicando possíveis novos marcadores moleculares. Uma análise estatística supervisionada com dados de 16 pacientes identificou uma assinatura de ncRNAs correlacionada com sobrevida de 5 anos, formada por 27 ncRNAs com significativa expressão alterada em pacientes livres da doença em comparação com pacientes que morreram em função da doença. Uma assinatura adicional de 64 genes codificadores de proteínas também foi identificada como significativamente correlacionada com o acompanhamento clínico dos pacientes. Com a plataforma de 44 mil sondas foram analisados 17 pacientes, com amostras pareadas de tecido renal tumoral e não-tumoral agrupadas em 8 pools, sendo 4 de amostras tumorais e 4 de não-tumorais. Um conjunto de 66 ncRNAs parcialmente intrônicos antisenso e outro de 52 ncRNAs totalmente intrônicos antisenso foram identificados como diferencialmente expressos. Identificamos um subconjunto de 28 ncRNAs totalmente intrônicos antisenso e senso cuja expressão do gene codificador de proteína do mesmo locus estava simultaneamente alterada. Estes dados apontam para possíveis redes de regulação da expressão gênica dos ncRNAs em câncer. A extensa lista de ncRNAs e de genes codificadores para proteína identificados neste estudo podem ser promissores marcadores moleculares de carcinoma renal subtipo célula clara. / Renal cell carcinoma (RCC) is the most common malignancy of the adult kidney, and the clear cell subtype is the most prevalent and lethal cancer of the urinary system. Late diagnosis for this type of cancer is frequent, usually as a consequence of the lack of symptoms. One of the objectives of the present work is the identification of new molecular markers for the early diagnosis, which would help decrease mortality that develops as a function of disease progression. Another objective is the identification of a set of prognosis molecular markers, so as to accurately predict the clinical outcome of the disease, and consequently, patient survival. Transcriptional changes associated to carcinogenesis and to kidney cancer progression have not been entirely elucidated. Besides oncogenes and tumor suppressor genes, non-coding RNAs (ncRNAs) have been recently indicated as important regulators of gene expression in humans, and could have an important role in the malignant transformation in renal cancer. In order to measure ncRNA and protein-coding gene expression we have used two microarray platforms developed by our group, which are enriched in ncRNA probes. One of the platforms has 4 thousand cDNA probes, of which 822 are for ncRNAs that map to intronic regions. Another has 44 thousand elements and combines 60-mer oligonucleotide probes for intronic and exonic regions from the same genomic locus. Statistical analyses have been performed with the Significance Analysis of Microarrays tool (q &#8804; 0.05) combined with a patient leave-one-out approach (genes present in 100% of the sub-sets), or alternatively with Golubs discriminant test (p &#8804; 0.01 or p < 0.05). 11 With the 4-thousand probes platform we studied 30 samples from renal tissue of 18 RCC patients with clear cell subtype. A set of 36 ncRNAs has been identified as differentially expressed between tumor and non-tumor tissue. An additional signature of 265 protein-coding genes has been identified, indicating possible new molecular markers. A supervised statistical analysis with data from 16 patients has identified a ncRNA signature correlated to 5-year survival outcome, comprised of 27 ncRNAs with significantly altered expression in diseasefree patients compared to patients who died from cancer within the 5-year follow-up. An additional 64-gene signature of protein-coding genes has been identified as significantly correlated to clinical outcome. With the 44-thousand probes platform we have analyzed 17 patients, with paired tumor and non-tumor samples grouped into 8 pools, of which 4 were from tumor and 4 from nontumor samples. A set of 66 partially intronic antisense ncRNAs and another of 52 totally intronic antisense ncRNAs have been identified as differentially expressed between tumor and non-tumor tissue. A sub-set of 28 totally intronic antisense or sense ncRNAs were identified as having a simultaneous change in expression of the protein-coding gene from the same locus. Overall, the data point to a possible ncRNA regulatory network in cancer. The extensive lists of ncRNAs and of protein-coding genes identified in the present study can be seen as promising molecular markers of RCC from the clear-cell subtype.

Introdução: com o uso generalizado de métodos de imagem, aumentou-se a detecção de lesões renais como achados incidentais. Tais lesões podem ser tanto benignas, tais como os angiomiolipomas, e outras malignas, como os carcinomas de células renais, portanto torna-se importante um método não-invasivo com boa acurácia para sua distinção . A ultrassonografia é pouco específica para este fim. A tomografia computadorizada e a ressonância magnética são os métodos mais utilizados na caracterização de lesões incidentais renais. Na Ressonância Magnética, o uso de sequências convencionais ponderadas em T2W e T1W, antes e após a administração endovenosa de contraste paramagnético, mostrou-se pouco eficaz para este fim. Técnicas quantitativas associadas às imagens convencionais, tais como a oposição de fase e a restrição a difusão da água, vêm sendo estudadas, devido ao potencial para melhorar a caracterização não-invasiva, evitando nefrectomias parciais ou totais, e outras intervenções invasivas por lesões benignas. Objetivos: avaliar a acuidade diagnóstica de técnicas combinadas de ressonância magnética para diferenciação de angiomiolipomas pobres em gordura de lesões malignas do rim. Métodos: pacientes que obtiveram o diagnóstico histológico das lesões renais entre os anos de 2010 e 2014 e que realizaram exame pré-operatório. As lesões foram estudadas, colocando-se um ROI (region of interest, no inglês) na maior parte da lesão e córtex renal normal, evitando-se área de não-lesões, calculando a intensidade de sinal nas seqüências T1W in e out-phase, T2W, o Wash in, Wash out relativo e absoluto das lesões e o cálculo absoluto do sinal da lesão no ADC (coeficiente de difusão aparente, do inglês). Os resultados foram obtidos na forma de índices padronizados pelo córtex renal e baço, pelas fases pré e pós-contraste, e de forma absoluta pelo ADC. Os resultados foram confrontados com o diagnóstico final e feito associações estatísticas para observar a relevância. Resultados:Foram estudadas 85 lesões em 74 pacientes, sendo 40 do gênero masculino e 34 do feminino. O cálculo do teor de gordura se mostrou ineficaz para distinção entre CCRs e AMLpg; o índice de intensidade de sinal em T2W, lesão/córtex normal foi útil