Dissertations / Theses: 'GATA-1'

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Whyatt, David John. "Erythroid development and GATA-1." Thesis, Open University, 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.239713.

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Halsey, Christina. "The role of GATA-1 isoforms in haematopoiesis." Thesis, University of Glasgow, 2009. http://theses.gla.ac.uk/870/.

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GATA-1 is a key haematopoietic transcription factor which plays a pivotal role in differentiation of the erythroid, megakaryocytic, eosinophilic, mast cell and dendritic cell lineages. Since its initial cloning and characterisation in 1989 a huge amount of information has been gathered on the molecular mechanisms of action of GATA-1. This knowledge has helped understanding of the processes by which cells enact differentiation programmes and suppress alternative lineage choices. GATA-1 produces at least two protein isoforms – the well characterised GATA-1 full-length (GATA-1FL) isoform and a truncated isoform – GATA-1 short (GATA-1s). GATA-1FL comprises two conserved Zinc fingers (which interact with DNA and essential co-factors), a C-terminal tail (of mostly unknown function) and an N-terminal domain (thought to confer activation properties to the molecule, but which may also be involved in transcriptional repression). GATA-1s lacks the N-terminal domain but is otherwise identical. The biological role of GATA-1s is unknown and this isoform received scant attention until the discovery that GATA-1FL mutations were linked to a rare, but highly informative, acute megakaryoblastic leukaemia seen in children with Down syndrome (constitutional trisomy 21). This discovery was particularly interesting, not only because the association between trisomy 21 and the X-linked GATA-1 mutation was extremely tight (being seen in 100% of the cases examined), but also because the GATA-1FL mutations were not randomly located, but rather clustered within the N-terminus, allowing unhindered production of the GATA-1s isoform. This finding led to interest in the pathological and physiological role of GATA-1s in haematopoiesis. Some insight has been gained into the pathological role of GATA-1s by creation of a GATA-1s knock-in transgenic mouse and by experiments looking at the ability of GATA-1s to rescue GATA-1 deficient embryonic stem (ES) cell lines. GATA-1s produces hyper-proliferation of fetal liver meg-erythroid progenitors but allows at least partial differentiation of these cells. However, a number of key questions remain. In particular what is the physiological role of GATA-1s and the reason for the tight association between trisomy 21 and GATA-1s mutations? Given this background, this thesis describes experiments designed to address the physiological role of GATA-1s, to establish whether additional GATA-1 isoforms exist, and to investigate the association between GATA-1 isoform expression and trisomy 21. Firstly a comprehensive expression analysis was performed in murine and human primary tissues and cell lines. This aimed to identify whether GATA-1s had a unique expression profile, either in particular lineages, or at distinct stages of haematological ontogeny. Reverse-transcriptase polymerase chain reaction (RT-PCR) and western blot analyses showed that the expression patterns of GATA-1s and GATA-1FL were virtually identical, with the possible exception of one human primary monocytic cell preparation which appeared to preferentially express GATA-1s. Before proceeding to further analysis of GATA-1s a search was made for additional GATA-1 isoforms using in silico analysis, RT-PCR and western blotting. This led to identification of a clone carrying a GATA-1 mutation involving the C-terminal tail, derived from a patient with chronic myeloid leukaemia. An analysis of the properties of this clone was performed, confirming its altered C-terminus and demonstrating that this conferred increased transactivation properties on the molecule as measured by luciferase assays. This observation suggests that the C-terminal tail may be an important, and previously under-recognised, functional region of the GATA-1 molecule. The discovery of this potentially hyper-functioning GATA-1 mutation led to investigation of whether GATA-1 mutations could be a widespread phenomenon in CML. However, GATA-1 mutational analysis in 21 patient samples from CML blast crisis did not reveal any additional coding mutations. To address the physiological role of GATA-1s, attempts were made to perform gene targeting in murine embryonic stem cells to produce isoform specific knock-out cells i.e. ES cells engineered so that they exclusively express the GATA-1FL isoform (a GATA-1s knock-out) or the GATA-1s isoform (a GATA-1FL knockout). These cells could then be used in in vitro haematopoietic differentiation assays and for transcriptional profiling. In this way it was hoped to establish whether GATA-1s fulfilled any unique roles in primitive or definitive haematopoiesis that could not be compensated for by the presence of the GATA-1FL isoform. Unfortunately, despite evidence of apparently successful targeting from PCR screening of ES cell clones, it was impossible to confirm the existence of endogenously targeted alleles on Southern blotting. Following exhaustive attempts at screening further clones and subclones (more than 1000 clones in total), this approach was abandoned in favour of transgenic expression of GATA-1 isoforms in cell lines. Transgenic expression studies in murine ES cells showed that whilst GATA-1FL expression led to an expansion in numbers and maturity of erythroid and non-erythroid haematopoietic colonies in vitro, GATA-1s was incapable of supporting colony formation in this assay. Studies then moved on to human cell lines. Two cell lines were identified, both capable of in vitro haematopoietic differentiation into megakaryocytic and erythroid cells, but one carrying trisomy 21 (Meg-01) and the other disomic for chromosome 21 (K562). GATA-1FL expression in these cells generally drove differentiation along the megakaryocytic or erythroid lineage as measured by DNA ploidy analysis, haemoglobinisation, upregulation of erythroid or megakaryocytic gene expression (by quantitative PCR) and suppression of alternative lineage genes (PU.1 and Ikaros) and genes associated with progenitor proliferation (cyclin D2 and c-myb). GATA-1s, in contrast, produced less evidence of differentiation with lower DNA ploidy, less up-regulation of erythroid genes and failure to repress other lineage and haematopoietic progenitor associated genes. Examination of the link with trisomy 21 confirmed that that the chromosome 21 candidate gene Erg3 was upregulated in trisomic cells and that expression of GATA-1s appeared to confer a selective advantage in the presence of trisomy 21. However, no clear mechanistic reasons for the selective advantage could be identified. Overall, these studies show widespread GATA-1s expression in haematopoietic cells, confirm the association with inadequate repression of genes associated with primitive progenitors, and suggest that the C-terminal tail of GATA-1 may be an important functional part of the molecule. Finally, these observations have generated a number of testable hypotheses which could form the basis for future work.

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Eisbacher, Michael School of Medical Science UNSW. "The regulation of megakaryocyte-specific genes by Fli-1 and GATA-1." Awarded by:University of New South Wales. School of Medical Science, 2003. http://handle.unsw.edu.au/1959.4/19171.

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The successive activation of tissue-specific genes during cellular differentiation is orchestrated by the formation of transcriptional complexes consisting of cellspecific and ubiquitous transcription factors. Understanding the molecular events associated with normal megakaryocyte (Mk) differentiation is an issue of central importance to haematology. The aims of this study were therefore to: (i) define the transcription factors responsible for regulating the expression of Mkspecific genes such as Glycoprotein IX, (ii) identify the protein partners of such important Mk-regulatory transcription factors and (iii) examine the mechanisms utilised by these factors to regulate gene expression. First, the regulatory elements in the GPIX promoter required for basal and inducible expression were examined in megakaryoblastic Dami cells stimulated to undergo differentiation. The resulting data suggested that an Ets site in the GPIX promoter binding the Ets-family member Fli-1 was crucial in regulating both constitutive and inducible GPIX expression. Second, a two-hybrid screen of a K-562 cDNA library was used to identify transcription factors that interacted with Fli-1 and were potential regulators of Mk development. Results of this screen identified a novel protein-protein interaction with GATA-1, a previously well-characterised zinc finger transcription factor also implicated in erythroid and Mk development. Mapping of the domains required for the interaction show that the zinc fingers of GATA-1 interact with the Ets domain of Fli-1. The biological significance of the Fli-1/GATA-1 interaction was demonstrated in transient transfection assays, which resulted in synergistic activation of Mkspecific promoters. Analysis of Fli-1 and GATA-1 expression in a series of erythroleukaemic and megakaryoblastic cell lines demonstrated that the Fli- 1/GATA-1 combination correlates with a Mk-phenotype. Moreover, expression of Fli-1 in K-562 cells (a line rich in GATA-1 but normally lacking Fli-1) induces endogenous GPIX expression. Quantitative mobility shift assays reveal that Fli- 1 and GATA-1 exhibit cooperative DNA-binding in which the binding of GATA-1 to DNA is increased approximately 26 fold in the presence of Fli-1. This data provides a mechanism for the observed transcriptional synergy. In conclusion, this work suggests that Fli-1 and GATA-1 work together through protein-protein interaction and cooperative DNA-binding to activate the expression of genes associated with the terminal differentiation of Mks.

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Lefevre, Carine. "Mécanismes de régulation de la balance prolifération/différenciation érythroïde par les facteurs de transcription GATA-1, FOG-1, E2F et la voie de signalisation Akt." Thesis, Paris 11, 2013. http://www.theses.fr/2013PA11T010/document.

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Avec plus de 100 milliards de globules rouges produits chaque jour, le lignage érythroïde présente la plus grande capacité de production cellulaire chez le mammifère adulte. Cette production requiert une balance fine entre la prolifération cellulaire, régulée principalement par la voie de signalisation érythropoïétine (Epo)/PI3K/Akt, et la différenciation érythroïde induite par le couple de facteurs de transcription GATA-1/FOG-1. Des interconnexions entre ces deux grands systèmes ont été décrites dans le laboratoire : 1) le facteur de transcription GATA-1 est phosphorylé par Akt en réponse à l’Epo et cette phosphorylation semble avoir un rôle dans la différenciation érythroïde ; 2) GATA-1 est capable d’interagir avec la protéine du rétinoblastome pRb, impliquée dans la régulation du cycle cellulaire, et le complexe formé est nécessaire à l’érythropoïèse terminale.L'objectif de ma thèse était d’étudier les mécanismes moléculaires impliqués dans la balance prolifération/différenciation cellulaire au cours de l’érythropoïèse, et en particulier de déterminer le rôle moléculaire et physiologique de la phosphorylation de GATA-1 par Akt en réponse à l’Epo. Nos travaux ont montré que cette phosphorylation est une des clefs de la dynamique de l’érythropoïèse. Dans sa forme non phosphorylée, GATA-1 ralentit le cycle cellulaire via le complexe GATA-1/pRb/E2F. Cette étape préliminaire est nécessaire à la mise en place de la différenciation érythroïde terminale. La phosphorylation de GATA-1 induit d’une part la dissociation de GATA-1/pRb/E2F favorisant l’expansion cellulaire, et d’autre part la formation du complexe GATA-1/FOG-1 nécessaire à l’activation des gènes érythroïdes. Ce modèle apporte une explication moléculaire au blocage de la différenciation érythroïde terminale induite par le mutant GATA-1V205G qui n’interagit pas avec FOG-1. Ainsi, la phosphorylation constitutive de GATA-1V205G et l’augmentation de la quantité relative de FOG-1 permettent de restaurer la différenciation érythroïde induite par ce mutant in vitro. Enfin, l’étude d’un modèle murin exprimant une protéine GATA-1 non phosphorylable par Akt montre l’apparition d’une anémie létale lorsque la voie IGF-1 est inhibée. Cela démontre l’importance de la dynamique moléculaire induite par la phosphorylation de GATA-1, et met en évidence le rôle majeur de l’IGF-1 dans l’érythropoïèse in vivo.En conclusion, nous proposons un nouveau modèle moléculaire de la régulation de la balance prolifération/différenciation érythroïde dans lequel la phosphorylation de GATA-1 par Akt coordonne la distribution de GATA-1 dans deux complexes protéiques fonctionnels différents : GATA-1/pRb/E2F versus GATA-1/FOG-1. Nous mettons également en évidence l’IGF-1 comme acteur central de la compensation mise en place in vivo pour pallier à l’absence de phosphorylation de GATA-1
With more than 100 billion red blood cells generated every day, the erythroid lineage has the largest output of cell production in adult mammals. This production requires a tight balance between cell proliferation, mainly controlled by erythropoietin (Epo)/PI3K/Akt signaling pathway, and erythroid differentiation induced by GATA-1 and FOG-1 transcription factors. Various links between these two processes have been previously demonstrated in the laboratory: 1) Epo-activated Akt directly phosphorylates GATA-1 transcription factors, and this phosphorylation seems to be involved in erythroid differentiation; 2) GATA-1 binds to the cell cycle regulator retinoblastoma protein (pRb), and the resulting complex is essential for terminal erythropoiesis.We investigated the molecular mechanisms involved in the cell proliferation/differentiation balance during terminal erythropoiesis; in particular, we studied the molecular and physiological role of Epo-induced GATA-1 phosphorylation. Our findings suggest that this phosphorylation is one of the key processes in erythropoiesis dynamics. In its unphosphorylated form, GATA-1 can break cell cycle progression via GATA-1/pRb/E2F complex. This preliminary step is necessary for terminal erythroid differentiation. GATA-1 phosphorylation promotes GATA-1/pRb/E2F dissociation, allowing cell cycle progression, and GATA-1/FOG-1 binding, necessary to activate erythroid genes. Our model provides a molecular explanation for the arrest of terminal erythroid differentiation observed in the non-FOG-1-binding mutant GATA-1V205G. We show that the constitutive phosphorylation of GATA-1V205G and the increase of FOG-1 protein amount rescue erythroid differentiation in vitro. Finally, knock-in expression of unphosphorylatable GATA-1 in mice leads to lethal anemia when the IGF-1 signaling pathway is inhibited. This shows the importance of the molecular dynamics of GATA-1 phosphorylation, and highlights the major role of IGF-1 in erythropoiesis, in vivo.In conclusion, we propose a new molecular model for the control of the balance between proliferation and erythroid differentiation. GATA-1 phosphorylation by Akt coordinates the involvement of GATA-1 in two different functional protein complexes: GATA-1/pRb/E2F and GATA-1/FOG-1. We also highlight the major role of IGF-1 in compensating for the lack of GATA-1 phosphorylation in vivo

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Itagaki, Tetsurō. "Interlayer organic modification of 1:1 type clay mineral kaolinite = 1:1-gata nendo kōbutsu kaorinaito no sōkan yūki shūshoku /." Electronic version of text Electronic version of summary Electronic version of examination, 2003. http://www.wul.waseda.ac.jp/gakui/honbun/3450/.

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Penglong, Tipparat. "Molecular Basis of Erythroid Cell Proliferation and Differentiation." Thesis, Paris 11, 2015. http://www.theses.fr/2015PA11T022.

