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Walsh, Ciara. "The regulation of STIM1 translocation to the plasma membrane." Thesis, University of Liverpool, 2010. http://livrepository.liverpool.ac.uk/1482/.
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A rise in intracellular Ca2+ concentration is key to controlling both short term and long term Ca2+ dependent processes which include secretion, metabolism and gene expression, cell growth and proliferation. Store operated Ca2+ channels (SOCs), which are activated by the depletion of Ca2+ from internal Ca2+ stores, the main store being the endoplasmic reticulum (ER), are the major route for Ca2+ influx in non-excitable cell types. Stromal interacting molecule 1 (STIM1) is a Ca2+ sensing protein located in the endoplasmic reticulum (ER). Depletion of ER calcium stores triggers oligomerisation and subsequent translocation of STIM1 from its reticular location to specialized endoplasmic reticulum-plasma membrane (ER-PM) junctions where it forms STIM1 puncta and interacts with the SOC channel, Orai1. This induces the clustering of Orai1 into a functional tetrameric pore which is permeable to Ca2+ ions, enabling Ca2+ entry into the cell. The precise mechanism by which STIM1 is recruited to the plasma membrane to activate SOCs and the plasma membrane components involved in targeting STIM1 to the plasma membrane are largely unknown. In this study the mechanisms underlying movement of STIM1 to the plasma membrane and its accumulation at ER-plasma membrane junctions was explored in HeLa cells. In the initial part of this study I investigated whether the movement of STIM1 to the plasma membrane is an ATP-dependent process. I found that depletion of cytosolic ATP can stimulate STIM1 puncta formation in HeLa cells and that the formation of STIM1-Orai1 complexes at the plasma membrane is unaffected in these conditions. Inhibition of ATP synthesis also initiated the loss of phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2) from the plasma membrane. ATP depletion did not affect the structure of the microtubule cytoskeleton. These results suggest that the translocation of STIM1 and the formation of STIM1-Orai1 complexes is an ATP independent process which is not due to the disruption of microtubules and support a diffusional model for STIM1 puncta formation. It has been suggested that an additional interaction of the C-terminal polybasic domain of STIM1 with plasma membrane phosphoinositides could contribute to STIM1 puncta formation prior to binding to Orai1. I investigated the role of phosphoinositides in the formation of STIM1 puncta and SOCE in response to store depletion. Treatment of HeLa cells with inhibitors of the phosphatidylinositol 3-kinase (PI3K) and phosphatidylinositol 4-kinase (wortmannin and LY294002) partially inhibited formation of STIM1 puncta. Additional rapid depletion of PtdIns(4,5)P2 resulted in more substantial inhibition of the translocation of STIM1-EYFP into puncta. The inhibition was extensive at a concentration of LY294002 (50 μM) that should primarily inhibit PI3K consistent with a major role for PtdIns(4,5)P2 and PtdIns(3,4,5)P3 in puncta formation. Depletion of phosphoinositides also partially inhibited SOCE. Overexpression of Orai1 resulted in a recovery of translocation of STMI1 into puncta following phosphoinositide depletion and under these conditions SOCE was increased to above control levels. These observations support the idea that phosphoinositides are not essential but contribute to STIM1 accumulation at ER-PM junctions with a second translocation mechanism involving direct STIM1/Orai1 interactions. It was recently reported that STIM1 and Orai1 may function within a macromolecular complex involving other unidentified proteins. In this study I have identified that Golli-BG21, a member of the myelin basic protein (MBP) family, can directly interact with STIM1. Golli interacts with the C-terminal domain of STIM1 in both in vitro and in vivo binding assays and this interaction may be modulated by intracellular Ca2+ concentration. Golli also colocalises with full length STIM1 and Orai1 complexes in HeLa cells following store depletion. Overexpression of Golli reduces SOCE in HeLa cells but this inhibition is overcome by overexpressing STIM1. We therefore suggest that Golli binds to STIM1-Orai1 complexes to negatively regulate the activity of SOCs.
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Troupes, Constantine. "The Role of STIM1 in Hypertrophy-Related Contractile Dysfunction." Diss., Temple University Libraries, 2016. http://cdm16002.contentdm.oclc.org/cdm/ref/collection/p245801coll10/id/403786.
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Biomedical SciencesPh.D.
Increases in cardiac afterload caused by disease conditions results in remodeling of heart structure by hypertrophy and alterations in the molecular regulation of contractile performance. These adaptations can be regulated by various Ca2+-dependent signaling processes. STIM1 is an important regulator of Ca2+ signaling in different cell types by sensing endoplasmic reticular Ca2+ levels and coupling to plasma membrane Orai channels. The role of STIM1 in heart is not well understood, given the robust Ca2+ regulatory machinery present within cardiac myocytes. Previous reports indicate that STIM1 may play a role in regulation of cardiac hypertrophy. The goal of this work is to understand how STIM1 can affect contractile Ca2+ regulation in normal and diseased myocytes. We induced cardiac hypertrophy by slow progressive pressure overload in adult cats. Isolated adult feline ventricular myocytes (AFMs) exhibited increased STIM1 expression and activity, which correlated with altered Ca2+ handling. Use of BTP2 to block Orai channels resulted in a reduction of action potential (AP) duration and diastolic spark rate of hypertrophied myocytes, without affecting myocytes from sham-operated animals. Overexpressed STIM1 in cultured AFMs caused persistent Ca2+ influx that resulted in increased diastolic spark rates and prolonged APs, similar to myocytes from banded animals. STIM1 mediated Ca2+ influx could load the sarcoplasmic reticulum and activated CaMKII, which increased spark rates and lead to spontaneous APs. Importantly, STIM1 operated by associating with Orai channels because these effects could be blocked with either BTP2 or with a dominant negative Orai construct. Prolonged Ca2+ entry through this pathway eventually causes cell death. In conclusion, the work presented in this thesis establishes a role for STIM1-Orai in contractile Ca2+ regulation.
Temple University--Theses
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Gueder, Nahla. "sp²-Iminosugar-glucosidases inhibitor 1-C-octyl-2-oxa-3-oxocastanospermine - induced antiproliferative, apoptotic and necrotic effects in breast cancer cells via targeting GRP78, Stim1 and Orai1." Thesis, Amiens, 2018. http://www.theses.fr/2018AMIE0033/document.
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L'altération de glycosylation est l'une des caractéristiques du cancer du sein. Ainsi le défaut de glycosylation affecte différentes protéines glycosylées responsables des différents processus cancéreux. Les canaux SOC (Store operated channels) constituent la voie majeure de l'entrée du calcium dans les cellules et sont impliqués dans la prolifération, la migration et la survie des cellules cancéreuses du sein. CO-OCS est un nouvel inhibiteur de la glycosylation avec plus de sélectivité vis-à-vis des α-glucosidases, et montre des activités anticancéreuses des cellules cancéreuses du sein, sans affecter les cellules mammaires normales. L'objectif de ma thèse est d'étudier les mécanismes moléculaires par lesquels CO-OCS induit ses effets anti-tumoraux. CO-OCS inhibe la migration des cellules cancéreuses à fort potentiel métastatique. Cet effet anti-migratoire est dû à une réduction de l'expression de la β1-intégrine, de Stim1, et de l'activation des voies de signalisation FAK et ERK1/2 par CO-OCS. Dans les cellules cancéreuses peu invasives, CO-OCS diminue la prolifération et augmente la mortalité de ces cellules en affectant l'expression de 3 protéines : Stim1 et Orai1 : protéines N-glycosylées au niveau du réticulum endoplasmique (RE), et GRP78, protéine de stress du RE. Ainsi en supprimant complétement l'expression de Stim1, CO-OCS réduit la prolifération en accumulant les cellules dans les phases G1 et G2/M du cycle cellulaire. Alors que la réduction de l'expression de GRP78 et d'Orai1 par le CO-OCS augmente respectivement l'apoptose et la nécrose. Par ailleurs, l'invalidation de Stim1 atténue l'effet apoptotique induit par CO-OCS. CO-OCS réduit aussi le contenu calcique du RE. Cette réduction du calcium réticulaire est due à une fuite de calcium par le Translocon. En effet, l'Anisomycine, inhibiteur du Translocon, restore de contenu calcique réticulaire et antagonise l'apoptose induite par le CO-OCS. En conclusion, CO-OCS induit une accumulation de protéines mal-repliées dans le RE induisant ainsi un stress réticulaire. Trois cibles du CO-OCS ont été identifiées : l'expression de Stim1 favorise la prolifération tandis que celle d'Orai1 et de GRP78 protègent respectivement les cellules de l'apoptose et de la nécrose induites par CO-OCS. De plus, en diminuant l'expression de GRP78, CO-OCS induit une fuite du calcium du RE par le TransloconAlteration in glycosylation pattern is one of the hallmarks of breast cancer. The levels and the abnormal expressions of glycan were found in breast cancer patients. Glycosylation defect can affect different glycosylated proteins which are implicated in cancerogenesis. Changes in intracellular Ca2+ levels can regulate different cellular processes. SOC channels are implicated in breast cancer proliferation, migration and survival. CO-OCS is a new glycosylation inhibitor with more selectivity toward theα- glucosidases exhibited anti-cancer activities in breast cancer cells without affecting the normal mammary cells. The objective of my thesis is investigating the related molecular mechanisms by which CO-OCS induced its anti-tumour effects.CO-OCS impaired breast cancer migration through decrease β1-integrin expression and the activation of FAK and ERK1/2 signalling pathways. CO-OCS also induced anti-migratory effect via Stim1 protein expression down-regulation leading to inhibition of SOCE. Additionally, CO-OCS affected the expression of both Orai1 and Stim1 proteins leading to anti-proliferative effects and cell cycle arrest in G1 and G2/M phase respectively. Moreover, CO-OCS affected the expression of Stim1 at the protein level without affecting its transcript level. GRP78 implicated in CO-OCS apoptotic death. The expression of Stim1 regulated the apoptosis induced by CO-OCS via modulating GRP78 expression. Orai1 down-regulation promoted CO-OCS necrotic effect. CO-OCS induced ER- calcium depletion due to increase in ER calcium leak via the Translocon; Anisomycin (Translocon inhibitor) decreased the apoptosis induced by CO-OCS. In conclusion, these results show that in breast cancer, by targeting Stim1, Orai1 and GRP78, CO-OCS reduced cell proliferation and induced apoptosis and necrosis cell death. Stim1 favours CO-OCS apoptotic effect while Orai1 protected from necrosis induced by CO-OCS. The inhibition of Translocon decreased CO-OCS apoptotic cell death via restoring the ER calcium homeostasis
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Frappier, Maude. "MURC est un partenaire d’interaction de STIM1 impliqué dans la signalisation calcique." Mémoire, Université de Sherbrooke, 2015. http://hdl.handle.net/11143/8332.
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Dans les cardiomyocytes, la concentration intracellulaire de Ca2+ doit être finement régulée pour maintenir l’homéostasie calcique. La protéine Stromal interaction molecule 1 (STIM1), qui joue un rôle important dans le maintien des niveaux intracellulaires de Ca2+ des cellules non excitables en opérant l’entrée capacitative de Ca2+ (SOCE), est aussi présente dans les cardiomyocytes. Plusieurs études démontrent que STIM1 et le SOCE jouent un rôle important dans le développement de l’hypertrophie des cardiomyocytes. De plus, récemment, un nouveau rôle de STIM1 a commencé à émerger. STIM1 pourrait moduler l’expression de certains canaux calciques à la membrane plasmique en régulant leur trafic intracellulaire. Le but de l’étude était d’identifier des partenaires d’interaction de STIM1 dans l’optique de révéler le mécanisme par lequel STIM1 induit l’internalisation d’un canal calcique voltage-dépendant. Les protéines recueillies par le pull-down à partir des lysats de coeurs de rats avec une colonne d’affinité composée du domaine ERM de STIM1, ont été analysées en spectrométrie de masse. La protéine Muscle related coiled-coil (MURC), une protéine de la famille des Cavin, a été retenue comme partenaire d’interaction potentiel de STIM1. Comme elle est exprimée dans les cardiomyocytes et dans les cellules musculaires squelettiques, qui sont des cellules où la régulation de la signalisation calcique est primordiale pour le bon fonctionnement des tissus et qu’elle semble interagir avec STIM1, qui est un acteur important de la signalisation calcique, notre objectif s’est élargi et nous avons investigué sur l’implication de MURC dans la signalisation calcique en général. Nous avons donc confirmé par co-immunoprécipitation que le domaine ERM de STIM1 interagissait avec MURC. Puis, par des essais d’imagerie calcique, nous avons démontré que la surexpression de MURC pouvait provoquer différentes réponses dans différents types cellulaires en fonction de l’activation de la mobilisation calcique. En effet, nous avons observé une augmentation du SOCE qui est indépendante de la voie RhoA/ROCK dans les cellules HEK293T, une diminution de l’entrée de calcium médiée par un récepteur (ROCE) qui est pourrait être dépendante de la voie RhoA/ROCK dans les cellules T6.11 et une diminution de l’activation de RhoA de façon dépendante de l’activation du SOCE dans les cardiomyocytes HL-1. Nous avons aussi montré que MURC pouvait interagir à la membrane plasmique avec les protéines Orai1, qui sont les protéines formant les canaux CRAC (Ca2+ release-activated Ca2+) du SOCE et ce de façon dépendante de l’activation de STIM1. Enfin, les résultats de cette étude suggèrent que MURC est un partenaire d’interaction de STIM1 impliqué dans la signalisation calcique. En effet, MURC peut moduler l’activation de RhoA, ce qui pourrait induire l’internalisation de canaux calciques. De plus, son interaction avec STIM1 et Orai1 pourrait notamment faire un pont facilitant l’interaction entre STIM1 et Orai1 ce qui aurait pour effet d’augmenter le SOCE et possiblement contribuer à augmenter l’hypertrophie des cardiomyocytes.