na diferenciação dos CCRs CC de AMLpg. Outro parâmetro importante foi a cálculo de wash out relativo que se mostrou mais acentuado no AMLpg que em todos os subtipos de CCRs estudados e da medida do ADC médio, que apresentou valores maiores nos casos de CCR CC, em comparação com os outros subtipos e com os AMLpg. Conclusão: As técnicas combinadas de RM, principalmente o índice de sinal T2W da lesão, Wash out relativo e IS do ADC, associados a dados epidemiológicos são viáveis, quando utilizados em conjunto, para a diferenciação de lesões malignas renais dos angiomiolipomas, podendo ter implicações na conduta terapêutica, com redução do número de nefrectomias por lesão benignas. / Introduction: With the widespread use of imaging methods, detection of incidental renal masses has steadily increased in recent years, and these may be either benign, such as angiomyolipoma, or malignant, such as renal cell carcinomas. Therefore, it is important to have a method that allows accurate characterization. Ultrassonography is not very specific for this purpose. CT and MRI are the methods used in the characterization of renal incidental lesions. In MRI, the use of conventional sequences, such as T1W and T2W before and after intravenous administration of paramagnetic contrast media, has proved ineffective for this purpose. Quantitative techniques associated with conventional images, such as chemical shift and diffusion weighted imaging (DWI), have played a key role in this differentiation, which aims to improve characterization, avoiding partial or total nephrectomy, and other invasive interventions for benign lesions. Objectives: to evaluate the diagnostic accuracy of combined techniques of MRI to differentiate fat-poor angiomyolipoma from renal cell carcinomas. Methods: Patients who had a histological diagnosis of renal lesions between 2010 and 2014 and underwent pre-operative exam. An ROI (region of interest)cwas placed in most of the lesion and normal renal cortex, avoiding area of non lesions, by calculating the signal intensity in all sequences, in and out phase T1W, T2W, and the wash in, wash out, relative and absolute, of the lesions and estimation of the ADC. The results were obtained in the form of standardized indices for renal cortex and spleen, the pre- and post-contrast phases, and absolute values for ADC. Results were confronted with the final diagnosis and statistical analysis to observe the relevancy. Results: the estimation of intracellular fat content was ineffective for characterization, while the T2W signal intensity index was used for differentiation between CCRs clear cells from fat-poor AML. Another important parameter was the \"wash out\", which was more prominent for AMLpg. ADC values was higher for CCR CC. Conclusion: We concluded that the combined techniques of MRI mainly T2W signal ratio, \"Wash out\" and ADC values, when used in association and correlated with epidemiological data may be feasible for the differentiation among fat-poor angiomyolipomas and renal malignancies, with important therapeutics implications, reducing unnecessary nephrectomies for benign lesions.

The role of communities associated to a substrate as fundamental components of an aquatic system has been recognized by many authors. Although periphyton has been considered of great importance in shallow lakes, its inclusion in different ecological models has been unusual. Phillips et al. (1978) were the first in considering epiphyton as an internal mechanism leading the system to a change in its state from greater water transparecy to more turbid conditions. In contrast, Scheffer et al. (1993) proposed the Alternative Equilibria Hypothesis (AEH), which considers that a shallow lake can alternate between two states, a Clear Water State (stabilized by the presence of submerged macrophytes) and a Turbid Water State (stabilized by an elevated phytoplankton biomass), excluding in this scheme the periphytic communities. Later only the conceptual model coined by Goldsborough and Robinson (1996, G&R model) emphasizes again the importance of epiphyton as well as that of epipelon and metaphyton by establishing the possibility of occurrence of four states according to the contributions to the total algal biomass of each of the algal taxocenosis. Some analyses of the system’s dynamics have been performed in pampean lakes in the context of the AEH. On the other hand, knowledge regarding periphytic and benthic communities is still poor. We propose as an hypothesis that phytoperiphyton will respond to the occurrence of different equilibria states, displaying, at the same time, strong interrelationships with the phytobenthos as well as with the phytoplankton. On the other hand, we consider the possibility that the phytoperiphytic and phytobenthic assemblages may function as internal mechanisms both, as state change or as state stability factors. Finally, we assess the importance of phytoperiphyton as regards its contributions to the total algal biomass in the context of the G&R model, comparing then the results with those obtained in the case of the AEH. The general objectives of the present investigation are to gather information about the structure and dynamics of the phytoperiphyton and its relationships with environmental factors as well as with the phytobenthon and phytoplankton in a pampean shallow lake; and to analyze phytoperiphytic dynamics under the AEH and the G&R model contexts, evaluating periphytic contributions to the total algal biomass. This investigation was carried out in an seepage shallow lake, Lacombe Lake, which is included in the “Grupo Marginal de lagunas” group in the pericoastal region of Provincia de Buenos Aires. Sampling was performed from July 2001 to June 2002 at four sites according to their exposure to environmental factors and the presence of macrophytes: bulrush beds, (JP and JC); and sites devoided of emergent vegetation but with stands of submerged macrophytes during the growing season (ALe and ALr). J sites and ALe were also representative of deeper zones and ALr of a site near the shore. Environmental variables were measured in situ and water samples were collected for chemical analysis and for plankton identification and quantification, following a vertical profile (excepting in ALr). In the case of phytoperiphyton, bulrush epiphyte samples (epiphyton) were taken following a vertical profile and those of submerged macrophytes were collected from the first 25 cm of the stems. Benthon (epipelon) was sampled using a corer. Aliquots for chemical analysis were also extracted from those samples. Vegetal substrate dynamics was also evaluated during sampling. At the laboratory, concentrations of algal nutrients, of the eight principal ions and of soluble polyphenols, as