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Pour assurer la production de milliards de globules rouges, l’érythropoièse doit parfaitement contrôler les processus de prolifération et de différenciation. Ces deux processus sont régulés par l’expression de gènes spécifiques dépendant d’une coordination entre l’activité des facteurs de transcription (FT) et les fonctions épigénétiques portées par exemple par les protéines à bromodomaine. Cette étude se concentre sur les conséquences de l’association ou la dissociation du FT clef de l’érythropoièse GATA-1 avec les FT déterminant pour le cycle cellulaire, pRb et E2F. Dans la première partie de ma thèse, j’ai participé à l’étude du rôle de l’association/dissociation de GATA-1 et FOG-2 avec pRb/E2F dans le contrôle la balance prolifération/différenciation cellulaire. Nos résultats montrent que les souris exprimant une mutation de GATA-1 sur la sérine 310 (GATA-1S310A), qui a la capacité accrue à séquestrer E2F-2, présentent une anémie létale lorsqu’un mécanisme de compensation de production de E2F-2 induit par l’IGF-1 est inhibé. Puis, nous avons trouvé que les propriétés décrites pour GATA-1 sont partagées par le FT FOG-2 et montré que l’abrogation de sa fixation avec pRb induit une perturbation de l’adiposité dans des souris FOG-2pRb-. Dans la deuxième partie, l’expression de c-Myc étant régulé différentiellement par GATA-1 et E2F, j’ai testé si la drogue « JQ1 », premier inhibiteur épigenétique chimique de l’expression de c-Myc, pouvait contrôler l’érythropoièse. Pour cela, j’ai utilisé la ligné érythroleucémique UT7 qui prolifère sans se différencier en présence d’érythropoiétine (stade proérythroblaste). Les résultats montrent que le traitement par JQ1 bloque la prolifération des cellules UT7 et permet de réinitier le programme de différentiation érythroide terminale. J’ai alors recherché les mécanismes moléculaires impliqués dans cette régulation et trouvé que l’inhibition transcriptionnelle de c-Myc par JQ1 est associée à l’inhibition de l’activité transcriptionnelle de STAT5 sans modification de son état de phosphorylation. Enfin, j’ai montré que JQ1 pouvait avoir une activité comparable à celle du TGF-b mais sans implication les voies Smad. Des études in vivo montre que JQ1 augmente la viabilité cellulaire et accélère la maturation des cellules érythroides à la fois chez les souris sauvages et thalassémiques. Cette différence d’action de JQ1 sur l’érythropoièse normale et pathologique implique des modifications épigénétiques différentielles entre ces deux types cellulaires et sont à la base de nouvelles stratégies du traitement du cancer. Le rôle clef de la régulation de l’association/dissociation de GATA-1 ou FOG-2 avec pRb/E2F dans l’érythropoièse et l’adipogénèse, nous a conduit, dans une troisième partie, à déterminer in vivo, les conséquences physiologiques de la séquestration de E2F par pRb. Pour cela nous avons crée une souris transgénique exprimant de façon conditionnelle un peptide contenant la partie N terminale de GATA-1 qui se fixe à pRb (GATA-1Nter). In vitro, ce peptide séquestre E2F dans le complexe GATA-1Nter/pRb et inhibe la prolifération cellulaire de façon irréversible. In vivo, aucune souris transgéniques exprimant le peptide GATA-1Nter n’a pu être sélectionnée et une mortalité au stade embryonnaire est observée. Une expression induite de ce peptide au stade adulte ne produit que des souris chimériques avec une fréquence de recombinaison du transgène GATA-1Nter importante. L’établissement de lignées stables de souris exprimant le peptide GATA-1Nter permettra de déterminer les conséquences physiologiques de la séquestration de E2F dans le complexe GATA-1Nter/pRb
To ensure the generation of billions of erythrocytes daily, erythropoiesis must be well controlled by proliferation and differentiation processes. These two processes are regulated by expressions of specific genes, coordinated by transcription factors (TFs) and epigenetic factors, such as bromodomain proteins. This study focused on the effects of the binding and dissociation of a key erythroid TF, GATA-1, to the crucial cell cycle TFs, pRb and E2F. In the first part of this thesis, the role of GATA-1 and FOG-2 binding to pRb/E2F in a control balances between cell proliferation and differentiation was studied. Mice bearing a GATA-1 mutation (GATA-1S310A) displayed higher levels of E2F2 sequestration and suffered from fatal anemia when the compensatory pathway of E2F2 production via IGF-1 signaling was also inhibited. The properties described for GATA-1 were found to be common to FOG-2, and the abolition of FOG-2 binding to pRb led to obesity resistance in FOG-2pRb- mice. In the second part of this work, as c-Myc is regulated by GATA-1 and E2F, the first chemical epigenetic inhibitor repressing c-Myc expression to be described, JQ1, was investigated to see if it could control erythropoiesis. The UT7 erythroleukemia cell line, which proliferates without differentiating was used. This cell line stops differentiation at the proerythroblast stage, in response to erythropoietin. JQ1 treatment inhibited UT7 proliferation and restored terminal erythroid differentiation. The molecular mechanism underlying this regulation by JQ1 was shown that the inhibition of c-Myc expression was associated with the inhibition of STAT5 transcription, with no change in the phosphorylation of this protein. It was found that JQ1 had a putative TGF--like activity, which did not involve the Smad pathway. It was shown in the ex vivo studies that JQ1 increased the viability of erythroid cells and accelerated the maturation of these cells in both WT and thalassemic mice. The observed differences between leukemic and normal erythropoiesis involved differential epigenetic modifications that could be at the basis of new strategies regarding cancer treatment.The key role of the association of GATA-1 or FOG-2 had with pRb/E2F, and the dissociation of these factors, in erythropoiesis and adipogenesis, respectively, led us to investigate, in vivo, the physiological consequences of E2F sequestration by pRb. As a result, transgenic mice displaying conditional expression of a peptide containing the N-terminal part of GATA-1 that binds to pRb (GATA-1Nter) were developed. In vitro, this peptide traps E2F in a GATA-1Nter/pRb complex, resulting in the irreversible inhibition of cell proliferation. The yield of transgenic mice expressing the GATA-1Nter peptide in vivo was unsuccessful, as this expression lead to lethality at the embryonic stage. Using an alternative approach, based on the inducible expression of the peptide in adults, chimeric mice with a high frequency of recombination of the GATA-1Nter transgene were obtained for this study. The establishment of a stable mouse line expressing the GATA-1Nter peptide should make it possible to determine the pathophysiological consequences of E2F sequestration in the GATA-1Nter/pRb complex

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Rio, Sarah. "Etude des métabolismes du fer et de l’hème au cours de l’érythropoïèse normale et pathologique (anémie de Blackfan-Diamond)." Thesis, Sorbonne Paris Cité, 2016. http://www.theses.fr/2016USPCB055/document.

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L’anémie de Blackfan-Diamond (ABD) est une maladie hématologique rare qui touche 4 à 7 individus/ million de naissances. Cette maladie se manifeste par une érythroblastopénie congénitale sévère (≤ 5% de précurseurs érythroïdes dans la moelle osseuse). L’anémie est arégénérative et souvent macrocytaire et associée à des malformations osseuses dans 40% des cas. 70% des patients sont porteurs d’une mutation hétérozygote pour un gène de protéine ribosomique impliquée dans la traduction cellulaire. Les gènes les plus fréquemment mutés sont les gènes RPS19 (25%), RPL11 (5%) et RPL5 (7%). La maladie est hétérogène et évolutive. Les liens entre la traduction cellulaire et l’érythropoïèse ne sont pas bien élucidés. Les objectifs de cette thèse ont été d’étudier les métabolismes de l’hème et du fer ainsi que l’expression des globines dans des cellules de patients atteints d'ABD et dans un modèle shARN ciblant l'expression de ces trois gènes afin de comprendre les causes du tropisme érythroïde de la maladie. Ce travail de recherche a permis de mettre en évidence un défaut majeur de synthèse des globines ayant pour conséquence une augmentation de la quantité d’hème libre et une production de formes réactives de l'oxygène toxiques dans les cellules des patients qui pourraient expliquer en partie l’apoptose cellulaire et le déficit de globules rouges. Alors que le métabolisme du fer ne semblait pas altéré dans l'ABD, l’étude de l’expression de différentes protéines importantes pour l’érythropoïèse au cours de la différenciation érythroïde in vitro dans des conditions contrôles et chez des patients a permis de confirmer et de caractériser le retard de différenciation cellulaire en cas de mutation des gènes RPL5 et RPL11. Ce travail montre que le retard de différenciation et le défaut d'hémoglobinisation mis en évidence s'expliquent par un déficit du facteur de transcription GATA-1 qui est primordial au cours de l'érythropoïèse. Ce déficit de GATA-1 dans des cellules déficitaires en RPL11 est dû à une dégradation de sa protéine chaperonne HSP70. La restauration de HSP70, permet d'augmenter l'expression de GATA-1 et d'améliorer la différenciation érythroïde et l'hémoglobinisation cellulaire pour le gène RPL11. Ces résultats permettent de mieux comprendre le tropisme érythroïde de l'ABD et de proposer HSP70 comme une cible thérapeutique prometteuse dans son traitement
Diamond-Blackfan anemia (DBA) is a rare hematologic disease that affects 4 to 7 individuals / million births. This disease is characterized by a severe congenital erythroblastopenia (less than 5% erythroid precursors in the bone marrow). Anemia is agerenative, often macrocytic and associated with bone malformations in 40% of cases. 70% of patients carry a heterozygous mutation for a ribosomal protein gene involved in cell translation. The most frequently mutated genes are RPS19 (25%), RPL11 (5%) and RPL5 (7%) genes. The disease is heterogeneous and can evolve. The link between cell translation and erythropoiesis is not well understood. The objectives of this thesis were to study haem and iron metabolisms as well as the expression of globins in DBA patients cells and CD34+ cells transduced with shRNA targeting the expression of these three genes in order to understand the causes of the erythroid tropism of the disease. This research has highlighted a major defect of globin synthesis resulting in an increase in the amount of free heme and a production of toxic ROS in patients' cells that could explain in part cell apoptosis and red blood cell deficiency. While iron metabolism did not appear to be altered in DBA, the study of the expression of various important proteins for erythropoiesis in normal CD34+ or DBA cells during erythroid differentiation in vitro confirmed a strong cell differentiation delay for RPL5 and RPL11 mutations. This work shows that the delay of differentiation and the lack of hemoglobinization can be explained by a deficiency of the transcription factor GATA-1, which is essential during erythropoiesis. This deficiency of GATA-1 in shRPL11 cells is due to a degradation of its chaperone protein HSP70. The restoration of HSP70 increases the expression of GATA-1 and improves erythroid differentiation and cellular hemoglobinization for the shRPL11 condition. These results provide a better understanding of the erythroid tropism of ABD and suggest a role for HSP70 as a promising therapeutic target in its treatment

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Arlet, Jean-Benoît. "Rôle de la chaperonne HSP 70 dans l'éythropoïèse inefficace des béta-thalassémies majeures." Phd thesis, Université Paris Sud - Paris XI, 2013. http://tel.archives-ouvertes.fr/tel-01059816.

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L'érythropoïèse inefficace joue un rôle central dans la physiopathologie de l'anémie des β-TM. Ses caractéristiques sont triple: accélération de la différenciation érythroïde, arrêt de maturation au stade d'érythroblaste polychromatophile et mort par apoptose à ce stade de différenciation. Les mécanismes précis de cette apoptose et de l'arrêt de la maturation n'ont pas encore été élucidés. Il a été montré, au cours de l'érythropoïèse physiologique, que la protéine chaperonne Hsp70, en se localisant dans le noyau des érythroblastes en cours de différenciation, protège GATA-1 (facteur de transcription érythroïde majeur) de sa destruction par la caspase-3. Cette enzyme clé de l'apoptose est en effet activée physiologiquement au cours de la différenciation érythroïde et peut cliver GATA-1. Notre travail se base sur l'hypothèse suivante : Hsp70 pourrait, au cours de l'érythropoïèse des β-TM, être séquestrée dans le cytoplasme des érythroblastes matures (stade d'une intense hémoglobinisation) afin d'exercer son rôle de chaperonne des chaînes d'-globine libres. Cela aurait comme conséquence néfaste l'absence de localisation nucléaire d'Hsp70 et, en conséquence, la destruction de GATA-1 à l'origine de l'arrêt de maturation et de la mort cellulaire. Nous avons montré dans ce travail qu'Hsp70 était localisée principalement dans le cytoplasme des érythroblastes matures dans la moelle de patients β-TM, avec un défaut d'expression nucléaire. Par ailleurs, GATA-1 n'est plus exprimé dans ces cellules. Nous avons confirmé ces résultats dans un système de culture cellulaire érythroïde humaine en milieu liquide reproduisant les étapes de la différenciation érythroïde terminale. Une intéraction physique directe entre Hsp70 et l'-globine a été identifiée par techniques de microscopie confocale, d'immunoprécipitation et de double hybride. Enfin, la transduction dans les érythroblastes de β-TM d'un mutant d'Hsp70-S400A, principalement nucléaire, ou d'un mutant de GATA-1 non clivable par la caspase-3 corrige l'érythropoïèse inefficace.Une modélisation mathématique du complexe Hsp70/-globine nous a permis de préciser les domaines impliqués dans l'intéraction, ce qui ouvre la voie à une possibilité de criblage de petites molécules permettant la rupture de ce complexe afin de ramener Hsp70 dans le noyau avec un espoir thérapeutique pour améliorer l'érythropoïèse inefficace des β-TM.

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Sharma, Sribava. "Deletion of ΔdblGata Motif Leads to Increased Predisposition and Severity of IgE-mediated Food-induced Anaphylaxis Response." University of Cincinnati / OhioLINK, 2018. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1535701847469787.

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Johnson, Lacey Nicole St George Clinical School UNSW. "Molecular regulation of Megakaryopoiesis: the role of Fli-1 and IFI16." Awarded by:University of New South Wales. St George Clinical School, 2006. http://handle.unsw.edu.au/1959.4/26819.

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Megakaryocytes (Mks) are unique bone marrow cells, which produce platelets. Dysregulated Mk development can lead to abnormal platelet number and the production of functionally defective platelets, causing bleeding, thrombotic events, and leukaemia. Understanding the molecular mechanisms driving megakaryopoiesis may yield insights into the molecular genetics and cellular pathophysiology of a diversity of disorders. The primary aim of this thesis was to gain insight into the molecular events required for normal Mk development. As transcription factors and cytokines play a central role in driving Mk development, both of these processes were investigated. Fli-1 and GATA-1 are key transcription factors regulating Mk-gene expression, alone and co-operatively. To understand the mechanism of transcriptional synergy exerted by Fli-1 and GATA-1, in vitro assays were carried out investigating the interactions between Fli-1, GATA-1 and DNA that mediate synergy. A novel mechanism of synergy was identified, where Fli-1 DNA binding is not required, although an interaction between Fli-1 and GATA-1, and GATA-1 DNA binding is required. Importantly, the results demonstrate that Fli-1 DNA binding is not essential for promoting Mk-gene expression in primary murine bone marrow cells. Thrombopoietin (TPO) is the primary cytokine responsible for Mk and platelet development. Identifying novel TPO gene-targets may provide invaluable information to aid the understanding of the complex and unique processes required for Mk development. Using microarray technology, IFI16 was identified as a TPO-responsive gene that has not previously been studied in the Mk lineage. This work demonstrated that IFI16 is expressed in CD34+ HSC-derived Mks, and that the Jak/STAT pathway is essential for the activation of IFI16 by both TPO and IFN-??. Of biological significance, IFI16 was found to regulate both the proliferation and differentiation of primary Mks, suggesting that IFI16 may control the balance between these two essential processes. In conclusion, the data in this thesis presents a novel mechanism through which Fli-1 and GATA-1 regulate the synergistic activation of Mk genes. The identification and functional characterisation of a novel TPO-inducible gene, IFI16, involved in regulating the proliferation and differentiation of Mks is also described. These findings have implications for several congenital and malignant conditions affecting Mk and platelet development, and possibly a mechanism for IFN-induced thrombocytopaenia.

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Trécul, Anne. "Effet de l'acide valproïque sur l'hématopoïèse : rôle du réseau de régulation "microARN/ facteurs de transcription"." Thesis, Université de Lorraine, 2014. http://www.theses.fr/2014LORR0143/document.

