Issu de remaniements chromosomiques, le gène de fusion TMPRSS2 :ERG a été identifié dans plus de 50% des cas des cancers de la prostate (CaP). Cette découverte a ouvert une nouvelle voie dans la compréhension du processus de cancérisation. De nombreuses études cliniques démontrent l’association de la présence de ce gène de fusion avec un mauvais pronostic. Erg (Ets-Related-Gene) est un facteur de transcription dont l’expression est associée, en particulier, à la mise en place du cartilage et plus largement du squelette. Une étude transcriptomique, comparant les transcriptomes des chondrocytes embryonnaires de souris sauvages et de souris transgéniques qui expriment une protéine ERG tronquée, à l’effet trans-dominant négatif, nous a permis d’identifier de nombreux marqueurs osseux et gènes associés à la métastase osseuse comme gènes cibles de ce facteur de transcription. Les CaP, lorsqu’ils évoluent, provoquent dans la plupart des cas des métastases osseuses. Ces résultats suggèrent un rôle potentiel du gène de fusion dans la formation de métastases osseuses du CaP.Pour notre étude, nous avons établi des lignées de cellules cancéreuses prostatiques PC3c qui expriment stablement le gène de fusion TMPRSS2 :ERG. Dans un premier temps, nous avons démontré que l’expression ectopique du gène de fusion renforce la migration et l’invasion des cellules PC3c de manière dose-dépendante. Une étude transcriptomique a révélé que l’expression de certains gènes associés à la migration et l’invasion est dérégulée suite à l’expression du gène de fusion. Grâce à cette étude, de nouveaux gènes cibles de TMPRSS2 :ERG ont été mis en évidence dans les CaPs. L’identification de ces nouveaux gènes cibles renforce l’hypothèse d’un rôle du gène de fusion dans les métastases du cancer de la prostate. Par ailleurs, ces lignées de cellules cancéreuses prostatiques qui expriment stablement TMPRSS2 :ERG ont été injectées dans les tibias des souris SCID pour établir un modèle de métastases osseuses induites. Nous avons démontré que les cellules PC3c exprimant stablement le gène de fusion sont capables d’induire des lésions ostéocondensantes, tandis que les cellules témoins induisent des lésions mixtes (ostéolytiques et ostéocondensantes). Ce modèle nous a permis, pour la première fois, d’associer ce gène de fusion avec la formation des métastases osseuses du CaP in vivo.L’ensemble des résultats contribue à comprendre l’influence qualitative et quantitative de l’expression de TMPRSS2 :ERG dans les métastases du CaP. / Prostate cancer (PCa) is one of the most common malignancies that affect men in western countries. Recurrent gene fusion, involving the ERG gene and the androgen-regulated TMPRSS2 gene promoter, occurs in over 50% of PCa. The TMPRSS2:ERG gene fusion results in aberrant ERG transcription factor expression in PCa. We and others have shown that the ERG transcription factor, a member of ETS family, is associated with embryonic skeleton development. Interestingly, some of the potential ERG-target genes, identified using high-throughput DNA microarray analysis, have implications in the physiological bone homeostasis or the pathological bone metastases development. Bone metastases are frequent and represent severe complications of PCa. This suggests a potential role of the TMPRSS2:ERG in PCa bone metastases.In this study, we used PC3c cells line (derived from PC3 cell line) to establish TMPRSS2:ERG-expressing clones cells. The ectopic expression of the fusion resulted in significant induction of cell migration and invasion in a dose-dependent manner. In agreement with this phenotype, high-throughput microarray analysis revealed that a set of genes, functionally associated with cell motility and invasiveness, were deregulated in a dose-dependent manner in TMPRSS2:ERG-expressing cells. Importantly, further analyses of these deregulated genes revealed that some of them are direct target genes of TMPRSS2:ERG in PCa. These results provide novel insights into the role of the TMPRSS2:ERG fusion in PCa metastasis. To test the hypothesis of the implication of ERG in bone metastases development, we used an experimental bone metastases induction model. The implantation of the TMPRSS2:ERG-expression PC3c cells in the tibia of the SCID mice induced osteoblastic bone lesions, whereas the control PC3c resulted in mixed bone lesions. Our results show, for the first time, the possible implication of TMPRSS2:ERG in CaP bone metastasis formation.To summarize, this study provided novel evidences of the role of TMPRSS2:ERG fusion in PCa metastasis.