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Lur, Gyorgy. "STIM1, Orai1 and store operated calcium entry in pancreatic acinar cells." Thesis, University of Liverpool, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.539501.
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Saliba, Youakim. "Identification des partenaires de STIM1 dans le cœur normal et hypertrophié." Paris 6, 2012. http://www.theses.fr/2012PA066541.
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Nous avons déjà démontré un rôle important de STIM1 dans l’induction de l’hypertrophie cardiaque, mais l’identité des canaux STIM1 dépendants responsables de ce flux calcique pro-hypertrophique dans les cellules ventriculaires du rat reste à déterminer. Dans cette étude, nous avons développé une nouvelle méthode de transfert myocardique de gène non viral, en utilisant l’énergie des ultrasons, des liposomes et des injections pressurisées dans le myocarde du rat. Grâce à sa simplicité, son efficacité et sa faible immunogénicité, cette technique a produit un nombre suffisant de cellules transfectées pour effectuer des expériences biochimiques et physiologiques sur cellules isolées. Nous avons ensuite caractérisé le profil d’expression des protéines ORAIs et TRPCs dans les cellules ventriculaires normales et hypertrophiées, et avons trouvé une augmentation de l’expression de TRPC1 dans l’hypertrophie cardiaque. Ensuite nous avons utilisé la méthode de transfert de gènes pour identifier les partenaires canalaires de STIM1 via l’ARN interférence par injection de siARN. Nous avons identifié TRPC5 comme un canal calcique non-sélectif qui fonctionne d’une façon constitutive en conditions basales, avec une activité accrue dans l’hypertrophie cardiaque ; ainsi que ORAI3 qui opère dans deux modes: entrée capacitative de calcium et entrée constitutive en concordance avec TRPC5. Nous avons développé une nouvelle méthode de transfert myocardique de gène non viral que nous avons ensuite utilisée pour identifier TRPC5 et ORAI3 comme nouveaux canaux voltage indépendants et STIM1 dépendants dans les myocytes ventriculaires de ratWe previously showed an important role of STIM1 in cardiac hypertrophy; however, the identity of the channels responsible for the STIM1 dependent pro-hypertrophic Ca2+ entry in the rat ventricular myocytes remains to be elucidated. In this study we developed a new method of myocardial non viral gene delivery, by using the combination of ultrasound energy (USE), liposomes and high pressure injections to the rat heart. Due to its simplicity, low toxicity and low immunogenicity, this method produced sufficient number of transfected cells to perform biochemical experiments and single cell physiological measurements. We then characterized the expression profile of TRPCs and ORAIs proteins in both normal and hypertrophied ventricular myocytes, and found an upregulation of TRPC1 in hypertrophied cells. We further used the new gene delivery method to identify and screen for the STIM1 associated channel candidates via RNA interference. We identified TRPC5 as a non-selective Ca2+ channel that operates constitutively in basal conditions with increased activity in cardiac hypertrophy, as well as ORAI3 that functions in both modes: SOCE and constitutive basal Ca2+ entry in concordance with TRPC5. We developed an efficient non-viral cardiac gene delivery which we used to elucidate TRPC5 and ORAI3 as new voltage independent STIM1 regulated Ca2+ channels in the ventricular rat myocytes
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Premsler, Thomas [Verfasser], Albert [Akademischer Betreuer] Sickmann, and Jan Georg [Gutachter] Hengstler. "Mass spectrometry based interaction study of the STIM1 and VASP proteins : (Massenspektrometrie-basierte Interaktionsstudie der Proteine STIM1 und VASP) / Thomas Premsler. Betreuer: Albert Sickmann. Gutachter: Jan Georg Hengstler." Dortmund : Universitätsbibliothek Dortmund, 2012. http://d-nb.info/1110892373/34.
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Cao, Hang [Verfasser]. "Effect of STIM1/2 and of Ceritinib on Platelet Function / Hang Cao." Tübingen : Universitätsbibliothek Tübingen, 2021. http://d-nb.info/1234450763/34.
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Mukherjee, Sreya. "Applications of Molecular Modelling and Structure Based Drug Design in Drug Discovery." Scholar Commons, 2016. http://scholarcommons.usf.edu/etd/6331.
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Calcium ions have important roles in cellular processes including intracellular signaling, protein folding, enzyme activation and initiation of programmed cell death. Cells maintain low levels of calcium in their cytosol in order to regulate these processes. When activation of calcium-dependent processes is needed, cells can release calcium stored in the endoplasmic reticulum (ER) into the cytosol to initiate the processes. This can also initiate formation of plasma membrane channels that allow entry of additional calcium from the extracellular milieu. The change in calcium levels is referred to as calcium flux. A key protein involved in initiation of calcium flux is Stromal Interaction Molecule 1 (STIM1), which has recently been identified as a sensor of ER calcium levels. STIM1 is an ER transmembrane protein that is activated by a drop in ER calcium levels. Upon activation, STIM1 oligomerizes with a plasma membrane protein, ORA1, to form calcium-selective plasma membrane channels. Dysregulation of calcium flux has been reported in cancers, autoimmune diseases and other diseases. STIM1 is a promising target in drug discovery due to its key role early in calcium flux. Here we review the involvement and importance of STIM1 in diseases and we discuss STIM1 as a viable target for drug discovery using computational chemistry methods to rapidly identify new molecules to target STIM1. Herein, computational techniques were used to understand the mechanistic role of STIM1 and virtual screening is in process to discover potential inhibitors of STIM1 activity. Also mutational analysis on STIM1 was performed computationally to see the effect it had on the protein computationally.
It has been found that tumor cells and tissues, compared to normal cells, have higher levels of copper and possibly other metal ions. This presents a potential vulnerability of tumor cells that can serve as a physiological difference between cancer cells and normal cells and allows design of compounds that selectively target tumor cells while sparing normal cells. Recently we have identified compounds that have potential to inhibit the proteasome in tumor cells and induce cell death by mobilizing endogenous tumor copper resulting in in cellulo activation of the compound. These compounds hence act as pro-drugs, becoming active drugs in tumor cells with high copper content but remaining essentially inactive in normal cells, thereby greatly reducing adverse effects in patients. Such use would be of significant benefit in early detection and treatment of cancers, in particular, aggressive cancers such as pancreatic cancer which is usually not detected until it has reached an advanced stage. Six compounds were identified following virtual screening of the NCI Diversity Set with our proteasome computer model followed by confirmation with a biochemical assay that showed significant inhibition of the proteasome by the compounds in the presence of copper ions. In a dose response assay, NSC 37408 (6, 7-dihydroxy-1-benzofuran-3-one), our best compound, exhibited an IC50 of 3µM in the presence of 100 nM copper.
Chagas’ Disease, a parasitic disease caused by the parasite Trypanosma Cruzi, is endemic to Latin America. The disease manifests itself in a short acute phase and a long chronic phase. Current treatments are effective only in the acute phase and are not used in the chronic phase due to toxicity of the drugs. Hence a new drug discovery approach was chosen for this disease. Cruzain is the major etiologic enzyme involved in the disease and is only present in the parasite. It is also an enzyme expressed by the parasite in both phases. Herein, a novel peptoid library containing hydromethylketones was constructed and screened against a virtual structure of cruzain. The peptoids thus found through this drug discovery effort can be used as potential drug candidates against cruzain. Computational techniques will help achieve a high degree of specificity and aid in proposing assays for determining compounds with high activity
Alzheimer disease is the most common form of dementia. Its pathogenesis incorporates many potential targets for treatment. Among the targets identified, Apolipoprotein E4 (apoE4) is especially interesting due to its catalytic role in the degradation and clearance of amyloid beta (Aβ), a risk factor for Alzheimer disease. ApoE exists in 3 isoforms which directly impact its functionality in the body. There are characteristic structural differences between them. In ApoE4 ionic interactions exist between Arg-61 and Glu-255 residues, unlike the other isoforms. Hence interruption of this interaction by inhibitors may change the structure of apoE4 to a more linear structure as observed in the other isoforms. Virtual screening of the NCI diversity set on an energy minimized protein virtual structure was performed to identify potential small molecule inhibitors and to gain further understanding of interactions that can be targeted to inhibit this protein. From the top ligands in the NCI diversity set, a peptide library was designed to target the protein.
Previous research has indicated that liquid assisted grinding (LAG) is efficient and reliable for cocrystal formation when compared to solvent crystallization and dimethyl formamide is the best solvent for grinding. Herein, we report the comparison of four screening processes: Slurry, solvent crystallization, LAG and dry grinding. Thirty-eight crystal forms containing the Narom··· COOH, Narom···OH supramolecular heterosynthons were screened in the process, and it was observed that slurry methodology is as efficient and reliable in forming cocrystals as solution crystallization. Twenty-four new crystal forms were also isolated herein. LAG was found to be more efficient as compared to dry grinding and was successful in the formation of twenty-five crystal forms of the thirty-eight screened. Dimethyl formamide still remains the best solvent for LAG. All our slurry experiments were performed in water and it was found that water can be used reliably for this method for compounds within a wide range of solubility, thereby increasing the versatility and usability of this method for future screening procedures.
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Maues, de Paula André. "Pour une meilleure compréhension de la myopathie à agrégats tubulaires." Thesis, Aix-Marseille, 2016. http://www.theses.fr/2016AIXM5056.
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La myopathie à agrégats tubulaire (MTA) est une maladie rare caractérisée par la présence, dans la biopsie musculaire, d’« agrégats tubulaires ». Nous avons retrouvé 15 cas de MTA parmi 13987 biopsies musculaires réalisées dans les 35 dernières années dans notre service. Parmi ces cas, se trouvait une famille de trois patients qui ne pouvaient être inclus dans aucun groupe clinique connu de myopathie à agrégats tubulaires, car ils étaient asymptomatiques avec une hyperCKémie isolée et un test de contracture positif.Nos travaux ont mis en évidence des mutations hétérozygotes du gène STIM1. Ces mutations localisées dans la partie du gène qui code le domaine intraluminal EF-hand, entrainent une activation constitutive de STIM1 avec exacerbation du mécanisme SOCE et en conséquence l’augmentation de l’influx de Ca2+, démontré dans des myoblastes transfectés. Nous avons également mis en évidence la mutation p.Arg304Trp du gène STIM1, comme cause du syndrome de Stormorken. Cela augmente le spectre phénotypique des mutations de ce gène.Nos résultats de l’analyse protéomique montrent que la protéostase dans la MTA est dérégulée, car le profil du protéome est différent chez les patients et dans les agrégats tubulaires microdisséqués par laser, en comparaison a des contrôles. Les protéines enrichies s’avèrent appartenir à des voies biologiques impliquées dans l’homéostase ionique, les systèmes de membranes de la triade et de l’exosome et dans le métabolisme mitochondrial.Nos travaux ouvrent des perspectives pour mieux comprendre la physiopathologie de la myopathie à agrégats tubulaires et ainsi pouvoir proposer des solutions thérapeutiques efficacesMyopathy with tubular aggregates (MTA) is a rare disease characterized by the presence of tubular aggregates in muscle biopsy. We found 15 cases of MAT in our registry including 13987 muscle biopsies performed over 35 years. Among them, there was a family of three patients that did not fit any of the previously described clinical groups of MTA, since they were asymptomatic with isolated hyperCKemia and positive contracture test. Our works revealed heterozygous mutations in the gene STIM1. These mutations localized in the gene portion that codes for the intraluminal EF-hand domain, leads to a constitutive activation of STIM1 with SOCE mecanism enhancement and consequent increase of the Ca2+ entry which was demonstrated using transfected myoblasts. We also revealed the mutation p.Arg304Trp in the coding sequencing of the CC1 domain of STIM1, as the cause of Stormorken syndrome. This fact increases the phenotypical spectra of the mutations for this gene.The results of our proteomic analysis show that the proteostasis in MTA is disturbed, because the proteome profile is different in total muscle of the patients and in their tubular aggregates when compared to controls. The enriched proteins belong to the biological pathways linked to ionic homeostasis, membrane systems of the triads and the exosome, and to the mitochondrial metabolism.Our works bring perspectives for the continuation of our studies, in order to better understand the physiopathology of the myopathy with tubular aggregates and propose efficacious therapeutic solutions
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Ritchie, Michael. "TRANSCRIPTIONAL AND MOLECULAR CONTROL OF CALCIUM SIGNALING." Diss., Temple University Libraries, 2012. http://cdm16002.contentdm.oclc.org/cdm/ref/collection/p245801coll10/id/211071.