well as alkalinity and total hardness were measured. Phytoplanktonic, epiphytic and epipelic samples were treated for active a chlorophyll and pheopigment determinations. Organism identification and quantification were made according to standardized and own developed methods. First, variations in biotic and abiotic variables were analyzed for the determination of different equilibrium states occurrence. Three perturbation events were registered: precipitations in August, October and March. All of them involved water level increases and conductivity diminutions. Especially in the first two events, nutrient input from runoff was observed; meanwhile in March the input of water determined the dilution in nutrients concentrations. On the other hand, light conditions improved, particularly from October onwards, when stands of submerged macrophytes developed and a decrease of phytoplankton density and a change in its specific composition was detected. The latter being generated by the dilution effects of water input, the shading of the aquatic plants and algal consumption by cladocerans. March event also involved an improvement of light underwater climate but the increase in the water level affected macrophyte stands which mainly withdrawed to the shallowest zones. As regards phytoplankton, changes in the specific composition were recorded again. The statistic evaluation of these facts in conjunction with the analysis of permanence, resiliency and resistance properties of the dominating taxocenosis leads to interpret in the context of the AEH that two equilibria states occurred in the shallow lake. During the first three months a Turbid Water State developed, given by the phytoplankton domination given by an oscilatorial cyanophyte, minor transparency and relatively higher nutrient concentrations. In October and November the passage between states occurred, thus considering these months as transitional. From December onwards a Clear Water State established given by extended development of stands of submerged macrophytes, greater water transparency and minor nutrient concentrations. Finally, from March to May, given the submerged macrophyte withdrawal to shallowest sectors it can be interpreted that the system oscillated around a clear water state, or else, taking into account changes in the planktonic community, it is suggested that a third different state established, the “mixed phytoplankton” state. It is also suggested that states developed in Lacombe could be alternative. Next, the structure and dynamics of phytoperiphyton on bulrush and submerged macrophytes was analyzed. Vertical spatial and temporal variations of bulrush epiphyton showed to be more important than differences between sites. These variations involved smaller biomass values and a simpler physiognomy during turbid water phase and along the vertical gradient in all sampling occasions. In October, related to better light underwater conditions and the nutrient input, epiphytic biomass increased, principally promoted by species typical of the turbid phase. Nonetheless, a structural change started in this month, driven by herbivores but also by the modification of internal relationships between members of the same taxocenosis (shading and less bare substrate to be colonized). This resulted in the establishment of the characteristic assemblages of the clear water phase. During the first three months domination by stalked diatoms structured the community in a unique stratum, resistant to the less favourable light conditions and resilient to August perturbations. From December onwards Spirogyra species dominated, constituting the upper stratum, followed by Oedogonium species in the middle stratum and diatoms in the lower one. These assemblages were stabilized by mechanisms mainly displayed by the zignematalean taxon (shading, epifitism inhibition), allowing to considerer that they were resilient to March perturbation event (de novo establishment on non previously colonized surfaces). In face of the permanence, resistance and resilience properties showed by the taxocenosis, two different states of bulrush phytoperiphyton are considered: turbid water and clear water assemblages. This temporal pattern appeared less well delimited with depth and showed up later in time. Epiphyton data suggested that both states could be alternative. Differences between sites could be observed in respect to phytoperiphyton growing on submerged macrophytes, but the same temporal pattern showed by bulrush periphyton couldn’t be detected due to a great variability in the data gathered. In the next section, the analysis of the structure and dynamics of the phytobenton was carried out. It was showed that differences between the deepest sampling points and the shallowest site were greater than temporal ones. The composition of the assemblages was scarcely variable in time. The taxocenosis was principally related to the poor light conditions recorded on the sediment surface throughout the entire sampling period, even during the clear water phase. The examination of the specific composition in function of its origin evidenced the influence that the epipelon receives from other taxocenosis, constituting a refuge site for allochthonous taxa during unfavourable situations. The scarce variability of the epipelic fraction is also showed. In this way, it is considered that the taxocenosis maintained itself in only one state, indifferent to state changes in the lake and principally resisting the darkness and bearing to live in the unstable sediments. Subsequently, first the roles of the phytoperiphyton and complementary those of phytobenton, as state change or state stability agents were evaluated. The harmful effect of the phytoperiphyton on submerged macrophytes was probably counteracted by plant ramification production and allelopathy. The evidence suggested that its role as nutrient sequestrator is of minor importance in this case, due to other factors that might have disturbed phytoplankton more intensely than competition with phytoperiphyton. Much evidence supported the role of phytobenton as a source of epiphytic and planktonic species belonging either to turbid and clear water state assemblages. The results did not support the hypothesis that phytobenton may play a role as a sediment-stabilizing factor. On the contrary, it may be perturbating agent. Next, an analysis of the contributions of each community to the total algal biomass in the context of the G&R model was performed. Comparisons with AEH results were also carried out. This leaded to the consideration of the occurrence of the following states in Lacombe Lake: a Lake State consistent with the Turbid Water phase, an Open State consistent with the Clear Water phase and