Yes / Purpose: We introduce a method for determining the impulse response function (IRF) of the ERG derived from responses to temporal white noise (TWN) stimuli. Methods: This white noise ERG (wnERG) was recorded in participants with normal trichromatic vision to full-field (Ganzfeld) and 39.38 diameter focal stimuli at mesopic and photopic mean luminances and at different TWN contrasts. The IRF was obtained by cross-correlating the TWN stimulus with the wnERG. Results: We show that wnERG recordings are highly repeatable, with good signal-tonoise ratio, and do not lead to blink artifacts. The wnERG resembles a flash ERG waveform with an initial negativity (N1) followed by a positivity (P1), with amplitudes that are linearly related to stimulus contrast. These N1 and N1-P1 components showed commonalties in implicit times with the a- and b-waves of flash ERGs. There was a clear transition from rod- to cone-driven wnERGs at ~1 photopic cd.m 2. We infer that oscillatory potentials found with the flash ERG, but not the wnERG, may reflect retinal nonlinearities due to the compression of energy into a short time period during a stimulus flash. Conclusion: The wnERG provides a new approach to study the physiology of the retina using a stimulation method with adaptation and contrast conditions similar to natural scenes to allow for independent variation of stimulus strength and mean luminance, which is not possible with the conventional flash ERG. Translational Relevance: The white noise ERG methodology will be of benefit for clinical studies and animal models in the evaluation of hypotheses related to cellular redundancy to understand the effects of disease on specific visual pathways.

Preterm birth may lead to complications during the neonatal period that can cause visual dysfunctions. Retinopathy of prematurity (ROP) and neurological complications are well known reasons for visual dysfunctions, but preterm children with no or only mild ROP and no evident neurological problems may also be affected visually when they grow up. Retinal development starts early after gestation and continues long after birth. Major processes are underway during the second half of pregnancy when preterm children are born, and a preterm birth could possibly have a negative effect on normal retinal development. The aims of the studies were to evaluate retinal morphology and function in former preterm children and compare the results with children born at term. Former preterm children aged 5 to 17 years and born in a gestational age (GA) of 32 weeks or less were included in the different study groups. Children of similar ages who were born at term and with normal visual acuity (VA) acted as controls. Best corrected VA and refraction in cycloplegia were assessed in all children. Macular thickness and retinal nerve fiber layer (RNFL) thickness were measured with optical coherent tomography (OCT). Total retinal function was assessed with fullfield electroretinography (ffERG) and central macular function was assessed with multifocal electroretinography (mfERG). Preterm children had thicker central maculae than controls. There was a positive correlation between central macular thickness and GA at birth. RNFL thickness was reduced in the preterm children with severe ROP and treated ROP, but children with mild or no ROP did not differ from the fullterm children. The photoreceptor function measured with ffERG and the macular function measured with mfERG were reduced in the preterm group compared to controls. Preterm birth affects the retina both morphologically and functionally, and ROP has been suggested to be a reason for retinal changes. However, the results of this thesis indicate that children with no ROP also have retinal changes, suggesting an effect of prematurity itself. There were no correlations between any retinal changes and VA, but it is possible that larger studies using improved techniques may elucidate this further.