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L’acide valproïque (VPA) est un inhibiteur des histones désacétylases (HDACi), qui présente des propriétés anti-tumorales nsur différents types de cancers. Son utilisation depuis plusieurs décennies comme médicament antiépileptique a révélé des effets secondaires, notamment sur le système hématopoïétique. Dans la présente étude, nous nous sommes intéressés à l’effet du VPA sur les réseaux microARN (miR)/Facteurs de transcription (FT) spécifiquement impliqués dans la régulation des voies de différenciation érythro-mégacaryocytaires. Nous montrons que le VPA est capable d’inhiber la différenciation érythroïde dans les cellules érythroleucémiques humaines TF1 et K562 et dans les cellules souches hématopoïétiques (CSH) CD34+, stimulées par l’érythropoïétine recombinante (Epo) ou par l’aclacinomycine A. Cette inhibition se traduit par une diminution de l’expression de la glycophorine A, de la γ-globine, des miR-144/451 et du FT GATA-1. L’inhibition du pré-miR-144 suggère que le VPA est capable de réguler l’expression du gène miR-144/451 au niveau transcriptionnel, via GATA-1. Dans les cellules Epo/CD34+, le VPA induit l’augmentation du FT PU.1 en accord avec l’inhibition du miR-155 et favorise son interaction avec GATA-1 pour inhiber son activité. L’utilisation d’un analogue du VPA, sans activité HDACi (Valpromide) et d’un inhibiteur d’HDAC de classe I, le MS-275, a montré que l’activité HDACi du VPA n’est pas requise pour l’inhibition de la différenciation érythroïde. Le VPA affecte également la voie mégacaryocytaire issue d’un progéniteur commun aux cellules érythroïdes. Dans la lignée mégacaryoblastique Meg-01, le VPA induit des modifications morphologiques du type mégacaryocytaire, une augmentation du marqueur CD61, du FT GATA-2 et du miR-27a. En revanche, l’expression du FT GATA-1 et des miR-144/451 diminuent. L’augmentation du miR-27a coïncide avec la diminution de l’expression de l’ARNm du FT RUNX1, en accord avec l’induction de la voie mégacaryocytaire. En conclusion, le VPA est capable de moduler le programme de différenciation érythro-mégacaryocytaire, à travers un micro réseau de régulation miR/FT
Valproic acid (VPA), a histone deacetylase inhibitor (HDACi), exhibits anti-cancer properties against several tumor types. Its use as an anti-epileptic drug for several decades reveled side effects at the hematological level. In this study, we analyzed the effect of VPA on an erythro-megakaryocyte-specific miR/transcription factors network. VPA inhibited erythroid differentiation in the erythroleukemia cell lines TF1 and K562 as well as in CD34+/hematopoietic stem cells (HSCs), induced by the recombinant erythropoietin (Epo) or aclacinomycin. This inhibition was characterized by glycophorin-A, γ-globin and GATA-1/miR-144/451 down-regulation. Inhibition of pre-miR-144 expression suggested that VPA regulates transcription of the miR-144/451 gene through GATA-1. In Epo-stimulated HSCs, VPA induced PU.1 expression in correlation with miR-155 inhibition and promoted GATA-1/PU.1 interaction. The use of valpromide, a VPA analogue without HDACi activity and the class-I HDACi MS-275, showed that HDAC inhibition by VPA was not required for its inhibitory activity on erythropoiesis. VPA also induced megakaryocyte features in Meg-01 cells, at both cellular and molecular levels. Notably, CD61, GATA-2 and miR-27a were over-expressed. RUNX1 mRNA expression and GATA-1/miR-144/451 axis decreased in accordance with megakaryocyte differentiation. In conclusion, VPA is able to modulate erythro-megakaryocytic differentiation program, through a regulatory micro-network involving miRs and TFs

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Xi, Zong-Fang. "Opposing effects of two zinc finger genes, EVI-1 and GATA-1, on all-trans retinoic acid-induced NB4 cell granulocytic differentiation." Diss., The University of Arizona, 2000. http://hdl.handle.net/10150/284255.

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In hematopoietic stem cells, lineage specific differentiation is determined in part by specific transcription factors which stabilize a particular lineage network and activate differentiation programs. In myeloid and erythroid lineages, distinct transcription factors are expressed in committed progenitors and more differentiated cells. Zinc finger transcription factors, EVI-1 and GATA-1, are expressed in hematopoietic stem cells. Abnormal EVI-1 expression is associated with some human acute myeloid leukemias (AML). It has been shown that enforced EVI-1 expression blocked erythroid specific transcription factor, GATA-1, mediated transcriptional activation and erythroid differentiation. The regulation and the function of the EVI-1 in myeloid differentiation and possible antagonism of erythroid/myeloid lineage transcription factors in lineage commitment and differentiation were studied in NB4, a human acute promyelocytic leukemia (APL) cell line. EVI-1 and GATA-1 were stably expressed in NB4 cells by transfection and effects evaluated on granulocyte differentiation induced by all-trans retinoic acid (ATRA). The results showed that EVI-1 expressing NB4 clones differentiate more rapidly and more completely than control clones. In contrast, GATA-1 expressing NB4 clones showed delayed differentiation. These findings demonstrate that EVI-1 expression promotes and GATA-1 expression delays ATRA-induced NB4 cell granulocytic differentiation, suggesting that EVI-1 plays an important role in myeloid differentiation and EVI-1 and GATA-1 antagonize each other in erythroid/myeloid commitment and differentiation.

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Rémillard, Anthony. "Le facteur de transcription GATA-4 et son rôle dans la réponse inflammatoire intestinale chez le rat." Mémoire, Université de Sherbrooke, 2009. http://savoirs.usherbrooke.ca/handle/11143/4041.

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GATA-4 est un facteur de transcription essentiel dans la formation du tube cardiaque primitif. Dans l'intestin, GATA-4 joue un rôle dans le développement et dans le maintien de l'identité jujéno-iléale. L'analyse par micropuce à ADN effectuée sur des cellules IEC-6 surexprimant GATA-4 suggère un rôle de GATA-4 dans la réponse inflammatoire et des études antérieures réalisées au laboratoire ont permis de confirmer l'implication de GATA-4 dans la modulation de la réponse inflammatoire C/EBPdépendante. L'objectif de mon projet était de déterminer si GATA-4 module la réponse inflammatoire par d'autres mécanismes que ceux déjà découverts au laboratoire et de générer des mutants de GATA-4 pour caractériser l'implication de certains domaines et sites spécifiques de GATA-4 dans la modulation de la réponse inflammatoire intestinale. Des cellules IEC-6 surexprimant GATA-4 ou des mutants de GATA-4 de façon stable, induites ou non à l'IL-1[béta] ont été utilisées comme modèle. L'activation des voies MAPK et Akt a été vérifiée par immunobuvardage de type Western. La liaison à l'ADN des facteurs de transcription AP-1, C/EBPs et NF-[kappa]B a été vérifiée par gel de rétention. L'expression de certains gènes de réponse inflammatoire a été vérifiée par RT-PCR. Finalement, des lignées cellulaires exprimant des mutants de GATA-4 ont été partiellement caractérisées. Mes résultats montrent qu'une surexpression de GATA-4 dans les IEC-6 traitées à l'IL-1[béta] réduit la phosphorylation de p42/p44 et d'Akt. Les facteurs de transcription AP-1, C/EBP et NF-[kappa]B sont davantage recrutés à l'ADN. La nature de leurs complexes semble être également affectée. Cette induction de liaison coïncide avec une baisse d'expression des gènes pro-inflammatoires CCL5 et iNOS. La caractérisation des mutants de GATA-4 suggère que le premier doigt de zinc (216-240) pourrait moduler la morphologie cellulaire et que la région basique (294-336) pourrait contribuer à la vitesse de prolifération augmentée des cellules IEC-6 surexprimant GATA-4. En somme, mes résultats suggèrent que GATA-4 pourrait participer à l'instauration d'une tolérance immunitaire prenant place lors de la différenciation entérocytaire.

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Ribeil, Jean-Antoine. "Hsp70 est un nouveau régulateur majeur de l'érythropoïèse empêchant le clivage du facteur de transcription GATA-1 par la caspase-3 au cours de la différenciation." Phd thesis, Université Paris-Diderot - Paris VII, 2010. http://tel.archives-ouvertes.fr/tel-00451047.

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La production de globules rouges dépend du taux apoptose des précurseurs érythroïdes et est principalement régulé par l'érythropoïétine (Epo). La privation en Epo aboutit à l'activation de la caspase-3 (casp-3) qui clive GATA-1 ce qui entraîne l'apoptose des érythroblastes immatures. L'activation de la casp-3 est également indispensable aux modifications morphologiques caractéristiques observées au cours de la différenciation érythroïde terminale humaine, sans qu'il n'y ait ni d'apoptose ni de clivage de GATA-1. L'objectif de cette thèse était de mettre en évidence si Hsp70 inductible, dont un des rôles principaux est la régulation de l'apoptose, est impliquée dans la protection sélective des substrats de la casp-3 activée au cours de la différenciation érythroïde terminale humaine. Nous avons mis en évidence que lors de la différentiation érythroïde terminale pendant la phase d'activation des caspases, Hsp70 a une expression nucléo-cytoplasmique constitutive et co-localise avec GATA-1 dans le noyau. La localisation nucléaire d'Hsp70 est régulée par l'Epo : après privation des cellules en Epo, il y a une importante diminution de la localisation nucléaire d'Hsp70 et GATA-1 est clivée. L'inhibition de l'expression d'Hsp70 par une approche siRNA a comme conséquence le clivage de GATA-1 lors de l'activation de la casp-3 avec un arrêt de différenciation et une augmentation de la mort cellulaire. Hsp70 est une nouvelle protéine anti-apoptotique de la différenciation érythroïde terminale. Nous proposons un modèle dans lequel l'Epo détermine le destin des érythroblastes (apoptose vs différenciation) en aval de la casp-3 en régulant la localisation nucléaire d'Hsp70.

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Frisan, Emilie. "Etude de la différenciation érythroïde des syndromes myélodysplasiques de faible risque : rôle des protéines GATA-1 et Hsp70." Paris 5, 2010. http://www.theses.fr/2010PA05T007.

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L'engagement des progéniteurs hématopoïétiques normaux vers la lignée érythroïde est contrôlé par le stem cell factor (SCF) et) l'érythropoïétine (Epo) alors que la différenciation érythroïde terminale dépend de l'Epo. La dysérythropoïèse des syndromes! myélodysplasiques (SMP), maladies clonales de la cellule souche hématopoïétique, associe un défaut de différenciation des| progéniteurs et un excès d'apoptose des précurseurs. Ce travail montre : (1) l'implication du réticulum endoplasmique dans l'apoptose en aval du récepteur à domaine de mort Pas, (2) le défaut d'activation des MAPK par le récepteur de l'Epo chez les patients résistants au traitement par l'Epo, et (3) le clivage du facteur de transcription érythroïde GATA-1 par la caspase-3 par défaut de localisation nucléaire de la protéine chaperon Hsp70. L'export nucléaire d'Hsp70 dépend de sa phosphorylation en réponse au| SCF et serait augmenté par une expression ectopique du récepteur au SCF dans les érythroblastes matures de SMD
Normal hematopoietic progenitors commitment into erythroid lineage is controlled by stem cell factor (SCF) and erythropoietin (Epo) while erythroid terminal differentiation is only Epo-dependent. DyserythropoYesis of myelodysplastic syndromes (MDS) which clonal disorders of the hematopoietic stem cell, combines a defective differentiation of the progenitors and an increased apoptosis of the precursors. This work shows (1) the implication of endoplasmic reticulum in MDS apoptosis downstream of death domain) receptor Fas, (2) defective Epo-dependent activation of the MAPK in patients resistant to Epo treatment, and (3) a caspase-3-mediated Cleavage of the erythroid transcription factor GATA-1 due to a defective nuclear localization of the chaperone protein Hsp70 in MDS. Nuclear export of HspVO is regulated by AKT phosphorylation in response to SCF and would be increased by an ectopic expression) of SCF receptor in MDS mature erythroblasts

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Lahlil, Rachid. "Facteurs de transcription et differenciation erythroide de la lignee leucemique humaine k562 : modulation de gata-1, gata-2 et nf-e2 par un inhibiteur (azt) ou par une strategie sens et antisens." Reims, 1997. http://www.theses.fr/1997REIMP206.

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Vandekerckhove, Julie. "Mécanismes de régulation de GATA-1 par les protéines de choc Hsp27 et Hsp70 au cours de la différenciation érythroïde terminale." Phd thesis, Paris 11, 2009. http://www.theses.fr/2009PA11T078.

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Gillet, Reynald. "Voies de transduction et activation du facteur de transcription gata-1 au cours de la differenciation erythroide induite par l'aclacinomycine (doctorat : biochimie et biologie moleculaire)." Reims, 1999. http://www.theses.fr/1999REIMP210.

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Dutescu, Ralf Michael [Verfasser], and H. G. [Akademischer Betreuer] Wahl. "Expressionsanalyse der nukleären Rezeptoren PPAR-α/γ-1/γ-2 [PPAR-alpha, gamma-1, gamma-2] und der Transkriptionsfaktoren T-bet und GATA-3 nach Stimulation von dermalen Endothelzellen mit den Weichmacher, Di(2-ethylhexyl)phthalat-Metaboliten-2-Ethylhexanol und 4-Heptanon / Ralf Michael Dutescu. Betreuer: H. G. Wahl." Marburg : Philipps-Universität Marburg, 2011. http://d-nb.info/1013288459/34.

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Mazzi, Stefania. "Study of the role of the methyltransferase EZH2 in normal and pathological megakaryopoiesis." Thesis, Sorbonne Paris Cité, 2018. https://theses.md.univ-paris-diderot.fr/MAZZI_Stefania_2_complete_20180926.pdf.