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BiochemistryPh.D.
The extensive relationship between modulation of intracellular Ca2+ content and the control of cell proliferation (Boynton et al. 1974; Whitfield et al. 1979; Berridge and Irvine 1984), differentiation (Bridges et al. 1981; Holliday et al. 1991) and death (Orrenius et al. 2003) has led to much examination into the relationship between Ca2+ signaling pathways and the onset of various pathological conditions, including cancer, cardiac hypertrophy, immunodeficiency, neurodegeneration. Control of Ca2+ signals is achieved via an extensive combination of pumps, channels and exchangers which regulate the concentration of Ca2+ within not only the cytosol but also all intracellular compartments. Accordingly, a great deal of research has focused on the mechanisms which regulate these channels and pumps, and recently the primary mechanism for Ca2+ influx in non-excitable cells has been identified. This process, termed Store-operated calcium entry (SOCe), is a key evolutionarily conserved mechanism whereby decreases in endoplasmic reticulum Ca2+ content (sensed by the ER Ca2+ sensor, STIM1) leads to the influx of Ca2+ across the plasma membrane through the Orai family of Ca2+ channels. However, many questions remain about how this Ca2+ signaling pathway is regulated. In this thesis, I provide evidence regarding the transcriptional and molecular mechanisms regulating SOCe. Initial studies in my thesis work aimed to identify some of the key events leading to dysregulation of Ca2+ homeostasis in the kidney specific pediatric malignancy, Wilms' Tumor. I found that STIM1 expression levels and SOCe signals are significantly reduced in Primary Wilms' Tumor samples. Subsequent analysis of these phenomena led me to the finding that STIM1 expression is under the control of the transcription factors Wilms' Tumor Suppressor 1 (WT1) and Early Growth Response 1 (EGR1). Subsequent investigations were carried out with the purpose to assess how activation of the EGR1 transcription factor alters long term Ca2+ signals. Indeed, I found that receptor-mediated activation of EGR1 leads to induction of STIM1 expression and increases in SOCe. However, unexpectedly through these analyses, I propose a novel role for STIM1 that STIM1 interacts with the Plasma Membrane Ca2+ ATPase (PMCA) through its C-terminal proline-rich domain and reduces PMCA-mediated Ca2+ clearance, effectively creating local, augmented Ca2+ gradients. This coordinated control of Ca2+ entry and exit from the cell has wide-ranging implications for Ca2+ signaling in multiple cell types.
Temple University--Theses
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Giachini, Fernanda Regina Casagrande. "Contribuição da via STIM1/Orai1 para as diferenças relacionadas ao sexo na entrada de cálcio em miócitos vasculares durante a hipertensão arterial." Universidade de São Paulo, 2010. http://www.teses.usp.br/teses/disponiveis/42/42136/tde-28092010-170302/.
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Os distúrbios na regulação da concentração de cálcio (Ca2+) citoplasmático contribuem para a patogênese da hipertensão arterial. Evidências sugerem que as moléculas de interação estromal (STIM) atuam como sensores dos estoques intracelulares de Ca2+, enquanto as proteínas Orai representam as subunidades que formam os canais de Ca2+ ativados pela liberação de Ca2+ (CRAC). Neste estudo avaliamos a participação de STIM1/Orai1 na regulação das concentrações de Ca2+ citoplasmático e na ativação da contração vascular em aortas de ratos hipertensos. Nossos resultados sugerem que a ativação de STIM1/Orai1 pode representar um novo mecanismo que modula alterações vasculares nos níveis de Ca2+ intracelular na hipertensão arterial e que contribui para as diferenças sexuais de reatividade vascular em animais hipertensos.Disturbance in the regulation of cytoplasmic calcium (Ca2+) concentration contributes to the pathogenesis of hypertension. Evidences suggest that the stromal interaction molecule (STIM) acts as a sensor of intracellular Ca2+ stores, whereas Orai proteins are the subunits that form CRAC channels. In this study, we evaluated the role of STIM1/Orai1 in the regulation of cytoplasmic Ca2+ concentrations and in the activation of contraction in aortas from hypertensive rats. We also studied how the differential activation of this pathway contributes to sex differences observed between hypertensive rats, as well as the protective effects of the female sex hormones in the vasculature. Our results suggest that activation of STIM1/Orai1 may represent a new mechanism that modulates intracellular Ca2+ concentration during hypertension and contributes to sex differences in the vascular reactivity of hypertensive animals.
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Zanotto, Camila Ziliotto. "Papel da O-glicosilação com N-acetil-glucosamina (O-GlcNAc) no influxo e recaptação de cálcio pelo retículo sarcoplasmático em aorta de ratos: análise funcional." Universidade de São Paulo, 2013. http://www.teses.usp.br/teses/disponiveis/17/17133/tde-09092013-133940/.
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A O-glicosilação com N-acetil-glucosamina (O-GlcNAc) é uma modificação pós-translacional altamente dinâmica que modula diversas vias de sinalização. O processo de O-GlcNAc é controlado por duas enzimas: a enzima OGT é responsável por catalisar a adição de N-acetil-glucosamina no grupo hidroxila dos resíduos de serina e treonina, enquanto a OGA catalisa a remoção de O-GlcNAc das proteínas modificadas. Proteínas com importante papel na função vascular são alvo de O-GlcNAc e o aumento da expressão de proteínas modificadas por O-GlcNAc promove aumento da reatividade vascular para estímulos contráteis. Um dos mecanismos de extrema importância no controle do tônus vascular está ligado à regulação da concentração de cálcio (Ca2+) intracelular, onde destacamos a participação do sistema STIM1/Orai1. As moléculas de interação estromal (STIM) atuam como sensores dos estoques intracelulares de Ca2+ e as proteínas Orai representam as subunidades que formam os canais de Ca2+ ativados pela liberação de Ca2+ (CRAC). Neste estudo investigamos a hipótese de que o aumento dos níveis vasculares de proteínas glicosiladas aumenta a resposta contrátil em aorta de ratos, por mecanismos relacionados ao controle da concentração intracelular de Ca2+.Em nossos experimentos, utilizamos aortas torácicas de ratos incubadas com PugNAc (inibidor seletivo da OGA, ), por 24h. Utilizando protocolo experimental que permite avaliar contrações induzidas pelo influxo de Ca2+ e liberação de Ca2+ intracelular, demonstramos que a incubação com PugNAc aumentou a resposta contrátil à PE bem como a contração durante o período de influxo de Ca2+, induzida pela reintrodução de solução fisiológica contendo Ca2+ (1,56 mM). O bloqueio dos canais CRAC com 2-APB (100 ) e gadolíneo (Gd3+, 100 ) diminuiu significativamente as contrações induzidas pelo influxo de Ca2+ em aortas incubadas com PugNAc. Além disso, estas aortas apresentaram aumento da expressão protéica de STIM1, o que resultaria em maior influxo de Ca2+. A contração induzida por cafeína (20 mM) e serotonina (10 ), a qual reflete a capacidade funcional do retículo sarcoplasmático (RS) em captar Ca2+, foi maior em aortas incubadas com PugNAc. O papel da Ca2+-ATPase (SERCA) foi avaliado com a utilização de tapsigargina, bloqueador da SERCA. O efeito da tapsigargina foi semelhante em artérias incubadas com PugNAc e veículo, apesar do aumento de expressão proteica da SERCA em aortas incubadas com PugNAc. Como a proteína cinase C (PKC) é ativada por aumentos de Ca2+ intracelular, determinamos se a atividade de proteínas alvo da PKC estavam aumentadas. A incubação com PugNAc aumentou a expressão das formas fosforiladas da CPI-17, MYPT-1 e MLC. Em conjunto, estes resultados sugerem que a ativação de STIM1/Orai1, aumento da liberação de Ca2+ intracelular e ativação da via de sinalização da PKC podem representar mecanismos que modulam as alterações vasculares em resposta ao aumento de proteínas glicosiladas por O-GlcNAc.Glycosylation with O-linked -N-acetyl-glucosamine (O-GlcNAc) is a highly dynamic post-translational modification. The process of O-GlcNAc is controlled by two enzymes: the OGT enzyme catalyses the addition of N-acetyl-glucosamine to the hydroxyl group of serine and threonine residues of a target protein, while OGA catalyzes the cleavage of O-GlcNAc from post-translationally-modified proteins. Proteins with an important role in vascular function are targets of O-GlcNAc and increased levels of proteins modified by O-GlcNAc increase vascular reactivity to contractile stimuli. The regulation of intracellular calcium (Ca2+) concentration, including the activation of STIM1/Orai1, is key in the control of vascular tone. The stromal interaction molecules (STIM) act as sensors of intracellular Ca2+ stores whereas the Orai proteins represent subunits of the Ca2+ release-activated Ca2+ channels (CRAC). We hypothesized that increased levels of vascular O-GlcNAc proteins augment vascular contractile responses by altering mechanisms that regulate the intracellular Ca2+. Rat thoracic aortas were incubated with PugNAc (OGA selective inhibitor, ) for 24h. Using an experimental protocol that evaluates contractions induced by Ca2+ influx and release, we demonstrated that incubation with PugNAc increases contractile responses to phenylephrine (PE) as well as the contraction induced by Ca2+ influx, after depletion of intracellular Ca2+ stores. The CRAC channel blockers, 2-APB (100 ) and gadolinium (Gd3+, 100 ), significantly reduced the contractions induced by Ca2+ influx in aortas incubated with PugNAc. Furthermore, these aortas showed increased STIM1 protein expression, which could result in increased influx of Ca2+ and, in turn, increase vascular contraction. The contraction induced by the release of intracellular Ca2+ stores, stimulated by caffeine (20 mM) and serotonin (10 ), was increased in aortas incubated with PugNAc. The Ca2+-ATPase (SERCA) inhibitor thapsigargin produced similar effects in arteries incubated with PugNAc or vehicle, despite the increased SERCA protein expression in aortas incubated with PugNAc. Since PKC is activated by increases in intracellular Ca2+ and arteries incubated with PugNAc show activation of PKC, we determined whether the activity of proteins that are targets of PKC was increased in PugNAc-treated aortas. Incubation with PugNAc increased the expression of phosphorylated forms of CPI-17, MYPT-1 and MLC. Together, these results suggest that activation of STIM1/Orai1, increased release of intracellular Ca2+ and PKC activation may represent mechanisms that modulate vascular responses upon increased O-GlcNAc proteins.
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Onodera, Kaoru [Verfasser]. "STIM1-regulierter Ca2+-Einstrom durch die apikale und die basolaterale Membran im Kolonepithel / Kaoru Onodera." Gießen : Universitätsbibliothek, 2013. http://d-nb.info/1064992153/34.
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Cabanas, Hélène. "Rôle de la signalisation calcique dans la leucémie myéloïde chronique." Thesis, Poitiers, 2016. http://www.theses.fr/2016POIT2302/document.