Einleitung: Steady-State- Muster-ERGs (PERGs) werden in der Glaukomfrühdiagnostik angewandt. Da die transienten PERG-Komponenten N35, P50 und N95 unter schnellen Reizbedingungen nicht zu erkennen sind, können bisher keine Aussagen über die Beteiligung einzelner Retinaschichten an der Reizantwort gemacht werden. Mit der Deconvolution-Methode (CLAD) wurden 2004 akustisch evozierte transiente Antworten aus hochfrequenten Messungen entfaltet. In dieser Arbeit wird CLAD beim PERG validiert um herauszufinden, ob bei Messungen mit hohen Reizfrequenzen deutliche transiente Antworten entfaltet werden. Methodik: Es wurden 3 Reizbedingungen mit nicht-isochronen Stimuli der mittleren Frequenz 16,7 rps verglichen. Der Abstand zwischen 2 Stimuli variierte zwischen 15 und 105 ms, 30 und 90 ms oder 45 und 75 ms, der mittlere Abstand betrug jedoch immer 60 ms. Als Referenz wurden konventionelle transiente (1,4 rps) und Steady-State-Messungen (16,7 rps) durchgeführt. Um sie zu validieren, wurden in einem zweiten Schritt aus den transienten CLAD-Steady-State-Antworten synthetisiert und diese mit der Steady-State-Referenzmessung verglichen. Ergebnisse: Die große Übereinstimmung der synthetisierten Steady-State-Antworten mit der Referenzmessung im Bezug auf Amplitude und –Kurvenform bestätigte, dass die entfalteten transienten Antworten „trotz“ Abweichungen von konventionellen PERG-Antworten plausibel sind. Allerdings hatte der konkrete Stimulusabstand Einfluss auf die PERG-Antwort: die Amplituden der 3 CLAD-Reizbedingungen wichen signifikant voneinander ab. Diskussion: Diese Studie zeigte, dass durch CLAD transiente Reizantworten aus Messungen hoher Frequenz gewonnen werden können. Ob der Einsatz von CLAD Vorteile in der Glaukomfrühdiagnostik erbringt und der exakte retinale Ursprung der Antwortkomponenten ist in künftigen Studien zu validieren. Durch CLAD könnte es in Zukunft möglich werden, auf elektrophysiologischem Wege den Beitrag einzelner Retinaschichten bei Retinapathologien genauer aufzuzeigen.

Yes / The purpose of this paper is to demonstrate: 1) that silent substitution stimuli can be used generate electro-retinograms (ERGs) that effectively isolate rod photoreceptor function in humans without the need for dark adaptation and 2) that this approach constitutes a viable alternative to current clinical standard testing protocols. Rod-isolating and non-isolating sinusoidal flicker stimuli were generated on a 4 primary LED ganzfeld stimulator to elicit ERGs from non-dark adapted participants with normal and compromised rod function. Responses were subjected to Fourier analysis and the amplitude and phase of the fundamental were used to examine temporal frequency and retinal illuminance response characteristics. ERGs elicited by rod isolating silent substitution stimuli exhibit low-pass temporal frequency response characteristics with an upper response limit of 30Hz. Responses are optimal between 5 – 8 Hz and between 10-100 photopic Td. There is a significant correlation between the response amplitudes obtained with the silent substitution method and current standard clinical protocols. Analysis of signal to noise ratios reveals significant differences between subjects with normal and compromised rod function. Silent substitution provides an effective method for the isolation of human rod photoreceptor function in subjects with normal as well as compromised rod function when stimuli are used within appropriate parameter ranges. Translational Relevance: This method of generating rod ERGs rod isolation can be achieved without time consuming periods of dark adaptation and provides improved isolation of rod- from cone-based activity and will lead to the development of faster clinical electro-physiological testing protocols with improved selectivity.

Oxidative stress has a degenerative effect on neuronal health. Mutations in the copper zinc superoxide dismutase (SOD1), an important antioxidant, have been found in patients suffering from amyotrophic lateral sclerosis (ALS). Classical EMS induced mutations to SOD1 in Drosophila show similar loss of motor coordination and shortened lifespan seen in humans. A study of newly created human ALS point mutants along with the classic alleles show similar phenotypes in their neurodegeneration. I examined markers of oxidative stress, neuronal health and behavioral phenotypes throughout the lifecycle of aging flies. Larvae were largely found to be unaffected by mutations in SOD1, with no measured increase in ROS level over wild type (WT) flies. Mutant pupae were found to have two major defects in their circadian eclosion rhythm and their fundamental ability to eclose from the pupal casing. Adults showed the classic reduced lifespan and motor abilities. To further examine the health on non-glutamatergic synapses electroretinograms (ERGs) were recorded at different levels of survivorship indicated by Kaplan-Meier Survival curves. These ERGs show that the histaminergic synapses they record have greater degeneration in aging SOD1 mutants than in WT flies. This is true for their chronological age as well as their biological age. There was coinciding disruption of the photo transduction pathway of the photoreceptors that coincided with degeneration at the synapse. This demonstrates the separate degenerative effect of high levels of oxidative stress impart separate for the normal aging process.