local Dry States in ALr site occurred both during the turbid water phase (July) or the clear water situation (March-April). The substantial contribution of epiphyton especially, that growing on submerged macrophytes, is emphasized. The importance of the discrimination of different fractions of epipelon material according to its origin when evaluating the contributions of each community to total algal biomass is also highlighted. Finally, a new interpretation is presented based on algal taxocenosis dynamics, which incorporates elements from both, the AEH and the G&R model. In conclusion, phytoperiphyton in this pampean shallow lake is strongly affected by the factors that determine the establishment or passage from one equilibrium state to another. These assemblages respond to changes through modifications in their own internal structure. These structures gradually alter with depth, remaining these changes more fully explained by the comparison with phytobenton dynamics. Besides the possible roles that the taxocenosis might undertake as an internal mechanism in the change or stabilization of systems states in different situations other than those encountered during this study, epiphyton importance in the systems context, is principally evidenced by the great contribution that these taxocenosis make to the total algal biomass, constituting the dominant compartment during clear water states. / El rol de las comunidades relacionadas a un sustrato como componentes fundamentales dentro de un sistema acuático ha sido reconocido por varios autores. A pesar de la importancia dada al perifiton en lagos someros, su inclusión dentro de diferentes modelos ecológicos ha sido inusual. Phillips et al. (1978) son los primeros en considerar al epifiton como un mecanismo interno en el pasaje de un estado de aguas transparentes a otro más turbio. En contraste, Scheffer et al. (1993), proponen la Hipótesis de Estados de Equilibrio Alternativos (HEEA) que considera que un lago somero puede alternar entre dos estados, uno de Aguas Claras, (estabilizado por la presencia de macrófitas sumegidas) y otro de Aguas Turbias (estabilizado por la elevada biomasa de fitoplancton), no tomando en consideración a las comunidades perifiticas. Sólo el marco conceptual propuesto por Goldsborough y Robinson (1996, modelo G&R) da nuevamente relevancia al epifiton, incluyendo al epipelon, al metafiton y al fitoplancton dentro de un modelo ecológico, discriminando cuatro estados de acuerdo a la dominancia en la biomasa algal total de alguna de estas taxocenosis. En las lagunas pampeanas se han realizado algunos análisis de la dinámica de los sistemas en el marco de la HEEA. Por otro lado, el conocimiento acerca de las comunidades perifíticas y bentónicas todavía es escaso. Se propone como hipótesis que el fitoperifiton variará de acuerdo a la ocurrencia de los diferentes estados en la laguna (según HEEA), interrelacionándose al mismo tiempo con el fitobenton y el fitoplancton. Por otro lado, también se considera la posibilidad de que tanto los ensambles fitoperifíticos como fitobentónicos sean mecanismos internos en el pasaje o en la estabilización de un estado. Por último, se valora la importancia del fitoperifiton en cuanto a sus aportes a la biomasa algal total en el marco del modelo G&R, realizando comparaciones con los resultados obtenidos en el caso de la HEEA. Los objetivos generales de esta investigación son: obtener información acerca de la dinámica y estructura del fitoperifiton y su relación con factores ambientales así como, también, acerca de sus interrelaciones con el fitobenton y el fitoplancton en una laguna pampeana; y analizar su dinámica en el marco de la HEEA y del modelo G&R valorando los aportes del fitoperifiton a la biomasa algal total. El estudio se realizó en la laguna Lacombe comprendida en el Grupo Marginal de lagunas en la región pericostera de la Provincia de Buenos Aires. Los muestreos se realizaron desde julio 2001 a junio 2002 en cuatro sitios de acuerdo a su grado de exposición a las variables ambientales y la presencia de macrófitas: Juncales (JP y JC); sitios de Aguas Libres con o sin macrófitas sumergidas durante la época de crecimiento (ALe y ALr). Los sitios J y ALe representan, puntos de mayor profundidad y ALr un sitio cercano a la orilla. Siguiendo un perfil vertical (con excepción del sitio cercano a la orilla, ALr) se midieron variables ambientales in situ y se tomaron muestras de agua para realizar análisis químicos del agua y la identificación y la cuantificación del plancton. En el caso del fitoperifiton se tomaron muestras de epifitos sobre junco (epifiton) también en un perfil vertical y en los 25 cm apicales del tallo de plantas sumergidas. El benton (epipelon) fue muestreado con un corer, extrayendo muestras también para realizar análisis químicos de los sedimentos. En el campo también se evaluaron las dinámicas de los sustratos vegetales. En el laboratorio se midieron las concentraciones de nutrientes algales, de los ocho iones principales, de los polifenoles solubles así como también la reserva alcalina y la dureza total. Las muestras de fitoplancton, epifiton y epipelon fueron tratadas para la estimación de clorofila a activa y feopigmentos, y la identificación y cuantificación de organismos según métodos estandarizados y propios. En principio, se analizaron las variables bióticas y abióticas para determinar la ocurrencia de estados de equilibrio en la laguna. Tres eventos principales de perturbación ocurrieron durante el período de estudio: las precipitaciones en agosto, octubre y marzo. Todas ellas involucraron aumentos del nivel hidrométrico y disminuciones en la conductividad. En los dos primeros, especialmente en octubre, se produjo la entrada de nutrientes por escorrentía, en tanto que en marzo el aporte de agua resultó en la dilución de los mismos. Por otro lado, las condiciones lumínicas mejoraron especialmente a partir de octubre, lo que se relacionó con el desarrollo de manchones de macrófitas sumergidas y una menor densidad y composición diferente del fitoplancton. Esto último se produjo por los efectos de dilución de la entrada de agua, el sombreado de las plantas acuáticas y por el pastoreo de cladóceros. El evento de marzo involucró también un mejoramiento de las condiciones de luz en la columna de agua pero, paralelamente, determinó la retracción de los manchones de macrófitas sumergidas hasta zonas más cercanas a la orilla. En el fitoplancton se operaron cambios en la composición específica. La evaluación estadística de estos hechos junto con el análisis de las características de permanencia, resiliencia y resistencia de las taxocenosis dominantes condujeron a interpretar que durante el período de estudio se establecen dos estados de