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La Leucémie Myéloïde Chronique (LMC) est une maladie clonale caractérisée par la présence du chromosome Philadelphie codant pour Bcr-Abl, une tyrosine kinase constitutivement active responsable de la leucémogenèse. Bien que très efficaces, les inhibiteurs de tyrosine kinase (ITKs) restent cependant inactifs sur les cellules souches leucémiques. Ce travail de thèse montre que la signalisation calcique, connue pour réguler de nombreux processus dans les cellules saines et cancéreuses, est importante dans la signalisation cellulaire au décours de la LMC. Le rôle des entrées calciques dépendantes des stocks (SOCEs) médiées par STIM1 (STromal Interaction Molecule 1) et les canaux Orai1 et TRPC1 ainsi que des entrées calciques induites par la thrombine a été étudié dans la leucémogenèse. Nous avons observé une diminution de ces entrées dans les cellules exprimant Bcr-Abl pouvant être expliquée par le changement de stœchiométrie Orai1/STIM1. Ceci entraîne la diminution de l'activation de NFAT (Nuclear Factor of Activated T-cells) ainsi que des conséquences sur la prolifération et la migration cellulaire mais pas sur l'apoptose. De plus, les SOCEs sont restaurées dans les cellules cancéreuses après traitement à l'Imatinib, le principal ITK. Nous proposons alors que l'expression de Bcr-Abl joue un rôle sur l'homéostasie calcique en entraînant une dérégulation générale des fonctions cellulaires dans les cellules leucémiques notamment via la voie PKC (Protein Kinase C). Ainsi, ces résultats montrent une dérégulation des entrées calciques dans les cellules exprimant Bcr-Abl, suggérant que la signalisation calcique puisse être une cible thérapeutique en parallèle avec les ITKsChronic Myeloid Leukemia (CML) is a clonal disease characterized by the presence of the Philadelphia chromosome encoding for Bcr-Abl, a constitutively active tyrosine kinase responsible for leukemogenesis. Although Bcr-Abl tyrosine kinase inhibitors (TKIs) have revolutionized the therapy of Ph+ leukemia, the complete eradication of CML is limited by the emergence of resistance in hematopoietic stem cells. This thesis proposes that calcium (Ca2+) signaling pathways, known to govern a large number of functions in normal and cancer cells, may be important in CML cell signaling. Therefore, we studied the role of Store Operated-Calcium entry (SOCE) (i.e. STromal Interaction Molecule 1 (STIM1), Orai1 and TRPC1 channels) and thrombin induced Ca2+ entry in leukemogenesis. We found a decrease in both calcium entries in Bcr-Abl-expressing cells compared to normal cells. The reduced SOCE seems related to a change in stoichiometry of Orai1/STIM1. This leads to a reduction of the Nuclear Factor of Activated T-cells (NFAT) translocation and functional consequences on cell proliferation and migration but not on apoptosis. Moreover, we showed that SOCE is restored in malignant cells after treatment with Imatinib, the main TKI. We proposed that Bcr-Abl expression could impact on Ca2+ homeostasis enhancing a general disorganization of cell functions in leukemia cells notably via Protein Kinase C (PKC) pathway. Altogether this work shows a deregulation of Ca2+ entry in Bcr-Abl-expressing cells, suggesting that the Ca2+ signaling pathway could be a therapeutic target in parallel with TKIs
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Kato, Kenta. "Characterization of bioactive molecules using genetically engineered ion channels." 京都大学 (Kyoto University), 2010. http://hdl.handle.net/2433/120897.
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Samakai, Elsie. "TRANSCRIPTIONAL CONTROL OF Ca2+ SIGNALING IN T CELLS." Diss., Temple University Libraries, 2017. http://cdm16002.contentdm.oclc.org/cdm/ref/collection/p245801coll10/id/466164.
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BiochemistryPh.D.
Antigen presentation to T cells results in their activation through T Cell Receptor (TCR) stimulation, resulting in sustained elevation of cytosolic Ca2+ concentration critical for T cell activation. Sustained Ca2+ signals are important for the activation of Nuclear Factor of Activated T cells (NFAT), which is a key regulator of T cell activation through its transcriptional control of genes in multiple process including cytokine production, proliferation and differentiation(Rao, Luo, & Hogan, 1997). Recently it was shown that Stromal Interaction Molecule 1 (STIM1) inhibits plasma membrane Ca2+/ATPase 4 (PMCA4) function during T cell activation resulting in sustained elevation of Ca2+ signals(Ritchie, Samakai, & Soboloff, 2012). This interaction requires upregulation of both STIM1 and PMCA4. In this thesis, I hypothesize that changes in Ca2+ signals arising from transcriptional changes of STIM1 and PMCA are important for the efficient activation of T cells. In the first part of this thesis, I assess the transcriptional regulation of STIM1 and PMCA4. My in vitro studies show that expression of both proteins is regulated by the EGR family members, EGR1 and EGR4. Additionally, transcriptional regulation of PMCA inhibition by EGR1 and EGR4 is required for efficient activation of T cells. Interestingly, whereas significant roles for EGR1, EGR2 and EGR3 in T cell development and function have been established, a role for EGR4 has not, hitherto been elucidated. In the second half of this thesis, using qPCR, I reveal that EGR4 expression is stimulated by TCR engagement in primary double positive, CD4 and CD8 positive murine T cells. Further, EGR4-null mice exhibit shifts in early thymic development, although this does not affect the relative number of double or single positive T cells in the thymus. Interestingly, EGR4-null primary T cells exhibit normal Ca2+ entry, but fail to exhibit activation-induced inhibition of Ca2+ clearance. Although not all subsets of EGR1 and EGR4 null primary T cells exhibited decreased STIM1 expression, significant defects in proliferation, migration and/or cytokine production were observed upon stimulation in all populations, albeit to different extents. These findings reveal a two-faceted role in which EGRs regulate T cell development and function through both Ca2+-dependent and independent methods. I believe that these findings have important implications towards the general understanding of transcriptional control of Ca2+ signaling, as well as having a possible impact in the quest to advance therapies targeting immunological disorders.
Temple University--Theses
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Hasegawa, Yoshinori, Masahiro Sokabe, Mitsuo Sato, Masashi Kondo, Hiromichi Aso, Satoru Ito, and Nobukazu Suganuma. "STIM1 Regulates Platelet-Derived Growth Factor-Induced Migration and Ca2+ Influx in Human Airway Smooth Muscle Cells." Thesis, PLOS ONE, 2012. http://hdl.handle.net/2237/18819.
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Tohidi-Esfahani, Ibrahim. "Molecular profiling and characterisation of the procoagulant platelet subpopulation." Thesis, The University of Sydney, 2022. https://hdl.handle.net/2123/29642.
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Platelets play a major role in thrombotic disorders, with current antiplatelet agents limited by bleeding complications. The procoagulant platelet, implicated in thrombosis, has little role in haemostasis and specifically targeting it could improve thrombotic outcomes. This work aimed to develop a method to isolate procoagulant platelets, interrogate the transcriptome to identify key biological pathways for the formation of the subpopulation, and evaluate the pathways in downstream experiments.
RNA-sequencing of healthy donor platelets identified >1000 genes differentially expressed between procoagulant and activated/non-procoagulant platelets from the same donor, confirming the molecularly distinct nature of the procoagulant platelet. Enriched biological pathways included actin cytoskeleton, endocytosis, calcium signalling and leukocyte interaction pathways, among others.
Three identified pathways were explored. The role of dynamin, involved in actin and endocytosis pathways, was assessed. Dynamin inhibitor MiTMAB reduced procoagulant platelets generated by thrombin and glycoprotein-VI stimulation from 30.5±3.0% to 2.9±0.3% (p<0.001), while platelet activation was preserved (>99% P-selectin+). The novel discovery of dynamin as a key player in procoagulant platelet formation may have a future clinical role. STIM1, which mediates cellular calcium entry, was also assessed, with inhibition preventing procoagulant platelet formation. Elevated procoagulant platelets could also be used to identify STIM1 gain-of-function mutations. Testing the importance of leukocyte interaction, platelet concentrates for transfusion had blunted agonist-induced procoagulant platelet formation which normalised upon return into whole blood.
This work, through deep molecular profiling, has generated a large amount of previously unavailable data for hypothesis generation and advancements in the field. This may facilitate discovery of novel anti-thrombotic therapeutics and diagnostics.
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Keeley, Tyler S. "Investigating the Roles of Fucosylation and Calcium Signaling in Melanoma Invasion." Scholar Commons, 2018. https://scholarcommons.usf.edu/etd/7535.
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Melanoma is the deadliest form of skin cancer. Prognosis for early stage melanoma patients is excellent, and surgery is often curative for these patients. However, once patients have presented with invasive disease, the average 5-year survival rate drops significantly from over 90% to between 10 and 15%. Several therapies have been developed to target a commonly mutated oncogene BRAF, or its downstream effectors. Unfortunately, while these treatments show robust initial response, most patients relapse within a year. Moreover, therapy-resistant tumors are often more invasive and metastatic. Therefore, it is important to investigate the molecular mechanisms underlying melanoma invasion and metastasis, and to prevent melanoma cell dissemination and metastatic progression. Invadopodia are proteolytic membrane protrusions used by metastatic cancer cells to degrade the extracellular matrix and to facilitate cancer cell invasion and metastasis. In my thesis research I have focused on protein fucosylation and store-operated calcium entry, two separate mechanisms involved in invadopodial regulation.
Post translational modifications of proteins are essential for their structure and function. Many cell surface proteins require modifications such as glycosylation for protein-protein interactions, cell adhesion, and signal transduction. Fucosylation is a form of glycosylation that adds L-fucose on glycan structures of proteins. There is evidence indicating that fucosylation plays an important but cancer-type and branching dependent role in cancer progression. Emerging evidence indicates that the fucose salvage pathway and protein fucosylation are altered during melanoma progression and metastasis. Here, we report that the fucose salvage pathway inhibits invadopodia formation and extracellular matrix degradation by promoting α(1,2) fucosylation of cell surface proteins. The activation of the fucose salvage pathway decreases invadopodia numbers and inhibits the proteolytic activity of invadopodia in WM793 melanoma cells. Inhibiting fucokinase, one of the critical enzymes in the fucose salvage pathway, in melanoma cells abrogates L-fucose-mediated inhibition of invadopodia, suggesting dependence on the fucose salvage pathway. The inhibition of invadopodia formation by L-Fucose treatment or fucokinase overexpression could be rescued by treatment with α(1,2), but not α(1,3/4) fucosidase, implicating an α(1,2) fucose linkage-dependent inhibitory effect. The ectopic expression of FUT1, an α(1,2) fucosyltransferase, is sufficient to inhibit invadopodia formation and ECM degradation. Our findings indicate that the fucose salvage pathway can inhibit invadopodia formation, and consequently, invasiveness in melanoma via α(1,2) fucosylation. Re-activation of this pathway in melanoma could be useful for preventing melanoma invasion and metastasis.
Calcium is a critical second messenger involved in a multitude of biological processes from cell proliferation to muscle contraction. In melanoma, previous studies have found that activation of the store operated calcium entry (SOCE) channel promotes tumor invasion and metastasis, in vitro and in xenograft models. The expression levels of STIM1, an essential component of the store operated calcium channels, has been found to increase with later stages of melanoma. In melanoma cell lines, the over expression of STIM1 enhances invadopodia number whereas STIM1 knockdown inhibits invadopodia formation. Similarly, gelatin degradation activity is enhanced with STIM1 overexpression and abrogated with STIM1 knockdown, implicating STIM1 as an important factor in the regulation of invadopodia formation and melanoma invasion. Though the studies published have shown a significant role of STIM1 in tumor progression, a robust transgenic animal model has not yet been established. Here, we developed a novel transgenic mouse model which, upon 4-hydroxytamoxifen (4OHT) treatment, induces the BRAFV600E mutation and PTEN, STIM1, and STIM2 deletions in melanocytes via an inducible Cre-lox system. Our investigation found that the loss of STIM1 exacerbates tumor growth and results in tumor formation significantly more quickly than STIM1 wild type mice. Whereas PCR analysis of 4OHT-treated skin showed deletion of STIM1 and PTEN, immunohistochemical staining of these genes in tumors did not convincingly demonstrate complete deletion. Therefore, it remains to be determined whether the effects we observed are due to STIM1 and STIM2 loss. These findings need to be corroborated in the future.
Our studies focus on two important mechanisms required for melanoma progression and metastasis. We found that α(1,2) fucosylation is able to inhibit invadopodia formation, and melanoma cell invasion. The reestablishment of α(1,2) fucosylation in melanoma could potentially be exploited to inhibit melanoma metastasis. Additionally, early evidence points to STIM1 having a tumor suppressive role in melanoma oncogenesis and tumor growth based on the transgenic mouse model. Although the phenotype is unexpected, further investigation of this model will likely provide important insight for the complicate roles of SOCE in melanoma initiation and progression.
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Chung, Shan-Hua [Verfasser], and Matthias [Akademischer Betreuer] Seedorf. "Mechanisms of STIM1-mediated endoplasmic reticulum-plasma membrane (ER-PM) contact formation / Shan-Hua Chung ; Betreuer: Matthias Seedorf." Heidelberg : Universitätsbibliothek Heidelberg, 2014. http://d-nb.info/1180032667/34.