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Le processus qui aboutit à la formation de plaquettes est appelé mégacaryopoïèse. Les mégacaryocytes (MK) sont de grandes cellules de la moelle osseuse qui par fragmentation dans la circulation sanguine produisent des plaquettes. La régulation extrinsèque ou intrinsèque de ce processus a été largement étudiée. Cependant la régulation épigénétique reste mal connue bien que de nombreuses mutations dans des gènes de régulateurs épigénétiques soient retrouvées dans les hémopathies malignes de la lignée MK. En particulier des mutations du gène de la méthyltransférase EZH2, composant catalytique du Polycomb Repressive Complex 2 (PRC2) ont été détectées dans plusieurs types d’hémopathies. Ces mutations sont soit gain soit perte de fonction suggérant qu’EZH2 peut être à la fois un oncogène ou un gène suppresseur de tumeur. Dans les TE (Thrombocythémie Essentielle) et les MFP (Myélofibrose Primaire), deux néoplasmes myéloprolifératifs (NMPs), qui affectent principalement la lignée MK, des mutations d’EZH2 perte de fonction ont été retrouvées ainsi que dans les DS-AMKL (Down syndrome acute megakaryoblastic leukemia). Cela suggère qu’EZH2 joue un rôle important dans la mégacaryopoïèse normale. La caractérisation de cette fonction pourrait être utile pour mieux appréhender le rôle des mutations d’EZH2 dans les pathologies malignes mégacaryocytaires. Cette thèse peut être divisée en deux parties : 1) Caractérisation du rôle joué par EZH2 dans la mégacaryopoïèse normale et pathologique 2) Développement d‘un outil permettant d’étudier la coopération entre mutations dans les DS-AMKL.1) Lors des temps précoces de la différenciation in vitro des cellules CD34+ de sang de cordon vers la lignée mégacaryocytaire l’inhibition d’EZH2 entraîne l’acquisition plus rapide des marqueurs MK de surface (CD41 et CD42) pour un nombre de mitoses égal. Ceci suggère qu’EZH2 régule la spécification MK des progéniteurs hématopoïétiques. Plus tard dans la différenciation, l'inhibition constante d’EZH2 via des inhibiteurs ou des shRNA, arrête la prolifération et diminue le niveau de ploïdie des MKs en arrêtant la réplication de l’ADN. Ceci est du à la surexpression de plusieurs CDKi (Cyclin dependent kinase inhibiteurs), dont CDKN2D. L'analyse par Chip-Seq a montré que la transcription de CDKN2D est régulée par H3K27me3 au niveau de son promoteur et donc que CDKN2D est une nouvelle cible de PRC2. Dans les MKs les plus matures, l’inhibition d’EZH2 diminue la formation des proplaquettes, ceci est corrélé à des modifications d’expression de gènes régulant le cytosquelette d’actine. L’ensemble de ces résultats a été confirmé sur des MKs de patients porteurs de la mutation JAK2V617F.2) Par la technique CRISPR-Cas9, nous avons introduit dans des iPSC (induced pluripotent stem cells) disomiques et trisomiques pour le chromosome 21, la mutation GATA1s présente chez tous les patients avec une DS-AMKL. Nous avons montré que ces mutations modifiaient le cadre de lecture dans l’exon 2 et entrainaient l’expression de la forme courte de GATA1 (GATA1s). Nous sommes en train d'effectuer des études fonctionnelles ainsi que d’introduire d’autres mutations, y compris celles d’EZH2 pour modéliser la maladie.Au cours de cette thèse nous avons montré que l’inhibition d’EZH2 régule les temps initiaux de la mégacaryopoïèse en accélérant la spécification cellulaire au niveau des progéniteurs et ensuite la maturation terminale en inhibant profondément la polyploïdisation par surexpression de plusieurs CDKi dont CDKN2D et en inhibant la formation des plaquettes par un effet sur le cytosquelette d’actine. Ces résultats pourront être utiles pour mieux comprendre le rôle de la perte de fonction d’EZH2 dans les hémopathies malignes de la lignée mégacaryocytaire
The process that leads to platelet production is called megakaryopoiesis. Megakaryocytes (MK) are the large bone marrow cells that produce platelets by fragmentation in the blood flow. The extrinsic and intrinsic regulation of megakaryopoiesis has been largely studied. However, the epigenetic regulation remains poorly known although numerous mutations in genes of epigenetic regulators have been found in patients with MK hematological malignancies. The methyltransferase EZH2, the catalytic component of Polycomb Repressive Complex 2 (PRC2) is among the most studied epigenetic regulators. EZH2 is also mutated in many malignant hematological disorders where it can be an oncogene or a tumor suppressor gene. Particularly in ET (Essential Thrombocythemia) and PMF (Primary Myelofibrosis), two myeloproliferative neoplasms (MPNs) that affect mainly the MK lineage, loss of function EZH2 mutations have been found as well as in DS-AMKL (Down syndrome acute megakaryoblastic leukemia)Altogether these observations suggest that EZH2 controls normal megakaryopoiesis and characterization of this function could be helpful to understand the role of EZH2 in MK malignant diseases.This thesis can be divided in two parts:1) Characterization of the role of EZH2 in normal and pathological megakaryopoiesis 2) Establishment of a cellular tool to study the cooperation between the different mutations of DS-AMKL. RESULTS1) Using CD34+ cells isolated from cord blood, we showed that at early stages of differentiation, EZH2 inhibition accelerates the acquisition of MK surface markers (CD41a and CD42a) without increasing proliferation suggesting that EZH2 regulates the specification towards the MK lineage. Later in differentiation the constant inhibition of EZH2 via inhibitors or shRNAs, produced a proliferation arrest and a decrease in ploidy level that was related to an arrest in DNA replication due to an upregulation of several CDKi (Cyclin dependent kinase inhibitors), more particularly CDKN2D. Chip-Seq analysis demonstrated that CDKN2D is effectively regulated by H3K27me3 and is a new target of PRC2. This inhibition of ploidization by EZH2 inhibition was confirmed in MK from JAK2V617F patients. Furthermore in the more mature MKs (normal or JAK2V617F) we observed a defect in proplatelet formation, which was associated with an abnormal expression of genes regulating the actin filament. 2) By CRISPR-Cas 9, in iPSCs either disomic or chromosome 21 trisomic, we introduced, the GATA1s mutation present in all DS-AMKL patients. We confirmed at the gene and protein level that this genome editing has been correctly performed and that it induces as previously observed a blockage in erythroid differentiation. We are now carrying out the complete functional characterization together with the introduction of other mutations of DS-AMKL including EZH2.CONCLUSIONThis study describes EZH2 as a regulator of megakaryopoiesis via an initial control of cell specification and then of MK maturation. These results will be useful to better understand the role that EZH2 plays in diseases affecting the MK lineage such as MPNs and DS-AMKL

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Gastou, Marc François Philippe. "Rôle de la protéine HSP70 au cours de l'anémie de Blackfan-Diamond." Thesis, Sorbonne Paris Cité, 2016. http://www.theses.fr/2016USPCC231/document.

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L’anémie de Blackfan-Diamond (ABD) est une érythroblastopénie congénitale rare, secondaire à un blocage de la maturation érythroïde entre les stades BFU-e et CFU-e. L’ABD est le plus souvent la conséquence d’une mutation germinale affectant un gène codant pour une protéine ribosomique (RP) de la petite ou de la grande sous-unité du ribosome. Quatorze gènes distincts ont été identifiés. Les gènes les plus fréquemment mutés sont les gènes RPL5, RPL11 et RPS19 (37% des patients). Plus rarement, l’ABD est la conséquence de mutations dans le gène TSR2 ou dans le gène GATA-1. Ce dernier code pour un facteur de transcription majeur de l’érythropoïèse. Chez les patients ABD, les mutations de GATA-1 induisent une perte quasi-totale de la forme longue de GATA-1 qui est nécessaire à la différenciation de la cellule érythroïde. Notre groupe a identifié deux phénotypes de l’ABD in vitro en fonction du gène muté. En cas d’haploinsuffisance RPS19, la prolifération érythroïde est moins réduite qu’en cas d’haploinsuffisance RPL5 ou de RPL11. Une haploinsuffisance RPS19 n’altère pas la différenciation érythroïde et n’induit pas d’apoptose contrairement à l’haploinsuffisance RPL5 ou RPL11 où il existe un retard de différenciation érythroïde et un excès net d’apoptose responsable au moins en partie de la diminution drastique de la prolifération érythroïde dans ces phénotypes.HSP70 est impliquée dans la survie cellulaire et la différenciation érythroïde en protégeant GATA-1 du clivage par la caspase-3, une protéase activée lors de la différenciation érythroïde terminale. Comme la différence entre les deux phénotypes d’ABD in vitro concernait la différenciation érythroïde et la survie cellulaire, nous avons émis l’hypothèse selon laquelle la mutation de certains gènes RP provoque un défaut d’expression d’HSP70 conduisant au blocage de la différenciation érythroïde et à l’excès d’apoptose retrouvés dans les phénotypes sévères d’ABD.Nous avons étudié différents patients atteints d’ABD, porteurs de mutations dans les gènes RPS19, RPL5 ou RPL11 et généré un modèle in vitro d’ABD en exprimant, dans des cellules CD34+ humaines issues de sang de cordon, des ARN interférents ciblant RPL5, RPL11 ou RPS19. Chez les patients comme dans le modèle reproduisant l’ABD, l’haploinsuffisance RPL5 ou RPL11 diminue drastiquement l’expression protéique de HSP70 et de GATA-1 (Western blot, microscopie confocale et en cytométrie couplée à des techniques d’imagerie, (technologie ImageStream) à la différence de 1’haploinsuffisance RPS19. Dans tous les cas, HSP70 est normalement transcrite et traduite. Les inhibiteurs du protéasome (MG132, lactacystine, bortezomib) restaurent l’expression10de HSP70. La diminution d’expression de HSP70 est donc liée à une dégradation protéasomale. L’invalidation de RPL11 induit une polyubiquitinylation importante de HSP70. La transduction lentivirale de l’ADN complémentaire d’HSP70 dans les cellules primitives invalidées pour RPL11 permet de restaurer l’expression de HSP70 et de GATA-1 à un niveau similaire aux contrôles et de rétablir la prolifération cellulaire et la différenciation érythroïde, confirmant le rôle clé de HSP70 dans le phénotype sévère RPL5+/Mut ou RPL11+/Mut. Les formes les plus sévères de l’ABD sont associées à la dégradation de HSP70 par le protéasome. La perte de la protéine chaperone de GATA-1 induit la perte de GATA-1, facteur de transcription majeur de la différenciation érythroïde. Une augmentation de l’expression de HSP70 pourrait ainsi constituer une nouvelle approche thérapeutique dans l’ABD
Diamond-Blackfan anemia (DBA) is the first ribosomopathy identified and is characterized by a moderate to severe, usually macrocytic aregenerative anemia associated with congenital malformations in 50% of the DBA cases. This congenital rare erythroblastopenia is due to a blockade in erythroid differentiation between the BFU-e and CFU-e stages. The link between a haploinsufficiency in a ribosomal protein (RP) gene that now encompass 15 different RP genes and the erythroid defect is still to be fully defined. Recently, mutations in TSR2 and GATA-1 genes have been identified in a few DBA families. The GATA-1 gene encodes for the major transcription factor critical for erythropoiesis and mutation in this gene that lead to loss of expression of the long form of the protein, necessary for the erythroid differentiation accounts for erythroblastopenia of DBA phenotype. Our group and others (Dutt et al., Blood 2011) have shown previously that p53 plays an important role in the DBA erythroblastopenia, inducing cell cycle arrest in G0/G1 and depending on the nature of RP gene mutation, a delayed erythroid differentiation and an increased apoptosis. Indeed, we identified two distinct DBA phenotypes (H. Moniz, M. Gastou, Cell Death Dis, 2012): a haploinsufficiency in RPL5 or RPL11 reduced dramatically the erythroid proliferation, delayed the erythroid differentiation, and markedly increased apoptosis, while RPS19 haploinsufficiency while reduced the extent of erythroid proliferation without inducing significant apoptosis. While p53 pathway has been found to be activated in RP haploinsufficient erythroid cells in DBA patients or shRNA-RPS19, -RPL5, or -RPL11 infected CD34+ erythroid cells, the intensity of the p53 activation pathway (p21, BAX, NOXA) is different depending on the mutated RP gene. Since the differences between the two phenotypes involved the eytrhoid differentiation and the degree of apoptosis we hypothesized that HSP70, a chaperone protein of GATA-1 may play a key role in the erythroid defect of DBA. Indeed, HSP70 protects GATA-1 from the cleavage by the caspase 3, a protease activated during erythroid differentiation. As such reduced levels of HSP70 related to a RP haploinsufficiency could account for increased apoptosis and delayed erythroid differentiation of erythroid cells in DBA. Indeed, a defect in RPL5 or RPL11 decreased dramatically the expression level of HSP70 and GATA-1 in primary human erythroid cells from DBA patients and following in vitro knockdown of the proteins in CD34+ cells by RPL5 or RPL11 shRNA. Importantly, RPS19 haploinsufficiency did not exhibit this effect in conjunction with normal levels of HSP70 expression. Furthermore, we found that the decreased expression level of12HSP70 was independent on the p53 activation. Strikingly, HSP70 was noted to be degraded by the proteasome since the bortezomib, the MG132, or the lactacystin were able to restore both the HSP70 expression level and intracellular localization in the cell. The lentiviral infection of depleted RPL11 cord blood CD34+ cells with a wild type HSP70 cDNA restored both the erythroid proliferation and differentiation, and reduced apoptosis, confirming a critical role for HSP70 in the erythroid defect in the RPL11+/Mut DBA phenotypes. The loss of HSP70 may explain the loss of GATA-1 in DBA and also the erythroid tropism of the DBA disease. Restoration of the HSP70 expression level may be a viable and novel therapeutic option for management of this debilitating and difficult to manage erythroid disorder

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22

Gross, Blaine Jeffrey. "1/f noise in MOSFETs with ultrathin gate dielectrics." Thesis, Massachusetts Institute of Technology, 1992. http://hdl.handle.net/1721.1/13192.

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Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Electrical Engineering and Computer Science, 1992.
Includes bibliographical references (p. 176-184).
by Blaine Jeffrey Gross.
Ph.D.

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23

Jayaraman, Rajsekhar. "Reliability and 1/f noise properties of MOSFETs with nitrided oxide gate dielectrics." Thesis, Massachusetts Institute of Technology, 1988. http://hdl.handle.net/1721.1/41582.

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Schrank, Martina. "Study on the regulation of mRNA expression of GABA transporters (GAT-1 and GAT-3) in rat brain." [S.l.] : [s.n.], 2000. http://ArchiMeD.uni-mainz.de/pub/2000/0097/diss.pdf.

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25

Kerkelä, R. (Risto). "Signaling pathways in myocyte hypertrophy:role of GATA4, mitogen-activated protein kinases and protein kinase C." Doctoral thesis, University of Oulu, 2003. http://urn.fi/urn:isbn:9514269950.

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Abstract Cardiac myocytes react to increased workload and hypertrophic neurohumoral stimuli by increasing protein synthesis, reinitiating expression of fetal forms of structural genes, α-skeletal actin (α-SkA) and β-myosin heavy chain (β-MHC), and by increasing expression and secretion of atrial natriuretic peptide (ANP) and B-type natriuretic peptide (BNP). Initially, the response is beneficial, but when prolonged, it leads to pathological cardiomyocyte hypertrophy. In this study, cardiomyocyte hypertrophy was initiated by hypertrophic agonists, endothelin-1 (ET-1) and phenylephrine (PE), and by increased stretching of atrial wall. Transcription factor GATA4 was studied to identify the mechanism leading to increased gene expression of BNP. In BNP promoter, GATA4 binds to cis elements mediating hypertrophic response. Eliminating GATA4 binding by using the decoy approach, basal BNP gene expression was reduced. To identify mechanisms regulating GATA4, the roles of mitogen-activated protein kinases (MAPKs) were studied. Activation of p38 MAPK increased GATA4 binding to BNP gene and led to increased GATA4 dependent BNP gene expression. p38 MAPK was required for ET-1 induced GATA4 binding, whereas extracellular signal-regulated kinase (ERK) was required for maintaining basal GATA4 binding activity. PE and ET-1 activated protein kinase C (PKC) signaling in cardiac myocytes. Antisense oligonucleotide inhibition of PKCα markedly reduced PE induced ANP secretion and ET-1 induced BNP secretion, whereas gene expression of natriuretic peptides was not affected. Antisense PKCα treatment inhibited PE induced expression of α-SkA, while increased protein synthesis or β-MHC gene expression were not affected. Sretching of the perfused rat atria increased BNP, c-fos and BNP gene expression via mechanism involving p38 MAP kinase activation of transcription factor Elk-1. In cultured neonatal rat atrial myocytes stretch induced BNP gene expression was dependent upon transcription factor Elk-1 binding sites within the BNP gene promoter. In conclusion, hypertrophic signaling in cardiac myocytes involves multiple signaling cascades. Activation of p38 MAPK is required for the development of ET-1 induced hypertrophic phenotype and GATA4 mediated BNP gene expression in cultured ventricular myocytes, and for stretch induced Elk-1 dependent BNP gene expression in atrial myocytes. PKCα is involved in PE induced hypertrophic response and PE induced switch in gene programming inducing expression of α-SkA, the fetal form of cardiac α-actin.