equilibrio según la HEEA. Durante los tres primeros meses se desarrolló un estado de aguas turbias definido por la dominancia del fitoplancton con el predominio de una oscilatorial, menor transparencia y concentraciones de nutrientes relativamente más elevadas. En octubre y noviembre se produjo el pasaje entre estados, considerándose a estos meses como transicionales. De diciembre hasta el final del estudio, se desarrolló un estado de aguas claras definido por la presencia de manchones de macrófitas, mayor transparencia del agua y menores concentraciones de nutrientes. Finalmente, de marzo hasta mayo, dada la retracción de los manchones de macrófitas sumergidas, se puede interpretar que ocurrió una oscilación del sistema alrededor de un estado de aguas claras o, dados los cambios operados en el plancton, un tercer estado “mixed phytoplankton”. Se evidenció que los estados establecidos podrían ser alternativos. Seguidamente se analizaron la estructura y dinámica del fitoperifiton sobre junco y las macrófitas sumergidas. Las variaciones espaciales verticales y temporales en la dinámica del perifiton sobre junco resultaron más marcadas que las espaciales horizontales (entre sitios). Estas variaciones implicaron una menor biomasa y una fisonomía más simple durante la fase de aguas turbias y a lo largo del perfil vertical en todas las fechas. Relacionado con el incremento de la luz en la columna de agua y por la entrada de nutrientes en octubre, la biomasa epifítica aumentó, impulsada por las especies características de la fase turbia. Sin embargo, a partir de ese mes se dió un cambio estructural, generado por el pastoreo pero, también, por la modificación en las relaciones entre los componentes de la taxocenosis (autosombreado y menor disponibilidad de sustrato libre) que desembocó en el establecimiento de los ensambles característicos de la fase clara. Durante los tres primeros meses el predominio de diatomeas pedunculadas conformó una comunidad uniestrato, resistente a una menor disponibilidad de la luz y resiliente frente a las perturbaciones de agosto. De diciembre a febrero se desarrolló un ensamble donde predominan las especies de Spirogyra formando un estrato superior, las de Oedogonium en un estrato medio y las diatomeas en un estrato inferior. Estos ensambles presentaron mecanismos de estabilización dados por los talos de la zignematal (sombreado, inhibición del crecimiento de epifitos secundarios), considerándose resilientes frente a las perturbaciones de marzo (desarrollo de los ensambles en sitios previamente no colonizados). En función de las características de permanencia, resistencia y resiliencia, se diferenciaron dos estados en el fitoperifiton de junco: ensambles de fase turbia y ensambles de fase clara. Este patrón temporal se hizo menos marcado en profundidad y aparece en forma retrasada en el tiempo. Los datos habrían indicado la posibilidad de que ambos estados fuesen alternativos. El fitoperifiton sobre macrófitas sumergidas presentó variaciones definidas en el sentido espacial horizontal (mayor biomasa en sitios más profundos) pero no se detectó un patrón temporal, similar al del perifiton sobre junco, dada la gran variabilidad de los datos obtenidos. En el apartado siguiente se analizaron la estructura y dinámica del fitobenton. Se estableció que las variaciones entre sitios de muestreo fueron más marcadas que las temporales. Los ensambles epipélicos presentaron en general una composición escasamente variable a lo largo del tiempo relacionándose principalmente con la disponibilidad de luz. Ésta fue escasa durante todo el período de estudio por lo que las condiciones de crecimiento para las algas béntonicas no fueron favorables, aún con los cambios de estado en la laguna. El análisis del material en función de su origen evidenció las influencias que esta comunidad recibe desde otros ensambles, siendo un sitio de refugio para organismos alóctonos durante épocas desfavorables. También se evidenció la escasa variabilidad de la fracción epipélica propiamente dicha durante todo el período de estudio. Así, se considera que esta taxocenosis se mantuvo en un solo estado, ajena al pasaje de estado en la laguna, principalmente resistiendo la falta de luz y la inestabilidad de los sedimentos. Posteriormente, se evaluó primeramente el papel del fitoperifiton y, en forma complementaria, el del fitobenton como impulsores del pasaje de estados o como estabilizadores de los mismos en la laguna. El efecto de sombreado del fitoperifiton sobre macrófitas sumergidas probablemente fue contrarrestado por la producción de nuevas ramificaciones de las macrófitas y por alelopatía. Las evidencias señalaron que su rol como secuestradores de nutrientes disponibles para el fitoplancton fue menor en este caso, dado que éste último se halla perturbado por otros factores más que por la competencia con el epifiton. Las evidencias señalaron al epipelon como simiente de especies del epifiton y del fitoplancton tanto pertenecientes a la fase turbia como a la clara. En cambio, los resultados no avalaron su rol como estabilizador de los sedimentos. Al contrario, podría ser una perturbación de éstos. A continuación, se examinaron las contribuciones de las taxocenosis a la biomasa algal total en el marco del modelo G&R. También se realizó la comparación de los resultados con aquellos obtenidos a partir de la HEEA. Se consideró la ocurrencia de un estado Lake State congruente con la fase turbia en la laguna, un estado Open State congruente con la fase clara y estados locales Dry State en ALr tanto en la fase turbia (julio) como en la clara (marzo-abril). Se destacó la substancial contribución del epifiton sobre macrófitas sumergidas. También, se subrayó la importancia de discriminar, en relación con el epipelon, los aportes de cada una de las taxocenosis cuando se evalúan las contribuciones a la biomasa algal total. Finalmente, se ofreció una interpretación basada sobre la dinámica de las taxocenosis algales considerando elementos de la HEEA y del modelo G&R. En conclusión, el fitoperifiton en las lagunas pampeanas fue fuertemente influido por los factores que determinan los estados de equilibrio, respondiendo al cambio de los mismos con la modificación en su estructura interna. Ésta se alteró gradualmente con la profundidad, quedando estos cambios más completamente explicados a través de la comparación con la dinámica del fitobenton. Además de los posibles roles que podría desempeñar como mecanismo interno de pasaje o como estabilizador de estados en situaciones diferentes a las halladas en Lacombe, su importancia en el contexto del sistema se manifiesta en el gran aporte que realiza a la biomasa algal total, constituyendo el compartimiento dominante en los estados de aguas claras.