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Luik, Riina M. "Molecular mechanisms of store-operated calcium signaling : local activation of CRAC channels by STIM1, the ER calcium sensor /." May be available electronically:, 2007. http://proquest.umi.com/login?COPT=REJTPTU1MTUmSU5UPTAmVkVSPTI=&clientId=12498.
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Latour, Simon. "Rôle des protéines Orai1 et STIM1 dans les lymphomes B non-Hodgkiniens, établissement d'un modèle d'étude en 3D." Thesis, Bordeaux, 2018. http://www.theses.fr/2018BORD0038.
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Les lymphomes B non-Hodgkiniens (LNHB) représentent le type d’hémopathie maligne le plus fréquent. Ces pathologies sont traitées par l’association de chimiothérapies conventionnelles et d’immunothérapies dirigées contre le CD20. Bien qu’efficace, 40% des patients résistent ou rechutent après le traitement. Deux raisons peuvent expliquer ces échecs thérapeutiques : 1) l’absence de cibles thérapeutiques impliquées dans plusieurs processus oncogéniques et 2) l’absence de modèles pré-cliniques de LNHB pertinents pour le test de molécules thérapeutiques et la compréhension de la lymphomagenèse. Le calcium est un messager ubiquitaire qui est impliqué dans de nombreux processus cellulaires en condition physiologique et pathologique. La principale voie d’entrée de calcium dans les lymphocytes B est l’entrée capacitive de calcium médiée par Orai1 et STIM1. Ces deux protéines ont été largement décrites pour être impliquées dans les processus tumoraux de nombreux cancers, cependant leurs rôles dans la lymphomagenèse restait à élucider. Nos travaux ont révélé l'implication de la signalisation calcique dans la mort induite par le GA101, un anti CD20 de nouvelle génération actuellement en essai clinique. De plus, nous avons mis en évidence l’implication des protéines Orai1 et STIM1 dans la migration des cellules cancéreuses de LNHB. De manière intéressante, l’implication de ces deux protéines dans la migration cellulaire est calcium indépendante, suggérant donc un nouveau rôle de ces protéines. Enfin, grâce à la technologie des capsules cellulaires nous avons établi un nouveau modèle 3D de lymphome mimant la niche tumorale en incluant des cellules du microenvironnement et de la matrice extracellulaire. Ce modèle semble particulièrement pertinent pour le screening de molécules et la compréhension des mécanismes de la lymphomagenèse. Ce travail de thèse révèle ainsi le ciblage de Orai1 et STIM1 comme potentiellement intéressant dans le traitement du LNHBB-cell non-Hodgkin lymphomas (BNHL) are the most common hematological malignancies, usually treated with a combination of chemotherapy and anti CD20 immunothérapie. However, 40% of patients are resistant or relapse after treatment. These therapeutic failures could be due to 1) lack of therapeutic targets implicated in several oncogenic processes, 2) lack of relevant preclinical BNHL models for drug screening and lymphomagenesis studies. Calcium is an essential second messenger involved in various cell functions. In B cells, calcium entry is mainly due to Orai1 and STIM1 proteins, both of which have been associated with oncogenesis on solid tumors. However, their role in lymphomagenesis still remains to be elucidated. Our work shows that calcium signaling in BNHL cells participates in cell death induced by GA101, a novel anti-CD20 monoclonal antibody. We also demonstrate that Orai1 and STIM1 play a role in BNHL cell migration. Interestingly, both proteins controlled cell migration in a calcium-independent manner, suggesting a new role for these proteins. Finally, using cellular capsule technology, we established a new BNHL 3D model mimicking tumoral niche by including extracellular matrix and stromal cells. This new model could be used for drug screening and understanding lymphomagenesis. In summary, this work suggests that targeting of Orai1 and STIM1 is promising for BNHL treatment
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Debant, Marjolaine. "Etude de la dérégulation des entrées calciques du lymphocyte B de leucémie lymphoïde chronique : mise en évidence d'une nouvelle piste thérapeutique." Thesis, Brest, 2017. http://www.theses.fr/2017BRES0150.
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La leucémie lymphoïde chronique (LLC) constitue l'hémopathie maligne la plus fréquente dans les pays occidentaux et résulte d’une accumulation de lymphocytes B (LB) monoclonaux matures porteurs de la glycoprotéine CD5. Les LB de LLC sont également caractérisées par une altération de l'homéostasie du calcium avec, d'une part, la mise en évidence de facteurs de survie contrôlés par le calcium tels que phospho-ERK, NFAT-2 et IL-10, et d'autre part par une progression de la maladie qui est associée à la réponse au calcium. Tout d'abord, afin de mieux comprendre l'impact de la molécule CD5 sur les dérégulations du calcium, il a été montré que l'introduction d'un plasmide d'expression pour CD5 dans les lignées de cellules B humaines s'accompagnait d'une entrée calcique à l’état basale. Ensuite, cette entrée, appelée entrée constitutive, a été recherchée dans les LB de LLC pour montrer qu'elle caractérisait les patients non traités en phase d'évolution. Comme pour les lignées de cellules B CD5, cette nouvelle voie d'entrée du calcium est autonome et indépendante de la voie classique de signalisation du LB de LLC : BCR-IP3R. L'étude des protéines responsables de cet influx a permis de mettre en évidence, premièrement trois partenaires STIM1, Orai1 et TRPC1, et deuxièmement l'importance de la fraction membranaire de STIM1 (STIMPM) puisque l'utilisation d'un anticorps monoclonal anti-STIM1 (Acm) est capable d’inhiber l'entrée constitutive du calcium qui à son tour agit sur la survie des LB, pour les patients STIMPM positifs, lorsque l'Acm anti-STIM1 est utilisé en association avec le rituximab, un Acm thérapeutique anti-CD20. Enfin, la modélisation de la partie Cterminale de STIM1 permet d'envisager plusieurs cibles potentielles pour le développement de nouveaux Acm anti-STIM1. En conclusion, la mise en place, par le clone malin de LLC, d'une entrée constitutive du calcium favorise son agressivité et constitue donc une nouvelle voie thérapeutique contrôlable par l'utilisation d’Acm anti-STIM1 ce qui ouvre de nouvelles perspectives comme outils diagnostiques et thérapeutiquesChronic lymphocytic leukemia (CLL) is the most common hematological malignancy in Western countries and is a result of the accumulation of mature monoclonal B lymphocytes (B-CLL) carrying the CD5 glycoprotein. The B-CLL are also characterized by an alteration of calcium homeostasis with, on the one hand, the demonstration of calcium-controlled survival factors such as phospho-ERK, NFAT- 2 and IL-10, and on the other hand by a progression of the disease which is associated with the response to calcium. Initially, in order to better understand the impact of the CD5 molecule on calcium deregulations, it has been shown that the introduction of an expression plasmid for CD5 into human B cell lines was accompanied by a calcium entry in the basal state. Then, this entry, called constitutive entry, was sought in the B-CLL to show that it characterized untreated patients in the evolution phase.As with CD5 B cell lines, this new calcium entry pathway is autonomous and independent of the classical B-CLL signaling pathway: BCR-IP3R. The study of the proteins responsible for this influx made it possible to highlight, firstly three partners (STIM1, Orai1 and TRPC1), and secondly the importance of the membrane fraction of STIM1 (STIMPM) since the use of a human monoclonal antibody (mAb) anti-STIM1 is able to inhibit the constitutive entry of calcium which in turn acts on the survival of B-CLL, for STIMPM positive patients, when the anti-STIM1 mAb was used in combination with rituximab, a therapeutic anti-CD20 mAb. Finally, the modeling of the C-terminal part of STIM1 makes it possible to envisage several potential targets for the development of new anti-STIM1 mAbs. In conclusion, the introduction by the CLL malignant clone of a constitutive entry of calcium favors its aggressiveness and thus constitutes a new therapeutic pathway controllable by the use of anti-STIM1 mAb, which opens new perspectives like diagnostic and therapeutic tools
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Dramane, Gado. "Etude de la signalisation calcique dans les cellules gustatives lipidiques chez la souris." Thesis, Dijon, 2012. http://www.theses.fr/2012DIJOS035/document.
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Les personnes en surcharge pondérale semblent préférer une alimentation riche en graisse. Face à l'épidémie d'obésité qui touche nos Sociétés tant urbaines que rurales, élucider les mécanismes de la détection des lipides alimentaires devient un enjeu majeur. Il avait précédemment été admis que la glycoprotéine CD36 exprimée dans les papilles caliciformes de souris, est impliquée dans la perception oro-gustative des lipides alimentaires. Dans ce travail, nous avons montré que l'acide linoléique (LA), en activant les phospholipases A2, sPLA2, cPLA2 et iPLA2 via CD36, produit de l'acide arachidonique (AA) et la lyso-phosphatidylcholine (lyso-PC). LA déclenche un influx calcique dans les cellules CD36-positives et induit la production du facteur CIF (Calcium Influx Factor). CIF, AA et lyso-PC exercent différentes actions sur l'ouverture des canaux SOC (Stored Operated Calcium Channel) constitués de protéines Orai et contrôlés par STIM1. Stim1 est un senseur calcique situé sur la membrane du réticulum endoplasmique activé par la déplétion du calcium intracellulaire. Nous avons utilisé la technologie siRNA et des modèles de souris transgéniques pour montrer que CIF et lyso-PC activent des canaux calciques homodimériques composés de protéines Orai1 tandis qu’AA active des canaux hétéro-pentamériques composés d’Orai1 et Orai3. Nous avons également montré que STIM1 régule la production de CIF dans les cellules stimulées par la thapsigargine et l’acide linoléique ainsi que l'ouverture de deux types de canaux calciques. Par ailleurs les souris au phénotype Stim1-/- perdent la préférence spontanée pour les lipides observé chez les animaux de type sauvage. D’un autre côté les cellules CD36-positive de souris Stim1-/- sont incapables de libérer la sérotonine dans l'environnement extracellulaire. Nos résultats suggèrent que des acides gras à longue chaine (AGLC) induisent la signalisation calcique régie par STIM1 via CD36. La perception oro-gustative des lipides alimentaires détermine la préférence spontanée pour les lipides observée chez les mammifèresThe lipid-binding glycoprotein CD36, expressed by circumvallate papillae (CVP) of the mouse tongue, has been shown to be implicated in oro-gustatory perception of dietary lipids. We demonstrate that linoleic acid (LA) by activating sPLA2, cPLA2 and iPLA2 via CD36, produced arachidonic acid (AA) and lyso-phosphatidylcholine (Lyso-PC) which triggered Ca2+ influx in CD36-positive taste bud cells (TBC), purified from mouse CVP. LA induced the production of Ca2+ influx factor (CIF). CIF, AA and Lyso-PC exerted different actions on the opening of store-operated Ca2+ (SOC) channels, constituted of Orai proteins and regulated by STIM1, a sensor of Ca2+ depletion in the endoplasmic reticulum. We used siRNA technology and transgenic mice models and observed that CIF and Lyso-PC opened Orai1 channels whereas AA-opened Ca2+ channels were composed of Orai1/Orai3. STIM1 was found to regulate LA-induced CIF production and opening of both kinds of Ca2+ channels. Furthermore, Stim1–/– mice lost the spontaneous preference for fat, observed in wild-type animals. The CD36-positive TBC from Stim1–/– mice also failed to release serotonin into extracellular environment. Our results suggest that fatty acid-induced Ca2+ signaling, regulated by STIM1 via CD36, might be implicated in oro-gustatory perception of dietary lipids and the spontaneous preference for fat
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Usui, Ryota. "GPR40 activation initiates store-operated Ca²⁺ entry and potentiates insulin secretion via the IP3R1/STIM1/Orai1 pathway in pancreatic β-cells." Kyoto University, 2020. http://hdl.handle.net/2433/253196.
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Gueder, Nahla. "sp²-Iminosugar-glucosidases inhibitor 1-C-octyl-2-oxa-3-oxocastanospermine - induced antiproliferative, apoptotic and necrotic effects in breast cancer cells via targeting GRP78, Stim1 and Orai1." Electronic Thesis or Diss., Amiens, 2018. http://www.theses.fr/2018AMIE0033.