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26

Rosa, Rita Mourão. "O papel da alfa-1 glicoproteína ácida na monitorização clínica da gengivoestomatite crónica no gato : um estudo exploratório." Master's thesis, Universidade de Lisboa, Faculdade de Medicina Veterinária, 2018. http://hdl.handle.net/10400.5/15894.

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Dissertação de Mestrado Integrado em Medicina Veterinária
A alfa-1 glicoproteína ácida (AGP) é uma proteína de fase aguda cuja concentração sérica se encontra elevada nas doenças sistémicas nos gatos. O objetivo do presente estudo consistiu em determinar os níveis de AGP numa amostra de animais com gengivoestomatite crónica (GECF), comparar os mesmos com um grupo saudável e verificar a sua evolução em dois momentos pós-cirúrgicos (dia 30 e dia 60). Foram selecionados 20 gatos: 10 controlos e 10 doentes, sem co-morbilidades diagnosticadas. Procedeu-se ao doseamento da AGP sérica com recurso a um kit AGP-8 de ensaio de imunoabsorção enzimática (ELISA). Todos os gatos do grupo doentes apresentam lesões clínicas graves de mucosite caudal e estomatite, alterações histológicas de inflamação máxima, bem como positividade para a presença de antigénio para o calicivírus felino reforçando a homogeneidade deste grupo. Foi observado um aumento significativo da concentração sérica de AGP nos gatos afetados, confirmando que existe inflamação com impacto sistémico. Observou-se ainda uma correlação positiva, estatisticamente significativa, entre os valores de AGP e a presença de mucosite caudal no dia 0, a presença de estomatite nos dias 30 e 60. Este estudo exploratório sugere que este biomarcador poderá ser útil como fator de mau prognóstico do tratamento cirúrgico.
ABSTRACT - THE ROLE OF ALPHA-1 ACID GLYCOPROTEIN IN CLINICAL MONITORING OF CHRONIC GINGIVOSTOMATITIS IN CAT: AN EXPLORATORY STUDY - Alpha-1 acid glycoprotein (AGP) is an acute phase protein found to be high in systemic diseased cat’s. The objective of the present study was to determine AGP seric levels in a sample of animals with chronic gingivostomatitis (FCGS), compare those with a healthy group and evaluate their levels in 2 post-operative moments (30 and 60 days). Twenty cats were selected: 10 controls and 10 diseased, without diagnosed co-morbilities. Serum AGP was determined using an AGP-8 enzyme linked immunosorbent assay (ELISA) kit. In this study all diseased cats presented severe clinical lesions of caudal mucositis and bucostomatitis, histological findings of maximal inflammation, and positive isolation feline calicivirus (FCV) antigen, reinforcing the homogeneity of the group. The serum concentration of AGP is significantly increased in the FCGS group, confirming that these cats are in a systemic inflammatory state. A positive statistically significant association was found between AGP values and the presence of caudal mucositis and stomatitis lesions, in the pre-operative and post operative moment. This exploratory study suggests that this biomarker may be useful as poor prognostic fator of surgical treatment.
CIISA
N/A

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Dinallo, Heloíse Rangel. "Perfil das proteínas de fase aguda em gatos com doença do trato urinário inferior obstrutiva." Botucatu, 2019. http://hdl.handle.net/11449/191153.

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Orientador: Priscylla Tatiana Chalfun Guimarães Okamoto
Resumo: A doença do trato urinário inferior em felinos (DTUIF) apresenta diversos fatores etiológicos, sendo a forma obstrutiva a mais grave. As proteínas de fase aguda (PFAs) são biomarcadores utilizados para avaliar processos inflamatórios sistêmicos. A Alfa-1 Glicoproteína Ácida e o Amilóide A Sérico são PFAs positivas e major, o fibrinogênio é PFA minor e a albumina é negativa em gatos. O objetivo deste estudo é determinar as concentrações séricas das proteínas de fase aguda, Amilóide A sérico, Alfa-1 Glicoproteína Ácida, fibrinogênio e albumina e utilizá-las como biomarcadores de inflamação no monitoramento do processo inflamatório de gatos com doença do trato urinário inferior obstrutiva. Foram avaliados 25 gatos, machos, sem predileção de raça e idade, divididos em dois grupos experimentais, GC - grupo controle com oito gatos hígidos e GO - grupo obstruído com 17 gatos diagnosticados com DTUIF obstrutiva. Foram coletadas amostras para determinação das PFAs, bioquímica sérica, urinálise e UP/C nos M0, M12, M24 e M48 no GO e no GC somente no primeiro momento. As determinações das PFAs foram realizadas com os kits de ELISA para SAA, Kit Cat Serum Amyloid A Elisa (LIFE-SAA-8) e AGP, Kit Cat Alpha-1-Acid Glycoprotein Elisa (LIFE-AGP-8), ambos marca: Life Diagnostics®. No M0 houve correlações positivas de SAA, AGP e fibrinogênio com ureia e creatinina e correlação negativa de albumina com hematúria, SAA e potássio. No M48, houve correlações positivas entre SAA e AGP, AGP e ureia, fi... (Resumo completo, clicar acesso eletrônico abaixo)
Abstract: The Feline Lower Urinary Tract Disease (FLUTD) presents several etiologic factors, with the obstructive form representing the most severe. The acute phase proteins (APPs) are biomarkers used to evaluate systemic inflammatory processes. In cats, Alpha-1-Acid Glycoprotein and Serum Amyloid A are major positive APPs, fibrinogen is a minor APP and albumin is a negative APP. This study aims at determining the serum concentrations of acute phase proteins Serum Amyloid A, Alpha-1-Acid Glycoprotein, fibrinogen and albumin, in addition of using them as biomarkers of inflammation during the monitoring of the inflammatory processes of cats with obstructive feline lower urinary tract disease. A total of 25 male cats were recruited for the study irrespective of breed and age, and were divided into two experimental groups: the control group (CG), comprised of eight healthy cats; and the obstruction group (OG), comprised of 17 cats diagnosed with obstructive FLUTD. Samples were collected for APP analysis, serum biochemical assay, urinalysis and UP/C determination at M0, M12, M24 and M48 in the OG, and at M0 in the CG. The concentrations of the APPs were determined using commercially-available ELISA kits for SAA (Kit Cat Serum Amyloid A Elisa, LIFE-SAA-8) and AGP (Kit Cat Alpha-1-Acid Glycoprotein Elisa, LIFE-AGP-8) (Life Diagnostics®). At M0, there were positive correlations of SAA, AGP and fibrinogen with urea and creatinine, as well as negative correlations between albumin and hematuria, ... (Complete abstract click electronic access below)
Mestre

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28

Ipek, Hakan. "Modelling Of Resin Transfer Molding For Composites Manufacturing." Master's thesis, METU, 2005. http://etd.lib.metu.edu.tr/upload/12606815/index.pdf.

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The resin transfer molding (RTM ) process, in which a thermosetting resin is injected into a mold cavity preloaded with a porous fiber preform, is a manufacturing method for producing advanced continuous fiber reinforced composite products with complex geometries. Numerical simulation of resin transfer molding process is an often needed tool in manufacturing design, in order to analyze the process before the mold is constructed. In this study, a numerical simulation of the resin impregnation process in RTM of composite materials is performed by using and modifying an existing simulation program. The parts that are molded in the simulations have their planar dimensions much larger than their thicknesses. Therefore, the mold filling process can be modeled as two dimensional by neglecting the variations along the thickness direction. The program is capable of simulating two-dimensional, isothermal impregnation processes through orthotropic fiber preforms of planar but complex geometries. The formulations of the physical problem, used in this study, were taken from the theory of macroscopic flow through anisotropic porous media. The formulated governing equation and boundary conditions are solved in a regular-geometry computational domain by transformation through boundary fitted coordinate system. The discretization for numerical solution is performed by the finite difference method. The current study extends the existing capabilities of the simulation program by enabling the simulation of impregnation through non-homogeneous fiber preforms. Furthermore, the capability to simulate injection from two gates (as opposed to a single gate injection that existed before) is developed and added to the program. Various one-dimensional impregnation simulations (as parametric studies) are performed to assess the influence of process parameters. Results are also compared with analytical solutions and found to be in agreement with them. Two-dimensional impregnation simulations are performed for a planar, complex geometry mold. The two-dimensional results are compared with experimental results from the literature and are found to be in acceptable agreement with them. In addition to the study of various parametric variations in two-dimensional impregnation, double-gate resin injection simulations are performed and discussed as well.

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Humphrey, Peter Saah. "Signal transduction mechanisms for stem cell differentation into cardiomyocytes." Thesis, University of Hertfordshire, 2009. http://hdl.handle.net/2299/3760.

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Cardiovascular diseases are among the leading causes of death worldwide and particularly in the developed World. The search for new therapeutic approaches for improving the functions of the damaged heart is therefore a critical endeavour. Myocardial infarction, which can lead to heart failure, is associated with irreversible loss of functional cardiomyocytes. The loss of cardiomyocytes poses a major difficulty for treating the damaged heart since terminally differentiated cardiomyocytes have very limited regeneration potential. Currently, the only effective treatment for severe heart failure is heart transplantation but this option is limited by the acute shortage of donor hearts. The high incidence of heart diseases and the scarcity donor hearts underline the urgent need to find alternative therapeutic approaches for treating cardiovascular diseases. Pluripotent embryonic stem (ES) cells can differentiate into functional cardiomyocytes. Therefore the engraftment of ES cell-derived functional cardiomyocytes or cardiac progenitor cells into the damaged heart to regenerate healthy myocardial tissues may be used to treat damaged hearts. Stem cell-based therapy therefore holds a great potential as a very attractive alternative to heart transplant for treating heart failure and other cardiovascular diseases. A major obstacle to the realisation of stem cell-based therapy is the lack of donor cells and this in turn is due to the fact that, currently, the molecular mechanisms or the regulatory signal transduction mechanisms that are responsible for mediating ES cell differentiation into cardiomyocytes are not well understood. Overcoming this huge scientific challenge is absolutely necessary before the use of stem cell-derived cardiomyocytes to treat the damaged heart can become a reality. Therefore the aim of this thesis was to investigate the signal transduction pathways that are involved in the differentiation of stem cells into cardiomyocytes. The first objective was the establishment and use of cardiomyocyte differentiation models using H9c2 cells and P19 stem cells to accomplish the specific objectives of the thesis. The specific objectives of the thesis were, the investigation of the roles of (i) nitric oxide (ii) protein kinase C (PKC), (iii) p38 mitogen-activated protein kinase (p38 MAPK) (vi) phosphoinositide 3-kinase (PI3K) and (vi) nuclear factor-kappa B (NF-kB) signalling pathways in the differentiation of stem cells to cardiomyocytes and, more importantly, to identify where possible any points of convergence and potential cross-talk between pathways that may be critical for differentiation to occur. P19 cells were routinely cultured in alpha minimal essential medium (α-MEM) supplemented with 100 units/ml penicillin /100 μg/ml streptomycin and 10% foetal bovine serum (FBS). P19 cell differentiation was initiated by culturing the cells in microbiological plates in medium containing 0.8 % DMSO to form embryoid bodies (EB). This was followed by transfer of EBs to cell culture grade dishes after four days. H9c2 cells were cultured in Dulbecco’s Modified Eagle’s medium (DMEM) supplemented with 10% FBS. Differentiation was initiated by incubating the cells in medium containing 1% FBS. In both models, when drugs were employed, they were added to cells for one hour prior to initiating differentiation. Cell monolayers were monitored daily over a period of 12 or 14 days. H9c2 cells were monitored for morphological changes and P19 cells were monitored for beating cardiomyocytes. Lysates were generated in parallel for western blot analysis of changes in cardiac myosin heavy chain (MHC), ventricular myosin chain light chain 1(MLC-1v) or troponin I (cTnI) using specific monoclonal antibodies. H9c2 cells cultured in 1% serum underwent differentiation as shown by the timedependent formation of myotubes, accompanied by a parallel increase in expression of both MHC and MLC-1v. These changes were however not apparent until 4 to 6 days after growth arrest and increased with time, reaching a peak at day 12 to 14. P19 stem cells cultured in DMSO containing medium differentiated as shown by the timedependent appearance of beating cardiomyocytes and this was accompanied by the expression of cTnI. The differentiation of both P19 stem cells and H9c2 into cardiomyocytes was blocked by the PI3K inhibitor LY294002, PKC inhibitor BIM-I and the p38 MAPK inhibitor SB2035800. However when LY294002, BIM-I or SB2035800 were added after the initiation of DMSO-induced P19 stem cell differentiation, each inhibitor failed to block the cell differentiation into beating cardiomyocytes. The NF-kB activation inhibitor, CAPE, blocked H9c2 cell differentiation into cardiomyocytes. Fast nitric oxide releasing donors (SIN-1 and NOC-5) markedly delayed the onset of differentiation of H9c2 cells into cardiomyocytes while slow nitric oxide releasing donors (SNAP and NOC-18) were less effective in delaying the onset of differentiation or long term differentiation of H9c2 cells into cardiomyocytes. Akt (protein kinase B) is the key downstream target of PI3K. Our cross-talk data also showed that PKC inhibition and p38 MAPK inhibition respectively enhanced and reduced the activation of Akt, as determined by the phosphorylation of Akt at serine residue 473. In conclusion, PKC, PI3K, p38 MAPK and NF-kB are relevant for the differentiation of stem cells into cardiomyocytes. Our data also show that the PKC, PI3K and p38 MAPK signalling pathways are activated as very early events during the differentiation of stem cells into cardiomyocytes. Our data also suggest that PKC may negatively regulate Akt activation while p38 MAPK inhibition inhibits Akt activation. Our fast NO releasing donor data suggest that nitric oxide may negatively regulate H9c2 cell differentiation.

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Sakr, Rafael Lima. "A cláusula da nação mais favorecida na ordem econômica internacional: uma investigação sobre o discurso jurídico do artigo I: 1 do GATT." Universidade de São Paulo, 2010. http://www.teses.usp.br/teses/disponiveis/2/2135/tde-26092011-140858/.