Sammanfattning Titel: Självständigt arbete, en studie ur fastighetsmäklarens vardag. Nivå: C-uppsats i ämnet företagsekonomi Författare: Oscar Hallbygård & Oscar Hammenstål Handledare: Jonas Kågström Datum: 2014 – Maj Syfte: Författarna till denna studie har valt att undersöka fastighetsmäklarens självständiga arbete och vilket stöd arbetstagaren får av arbetsgivaren. Förmånen att ha kontroll och kunna styra över sitt eget arbete, möjlighet till självständighet samt varierande arbetsuppgifter är viktiga parametrar för att trivas på arbetet. Uppsatsen syfte lyder: Syftet med detta examensarbete är att ur ett arbetstagarperspektiv analysera huruvida chefer i praktiken stödjer/uppmuntrar sina anställda att bli mer självgående på arbetsplatsen. Metod: Uppsatsen har tillämpat ett kvantitativt metodval med hjälp utav en enkätundersökning. Inhämtad rådata har analyserats med hjälp av statistikprogrammet SPSS och redovisas i kapitel 3, 4, 5 & 6. Resultat & slutsats: Författarna till studien vill uppmärksamma följande slutsatser. Studien visar att fastighetsmäklarens personliga utveckling påverkas av arbetsgivarens autonoma stöd. Genom denna slutsats kan vi påstå att arbetsgivarens autonoma stöd kan påverka arbetstagarens upplevda autonomi och benägenheten till att ta egna beslut på arbetsplatsen Förslag till fortsatt forskning: Författarna till studien skulle vilja se en framtida studie i en bransch som är känd för att inte förespråka självständighet i yrkesrollen och se om arbetsgivaren bidrar med stöd för självständighet. Uppsatsens bidrag: Författarna till denna studie har en förhoppning att kunna bidra med kunskap kring arbetsgivarens stöd vid självständigt arbete i fastighetsmäklarbranschen. Nyckelord: Självständig, SDT, Autonomi, Utveckling, Autonomt stöd, ERG

World-class outcrops of an outermost erg-margin can be observed within the Middle Jurassic Entrada Sandstone near Capitol Reef National Park, Utah. These erg-margin deposits contain isolated reservoir quality sandstone bodies that transition into a muddy tidal flat facies. These high quality reservoirs are dominated by eolian-influenced facies interbedded with sandy interdune facies. They are sealed vertically by muddy and silty facies of associated tidal flat deposits that act as excellent stratigraphic traps in the subsurface. A variety of approaches were used to characterize these Entrada erg-margin reservoirs including: annotated panoramas of outcrops, measured sections, scintillometer measurements of field sections, facies analysis, 2D high-resolution shallow seismic surveys, porosity and permeability analysis, and sedimentary petrography. Logs from the North Hill Creek/Flat Rock gas field were analyzed and correlated to the outcrop study. Eolian dune facies, along with an upper ripple laminated facies representing interdune deposits, display the highest porosities and permeabilities and are volumetrically the most important facies of the reservoir quality sandstones. Baffles and possible barriers within the sandstone bodies are limited to quartz filled fractures, deformation bands, silty and muddy interdune facies, and first order bounding surfaces. Many of the sandstone bodies within the outcrop belt are genetically related and in communication with each other. This relationship results from dune complex migration to the south and up section over time. Stratigraphic climb can potentially be imaged seismically and may serve as a key indicator of eolian dune complexes in the subsurface. The volumetric size of one of these complexes is estimated around 470 million cubic feet. Smaller outcrop sandstone bodies were often found to be isolated from the large dune complexes and ranged down to 1 million cubic feet in size.