Konkurrenssituationen för dagens tillverkande företag har blivit allt hårdare, nya krav på högre kvalitet och lägre produktionskostnader krävs för att överleva på marknaden. Ett företag som slutar att utvecklas kommer snart att upptäcka att de inte är tillräckligt bra, Bergman &amp; Klefsjö (2001). Detta påstående är allmänt känt, vilket gör att ständiga förbättringar blir en viktig hörnsten inom produktionsfilosofier såsom Lean Manufacturing och Toyota Production System. Ständiga förbättringar är en väldigt viktig aktivitet som ett företag inte kan lyckas med över en natt, problem med både implementation och bibehållande av ett ständigt förbättringsprogram kommer att dyka upp. Det existerar ett stort antal artiklar och böcker om hur man skall klara av dessa svårigheter och lyckas med ett förbättringsprogram, men det saknas bra modeller. Magisteruppsatsen tar upp de faktorer som är viktiga inom förbättringsarbete och en modell föreslås. Denna modell innehåller följande delar: ledningsengagemang, utbildning, design, kommunikation och feedback, mätningar och benchmarking, belöning, klara mål, samt förnyelse. En undersökning utvecklades baserad på modellen och utfördes sedan på sju företag inom Swedwood-koncernen (en av världens största möbeltillverkare) och två svenska företag; ITT Flygt AB och Autoliv Sverige AB. Syftet med undersökningen var att kartlägga hur vardera företag arbetar med de olika delarna i modellen idag; samt hitta bra sätt att arbeta på, s.k. good practice. Undersökningen var indelad i tre delar; först intervju med anställda inom ledningen, därefter en enkätundersökning med ledning respektive operatörer. Undersökningen verifierade de flesta delar av modellen och kan användas för att belysa de delar inom en organisation som behöver förbättras för att lyckas med ett förbättringsprogram. Då företagen kartlades hittades ett antal good practice: 1. Traditionell utbildningsteknik kan alterneras med spel för att öka motivation och förståelse. 2. Mätningar och benchmarking inom ständiga förbättringar skall utföras så att utvecklingen av ett förbättringsprogram kan följas. 3. Eftersom belöning är en av de viktigaste motivationsfaktorerna för operatörer är det väldigt viktigt att definiera och strukturera ett belöningssystem. Genom att lägga ner tid och arbete på att visa och informera de anställda om systemet vill man skapa en allmän kunskap inom företaget. 4. Inom ett förbättringsprogram är det väldigt viktigt med en snabb feedback. 5. De flesta av fallföretagen använder sig av ett individuellt förbättringsprogram. Teamwork rekommenderas eftersom en grupp av individer kan åstadkomma mer tillsammans än individuellt. / The competition within the manufacturing branch becomes harder and harder, and demands such as higher quality and lower production costs has to be attained to stay on the market. A company who stops becoming better, will soon end up just not being good enough, Bergman &amp; Klefsjö (2001). This is a statement commonly known and therefore Continuous Improvement (CI) is one of the cornerstones within production philosophies such as Lean Manufacturing and Toyota Production System. CI is a very important activity but will not be something that a company acquires over a night, and there will be problems both with the implementation and the long-term sustainability of the programme. There exists a large number of articles and books of how to overcome these difficulties and draw up a successful Continuous Improvement Programme (CIP), but there is a lack of good models. In this thesis the factors that are important for CI work are emphasized and a model is suggested. The model includes the following constituents; Management commitment, education, design, communication and feedback, measurements and benchmarking, recognition, clear goals, and regeneration. A survey based on the model was constructed and performed at seven companies within the Swedwood group (one of the world’s biggest furniture manufacturers) and two Swedish companies; ITT Flygt AB and Autoliv Sverige AB. The intention was to map the case companies’ present work within the constituents of the model and to find good practice. The survey was divided into three parts; first interviews with the management and then questionnaires to both management and operators. The survey verified many parts of the suggested model, and can be used with the intention to highlight which area of the organisation that needs to be improved in order to run a successful CIP. During the mapping of the case companies some good practice was found: 1. Traditional education technique can be mixed with games to increase motivation and understanding. 2. In order to follow the progress with a CIP, CI has to be measured and benchmarked. 3. Recognition is one of the most important motivation factors for operators. Therefore it is important to define and structure the recognition system and make a real effort to show and inform employees about the recognition given, so that this will become common knowledge. 4. It is of great importance that there is a rapid feedback within a CIP. 5. Most of the case companies have implemented an individual CIP, teamwork is advocated as a group of individuals can achieve more together than working separately.

The population dynamics and demography of Daphnia galeata was analysed in a five year study in the biomanipulated Bautzen Reservoir. Samples were taken two times a week during the period May-July in the pelagic zone of this highly eutrophic water. Major bottom-up and top-down factors were determined during the study period and analysed with regard to their influence on Daphnia dynamics and life history. Field data on fecundity and population structure of D. galeata were combined with results from life table and growth experiments performed under approximately in situ conditions to gain insight into the mechanisms leading to a midsummer decline of this cladoceran species which dominates the zooplankton community in Bautzen Reservoir. Two main patterns of Daphnia dynamics emerged: In years without a midsummer decline the population increased slowly in spring, starting from low densities. High water transparency was observed already during the build-up of the population of D. galeata. Despite considerable fluctuations, Daphnia abundance remained on a high level throughout summer. In years with a midsummer decline the population started from relatively high densities in early May and more than doubled during one week. Peak densities were reached before the clear-water stage emerged. At the end of this period the population declined to low values which lasted for the rest of the summer. Fecundity of the Population of D. galeata declined, whereas the mean egg volume increased at the beginning of the clear-water stage as a result of declining food levels. The size at maturity (SAM) remained high during this period. Additionally, juvenile growth was reduced and the age at maturity was retarded. Survival probability was low for those daphnids born shortly before or during the clear-water stage compared to those born later. It can be concluded from these results that recruitment to adult stages is strongly reduced during the clear-water stage. The end of this period is marked by an alternation in generations. Only at that time can SAM be reduced because the new generation of adults matures at a smaller size, carrying small eggs. A high impact of non-predatory adult mortality can be expected when the population is dominated by a strong peak-cohort during the clear-water stage according to recruitment patterns during the build-up of the population. The most drastic decline both of Daphnia abundance and SAM was observed in those years when the biomass of juvenile fish exceeded 20 kg ha-1 at the end of the clear-water stage. Due to gape-size limitation juvenile fish mainly feed on juvenile daphnids during this period and thus, they reinforce bottom-up effects on the Daphnia population. When fish change their size selection towards adult daphnids at the time when the new generation takes over, this seems to represent the worst case for the Daphnia population. Consequently, the timing between bottom-up effects and the feeding pressure of juvenile fish determines the extent of the decline. - (This manuscript is also available - in the form of a book - from Shaker Verlag GmbH, Postfach 101818, 52018 Aachen, Germany world-wide web address: http://www.shaker.de, electronic-mail address: info@shaker.de. It has been posted on the web sites of Dresden University of Technology with the permission of the publisher)