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Como produto da prática mercantil, a cláusula da nação mais favorecida (CNMF) é um fenômeno jurídico complexo. Enquanto sua estrutura variante não é passível de padronização, por se adaptar às necessidades da sociedade internacional em cada momento histórico, seu núcleo funcional permanece imutável. Na ordem econômica internacional, a descentralização do poder político provoca desconfianças nos agentes econômicos, resultando em um permanente estado de guarda e competitividade predatória. Para assegurar maior estabilidade às expectativas normativas, os Estados celebram tratados, a fim de alterar tais percepções, conferindo durabilidade às relações econômicas internacionais. Resultado da configuração contemporânea da governança econômica internacional, a Organização Mundial do Comércio (OMC) simboliza a consolidação das expectativas normativas dos atores internacionais em torno do sistema multilateral de comércio (SMC). A OMC tem a função de consolidar o SMC, garantindo a posição de autoridade para corrigir as numerosas lacunas e antinomias jurídicas e reforçar a eficácia social, mediante a atuação de seu Órgão de Solução de Controvérsias (OSC). O SMC é um ordenamento jurídico, com lógica própria e princípios e regras específicos, que disciplina o mercado globalizado, cuja origem remonta à celebração do Acordo Geral sobre Tarifas e Comércio (GATT) em 1947. Prevista no artigo I:1 do GATT, a CNMF positiva o princípio da não discriminação, tendo por finalidade sistêmica desempenhar um papel dinâmico e integrado, ao: (i) assegurar transparência e dispersão de conhecimento; (ii) promover a cooperação internacional, a fim de eliminar ou reduzir, reciprocamente, as barreiras às trocas comerciais; (iii) vedar as práticas e instrumentos discriminatórios e protecionistas, tendo por função estender, automática, multilateral e incondicionalmente, as vantagens concedidas; e (iv) conservar as expectativas normativas, mediante a incorporação dos compromissos negociados ao SMC. Contudo, a proliferação de acordos preferenciais de comércio e de medidas protecionistas e discriminatórias pelos Estados-membros tem ameaçado o SMC de desautorização. Por recorrerem a exceções válidas à CNMF, esses fenômenos permitem a formação de relações discriminatórias e protecionistas, o que impacta negativamente as expectativas normativas dos agentes econômicos, ameaçando a função unificadora de sentido da CNMF, cujo resultado é a erosão da ideia de livre-mercado mundial. As reiteradas quebras de expectativas implicam problemas de coesão e eficácia normativa ao SMC, os quais são denominados desafios sistêmicos. Com efeito, o SMC sofre um processo de desestruturação, causado pela tensionada interação das dimensões ideacional e fática. Isso exige um controle de legalidade e de licitude dos atos jurídicos e das práticas dos Estados-membros. Em face desses desafios sistêmicos, a dissertação verifica se o artigo I:1 permanece como regra determinante para a decidibilidade do OSC. Para responder adequadamente, empregam-se os métodos analítico, hermenêutico e argumentativo, com um enfoque essencialmente dogmático, dentro de um ângulo crítico zetético. Ao fim da investigação, constata-se que a CNMF vem se consolidando como regra determinante para a construção do discurso jurídico-decisório pelo OSC. A confirmação jurisprudencial da imperatividade e da eficácia normativa do artigo I:1 reverbera reflexamente sobre os desafios sistêmicos, tendo o poderoso efeito de simbolizar a preferibilidade da incidência da CNMF sobre as relações econômicas internacionais.
As a product of commercial practice, the most-favored-nation clause (\"MFN\") is a complex legal phenomenon. While its variable structure is not subject to standardization, since it adapts to the needs of international society in each historical moment, its functional core remains unchanged. In the international economic order, the decentralization of political power leads to distrust of the economic agents, resulting in a permanent state of awareness and predatory competition. To ensure greater stability to the normative expectations, States enter into treaties in order to change such perceptions, providing durability to international economic relations. Result of the contemporary configuration of international economic governance, the World Trade Organization (\"WTO\") symbolizes the consolidation of the normative expectations of international actors around the multilateral trading system (\"MTS\"). The WTO has the mission of consolidating the MTS, ensuring a position of authority to correct the many shortcomings and antinomies of law and strengthen the social effectiveness through its Dispute Settlement Body (\"DSB\"). The MTS is a legal system, with its own logic and specific principles and rules, which regulates the globalized market, and has its origins in the General Agreement on Tariffs and Trade (GATT) in 1947. Set forth in Article I:1 of the GATT, the MFN establishes the principle of non-discrimination and has the systemic purpose of playing an integrated and dynamic role as it: (i) ensures transparency and dissemination of knowl edge,(ii) promotes international cooperation, by eliminating or reducing reciprocal barriers to trade, (iii) deters discriminatory and protectionist practices and instruments, being its function to extend, automatically, multilaterally and unconditionally, the benefits provided, and (iv) maintains the normative expectations, through the incorporation of negotiated concessions to the MTS. However, the proliferation of preferential trade agreements and protectionist and discriminatory measures by the member states has threatened the MTS of disempowerment. By resorting to MFNs valid exceptions, these phenomena allow the formation of discriminatory and protectionist relationships, which negatively impacts the normative expectations of economic agents, and threatening the harmonizing function of MFN; the result of which is the erosion of the global free market idea. Repeated breaches of expectations result in problems of cohesion and normative effectiveness of the MTS, which are called systemic challenges. Indeed, the MTS undergoes a process of disintegration, caused by the tensioned interaction of ideational and factual dimensions. This requires a control of legality and legitimacy of legal acts and practices of the member States. Given these systemic challenges, the dissertation verifies if Article I:1 remains the rule for determining the decidability of the DSB. In order to properly answer that, analytical, hermeneutic and argumentative methods are employed, with a primarily dogmatic focus, within a zetetic critical angle. By the end of the investigation, its stated that the MFN is becoming the consolidated rule for determining the construction of the legal and decision making discourse of the DSB. The confirmation from case law of the imperative nature and of the normative effectiveness of Article I:1 reverberates reflexively on the systemic challenges, having the powerful effect of symbolizing the desirability of MFN impact on international economic relations.

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31

Simões, Celina Bertelli. "Detecção do herpes vírus felino tipo 1 (HVF-1) pela técnica de PCR em tempo real e sua associação com sinais oculares em uma poipulação de gatos domésticos /." Araçatuba, 2013. http://hdl.handle.net/11449/128154.

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Resumo: O presente estudo buscou detectar o herpesvirus felino tipo 1 (HVF-1) em fragmentos de conjuntiva de uma população de gatos pela técnica de PCR em tempo real. Além disso, procurou-se associar estes resultados aos sinais oculares verificados nestes animais. Para isso, foram utilizados 70 gatos, que conviviam em contato direto, provenientes de uma residência da cidade de Araçatuba, SP. Por meio de PCR em tempo real, foi detectado DNA de HVF-1 em 78,1% (25/32) dos gatos com ao menos um sinal ocular e em 26,3% (10/38) dos assintomáticos, totalizando uma prevalência de 50% (35/70) na amostra global. Nos animais com sinais de conjuntivite, em 60% (21/35) dos gatos positivos havia ao menos um destes sinais e nenhum destes nos 40% (14/35) restantes. Nos gatos com sinais de ceratite, em 49% (17/35) dos positivos havia ao menos um destes sinais e nenhum deste nos 51% (18/35) restantes. Foi detectada a presença de HVF-1 em todos (17/100%) os gatos com defeito epitelial corneal. Houve associação significativa entre a presença de ao menos um sinal ocular, ao menos um sinal de conjuntivite e de ceratite com os resultados do PCR. Em relação a cada sinal ocular, somente o defeito epitelial corneal e o blefarospasmo tiveram associação significativa com estes resultados e também estavam associados entre si, sugerindo que, nos gatos com sinais de ceratoconjuntivite, o defeito epitelial corneal pode ser um fator de influência ao surgimento do blefarospasmo. A elevada prevalência da infecção ocular por HVF-1 encontrada nos animais com sinais oculares sugere o agente como possível causador destas lesões
Abstract: This study aimed to detect feline herpesvirus type 1 (FHV-1) in the conjunctival fragments of a cat population by PCR real-time. In addition, we sought to associate these results to ocular signs observed in these animals. For this, we used 70 cats that lived in direct contact, from a residence from Araçatuba, SP. By means of real-time PCR, DNA was detected FHV-1 in 78.1% (25/32) of cats with at least one ocular sign and 26.3% (10/38) of asymptomatic patients, a total prevalence 50% (35/70) in the sample. In animals with signs of conjunctivitis in 60% (21/35) cats were positive at least one of these signals and none of the 40% (14/35) remaining. In cats with signs of keratitis in 49% (17/35) were positive from at least one of the signals and none of the 51% (18/35) remaining. We have detected the presence of FHV-1 at all (17/100%) cats with corneal epithelial defect. There was a significant association between the presence of at least one eye sign, at least a sign of conjunctivitis and keratitis with PCR results. For each ocular sign, only the corneal epithelial defect and blepharospasm were significantly associated with these outcomes and also were associated with each other, suggesting that, in cats with signs of keratitis, corneal epithelial defect may be a factor influencing the emergence of blepharospasm. The high prevalence of ocular infection by FHV-1 found in animals with ocular signs suggested as a possible causative agent of these injuries
Orientador: Alexandre Lima de Andrade
Banca: Cristiane dos Santos Honsho
Banca: Flávia Resende Eugênio
Mestre

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32

Simões, Celina Bertelli [UNESP]. "Detecção do herpes vírus felino tipo 1 (HVF-1) pela técnica de PCR em tempo real e sua associação com sinais oculares em uma poipulação de gatos domésticos." Universidade Estadual Paulista (UNESP), 2013. http://hdl.handle.net/11449/128154.

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Made available in DSpace on 2015-10-06T13:03:35Z (GMT). No. of bitstreams: 0 Previous issue date: 2013-09-03. Added 1 bitstream(s) on 2015-10-06T13:18:21Z : No. of bitstreams: 1 000819068.pdf: 952868 bytes, checksum: 7410b5526acec730446d60283aed0de1 (MD5)
Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
O presente estudo buscou detectar o herpesvirus felino tipo 1 (HVF-1) em fragmentos de conjuntiva de uma população de gatos pela técnica de PCR em tempo real. Além disso, procurou-se associar estes resultados aos sinais oculares verificados nestes animais. Para isso, foram utilizados 70 gatos, que conviviam em contato direto, provenientes de uma residência da cidade de Araçatuba, SP. Por meio de PCR em tempo real, foi detectado DNA de HVF-1 em 78,1% (25/32) dos gatos com ao menos um sinal ocular e em 26,3% (10/38) dos assintomáticos, totalizando uma prevalência de 50% (35/70) na amostra global. Nos animais com sinais de conjuntivite, em 60% (21/35) dos gatos positivos havia ao menos um destes sinais e nenhum destes nos 40% (14/35) restantes. Nos gatos com sinais de ceratite, em 49% (17/35) dos positivos havia ao menos um destes sinais e nenhum deste nos 51% (18/35) restantes. Foi detectada a presença de HVF-1 em todos (17/100%) os gatos com defeito epitelial corneal. Houve associação significativa entre a presença de ao menos um sinal ocular, ao menos um sinal de conjuntivite e de ceratite com os resultados do PCR. Em relação a cada sinal ocular, somente o defeito epitelial corneal e o blefarospasmo tiveram associação significativa com estes resultados e também estavam associados entre si, sugerindo que, nos gatos com sinais de ceratoconjuntivite, o defeito epitelial corneal pode ser um fator de influência ao surgimento do blefarospasmo. A elevada prevalência da infecção ocular por HVF-1 encontrada nos animais com sinais oculares sugere o agente como possível causador destas lesões
This study aimed to detect feline herpesvirus type 1 (FHV-1) in the conjunctival fragments of a cat population by PCR real-time. In addition, we sought to associate these results to ocular signs observed in these animals. For this, we used 70 cats that lived in direct contact, from a residence from Araçatuba, SP. By means of real-time PCR, DNA was detected FHV-1 in 78.1% (25/32) of cats with at least one ocular sign and 26.3% (10/38) of asymptomatic patients, a total prevalence 50% (35/70) in the sample. In animals with signs of conjunctivitis in 60% (21/35) cats were positive at least one of these signals and none of the 40% (14/35) remaining. In cats with signs of keratitis in 49% (17/35) were positive from at least one of the signals and none of the 51% (18/35) remaining. We have detected the presence of FHV-1 at all (17/100%) cats with corneal epithelial defect. There was a significant association between the presence of at least one eye sign, at least a sign of conjunctivitis and keratitis with PCR results. For each ocular sign, only the corneal epithelial defect and blepharospasm were significantly associated with these outcomes and also were associated with each other, suggesting that, in cats with signs of keratitis, corneal epithelial defect may be a factor influencing the emergence of blepharospasm. The high prevalence of ocular infection by FHV-1 found in animals with ocular signs suggested as a possible causative agent of these injuries

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33

Turkcu, Ozlem. "Development Of An Electronic Attack (ea) System In Multi&amp." Master's thesis, METU, 2007. http://etd.lib.metu.edu.tr/upload/12609045/index.pdf.

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In this study, an expert system based EA and tracking system is developed and the performances of these systems are optimized. Tracking system consists of a monopulse tracking radar and a Multiple Hypothesis Tracking (MHT) algorithm. MHT is modelled as a measurement&
#8208
oriented approach, which is capable of initiating tracks. As each measurement is received, probabilities are calculated for the hypotheses and target states are estimated using a Kalman filter. Range Gate Pull-Off (RGPO) is selected as an EA technique to be developed because it is accepted to be the primary deception technique employed against tracking radar. Two modes of RGPO technique
linear and parabolic, according to time delay controller are modelled. Genetic Algorithm (GA) Toolbox of MATLAB is used for the optimization of these systems over some predetermined scenarios. It is observed that the performance of the tracking radar system is improved significantly and successful tracking is achieved over all given scenarios, even for closely spaced targets. RGPO models are developed against this improved tracking performance and deception of tracking radar is succeeded for all given target models.

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34

Müller, Felix. "Schutzmassnahmen gegen Warenimporte unter der Rechtsordnung der WTO : die materiell-rechtlichen Anwendungsvoraussetzungen der "Safeguard Measures" gem. Art. XIX:1(a) GATT 1994 und Art. 2.1 des Agreement on Safeguards /." Tübingen Mohr Siebeck, 2006. http://deposit.ddb.de/cgi-bin/dokserv?id=2816705&prov=M&dok_var=1&dok_ext=htm.

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35

Pereira, Marco Aurélio Amador. "Avaliação da eficácia analgésica e da inibição ex vivo da atividade das cicloxigenases 1 e 2 após o emprego da dipirona ou do meloxicam em gatas submetidas à ovariosalpingohisterectomia eletiva." Universidade de São Paulo, 2017. http://www.teses.usp.br/teses/disponiveis/10/10137/tde-13042018-163802/.