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L'altération de glycosylation est l'une des caractéristiques du cancer du sein. Ainsi le défaut de glycosylation affecte différentes protéines glycosylées responsables des différents processus cancéreux. Les canaux SOC (Store operated channels) constituent la voie majeure de l'entrée du calcium dans les cellules et sont impliqués dans la prolifération, la migration et la survie des cellules cancéreuses du sein. CO-OCS est un nouvel inhibiteur de la glycosylation avec plus de sélectivité vis-à-vis des α-glucosidases, et montre des activités anticancéreuses des cellules cancéreuses du sein, sans affecter les cellules mammaires normales. L'objectif de ma thèse est d'étudier les mécanismes moléculaires par lesquels CO-OCS induit ses effets anti-tumoraux. CO-OCS inhibe la migration des cellules cancéreuses à fort potentiel métastatique. Cet effet anti-migratoire est dû à une réduction de l'expression de la β1-intégrine, de Stim1, et de l'activation des voies de signalisation FAK et ERK1/2 par CO-OCS. Dans les cellules cancéreuses peu invasives, CO-OCS diminue la prolifération et augmente la mortalité de ces cellules en affectant l'expression de 3 protéines : Stim1 et Orai1 : protéines N-glycosylées au niveau du réticulum endoplasmique (RE), et GRP78, protéine de stress du RE. Ainsi en supprimant complétement l'expression de Stim1, CO-OCS réduit la prolifération en accumulant les cellules dans les phases G1 et G2/M du cycle cellulaire. Alors que la réduction de l'expression de GRP78 et d'Orai1 par le CO-OCS augmente respectivement l'apoptose et la nécrose. Par ailleurs, l'invalidation de Stim1 atténue l'effet apoptotique induit par CO-OCS. CO-OCS réduit aussi le contenu calcique du RE. Cette réduction du calcium réticulaire est due à une fuite de calcium par le Translocon. En effet, l'Anisomycine, inhibiteur du Translocon, restore de contenu calcique réticulaire et antagonise l'apoptose induite par le CO-OCS. En conclusion, CO-OCS induit une accumulation de protéines mal-repliées dans le RE induisant ainsi un stress réticulaire. Trois cibles du CO-OCS ont été identifiées : l'expression de Stim1 favorise la prolifération tandis que celle d'Orai1 et de GRP78 protègent respectivement les cellules de l'apoptose et de la nécrose induites par CO-OCS. De plus, en diminuant l'expression de GRP78, CO-OCS induit une fuite du calcium du RE par le TransloconAlteration in glycosylation pattern is one of the hallmarks of breast cancer. The levels and the abnormal expressions of glycan were found in breast cancer patients. Glycosylation defect can affect different glycosylated proteins which are implicated in cancerogenesis. Changes in intracellular Ca2+ levels can regulate different cellular processes. SOC channels are implicated in breast cancer proliferation, migration and survival. CO-OCS is a new glycosylation inhibitor with more selectivity toward theα- glucosidases exhibited anti-cancer activities in breast cancer cells without affecting the normal mammary cells. The objective of my thesis is investigating the related molecular mechanisms by which CO-OCS induced its anti-tumour effects.CO-OCS impaired breast cancer migration through decrease β1-integrin expression and the activation of FAK and ERK1/2 signalling pathways. CO-OCS also induced anti-migratory effect via Stim1 protein expression down-regulation leading to inhibition of SOCE. Additionally, CO-OCS affected the expression of both Orai1 and Stim1 proteins leading to anti-proliferative effects and cell cycle arrest in G1 and G2/M phase respectively. Moreover, CO-OCS affected the expression of Stim1 at the protein level without affecting its transcript level. GRP78 implicated in CO-OCS apoptotic death. The expression of Stim1 regulated the apoptosis induced by CO-OCS via modulating GRP78 expression. Orai1 down-regulation promoted CO-OCS necrotic effect. CO-OCS induced ER- calcium depletion due to increase in ER calcium leak via the Translocon; Anisomycin (Translocon inhibitor) decreased the apoptosis induced by CO-OCS. In conclusion, these results show that in breast cancer, by targeting Stim1, Orai1 and GRP78, CO-OCS reduced cell proliferation and induced apoptosis and necrosis cell death. Stim1 favours CO-OCS apoptotic effect while Orai1 protected from necrosis induced by CO-OCS. The inhibition of Translocon decreased CO-OCS apoptotic cell death via restoring the ER calcium homeostasis
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Peche, Georges Arielle. "Physiopathologie de la myopathie à agrégats tubulaires." Thesis, Strasbourg, 2017. http://www.theses.fr/2017STRAJ008/document.
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La myopathie à agrégats tubulaires (TAM) est une maladie génétique qui se caractérise par la présence d’agrégats tubulaires dans les biopsies musculaires de patients. Notre équipe a identifié pour la première fois des mutations dans STIM1 comme étant à l’origine de cette maladie. STIM1 (stromal interaction molecule 1) est le senseur calcique du réticulum sarco/endoplasmique (RE/RS). En effet, en cas de diminution du calcium (Ca2+) dans le RE/RS, STIM1 se déplie, oligomérise et migre à proximité de la membrane plasmique (MP) pour activer le canal calcique ORAI1 et permettre le remplissage des stocks. Ce mécanisme est le «store-operated Ca2+ entry» (SOCE). D’autres équipes ont rapporté une mutation dans STIM1 (p.R304W) conduisant à une TAM associée à d’autres symptômes, ou encore syndrome de Stormorken. Ainsi, ce travail a eu pour but d’étudier et de comparer l’impact des mutations TAM et Stormorken à différents niveaux du SOCE. Nous avons ainsi montré que les mutations TAM et Stormorken conduisent à une augmentation de l’expression de STIM1, à la formation de clusters constitutifs de STIM1 à proximité de la MP, ainsi qu’au recrutement du canal ORAI1 et à l’activation de la voie du NFAT, dépendante du Ca2+Tubular aggregate myopathy (TAM) is a genetic disorder characterized by tubular aggregates in muscle biopsies of patients. Our team identified for the first time mutations in STIM1 as causative of this disease. STIM1 (stromal interaction molecule 1) is the main calcium (Ca2+) sensor of the endo/sarcoplasmic reticulum (ER/SR). Following Ca2+ depletion of the ER/SR, STIM1 unfolds, oligomerizes and migrates close to the plasma membrane (PM) to activate the Ca2+ channel ORAI1, leading to Ca2+ entry. This mechanism is the «store-operated Ca2+ entry» (SOCE). Several teams report a mutation in STIM1 (p.R304W) leading to TAM associated with other symptoms, described as Stormorken syndrome. Therefore, this work aims to assess and compare the impact of TAM and Stormorken mutations at different stages of the SOCE pathway. We show that TAM and Stormorken mutations lead to an increase expression of the protein, a constitutive STIM1 clustering near the PM, to ORAI1 constitutive recruitment and to the activation of a Ca2+ -dependent pathway: the NFAT pathway
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Huang, Yun. "Integration of Extracellular and Intracellular Calcium Signals: Roles of Calcium-Sensing Receptor (CASR), Calmodulin and Stromal Interaction Molecule 1 (STIM1)." Atlanta, Ga. : Georgia State University, 2008. http://digitalarchive.gsu.edu/chemistry_diss/28/.
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Thesis (Ph. D.)--Georgia State University, 2008.Title from title page (Digital Archive@GSU, viewed July 1, 2010) Jenny J. Yang, committee chair; Edward Brown, Giovanni Gadda, Zhi-ren Liu, committee members. Includes bibliographical references (p. 230-258).
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Ramanujam, Deepak Prabhu [Verfasser], Martin [Akademischer Betreuer] Klingenspor, and Stefan [Akademischer Betreuer] Engelhardt. "Characterization of stromal interaction molecule 1 (STIM1) in the myocardium / Deepak Prabhu Ramanujam. Betreuer: Martin Klingenspor. Gutachter: Stefan Engelhardt ; Martin Klingenspor." München : Universitätsbibliothek der TU München, 2013. http://d-nb.info/1079001891/34.
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Cordero, Sanchez Celia. "Characterization of a mouse model for Tubular Aggregate Myopathy and development of small molecules." Doctoral thesis, Università del Piemonte Orientale, 2022. https://hdl.handle.net/11579/148545.
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Tubular aggregate myopathy (TAM) is one of a cluster of rare genetic diseases, together with Stormorken syndrome and York platelet syndrome (YPS). The aetiology of these diseases is the mutation in one of two key proteins, ORAIi and STIMÍ. Both proteins are the principai protagonists in Store-Operated Calcium Entry (SOC Entry), a mechanism of calcium homeostasis. Up to now, no mouse model has been designed bearing a luminal STIM1 mutation associated with the clinical diagnosis of any of the diseases. A mouse model bearing 11 15F mutation, which is associated with TAM and YPS, is extremely needed, to guarantee the effectiveness of putative treatment in patients bearing luminal STIM1 mutations. This thesis project demonstrates that the KI-STIM" mouse model is valid for both TAM and YPS, confirming at the same time the hypothesis of some authors, who defend that TAM, Stormorken, and YPS are indeed the spectra of the same disease whose symptoms differences are base in the position and effect of the point mutation.
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Mariotti, Valeria. "Molecular pathophysiology of cholangiopathies: lessons from polycystic and fibropolycystic liver disease." Doctoral thesis, Università degli studi di Padova, 2019. http://hdl.handle.net/11577/3422326.
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Cholangiocytes, the epithelial cells lining the intra- and extrahepatic biliary tree, play an essential role in regulating the production of bile and digestive functions. Cholangiocytes are also involved in the repair from liver damage and are also the target of cholangiopathies a group of acquired, congenital, or genetic complex chronic liver diseases , that represents a substantial cause of morbidity and mortality and a frequent indication for liver transplant. Treatment of cholangiopathies is unsatisfactory and these diseases are among the main unmet needs in modern hepatology.
Cholangiocytes in addition of being the target of the disease, also contribute to disease development. After biliary damage cholangiocytes, become “reactive”, acquire the ability to produce a number of cytokines and inflammatory mediators and growth factors and to promote further inflammation and portal fibrosis that ultimately may lead to cirrhosis and chronic liver failure. A hallmark of this process is the activation of intracellular morphogenetic signaling, such as the Notch, Wnt/β-catenin and VEGF pathways that are transiently involved in the development of the biliary epithelium during embryogenesis. The understanding of the mechanism underlying the reactivation of these intracellular signaling pathways has been boosted thanks to the availability of animal models that phenocopy the congenital and genetic cholangiopathies. These translational tools improved the knowledge of the pathophysiology of cholangiocytes and provided experimental therapeutic strategies.
Acquired cholangiopathies are complex diseases for which only unsatisfactory animal and cellular models are available. Therefore, the approach taken by our laboratory has been to study the pathogenesis of genetic cholangiopathies with an identified causative gene and derive “lessons” that can be applied to the more general field of biliary and liver diseases.
During the Ph.D. program, I mainly focused on investigating the molecular mechanism that drives the progression of two inherited cholangiopathies: 1) the polycystic liver disease associated with autosomal dominant polycystic kidney disease (PLD-ADPKD), in which the abnormal proliferation of cystic cholangiocytes is associated with changes in intracellular Ca2+ homeostasis and with the reactivation of biliary angiogenetic signals and, 2) the fibro-polycystic disease (Caroli Disease and Congenital Hepatic Fibrosis), where the aberrant lack of fibrocystin leads to activation of β-Catenin, chemokine secretion and recruitment of macrophages and fibrogenic mesenchymal cells , leading to liver fibrosis and portal hypertension. The thesis is divided in two parts; for both parts, the experimental methodology used is reported in the method section.
In part I, we show that polycystin 2 (PC2), the ciliary protein affected in PLD-ADPKD, is a regulator of Ca2+ homeostasis and cAMP signaling in cholangiocytes. Expression of PC2 is essential for cytoplasmic and endoplasmic reticulum (ER) calcium handling. As PC2-deficient cells are also characterized by increased cAMP/PKA signaling and by Ras/Raf/ ERK/mTOR/HIF1α-dependent VEGF secretion, we aimed at understanding the relationships between altered Ca2+ homeostasis and increased cAMP production. We found that increased cAMP production is sustained by Adenylyl Cyclase 5 (AC5), an AC that is inactive at the normal intracellular Ca2+ concentrations,but it is activated at the decreased intracellular Ca2+ concentrations measured in PC2-deficient cystic cholangiocytes. As activation of AC5 was also dependent on STIM1 (a ER calcium sensor protein responsible for activation of the store-operated Ca2+ entry) we also concluded that the absence of PC2, intracellular Ca2+ store depletion promotes the interaction of STIM1 with AC5, resulting in aberrant cAMP production, stimulation of cystic cholangiocyte proliferation and cyst growth via Ras/Raf/ERK/mTOR/HIF1α-dependent VEGF secretion. These findings indicate that AC5 is an attractive target for therapy, as also demonstrated by inhibition of cyst growth in mice treated with the AC5 inhibitor SQ22,536. (Part of this work has been published. Spirli C., et al., Journal of Hepatology, 2017)
In part II, we studied how fibrosis develops in Caroli Disease and Congenital Hepatic Fibrosis, two variants of the fibropolycystic diseases resulting from congenital deficiency of fibrocystin and characterized by biliary cysts and severe liver fibrosis. In prior studies we showed that in this condition, fibrosis was not a consequence of necroinflammatory damage to the biliary tree, but rather of a low grade chronic inflammation originating from the fibrocystin-deficient cholangiocytes and the consequent recruitment of macrophages. As macrophages recruitment depended on increased secretion of CXCL10 by fibrocystin-defective cholangiocytes we aimed to better understand the link between fibrocystin deficiency and increased CXCL10 secretion. We found that in the absence of fibrocystin, CXCL10 secretion is mediated by β-catenin nuclearization through phosphorylation at the Serine675 and by the abnormal secretion of IL-1β through the activation of NLRP3 inflammasome complex. These findings suggest that periportal fibrosis can be generated by a chronic low level inflammation originating from the fibrocystin deficient cholangiocytes and open new therapeutic avenues aiming at targeting β-catenin signaling and the inflammasome. (This work has been published. Kaffe E. et al., Hepatology, 2018)
The identification of these pathobiological mechanisms has clarified some general aspects of the sequence of events leading from cholangiocyte dysfunction to hyperplasia and biliary fibrosis. This process is the result of increased cAMP levels, secretion of pro-inflammatory chemokines and angiogenetic factors, and macrophage recruitment; all these steps which may be targeted also for the treatment of acquired cholangiopathies that share common pathobiological traits.