Ce travail décrit un criblage systématique du système ubiquitine-protéasome (UPS) basé sur une organisation en cascade. Nous avons évalué l’effet de l’inhibition par ARN interférent de composants individuels d’UPS sur la viabilité de cellules cancéreuses de la prostate, avec un accent particulier sur les cellules TMPRSS:ERG-positive (VCaP), comme un modèle de phénotype prévalent du cancer. Sept gènes ont été identifiés comme étant particulièrement importants pour le fonctionnement des cellules cancéreuses de la prostate. Parmi eux, le gène-candidat le plus prometteur était UBE2U. Cette thèse met en évidence l’implication d’UBE2U dans la carcinogénèse de la prostate et décrit les premières caractérisations d’UBE2U comme une cible thérapeutique potentielle.La prévalence des composants de la voie CRL/NEDD8 parmi les hits (4 sur 7) suggère que la neddylation est importante dans la biologie des cellules cancéreuses de la prostate. Deux de ces gènes, CUL2 et RBX1, n’ont des effets spécifiques que dans des cellules TMPRSS2:ERG-positives, et, donc, sont potentiellement ERG-dépendantes. Nous avons également révélé un rôle crucial du facteur d’échange de CRL (CAND1), en particulier lorsque la neddylation est compromise. L’inhibition de CAND1 induit l’apoptose dans des cellules VCaP, qui est renforcé par l’inhibiteur spécifique de la neddylation MLN4924. CAND1 est donc une nouvelle cible thérapeutique potentielle. Par ailleurs, nous avons démontré que l’inhibition de la voie CRL/NEDD8 dans les cellules cancéreuses de prostate a des conséquences qui dépendent fortement du contexte cellulaire. L’inhibiteur MLN4924 induit l’apoptose dans toutes les lignées cellulaires testées, bien que les cellules TMPRSS2:ERG-positives se soient révélées significativement plus résistantes. Nous avons démontré que la résistance accrue des cellules VCaP reflète la plasticité des cellules cancéreuses régulée par un réseau d’interactions ERG:NF-kB:c-Myc:Wnt/β-cat:AR. L’inhibition partielle de la neddylation enclenche une reprogrammation transcriptionnelle de cellules VCaP, amenant à la quiescence des cellules et à l’inhibition de l’apoptose dépendant de la prolifération. Cet effet est le résultat de la réactivation du programme AR. Nous avons conclus que la voie CRL/NEDD8 régule le réseau transcriptionnel qui contrôle la plasticité des cellules cancéreuses. Ces résultats peuvent aider à trouver des traitements plus efficaces de cancers TMPRSS2:ERG-positifs.Finalement, nous avons observé que l’inhibition de la neddylation modifie les propriétés de la membrane et la morphologie des cellules VCaP. Cet effet est accompagné par des changements du taux et de la localisation de plusieurs protéines associées à la membrane, y compris l’occludine, la N-cadherine, la paxilline and FAK. Nous en avons conclus que la voie CRL/NEDD8 pourrait être impliquée dans le tri/trafic des protéines membranaires. Cette partie du projet nécessite de plus amples études, étant donné que la compréhension des mécanismes sous-jacents est importante et peut mettre à jour un nouveau rôle de la voie CRL/NEDD8 dans la régulation des fonctions cellulaires.Conclusion générales :1. Nous avons caractérisé l’implication de tous les composants E1-E2 d’UPS dans la régulation de la viabilité des cellules cancéreuses de prostate (avec cinq différentes lignées cellulaires).2. Nos travaux ont mise en évidence de nouvelles cibles thérapeutiques potentielles pour le traitement du cancer, telles qu’UBE2U et CAND1.3. Nous avons démontré le rôle de la voie CRL/NEDD8 dans la régulation de la plasticité et de la morphologie des cellules cancéreuses. / In this work we describe a systematic approach for screening of ubiquitin-proteasome system (UPS) based on cascade organization. We have evaluated the effect of RNAi knockdown of individual UPS components on viability of PCa cells with major focus on TMPRSS:ERG-positive cell line, VCaP, as a model of prevalent phenotype of prostate cancer. Seven genes have been identified to be particularly important for the functioning