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Os AINE\'s são frequentemente empregados para o tratamento da dor aguda em gatos, porém, podem ser contraindicados pela propensão em causar efeitos adversos. A dipirona é um antigo analgésico não-opioide extensamente utilizado cujo mecanismo de ação ainda não foi completamente elucidado. O presente estudo prospectivo, randomizado e cego teve como objetivo avaliar o efeito analgésico e o mecanismo de ação via cicloxigenases (COX-1 e 2) da administração por via intravenosa (IV) de dipirona (12,5 mg/kg a cada 12 horas D12,5 ou 25 mg/kg a cada 24 horas D25) ou de meloxicam (0,1 mg/kg a cada 24 horas M) em gatas submetidas à ovariosalpingohisterectomia (OSH) eletiva. Trinta gatas (13 ± 5 meses e 2,7 ± 0,5 kg) foram avaliadas durante 24 horas após o início do tratamento a partir de ferramentas objetivas e subjetivas da dor. Medicação resgate com cloridrato de tramadol (2 mg/kg, IV) foi instituída quando escores ≥ 5 pela Escala Glasgow. A atividade das COX-1 e 2 foi avaliada a partir da mensuração das concentrações de tromboxano B2 (TXB2) e prostaglandina E2 (PGE2). Efeitos adversos foram registrados e exames laboratoriais, incluindo concentrações séricas de dimetilarginina simétrica (SDMA), foram realizados. As análises estatísticas foram efetuadas com o software GraphPad Prism versão 7.03. O grau de significância estabelecido para os testes foi de 5% (P < 0,05). Mudanças nos parâmetros fisiológicos cardiovasculares não foram clinicamente relevantes, porém as gatas do grupo M apresentaram aumento de frequência cardíaca em relação ao basal (P = 0,0331). A temperatura retal reduziu no momento T1h em todos os grupos (P = 0,0001). Houve um aumento da glicemia no momento T4h no grupo D25 (P = 0,0178) e em T1h no grupo M (P = 0,0205). Apesar de os escores de dor e sedação 9 não diferirem entre grupos, a escala analógica visual revelou aumento em T4h em relação ao basal no D12,5 (P = 0,0415) e os escores de sedação em T1h foram superiores ao basal em todos os tratamentos (P < 0,0001). Não houve diferença quanto ao resgate analgésico, porém duas gatas dos grupos D12,5 (20%) e M (20%) e quatro do D25 necessitaram de medicação resgate. As concentrações de TXB2 foram superiores no grupo M em relação ao D12,5 e D25 em T4h (P = 0,0032 e P < 0,0001, respectivamente) e T24h (P = 0,0070 e 0,0111, respectivamente). Houve redução muito significativa em T1/2h, T4h e T24h quando comparados ao T0h em todos os grupos (P < 0.0001) e ocorreu aumento entre T1/2h e T4h no grupo M (P = 0,0004). As concentrações de PGE2 estimulada por lipopolisacarídeos (LPS) foram superiores em D25 em relação ao M em T4h (P = 0,0479). No grupo D12,5, em T1/2h, esta foram inferiores as de T0h (P = 0,0001) e T4h (P = 0,0112). O mesmo ocorreu no grupo D25 em T0h, T4h e T24h (P < 0,0001, P = 0,001 e 0,0004, respectivamente) enquanto que no M, os momentos T1/2h e T4h apresentaram valores inferiores ao T0h (P = 0,0016 e 0,0075). As concentrações séricas de SDMA do grupo D25 reduziram em T24h quando comparadas as de T0h (P = 0,0322), porém apenas uma gata do grupo M apresentou concentração acima do limite para a espécie. A partir dos resultados observados conclui-se que os protocolos analgésicos instituídos foram efetivos para o controle da dor pós-operatória neste contexto, apresentando inibição não seletiva COX-2 sem causar efeitos adversos e alterações hematológicas, na atividade das enzimas hepáticas e na taxa de filtração glomerular.
NSAIDs are often used for treatment of acute pain in cats, but it may be contraindicated for propensity to cause adverse effects. Dipyrone ia a widely used non-opioid analgesic whose mechanism of action has not yet been fully elucidated. The present prospective, randomized, blind study aimed to evaluate the analgesic effect and mechanism of action of inhibition of cycloxigenases (COX-1 and 2) of intravenous (IV) administration of dipyrone (25 mg/kg q 24 hours or 12.5 mg/kg q 12 hours) or meloxicam (0.1 mg/kg q 24 hours) in cats underwent elective ovariohysterectomy. Thirty cats (13 ± 5 months and 2,7 ± 0,5 kg) were evaluated for 24 hours after surgical procedure using objective and subjective pain tools. Rescue medication with tramadol hydrochloride (2 mg/kg IV) was administrated when scores ≥ 5 by the Glasgow scale. The activity of COX-1 and 2 was assessed by measuring the concentrations of thromboxane B2 (TXB2) and prostaglandin E2 (PGE2). Adverse effects were recorded and laboratory tests, including serum concentrations of symmetrical dimethylarginine (SDMA), were performed. Data was analyzed with GraphPad Prism version 7.03. Values of P < 0.05 were considered significant. Changes in cardiovascular parameters were not clinically relevant, but the M group presented higher heart rate than basal (P = 0.0331). The rectal temperature reduced at time T1h in all groups (P = 0.0001). There was an increase in blood glucose at time T4h in group D25 (P = 0.0178) and in T1h in group M (P = 0.0205). Although pain and sedation scores did not differ between groups, the visual analogue scale 11 showed an increase in T4h over baseline in D12.5 (P = 0.0415) and sedation scores in T1h were higher than baseline in all treatments (P < 0.0001). There was no difference in the analgesic rescue, but two cats of D12.5 (20%) and M (20%) groups and four from the D25 required rescue medication. The concentrations of TXB2 were higher in M group compared to D12.5 and D25 at T4h (P = 0.0032 and P < 0.0001, respectively) and T24h (P = 0.0070 and 0.0111, respectively) and there was a very significant reduction in T1/2h, T4h and T24h when compared to T0h in all groups (P < 0.0001) and there was an increase between T1/2h and T4h in group M (P = 0.0004). Concentrations of lipopolysaccharide-stimulated PGE2 (LPS) were higher in D25 compared to M in T4h (P = 0.0479). Those in the D12.5 group, in T1/2h, were lower than in T0h (P = 0.0001) and T4h (P = 0.0112). The same occurred in group D25 at T0h, T4h and T24h (P <0.0001, P = 0.001 and 0.0004, respectively) whereas in M, T1/2h and T4h moments presented values lower than T0h (P = 0.0016 and 0.0075). Serum concentrations of SDMA of D25 group decreased in T24h when compared to T0h (P = 0.0322) and only one cat (group M) showed serum concentration above the feline cut-off. In conclusion, the analgesic protocols were effective for the control of postoperative pain in this context, presenting COX-2 non-selective inhibition without causing adverse effects and hematological, liver enzyme activity and glomerular filtration rate alterations.

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36

Dong, Quan. "HEMTs cryogéniques à faible puissance dissipée et à bas bruit." Thesis, Paris 11, 2013. http://www.theses.fr/2013PA112035.

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Les transistors ayant un faible niveau de bruit à basse fréquence, une faible puissance de dissipation et fonctionnant à basse température (≤ 4.2 K) sont actuellement inexistants alors qu’ils sont très demandés pour la réalisation de préamplificateurs à installer au plus près des détecteurs ou des dispositifs à la température de quelques dizaines de mK, dans le domaine de l’astrophysique, de la physique mésoscopique et de l’électronique spatiale. Une recherche menée depuis de nombreuses années au LPN vise à réaliser une nouvelle génération de HEMTs (High Electron Mobility Transistors) cryogéniques à haute performance pour répondre à ces demandes. Cette thèse, dans le cadre d’une collaboration entre le CNRS/LPN et le CEA/IRFU, a pour but la réalisation de préamplificateurs cryogéniques pour des microcalorimètres à 50 mK.Les travaux de cette thèse consistent en des caractérisations systématiques des paramètres électriques et des bruits des HEMTs (fabriqués au LPN) à basse température. En se basant sur les résultats expérimentaux, l’une des sources de bruit à basse fréquence dans les HEMTs a pu être identifiée, c’est-à-dire la part du courant tunnel séquentiel dans le courant de fuite de grille. Grâce à ce résultat, les hétérostructures ont été optimisées pour minimiser le courant de fuite de grille ainsi que le niveau de bruit à basse fréquence. Au cours de cette thèse, différentes méthodes spécifiques ont été développées pour mesurer de très faibles valeurs de courant de fuite de grille, les capacités du transistor et le bruit 1/f du transistor avec une très haute impédance d’entrée. Deux relations expérimentales ont été observées, l’une sur le bruit 1/f et l’autre sur le bruit blanc dans ces HEMTs à 4.2 K. Des avancées notables ont été réalisées, à titre d’indication, les HEMTs avec une capacité de grille de 92 pF et une consommation de 100 µW peuvent atteindre un niveau de bruit en tension de 6.3 nV/√Hz à 1 Hz, un niveau de bruit blanc de 0.2 nV/√Hz et un niveau de bruit en courant de 50 aA/√Hz à 10 Hz. Enfin, une série de 400 HEMTs, qui répondent pleinement aux spécifications demandées pour la réalisation de préamplificateurs au CEA/IRFU, a été réalisée. Les résultats de cette thèse constitueront une base solide pour une meilleure compréhension du bruit 1/f et du bruit blanc dans les HEMTs cryogéniques afin de les améliorer pour les diverses applications envisagées
Transistors with low noise level at low frequency, low-power dissipation and operating at low temperature (≤ 4.2 K) are currently non-existent, however, they are widely required for realizing cryogenic preamplifiers which can be installed close to sensors or devices at a temperature of few tens of mK, in astrophysics, mesoscopic physics and space electronics. Research conducted over many years at LPN aims to a new generation of high-performance cryogenic HEMTs (High Electron Mobility Transistors) to meet these needs. This thesis, through the collaboration between the CNRS/LPN and the CEA/IRFU, aims for the realization of cryogenic preamplifiers for microcalorimeters at 50 mK.The work of this thesis consists of systematic characterizations of electrical and noise parameters of the HEMTs (fabricated at LPN) at low temperatures. Based on the experimental results, one of the low-frequency-noise sources in the HEMTs has been identified, i.e., the sequential tunneling part in the gate leakage current. Thanks to this result, heterostructures have been optimized to minimize the gate leakage current and the low frequency noise. During this thesis, specific methods have been developed to measure very low-gate-leakage-current values, transistor’s capacitances and the 1/f noise with a very high input impedance. Two experimental relationships have been observed, one for the 1/f noise and other for the white noise in these HEMTs at 4.2 K. Significant advances have been made, for information, the HEMTs with a gate capacitance of 92 pF and a consumption of 100 µW can reach a noise voltage of 6.3 nV/√ Hz at 1 Hz, a white noise voltage of 0.2 nV/√ Hz, and a noise current of 50 aA/√Hz at 10 Hz. Finally, a series of 400 HEMTs has been realized which fully meet the specifications required for realizing preamplifiers at CEA/IRFU. The results of this thesis will provide a solid base for a better understanding of 1/f noise and white noise in cryogenic HEMTs with the objective to improve them for various considered applications

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37

von, Haartman Martin. "Low-frequency noise characterization, evaluation and modeling of advanced Si- and SiGe-based CMOS transistors." Doctoral thesis, KTH, Mikroelektronik och Informationsteknik, IMIT, 2006. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-3888.

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A wide variety of novel complementary-metal-oxide-semiconductor (CMOS) devices that are strong contenders for future high-speed and low-noise RF circuits have been evaluated by means of static electrical measurements and low-frequency noise characterizations in this thesis. These novel field-effect transistors (FETs) include (i) compressively strained SiGe channel pMOSFETs, (ii) tensile strained Si nMOSFETs, (iii) MOSFETs with high-k gate dielectrics, (iv) metal gate and (v) silicon-on-insulator (SOI) devices. The low-frequency noise was comprehensively characterized for different types of operating conditions where the gate and bulk terminal voltages were varied. Detailed studies were made of the relationship between the 1/f noise and the device architecture, strain, device geometry, location of the conduction path, surface cleaning, gate oxide charges and traps, water vapour annealing, carrier mobility and other technological factors. The locations of the dominant noise sources as well as their physical mechanisms were investigated. Model parameters and physical properties were extracted and compared. Several important new insights and refinements of the existing 1/f noise theories and models were also suggested and analyzed. The continuing trend of miniaturizing device sizes and building devices with more advanced architectures and complex materials can lead to escalating 1/f noise levels, which degrades the signal-to-noise (SNR) ratio in electronic circuits. For example, the 1/f noise of some critical transistors in a radio receiver may ultimately limit the information capacity of the communication system. Therefore, analyzing electronic devices in order to control and find ways to diminish the 1/f noise is a very important and challenging research subject. We present compelling evidence that the 1/f noise is affected by the distance of the conduction channel from the gate oxide/semiconductor substrate interface, or alternatively the vertical electric field pushing the carriers towards the gate oxide. The location of the conduction channel can be varied by the voltage on the bulk and gate terminals as well by device engineering. Devices with a buried channel architecture such as buried SiGe channel pMOSFETs and accumulation mode MOSFETs on SOI show significantly reduced 1/f noise. The same observation is made when the substrate/source junction is forward biased which decreases the vertical electric field in the channel and increases the inversion layer separation from the gate oxide interface. A 1/f noise model based on mobility fluctuations originating from the scattering of electrons with phonons or surface roughness was proposed. Materials with a high dielectric constant (high-k) is necessary to replace the conventional SiO2 as gate dielectrics in the future in order to maintain a low leakage current at the same time as the capacitance of the gate dielectrics is scaled up. In this work, we have made some of the very first examinations of 1/f noise in MOSFETs with high-k structures composed by layers of HfO2, HfAlOx and Al2O3. The 1/f noise level was found to be elevated (up to 3 orders of magnitude) in the MOSFETs with high-k gate dielectrics compared to the reference devices with SiO2. The reason behind the higher 1/f noise is a high density of traps in the high-k stacks and increased mobility fluctuation noise, the latter possibly due to noise generation in the electron-phonon scattering that originates from remote phonon modes in the high-k. The combination of a TiN metal gate, HfAlOx and a compressively strained surface SiGe channel was found to be superior in terms of both high mobility and low 1/f noise.
QC 20100928

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38

Böttner, Stefan [Verfasser], Wolfgang [Akademischer Betreuer] Dröge-Laser, and Christiane [Akademischer Betreuer] Gatz. "Funktionale Bedeutung der Protein-Protein-Interaktion zwischen dem Tabak-Ankyrin-Repeat-Protein ANK1 und dem bZIP-Transkriptionsfaktor BZI-1 im Rahmen der pflanzlichen Auxin- und Pathogenantwort / Stefan Böttner. Gutachter: Wolfgang Dröge-Laser ; Christiane Gatz. Betreuer: Wolfgang Dröge-Laser." Göttingen : Niedersächsische Staats- und Universitätsbibliothek Göttingen, 2008. http://d-nb.info/1042842205/34.

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Lorusso, Marco. "FPGA implementation of muon momentum assignment with machine learning at the CMS level-1 trigger." Master's thesis, Alma Mater Studiorum - Università di Bologna, 2021. http://amslaurea.unibo.it/23211/.

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With the advent of the High-Luminosity phase of the LHC (HL-LHC), the instantaneous luminosity of the Large Hadron Collider at CERN is expected to increase up to around 7.5 x 10^34 cm^-2s^-1. Therefore, new strategies for data acquisition and processing will be necessary, in preparation for the higher number of signals produced inside the detectors, that would eventually make the trigger and readout electronics currently in use at the LHC experiments obsolete. In the context of an upgrade of the trigger system of the Compact Muon Solenoid (CMS), new reconstruction algorithms, aiming for an improved performance, are being developed. For what concerns the online tracking of muons, one of the figures that is being improved is the accuracy of the transverse momentum (pT) measurement. Machine Learning techniques have already been considered as a promising solution for this problem, as they make possible, with the use of more information collected by the detector, to build models able to predict the pT with an improved precision. In this Master Thesis, a step further in increasing the performance of the pT assignment is taken by implementing such models onto a type of programmable processing unit called Field Programmable Gate Array (FPGA). FPGAs, indeed, allow a smaller latency with a relatively small loss in accuracy with respect to traditional inference algorithms running on a CPU, both important aspects for a trigger system. The analysis carried out in this work uses data obtained through Monte Carlo simulations of muons crossing the barrel region of the CMS muon chambers, and compare the results with the pT assigned by the current (Phase-1) CMS Level 1 Barrel Muon Track Finder (BMTF) trigger system. Together with the final results, the steps needed to create an accelerated inference machine for muon pT are also presented.

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Roth, Claude. "Analyse du repertoire des cellules b impliquees dans la reponse au terpolymer poly-(acide glutamique#6#0-alanine#3#0-tyrosine#1#0) gat et contribution de la lymphokine "b cell activating factor" bcaf dans l'activation precoce du lymphocyte b." Paris 7, 1988. http://www.theses.fr/1988PA077203.