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Castro, Kraftchenko Joel, and kraf0005@flinders edu au. "STORE OPERATED Ca2+ CHANNELS IN LIVER CELLS: REGULATION BY BILE ACIDS AND A SUB-REGION OF THE ENDOPLASMIC RETICULUM." Flinders University. Medicine, 2008. http://catalogue.flinders.edu.au./local/adt/public/adt-SFU20080826.135311.
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Cholestasis is an important liver pathology. During cholestasis bile acids accumulate in the bile canaliculus affecting hepatocyte viability. The actions of bile acids require changes in the release of Ca2+ from intracellular stores and in Ca2+ entry. The target(s) of the Ca2+ entry pathway affected by bile acids is, however, not known. The overall objective of the work described in this thesis was to elucidate the target(s) and mechanism(s) of bile acids-induced modulation of hepatocytes calcium homeostasis.
First, it was shown that a 12 h pre-incubation with cholestatic bile acids (to mimic cholestasis conditions) induced the inhibition of Ca2+ entry through store-operated Ca2+ channels (SOCs), while the addition of choleretic bile acids to the incubation medium caused the reversible activation of Ca2+ entry through SOCs. Moreover, it was shown that incubation of liver cells with choleretic bile acids counteracts the inhibition of Ca2+ entry caused by pre-incubation with cholestatic bile acids. Thus, it was concluded that SOCs are the target of bile acids action in liver cells.
Surprisingly, despite the effect of choleretic bile acids in activating SOCs, the Ca2+ dye fura-2 failed to detect choleretic bile acid-induced Ca2+ release from intracellular stores in the absence of extracellular Ca2+. However, under the same conditions, when the sub-plasma membrane Ca2+ levels were measured using FFP-18 Ca2+ dye, choleretic bile acid induced a transient increase in FFP-18 fluorescence. This evidence suggested that choleretic bile acids-induced activation of Ca2+ entry through SOCs, involving the release of Ca2+ from a region of the endoplasmic reticulum (ER) located in the vicinity of the plasma membrane.
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Castiblanco, Adriana P. "Expression and Purification of Engineered Calcium Binding Proteins." Digital Archive @ GSU, 2009. http://digitalarchive.gsu.edu/chemistry_theses/20.
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Previous studies in Dr. Yang’s laboratory have established a grafting, design, and subdomain approach in order to investigate the properties behind Ca2+-binding sites located in Ca2+-binding proteins by employing engineered proteins. These approaches have not only enabled us to isolate Ca2+-binding sites and obtain their Ca2+-binding affinities, but also to investigate conformational changes and cooperativity effects upon Ca2+ binding. The focus of my thesis pertains to optimizing the expression and purification of engineered proteins with tailored functions. Proteins were expressed in E. coli using different cell strains, vectors, temperatures, and inducer concentrations. After rigorous expression optimization procedures, proteins were further purified using chromatographic and/or refolding techniques. Expression and purification optimization of proteins is essential for further analyses, since the techniques used for these studies require high protein concentrations and purity. Evaluated proteins had yields between 5-70 mg/L and purities of 80-90% as confirmed by SDS-PAGE electrophoresis.
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Nguyen, Nathalie. "Régulation négative de l'activité des canaux calciques : identification de deux nouveaux frins biologiques." Thèse, Université de Sherbrooke, 2013. http://hdl.handle.net/11143/6248.
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Le Ca[indice supérieur 2+] joue un rôle primordial dans plusieurs processus physiologiques comme la contraction, la sécrétion, la croissance et la prolifération cellulaire et même l’apoptose. Ainsi, une régulation adéquate des niveaux de Ca[indice supérieur 2+] dans la cellule est essentielle à son bon fonctionnement. Plusieurs protéines-clés sont impliquées dans cette homéostasie calcique. Cette étude a permis d’identifier deux mécanismes distincts responsables de friner la signalisation calcique dans un modèle de cellules non-excitables et dans un modèle de cellules excitables, en mettant l’emphase sur la régulation des canaux calciques par leur interaction avec des protéines clés. Le premier mécanisme implique la régulation du récepteur à l’inositol 1,4,5-trisphosphate (IP[indice inférieur 3]R) par la protéine chaperonne Hsp90 dans les cellules HEK293T. J’ai montré que Hsp90 interagit avec l'IP[indice inférieur 3]R dans le but de diminuer son activité. De plus, cette interaction dépend de la voie de signalisation de l’insuline, plus particulièrement des protéines kinases mTOR et Src, qui sont impliquées dans cette interaction. Ainsi, l’interaction entre Hsp90 et l'IP[indice inférieur 3]R fait partie d’un mécanisme de rétroaction négative de la voie de signalisation de l’insuline. Le second mécanisme implique la régulation des canaux calciques dépendants du voltage de type T (T-VDCC) par la protéine STIM1 dans les cardiomyocytes HL-1. J'ai montré que STIM1, qui est le principal senseur de Ca[indice supérieur 2+] du réticulum sarco/endoplasmique, interagit avec les T-VDCC afin de diminuer leur expression à la membrane cellulaire et ainsi atténuer leur activité. Cette interaction permet de prévenir une entrée excessive de Ca[indice supérieur 2+] dans la cellule menant à une surcharge de Ca[indice supérieur 2+] dans le réticulum sarcoplasmique et, par le fait même, à des événements arythmiques associés à la surcharge. Bien que ces deux études aient été effectuées dans des environnements différents, soit un modèle de cellules non-excitables et un modèle de cellules excitables, elles mettent en évidence deux mécanismes pouvant friner la signalisation calcique dans le but d’assurer une fonction normale à la cellule.
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Eylenstein, Anja [Verfasser], and Florian [Akademischer Betreuer] Lang. "Regulierung des speicherabhangigen Ca2+-Kanals Orai1 und des Ca2+-sensitiven Proteins STIM1 durch die Serum- und Glukokortikoid induzierbare Kinase 1 / Anja Eylenstein ; Betreuer: Florian Lang." Tübingen : Universitätsbibliothek Tübingen, 2011. http://d-nb.info/1160309620/34.
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Henke, Nadine Verfasser], Axel [Akademischer Betreuer] Methner, and Dieter [Akademischer Betreuer] [Willbold. "How Stromal Interaction Molecule 1 (STIM1) and Store Operated Calcium Entry (SOCE) affect mitochondrial energy metabolism and neuronal function / Nadine Henke. Gutachter: Axel Methner ; Dieter Willbold." Düsseldorf : Universitäts- und Landesbibliothek der Heinrich-Heine-Universität Düsseldorf, 2013. http://d-nb.info/1031074910/34.
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Henke, Nadine [Verfasser], Axel Akademischer Betreuer] Methner, and Dieter [Akademischer Betreuer] [Willbold. "How Stromal Interaction Molecule 1 (STIM1) and Store Operated Calcium Entry (SOCE) affect mitochondrial energy metabolism and neuronal function / Nadine Henke. Gutachter: Axel Methner ; Dieter Willbold." Düsseldorf : Universitäts- und Landesbibliothek der Heinrich-Heine-Universität Düsseldorf, 2013. http://d-nb.info/1031074910/34.
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Cohen-Aubart, Fleur. "Rôle de la protéine STIM1 sur la fonction des cellules musculaires vaculaires et cardiaques par modulation de la voie de signalisationcalcineurine-NFAT : implications physiopathologiques et pharmacologiques." Paris 6, 2012. http://www.theses.fr/2012PA066169.
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Le calcium est un second messager universel régulant de multiples fonctions intra-cellulaires grâce à des mouvements compartimentés dans l'espace et de dynamique variable. Les mouvements rapides localisés régulent les fonctions nécessitant des réponses rapides (contraction, exocytose) alors que les mouvements répétés ou larges dans le temps ou l'espace régulent des fonctions plus lentes (régulation de l'expression génique). Les mouvements calciques sont altérés de façon adaptative ou pathologique dans les situations pathologiques.
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Tian, Geng. "On the Generation of cAMP Oscillations and Regulation of the Ca2+ Store-operated Pathway in Pancreatic Islet α- and β-cells." Doctoral thesis, Uppsala universitet, Institutionen för medicinsk cellbiologi, 2013. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-191852.
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Insulin and glucagon are released in pulses from pancreatic β- and α-cells, respectively. Both cell types are electrically excitable, and elevation of the cytoplasmic Ca2+ concentration ([Ca2+]i) due to depolarization with voltage-dependent entry of the cation is the main trigger of hormone secretion. Store-operated Ca2+ entry (SOCE) also contributes to the [Ca2+]i elevation and this process has been suggested to be particularly important for glucagon secretion. cAMP is another important messenger that amplifies Ca2+-triggered secretion of both hormones, but little is known about cAMP dynamics in islet cells. In type-2 diabetes, there is deteriorated β-cell function associated with elevated concentrations of fatty acids, but the underlying mechanisms are largely unknown. To clarify the processes that regulate insulin and glucagon secretion, cAMP signalling and the store-operated pathway were investigated in β- and α-cells, primarily within their natural environment in intact mouse and human islets of Langerhans. Fluorescent biosensors and total internal reflection microscopy were used to investigate signalling specifically at the plasma membrane (PM). Adrenaline increased and decreased the sub-PM cAMP concentration ([cAMP]pm) in immuno-identified α-cells and β-cells, respectively, which facilitated cell identification. Glucagon elicited [cAMP]pm oscillations in α- and β-cells, demonstrating both auto- and paracrine effects of the hormone. Whereas glucagon-like peptide 1 (GLP-1) consistently elevated [cAMP]pm in β-cells, only few α-cells responded, indicating that GLP-1 regulates glucagon secretion without changes of α-cell [cAMP]pm. Both α- and β-cells responded to glucose with pronounced oscillations of [cAMP]pm that were partially Ca2+-dependent and synchronized among islet β-cells. The glucose-induced cAMP formation was mediated by plasma membrane-bound adenylyl cyclases. Several phosphodiesterases (PDEs), including the PDE1, -3, -4, and -8 families, were required for shaping the [cAMP]pm signals and pulsatile insulin secretion. Prolonged exposure of islets to the fatty acid palmitate deteriorated glucose-stimulated insulin secretion with loss of pulsatility. This defect was associated with impaired cAMP generation, while [Ca2+]i signalling was essentially unaffected. Stromal interacting molecule 1 (STIM1) is critical for activation of SOCE by sensing the Ca2+ concentration in the endoplasmic reticulum (ER). ER Ca2+ depletion caused STIM1 aggregation, co-clustering with the PM Ca2+ channel protein Orai1 and SOCE activation. Glucose, which inhibits SOCE by filling the ER with Ca2+, reversed the PM association of STIM1. Consistent with a role of the store-operated pathway in glucagon secretion, this effect was maximal at the low glucose concentrations that inhibit glucagon release, whereas considerably higher concentrations were required in β-cells. Adrenaline induced STIM1 translocation to the PM in α-cells and the reverse process in β-cells, partially reflecting the opposite effects of adrenaline on cAMP in the two cell types. However, cAMP-induced STIM1 aggregates did not co-cluster with Orai1 or activate SOCE, indicating that STIM1 translocation can occur independently of Orai1 clustering and SOCE.
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Dramane, Gado, and Gado Dramane. "Etude de la signalisation calcique dans les cellules gustatives lipidiques chez la souris." Phd thesis, Université de Bourgogne, 2012. http://tel.archives-ouvertes.fr/tel-00833888.