of PCa cells. Among them, UBE2U was the strongest hit. This thesis provides the first evidence for UBE2U involvement in prostate carcinogenesis and describes initial characterization of UBE2U as a potential drug-target.The prevalence of the components of CRL/NEDD8 pathway in the hits (four out of seven) suggested the importance of neddylation for PCa biology. Two of these hits, CUL2 and RBX1, being specific to TMPRSS2:ERG-positive cells, are potentially ERG-dependent. We have also revealed the crucial role of CRL-exchange factor CAND1, in particular, when the neddylation is compromised. Knockdown of CAND1 induces apoptosis in VCaP cells that is further potentiated by neddylation-specific inhibitor MLN4924. CAND1 is, therefore, a novel potential drug target. Furthermore, we have demonstrated that the inhibition of CRL/NEDD8 pathway in prostate cancer cells has a complex outcome that strongly depends on cellular context. MLN4924 inhibitor induced apoptosis in all tested cell lines, though TMPRSS2:ERG positive cells were significantly more resistant. We have demonstrated that the increased resistance of VCaP cells reflects the plasticity of cancer cells ensured by sophisticated interaction network ERG:NF-kB:c-Myc:Wnt/β-cat:AR. We found that partial inhibition of neddylation triggered transcriptional reprogramming of VCaP cells leading to cell quiescence and inhibition of proliferation-dependent apoptosis. This was a result of re-activation of AR program and induction of differentiation-like state. We conclude that CRL/NEDD8 pathway regulates cancer transcriptional network that underlies cancer cells plasticity. This knowledge would help to find better treatments for TMPRSS2:ERG-positive cancers.Finally, we observed that neddylation inhibition changed membrane properties and morphology of VCaP cells. This was accompanied by dose-dependent changes in the level and the localization of several membrane-associated proteins, including occludin, N-cadherin, paxillin and FAK. We thus conclude that CRL/NEDD8 pathway might be involved in sorting/trafficking of membrane proteins. This part of the work requires further investigation, as understanding of the underlying mechanisms is of general importance and may uncover a new role of CRL/NEDD8 pathway in regulation of cellular functions.General conclusions:1. We have obtained a comprehensive dataset on the involvement of all human E1-E2 UPS components in the regulation of viability of PCa cells, represented by five different cell lines.2. Our work has revealed new potential drug targets for PCa treatment: UBE2U and CAND1.3. We have demonstrated the role of CRL/NEDD8 pathway in the regulation of cancer cell plasticity and morphology.

We did not find mutation in coding areas of genes for components of TGFbeta1 signaling pathway but we detected decreased or undetectable expression of these analysed genes.The decreased expression is probably caused by epigenetic changes, so by hypermethylation and deacetylation of promoter regionsof these genes.Antiproliferative and apoptotic effect of TGF1 was analysed in AML cell lines (ML1, ML2, CTV1 and Kasumi1). ML2 cells rezistence to inhibition of DNA synthesis by TGFβ1 is not caused by mutations of genes for components of TGFβ1 signaling pathway. We found that increased SnoN (Ski-like novel gene) expression on the level of coresponding mRNA and protein is probably accountable for this rezistence. Kasumi1 and M2 cells were sensitive to induction of apoptózis caused by TGFβ1 treatment but in less extent than by proteazome inhibitor bortezomib. The difference of AML cells of different lines answers shows a great heterogeneity AML in AML patients. Prognostic factors analysis in AML with normal karyotype confirmed that CEBPA (CCAAT/enhancer binding protein alpha) mutations predict favourable prognosis but the elevated EVI1 ("Ecotropic Virus Integration Site 1") and ERG ("ETS-related gene") expression are connected with unfavourable prognosis. EVI1 is a negative marker for MDS as well. We did not confirm...