HYAL-1 (hyaluronidase-1) appartient à la famille des hyaluronidases connues pour leur rôle dans la dégradation de l’acide hyaluronique. L’expression de HYAL-1 est élevée dans de nombreux type de cancers, notamment dans le cancer de la prostate, de la vessie, des reins et du sein où il est impliqué dans la croissance tumorale et les métastases. Récemment notre laboratoire a aussi démontré une expression élevée de HYAL-1 dans le cancer épithélial de l’ovaire (CEO) de type mucineux et à cellules claires, expression qui est inversement corrélée à celle du récepteur de l’oestrogène alpha (REα). Cependant, malgré le fait que le rôle de HYAL-1 dans le cancer soit bien établit, le mécanisme de sa régulation reste encore inconnu. Le REα est un facteur de transcription qui suite à sa liaison avec son ligand va réguler l’expression de plusieurs gènes. Le REα ainsi stimulé par l’hormone va activer la transcription de ces gènes cibles mais il est connu maintenant qu’une grande partie des gènes régulés par le REα sont en réalité réprimés par ce récepteur. Dans ce travail nous proposons d’étudier le mécanisme de la régulation du gène HYAL-1 par le REα dans le CEO à cellules claires et dans le cancer du sein. L’expression ectopique du REα dans la lignée TOV21G (RE-) de même que le traitement de la lignée MCF-7 (RE+) avec de l’oestrogène a induit une diminution du niveau d’expression de l’ARN m de HYAL-1. Ces résultats nous ont permis de confirmer que HYAL-1 est un gène cible du REα. Il est aussi connu que le REα peut exercer son action par différents mécanismes d’action, entre autres en interagissant avec une séquence d’ADN appelée élément de réponse à l’oestrogène (ERE), retrouvé sur le promoteur des gènes cibles ou bien indirectement par des interactions protéine-protéine en se liant à d’autres facteur de transcription tels que Sp1. Après avoir identifiés de telles séquences sur le promoteur proximal de HYAL-1, (1 ERE proximal à -900 pb, 3 distaux à -32350 pb, 48430, -50130 pb du site d’initiation de la transcription) en plus des 2 Sp1 connus (-60 et – 1020pb), nous avons démontrés par immunoprécipitation de la chromatine que le REα est recruté sur le promoteur de HYAL-1 au niveau de l’ERE proximal -900 pb et du distal -32350 pb de même que sur le site Sp1 -1020 pb. De plus, l’activité biologique de l’ERE -900 pb et du ii Sp1-1020pb à été confirmée par des essais de gènes rapporteurs à la luciférase. Avec son rôle connu dans la tumorigenèse, l’identification de HYAL-1 comme gène cible du REα pourrait être une avenue intéressante pour le traitement des cancers hormono-indépendants. / HYAL-1 (hyaluronidase-1) belongs to the hyaluronidase family of enzymes that degrade hyaluronic acid. HYAL-1 expression is elevated in many types of cancers including prostate, bladder, liver and breast cancer where it is involved in tumor growth and metastasis. In accordance to these observations, our group has also demonstrated high expression of HYAL-1 in clear cell and mucinous epithelial ovarian cancer (EOC) subtypes which was inversely correlated to that of estrogen receptor alpha (ERα). However, despite the fact that the role of HYAL-1 in cancer is well established, the mechanism of its regulation is still unknown. ERα is a transcriptional factor that regulates target-gene expression following ligand binding. Upon hormone stimulation, activated ERα will upregulate transcription of many target genes. However, it has been recently well documented that a large number of ERα responsive genes are in fact repressed. In this work we propose to study the mechanism by which ERα regulates HYAL-1 expression in clear cell EOC subtype as well as in breast cancer. The ectopic expression of ERα in TOV21G cell line (ERα -) and estrogen treatment of MCF-7 cells (ERα +) decreased HYAL-1 mRNA expression and allowed us to confirm that HYAL-1 is an ERα target gene. It is also known that ERα may exert its action through different mechanisms of action including interacting with a DNA sequence called estrogen response element (ERE) found in the promoter of target genes or indirectly by protein-protein interactions by binding to other transcription factor such as Sp1. Having identified such sequences in the proximal promoter of HYAL-1, (one proximal ERE -900 bp, 3 distals at -32350, -48430, -50130 bp from the start site of transcription) in addition to the two known Sp1 (-60 and -1020pb), we have demonstrated by chromatin immunoprecipitation that ERα is recruited at the HYAL-1 promoter at the ERE sites -900 pb and -32350 pb as well as at the Sp1 site -1020. Furthermore, the biological activity of the proximal ERE -900 and Sp1 -1020 sites were further confirmed by luciferase reporter gene assay. Given its known role in tumorigenesis, identification of HYAL-1 as an ERα target may provide an interesting approach for the treatment of hormono-independent cancer.

Die ekstrinsieke getuienis-reël word in die algemeen beskou as een van die mees problematiese areas van die Suid-Afrikaanse bewysreg. Die Suid-Afrikaanse Regskommissie het dan ook redelik onlangs ondersoek ingestel na die wenslikheid van die behoud of afskaffing van die reël. Ondersoek word ingestel na die ware aard van die ekstrinsieke getuienis-reël en of die reël op ’n korrekte wyse in die Suid-Afrikaanse reg ingevoer is. ’n Deeglike kritiese ontleding word verder gedoen van die twee komponente waaruit die ekstrinsieke getuienis-reël bestaan, naamlik die integrasiereël en die interpretasiereël, met behoorlike verwysing na relevante regspraak en die standpunte van akademiese skrywers op hierdie gebied. Kritiese ondersoek word verder ingestel na die verslag en aanbevelings van die Suid-Afrikaanse Regskommissie ten aansien van die ekstrinsieke getuienis-reël. Die impak van die 1996 Grondwet, asook die moderne ontwikkeling van die rekenaar en elektroniese dokumente op die ekstrinsiekegetuienis-reël word ook kortliks ondersoek. Daar word verder grondig ondersoek ingestel na die ekstrinsieke getuienis-reël in ’n regsvergelykende perspektief met spesifieke verwysing na die Engelse, Amerikaanse, Kanadese, Nederlandse, Duitse, Franse en Switserse reg, asook die moontlike waarde wat sodanige benaderings kan toevoeg tot die reël in ’n Suid-afrikaanse konteks. Die gevolgtrekking waartoe gekom word in hierdie proefskrif is dat die ekstrinsieke getuienis-reël aansienlik gemodifiseer moet word in die Suid-Afrikaanse reg deur middel van die promulgering van spesifieke wetgewing op hierdie gebied. Die essensie van hierdie wetgewing moet daaruit bestaan dat alle relevante ekstrinsieke getuienis toegelaat behoort te word ten einde die ware bedoeling van die kontrakspartye vas te stel, behalwe waar dit blyk dat hulle ooreengekom het om hulle algehele ooreenkoms in die skriftelike kontrak te integreer. Daar behoort egter terselftertyd ’n aansienlike bewyswaarde aan die skriftelike dokument, asook die terme daarin vervat, geheg te word ten einde die onskendbaarheid van die skriftelike dokument te preserveer. / Criminal and Procedural Law / LL.D.