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Etude du repertoire des lymphocytes b specifiques de l'antigene gat et de la contribution des differents segments genetiques codant pour les anticorps dans la reconnaissance de l'antigene et l'expression des idiotypes. Etude des signaux delivres par les lymphocytes t helper. Preparation de la lymphokine bcaf et demonstration de son activite dans le processus d'activation et de croissance des cellules b

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Lawson, Michael Nunes. "A reclamação de não violação no GATT/OMC." reponame:Biblioteca Digital de Teses e Dissertações da UFRGS, 2009. http://hdl.handle.net/10183/18273.

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A presente dissertação objetiva contribuir para a compreensão do instituto da reclamação de não violação, previsto no art. XXIII:1(b) do Acordo General de Tarifas e Comércio (GATT) e art. 26:1 do Entendimento sobre Solução de Controvérsias (ESC) da Organização Mundial do Comércio (OMC). Parte-se da premissa de que a reclamação de não violação deve ser abordada à luz da sua contraparte, a reclamação de violação (art. XXIII:1(a) do GATT). O estudo é levado a cabo recorrendo-se a conceitos desenvolvidos em direito internacional geral. A reclamação de violação conduz a responsabilidade por atos internacionalmente ilícitos, enquanto a reclamação de não violação conduz a responsabilidade por atos não proibidos pelo direito internacional (do comércio). O contraste entre as duas reclamações manifesta-se, ademais, no que tange ao regramento respectivo, especialmente no que diz com os remédios jurídicos disponíveis. A investigação é completada pela análise da jurisprudência do GATT/OMC que cuidou da reclamação de não violação, essencial face à amplitude da terminologia do art. XXIII:1(b). Assim, examina-se a interpretação conferida ao art. XXIII:1, caput - benefício, princípio da expectativa legítima (implícito), anulação ou prejuízo e nexo de causalidade -; art. XXIII:1(b) - medida -; por fim, art. XXIII:2, que versa sobre remédios jurídicos.
The present dissertation seeks to contribute to the understanding of the non-violation complaint, regulated in GATT art. XXIII:1(b) and WTO's DSU art. 26:1. It departs from the premise that the non-violation complaint must be approached in light of its counterpart, the violation complaint (GATT art. XXIII:1(a)). The study is carried on with resort to concepts developed in general international law. The violation complaint leads to responsibility for internationally wrongful acts, whilst the non-violation complaint leads to liability for acts not prohibited by international (trade) law. The contrast between the two complaints is manifested, in addition, with regard to their respective disciplines, particularly the remedies available. The investigation is completed by the analysis of the GATT/WTO case-law which dealt with the non-violation complaint, essential in view of the breadth of the terminology contained in art. XXIII:1(b). In this sense, it is examined the interpretation of art. XXIII:1, caput - benefit, the principle of legitimate expectations (implicit), nullification or impairment, nexus of causality -; art. XXIII:1(b) - measure -; lastly, art. XXIII:2, which concerns itself of remedies.

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ROZA, Marcello Rodrigues da. "Tomografia computadorizada de feixe cônico na odontologia de cães e gatos." Universidade Federal de Goiás, 2009. http://repositorio.bc.ufg.br/tede/handle/tde/1187.

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Made available in DSpace on 2014-07-29T15:13:52Z (GMT). No. of bitstreams: 1 Tese_Marcelo_Roza.pdf: 1255109 bytes, checksum: 0fed736dcde7d2550d7d1cd31a009acb (MD5) Previous issue date: 2009-06-22
Eleven dogs and four cats with buccodental alterations, treated in the Centro Veterinário do Gama, in Brasilia, DF, Brazil, were submitted to cone beam computed tomography. The exams were carried out in a i-CAT tomograph, using for the images acquisition six centimeters height, 40 seconds time, 0,2 voxel, 120 kilovolts (kV) and 46,72 milliampéres per second (mAs). The ideal positioning of the animal for the exam was also determined in this study and it proved to be fundamental for performing the exam, which required a simple and safe anesthetic protocol due to the relatively short time necessary in obtaining the images. Many alterations and diseases had been identified, with extreme accuracy of the images, becoming the cone beam computed tomography as a safe, accessible and feasible imaging method which could be included in the small animals dentistry routine diagnosis
A tomografia computadorizada é uma modalidade de diagnóstico utilizada na odontologia desde a década de 90, que apresenta uma série de vantagens sobre outros exames de imagem, como menor exposição à radiação ionizante, menor tempo na aquisição das imagens, dispensa do processamento químico e menor custo. O exame tomográfico de feixe cônico é empregado na odontologia humana, mas não foram encontrados na literatura consultada trabalhos científicos sobre o assunto. Este estudo teve por objetivo padronizar a tomografia de feixe cônico visando sua utilização na rotina da clínica veterinária de pequenos animais, utilizando pacientes atendidos na rotina clinica odontológica. Na padronização desenvolveu-se primeiramente um dispositivo que possibilitou o posicionamento dos pacientes no tomógrafo de feixe cônico e permitiu a aquisição das imagens de forma correta e sem repetições. Na seqüência estudaram-se os aspectos morfofuncionais da cabeça em cães e gatos e, num segundo momento avaliaram-se alterações dentárias em cães e gatos e temporomandibulares em gatos. O dispositivo desenvolvido e o exame tomográfico de feixe cônico mostraram-se apropriados para avaliar alterações buco dentarias em cães e gatos, podendo ser recomendado para essa finalidade diagnóstica

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Kramoliš, Ondřej. "Extrakce a modifikace vlastností číslicových zvukových signálů v dynamické rovině." Master's thesis, Vysoké učení technické v Brně. Fakulta elektrotechniky a komunikačních technologií, 2010. http://www.nusl.cz/ntk/nusl-218782.

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This thesis deals with basic methods of root mean square and peak value measurement of a digital acoustic signal, algotithms to measure audio programme loudness and true-peak audio level according to recommendation ITU-R BS.1770-1 and digital systems for control of signal dynamic range. It shows achieved results of root mean square and peak value measurement and results of implementation of dynamic processor with general piecewise linear non-decreasing static curve and algorithms according to recommendation ITU-R BS.1770-1.

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Burda, Pavel. "Role transkripčních faktorů PU.1 a GATA-1 v leukemické diferenciaci." Doctoral thesis, 2011. http://www.nusl.cz/ntk/nusl-297741.

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Hematopoiesis is coordinated by a complex regulatory network of transcription factors among them PU.1 (Spi1, Sfpi1) and GATA-1 represent key molecules. GATA-1 and PU.1 bind each other on DNA to block each others transcriptional programs to prevent development of undesired lineage during hematopoietic commitment. Murine erythroleukemia (MEL) cells, transformed erythroid precursors that are blocked from completing the late stages of erythroid differentiation, co-express GATA-1 and PU.1 and as my and others data document, are able to respond to molecular removal (down-regulation) of PU.1 or addition (up-regulation) of GATA-1 by inducing terminal erythroid differentiation. We provide novel evidence that downregulation of GATA-1 or upregulation of PU.1 induces incompletely differentiation into cell cycle arrested monocytic-like cells. Furthermore, PU.1- dependent transcriptome is negatively regulated by GATA-1 in MEL cells, including CCAAT/enhancer binding protein alpha (Cebpa) and Core-binding factor, beta subunit (Cbfb) that encode additional key hematopoietic transcription factors. Chromatin immunoprecipitation and reporter assays identified PU.1 motif sequences near Cebpa and Cbfb that are co-occupied by PU.1 and GATA-1 in the leukemic blasts. Furthermore, transcriptional regulation of these loci by...

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Eisbacher, Michael. "The regulation of megakaryocyte-specific genes by Fli-1 and GATA-1 /." 2003. http://www.library.unsw.edu.au/~thesis/adt-NUN/public/adt-NUN20040206.105222/index.html.

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Pan, Shu St George Clinical School UNSW. "Functional studies of transcription factors GATA-1, Fli-1 and FOG-1 in Megakaryocyte development." 2007. http://handle.unsw.edu.au/1959.4/40591.

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Transcription factors GATA-1, Fli-1 and FOG-1 are essential proteins for normal megakaryopoiesis, however, the detailed analyses of their functions within developmental stages of megakaryopoiesis are lacking. In my thesis, over expression of gene in target cells was adopted as the main strategy to study the biological functions of these proteins, therefore, an efficient gene delivery method was first developed by using retrovirus.This approach was then utilized to over express GATA-1, Fli-1 and FOG-1 in murine leukemia M1 cells and mouse hematopoietic stem cells (HSCs), and their effects on different developmental stages of megakaryopoiesis were investigated. In the transduced M1 cells, enforced expression of GATA-1 and Fli-1 was found to induce the megakaryocytic development, which was associated with the formation of megakaryocyte (Mk) and the increased expression of Mk specific genes c-Mpl and GPIX. In the transduced mouse HSCs, it was found that the expression of endogenous GATA-1, Fli-1 and FOG-1 was up-regulated throughout Mk differentiation; enforced expression of these transcription factors led to the significantly enhanced Mk development. Megakaryocytes over expressing GATA-1, Fli-1 and FOG-1 were characterized by the increased expression of various Mk-specific genes including GPIX, c-Mpl, platelet factor 4 (PF4), acetylcholinesterase (AChE) and NF-E2, an important transcription factor for terminal megakaryopoiesis; however, GATA-1, Fli-1 and FOG-1 displayed the different abilities in promoting the proliferation of hematopoietic cells and MK differentiation, as well as regulating other transcription factors involved in hematopoiesis. To further elucidate the role of the functional domains of Fli-1, various mutants of Fli-1 were also over expressed in mouse HSCs. The results demonstrated that first, the combination of the activation domain of Fli-1 and its Ets domain is required for early megakaryopoiesis but not sufficient for terminal megakaryopoiesis; second, DNA binding of Fli-1 was not the only requirement for early Mk enhancement, moreover, the interaction between Fli-1 and GATA-1 through the Ets domain and the resultant transcriptional synergy was the essential determinant for Fli?1 ability in Mk development. Taken together, the studies presented in this thesis provided strong in vitro evidence that GATA-1, Fli-1 and FOG-1 indeed play the critical roles in normal megakaryopoiesis.

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Lamoureux, Lise Marie. "Regulation of calreticulin gene by bHLH and GATA-1 proteins." 2004. http://hdl.handle.net/1993/16296.

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Ciou, Jyuan-Kai, and 邱雋凱. "Hif-1α Regulates GATA-1/2 Switch at Zebrafish Primitive Erythropoiesis." Thesis, 2015. http://ndltd.ncl.edu.tw/handle/80803236165232099473.

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碩士
國立臺灣海洋大學
生命科學暨生物科技學系
103
During erythropoiesis, transcription factor GATA2 is ciritcal for the survival and proliferation of immature hematopoietic stem cells and GATA1 is the key transcription factor required for the survival of erythroid progenitors and terminal differentiation of erythroid cells. .. After commitment, the GATA factor that regulates erythroid differentiation is switched from GATA2 to GATA1, a process referred to as GATA factor switching. a process referred to as GATA factor switching. As a result, the primitive erythroid progenitors differentiate into mature erythrocytes after the onset of blood circulation at 24 hpf. In adult vertebrates, the erythroid cell production can be increased under hypoxic stress by elevating EPO expression and enhancing iron utilization via the actions of Hypoxia Inducible Factors (HIFs). Previously it was demonstrated that depletion of HIF1α abrogated gata1expression in zebrafish embryos and blocked primitive erythropoiesis. Here I demonstrated that HIF1α is required for GATA factor switching during erythropoiesis. Knockdown HIF1α translation inhibited gata1 expression and sustained high level of gata2 transcription. Ectopic gata1 mRNA diminished gata2 transcription and rescued primitive erythropoiesis. It suggested that HIF1α plays critical role in GATA factor switching during erythrocyte maturation. This is the first time to demonstrate that HIF1α is directly involved in erythrocyte differentiation by controling GATA factor switching. It will help us to undestand the mechanism of hypoxia-mediated enhancement of erythrocyte proliferation.

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Chen, Chung-Te, and 陳俊德. "Negative regulation of the persistence-associated gene 1 of Hz-1 virus by GATA cis-element." Thesis, 1999. http://ndltd.ncl.edu.tw/handle/31069855981758098939.

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碩士
國立成功大學
生物學系
87
ABSTRACT Temporal viral gene expression has been reported in Hz-1 virus. During its persistent infection, the persistent-associated gene (pag1) is actively expressed, while rests of the viral genes are shut down. Previous studies have shown that the promoter regions from nucleotide -312 to -212 and from -158 to -90 have negative regulatory effects on pag1 gene expression. Here, We confirm the negative regulatory effect of GATA cis-element in these two regions by a substitution of 4 core nucleotides in the GATA-1 binding motif of pag-312/-90 fragment. Further confirmation of the effect of GATA element in pag-158/-90 and pag-107/+29 fragments were carried out. Furthermore, our results showed that the binding of GATA-1 like proteins on the GATA cis-element negatively regulates the PAT1 transcription rate. It has been reported that an intragenic pag1 promoter is lied from nucleotide +9 to +29. A set of mutants with tetranucleotide substitution within the fragment were constructed to test their effects on promoter activity. Results showed that none of the mutant abolished the promoter activity. From the GCG computational analysis, we found that the increase of pag1 promoter activity may be due to the introduction of Hind III cloning site which generates a C/EBP binding motif. In conclusion, the expression of pag1 is regulated by its extragenic promoter. Two GATA cis-elements within nuclotide -312/-90 region are responsible for the negative regulation of pag1 expression by regulation

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Lee, Jia-Han, and 李佳翰. "Studies of the expression and functional significance of GATA-1 in osteogenesis." Thesis, 2011. http://ndltd.ncl.edu.tw/handle/99237656141758425947.

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碩士
國立陽明大學
生化暨分子生物研究所
99
GATA-1 is a key transcription factor essential for erythroid and megakaryocyte development. Previous studies have shown that mice with low expression levels of GATA-1 (GATA-1low) exhibits increased bone densities due to paracrine actions from increased megakaryocyte precursors. Using osteogenic model cell systems, we have earlier demonstrated that GATA-1 could directly influence osteogenesis by interacting with Runx2 and inhibiting osteocalcin promoter activity. In this study, we investigated the effect of endogenous GATA-1 on osteogenesis using mesenchymal stem cells derived from bone marrow of GATA-1low mice. The cells were induced for osteoblast differentiation by treatment with BMP-2, analysis of alkaline phosphatase expression showed that depletion of GATA-1 accelerated osteoblastic differentiation, supporting that GATA-1 may be directly involved in osteogenesis. Examination of the 3’-UTR region of GATA-1 revealed the presence of putative binding sites for let-7 microRNAs, the expression of which has been shown to be elevated during osteogenesis. Consistent with this finding, we showed that the mRNA levels of Lin28, a negative regulator of let-7 microRNAs, decreased during induced osteogenesis. Using the reporter plasmids containing the wild-type and the let-7 site-deleted 3’-UTR of GATA-1 ligated downstream to the luciferase gene, we further show that overexpression of let-7 microRNAs significantly inhibited the luciferase activity induced by the wild-type, but not the let-7 site-deleted reporter construct. Together our finding support that GATA-1 may function as a negative regulator during the induced osteogenesis of mesenchymal stem cells, and suggest a possibility that the expression of GATA-1 may be suppressed via Lin28/let-7 microRNA axis.

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Dissertations / Theses on the topic 'GATA-1'

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