A Síndrome de von Hippel-Lindau é uma doença hereditária multissistêmica, causada por mutações germinativas no gene VHL que predispõe o portador a manifestações benignas e malignas em diversos órgãos. Entre esses eventos, o carcinoma de células claras renais (CRC) é o de pior prognóstico, com uma penetração média de 25% e sendo a principal causa de morte nestes pacientes. Os CRCs são tumores agressivos, pouco responsivos à quimioterapia e imunoterapia, e muitas vezes são diagnosticados em estágios avançados. Podem estar associados a síndromes hereditárias como o VHL ou apresentar a forma esporádica. Caracteristicamente, o CRC é provocado pela inativação dos dois alelos do gene VHL. Nos casos associados ao VHL, um alelo do gene VHL sofre uma mutação germinativa e um segundo evento mutacional somático nas células do tumor. Por outro lado, na forma esporádica, o CRC é resultado de dois eventos somáticos adquiridos, que incluem uma combinação de metilação do promotor, mutações pontuais que afetam a sequência de leitura aberta (ORF) e rearranjos cromossômicos, principalmente perda de heterozigosidade (LOH). Embora os eventos somáticos nos CRCs esporádicos já tenham sido explorados em outros estudos, os mecanismos de inativação somáticos do gene VHL nos CRCs associados à síndrome ainda não foram bem descritos. Este estudo avaliou os eventos somáticos no gene VHL em CRCs retirados em procedimentos cirúrgicos de pacientes portadores da síndrome. Os eventos somáticos em vários tumores de um mesmo paciente foram comparados a fim de verificarmos se essas mutações são independentes e não clonais. Oito pacientes com amostras CRCs previamente armazenadas no BNT tiveram sua mutação germinativa no gene VHL caracterizada por sequenciamento ou MLPA. Todas as amostras foram submetidas a uma revisão da patologia e macrodissecadas sempre que necessário. Para a análise das manifestações somáticas do gene VHL, o DNA foi extraído de 30 CRCs conservados em RNA latter ou formaldeído (parafina). As amostras foram analisadas quanto à metilação da região promotora do gene pelo método MS-PCR e para mutações pontuais por sequenciamento. Fomos capazes de detectar a mutação somática em 25 dos 30 tumores, incluindo uma mutação pontual e dois tumores diferentes de um mesmo paciente, nenhuma microdeleção e 23 grandes deleções. Em contraste com a literatura, nenhum dos tumores apresentou metilação no promotor do VHL. Devido ao grande número de achados LOH e da resolução limitada da técnica de MLPA para avaliar a extensão dos rearranjos cromossômicos em 3p, não foi possível concluir a análise de clonalidade dos tumores. Um estudo exploratório para caracterizar ganhos e perdas genômicas utilizando a técnica CNV array está em andamento em nosso laboratório. / The von Hippel-Lindau syndrome (VHL) is a multissystemic hereditary disease, caused by germline mutations in the VHL gene that predisposes the carrier to benign and malignant manifestations in different organs. Among these events, the clear cell renal carcinoma (RCC) is the most fearful, with an average penetration of 25% being the leading cause of death in these patients. RCCs are aggressive tumors, poorly responsive to chemo- and immunotherapy that are often diagnosed in advanced stages. They can be associated with hereditary syndromes such as VHL or present in a sporadic form. Characteristically, RCCs carrier the inactivation of the two alleles of VHL gene. In cases associated with VHL, one allele of the VHL gene is mutated in the germline, and the second mutational event occurs in the somatic cells of the tumor. On the other hand, in the sporadic form, RCCs results of two acquired somatic events, which includes a combination of methylation of the promoter, point mutations affecting the ORF, and rearrangements mainly loss of heterozigosity (LOH). Although somatic events in sporadic RCC have been explored before by others, the mechanisms of somatic VHL gene inactivation in VHL-associated RCCs have been poorly characterized. This study evaluated the somatic mutational events in the VHL gene of RCCs removed from VHL patients in therapeutic surgical procedures. The somatic events in multiple tumors from the same patient were compared in order to analyze whether these mutations are independent and not clonal. Eight patients with RCCs samples previously stored at BNT had their germline VHL gene mutation characterized by sequencing or MLPA. All samples were submitted to a pathology review and macrodissected whenever necessary. For the analysis of somatic events of VHL gene, DNA from 30 RCCs were extracted from either RNA later or archival formalin-fixed, paraffin-embedded tissue sections. Samples were analyzed for VHL gene promoter methylation by MS-PCR, and for point mutation in the coding DNA by sequencing. We were able to detect the somatic mutation in 25 of the 30 tumors, including one point mutations in two different tumors of the same patient, no micro-deletions, and 23 large deletions. In contrast to the literature, none of the tumors have shown methylation on the VHL promoter. Because of the large number of LOH findings, and the limited resolution of MLPA to evaluate the extension of 3p chromosomal rearrangements, we could not conclude the analysis of tumor clonality. An exploratory study to characterize genomic gains and losses using CNV-array technique are ongoing in our laboratory.