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Les personnes en surcharge pondérale semblent préférer une alimentation riche en graisse. Face à l'épidémie d'obésité qui touche nos Sociétés tant urbaines que rurales, élucider les mécanismes de la détection des lipides alimentaires devient un enjeu majeur. Il avait précédemment été admis que la glycoprotéine CD36 exprimée dans les papilles caliciformes de souris, est impliquée dans la perception oro-gustative des lipides alimentaires. Dans ce travail, nous avons montré que l'acide linoléique (LA), en activant les phospholipases A2, sPLA2, cPLA2 et iPLA2 via CD36, produit de l'acide arachidonique (AA) et la lyso-phosphatidylcholine (lyso-PC). LA déclenche un influx calcique dans les cellules CD36-positives et induit la production du facteur CIF (Calcium Influx Factor). CIF, AA et lyso-PC exercent différentes actions sur l'ouverture des canaux SOC (Stored Operated Calcium Channel) constitués de protéines Orai et contrôlés par STIM1. Stim1 est un senseur calcique situé sur la membrane du réticulum endoplasmique activé par la déplétion du calcium intracellulaire. Nous avons utilisé la technologie siRNA et des modèles de souris transgéniques pour montrer que CIF et lyso-PC activent des canaux calciques homodimériques composés de protéines Orai1 tandis qu'AA active des canaux hétéro-pentamériques composés d'Orai1 et Orai3. Nous avons également montré que STIM1 régule la production de CIF dans les cellules stimulées par la thapsigargine et l'acide linoléique ainsi que l'ouverture de deux types de canaux calciques. Par ailleurs les souris au phénotype Stim1-/- perdent la préférence spontanée pour les lipides observé chez les animaux de type sauvage. D'un autre côté les cellules CD36-positive de souris Stim1-/- sont incapables de libérer la sérotonine dans l'environnement extracellulaire. Nos résultats suggèrent que des acides gras à longue chaine (AGLC) induisent la signalisation calcique régie par STIM1 via CD36. La perception oro-gustative des lipides alimentaires détermine la préférence spontanée pour les lipides observée chez les mammifères
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Arunachalam, Sasi. "The Role of store operated calcium channels in human carcinoid cell lines." University of Toledo Health Science Campus / OhioLINK, 2010. http://rave.ohiolink.edu/etdc/view?acc_num=mco1279216983.
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Abdoul-Azize, Souleymane. "Implication de la signalisation calcique et des MAP kinases dans la perception gustative lipidique." Phd thesis, Université de Bourgogne, 2013. http://tel.archives-ouvertes.fr/tel-01018378.
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Dans ce travail, nous démontrons que STIM1, un senseur calcique activé par la déplétion du Ca2+ intracellulaire du réticulum endoplasmique, est indispensable pour la signalisation calcique et la préférence oro-sensorielle du gras. Nous observons que l'acide linoléique (LA), en activant les phospholipases A2 via CD36, produit de l'acide arachidonique (AA) et de la lyso-phosphatidylcholine (lyso-PC). Cette activation déclenche un influx calcique dans les cellules CD36-positives, et induit la production du facteur CIF (Ca2+ Influx Factor). CIF, AA et lyso-PC exercent différentes actions sur l'ouverture des canaux SOC (Stored Operated Calcium Channel) constitués de protéines Orai et contrôlés par STIM1. Par ailleurs, les souris au phénotype Stim1-/- perdent la préférence spontanée pour les lipides et la libération de la sérotonine à partir des cellules gustatives dans le milieu extracellulaire chez les animaux sauvages. Nous demontrons aussi que la signalisation calcique médiée via CD36 est doublement modulée lors de l'obésité. L'augmentation de la [Ca2+]i dans les cellules gustatives observée chez le Psammomys obesus, un modèle d'obésité nutritionelle, est fortement diminuée chez les souris rendues obèses par un regime hyperlipidique. Nous avons constaté également que l'interaction de LA avec le CD36 induit l'activation des MAP Kinases de la voie MEK1/2/ERK1/2/Elk-1 qui est non seulement à l'origine de l'activation des aires cérébrales telles que le NTS, le noyau arqué, l'hippocampe mais aussi indispensable pour la préférence spontanée pour les lipides alimentaires. Nos résultats suggèrent pour la prémière fois, que la voie ERK1/2 des MAPK et la signalisation calcique lipidique controlée par STIM1 sont impliquées dans la perception oro-gustative des lipides
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Terrie, Élodie. "Rôle de la signalisation calcique dépendante des Store-Operated Channels (SOC) dans les cellules souches neurales adultes et les cellules souches cancéreuses de glioblastomes." Thesis, Poitiers, 2019. http://www.theses.fr/2019POIT2322.
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Des cellules souches neurales (CSN) persistent dans le cerveau adulte et produisent des neurones et des cellules gliales tout au long de la vie de l’individu. Les CSN suscitent un intérêt considérable pour la médecine régénératrice mais leur utilisation thérapeutique potentielle nécessite au préalable d’approfondir les connaissances sur leurs mécanismes de régulation. Les glioblastomes, quant à eux, sont les tumeurs cérébrales les plus fréquentes chez l’adulte et les plus mortelles. Au sein de ces tumeurs, les cellules souches de glioblastomes (CSG) seraient issues de la transformation maligne des CSN et seraient responsable de l’initiation, de la propagation et de la résistance aux traitements des tumeurs. Des analyses transcriptomiques ont suggéré un rôle majeur de la signalisation calcique au sein des CSN et des CSG. Représentant une des voies principales d’entrée du calcium dans la cellule, les canaux calciques SOC (Store-Operated Channels) régulent de nombreux processus cellulaires, y compris la progression tumorale. L’objectif des travaux de cette thèse est d’évaluer le rôle des SOC dans les CSN et les CSG.Nous avons établi par des approches in vitro et in vivo, que les CSN de souris adulte expriment des SOC fonctionnels et que leur inhibition pharmacologique diminue la prolifération et l’autorenouvellement des CSN, propriété indispensable au maintien de la population souche. La deuxième partie de nos travaux montre que les CSG issues de cultures primaires de patients expriment des SOC dont l’inhibition altère la prolifération et l’autorenouvellement de ces cellules.Ainsi, les résultats obtenus lors de cette thèse mettent en évidence un rôle essentiel des SOC dans la régulation de l’autorenouvellement des CSN et des CSG. Les CSG étant responsables de la résistance aux traitements dans le glioblastome, ces travaux ouvrent des perspectives thérapeutiques ciblant les canaux calciques pour contrer cette pathologie au pronostic sombreNeural stem cells (NSC) persist in the brain of adult mammals and fuel the brain with new neurons and glial cells all lifelong. Recruited by brain injuries, NSC are considered with great interest by regenerative medicine. However, the development of new therapeutic approaches based on the use of NSC requires an in-depth knowledge of the mechanism regulating these cells. Glioblastomas are the most frequent and deadliest form of adult brain tumors. Within the tumor, glioblastoma stem cells (GSC) form a subpopulation of cells that is considered as responsible of tumor initiation, propagation and relapse, as these cells are particularly resistant to anti-tumoral treatments. GSC and NSC share key characteristics and numerous studies suggest that GSC arise from transformed NSC. Transcriptomic analysis of NSC and of GSC revealed an enrichment of calcium signaling transcripts in these two cell populations. Representing a major way of calcium influx into cells, Store-Operated Channels (SOC) are mobilized in response to a wide range of extracellular factors. SOC regulate many cellular processes and are often hijacked in cancer to promote tumor progression.The aim of this thesis is to evaluate potential SOC involvement in NSC and GSC regulation.The first part of this work, relying on in vitro and in vivo studies, demonstrates that NSC from adult mice express functional SOC whose inhibition by pharmacological agents reduces NSC proliferation and self-renewal. In the second part of this thesis, we demonstrate that GSC from primary cultures derived from patients express SOC, as do NSC, and that SOC inhibition reduces GSC ability to proliferate and self-renew.Accordingly, the results of this thesis demonstrate that SOC regulate NSC and GSC self-renewal, a property that is essential to maintain stem cells pool. As GSC are responsible for glioblastomas treatment resistance, our studies point to a potential new therapeutic way, via calcium channels, against this deadly pathology
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Pancaldi, Marco. "Stime satellitari di stabilità atmosferica." Bachelor's thesis, Alma Mater Studiorum - Università di Bologna, 2019. http://amslaurea.unibo.it/18271/.
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In questo lavoro si è affrontato lo studio degli indici di stabilità atmosferica prodotti da Eumetsat attraverso un algoritmo che sfrutta i dati del sensore SEVIRI a bordo del satellite geostazionario europeo MSG. L'obiettivo del lavoro è stato di valutare la loro efficacia nel prevedere fenomeni di precipitazione.
Sono stati considerati i prodotti di giornate in cui intense precipitazioni hanno interessato l'Emilia-Romagna nell'estate del 2018 così da poter confrontare le stime da satellite di stabilità con i dati dei radar e delle stazioni meteorologiche. Quattro diversi indici sono stati considerati: Lifted Index, K-Index, Konvektiv Index e Maximum Buoyancy.
L'analisi dei cinque casi di precipitazione è stata condotta analizzando l'evoluzione temporale delle mappe degli indici nelle ore antecedenti l'inizio della precipitazione, come misurata da alcune stazioni pluviometriche. In genere si è trovato che tutti gli indici testati evolvono durante la giornata su valori che descrivono un aumento della instabilità. Al primo formarsi della nuvolosità, tuttavia, il calcolo degli indici non viene più effettuato, e l'informazione non è più disponibile. In genere, comunque, si è notato che la precipitazione inizia dalle 3 alle 6 ore dopo che gli indici di stabilità hanno raggiunto il loro valore più critico. Invece si è notato che non vi è correlazione tra il valore degli indici prima della precipitazione e l'intensità della precipitazione.
Questo lavoro ha mostrato l'utilità delle stime da satellite di indici di stabilità, in quanto consentono un monitoraggio della stabilità atmosferica con maggiore dettaglio spazio temporale rispetto agli indici calcolati da dati da radiosonde. Gli aspetti negativi sono l'impossibilità di aver il dato al primo presentarsi della nuvolosità e l'incertezza nella stima dei profili di temperatura e umidità.
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Cesarini, Ettore. "Stima streaming di sottospazi principali." Master's thesis, Alma Mater Studiorum - Università di Bologna, 2019. http://amslaurea.unibo.it/17897/.
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Nell'attuale stato tecnologico in cui la dimensionalità dei dati vede un incremento esponenziale, si rende necessario l'utilizzo di algoritmi che permettano un'efficiente signal processing anche ai dispositivi con potenza di calcolo e disponibilità di memoria limitati. In questo lavoro è stato preso in considerazione un setting reale, per il quale si sono cercati due metodi algoritmici in grado di garantirne un'efficiente stima dell'energia del segnale, in modo streaming e riducendo la dimensionalità dei dati. Traendo ispirazione dallo stato dell'arte, sono stati decisi i due metodi di stima streaming di sottospazi principali HPCA e SEPS. In sede di simulazione si è fatto uso di dataset sintetici appositamente generati per ottenere un opportuno tuning dei parametri per ciascun algoritmo. In seguito si è proceduto a valutare l'efficacia delle scelte fatte sui parametri applicando i singoli algoritmi al setting reale. Nel medesimo setting, al passo successivo si è proceduto al confronto diretto dei due metodi valutandone le performance in termini di dinamica e valore di convergenza, costi computazionali e robustezza in seguito a fenomeni di carattere non stazionario. I risultati hanno favorito l'algoritmo HPCA in termini di prestazioni e robustezza, a scapito di una maggiore incidenza sulla complessità computazionale e memory footprint rispetto al secondo algoritmo. SEPS ha reso la fase di tuning un passo molto più delicato, dovuta all'alta sensibilità del suo learning rate; tuttavia a seguito dell'opportuna paramettrizzazione, ha primeggiato relativamente ai costi computazionali e al memory footprint, dando prova di alta flessibilità, semplicità implementativa ma minore velocità di convergenza. Ai fini di questa elaborazione, sia HPCA che SEPS si sono rivelati una scelta corretta e soddisfacente nell'ottica della stima streaming di sottospazi principali, facendo prediligere il secondo per le performance ottenute nello specifico setting qui analizzato.
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Dotti, Michele. "Stima dell'assetto di un UAV." Bachelor's thesis, Alma Mater Studiorum - Università di Bologna, 2014. http://amslaurea.unibo.it/6362/.
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Lo scopo del lavoro è simulare il comportamento di un sistema di misura dell'assetto detto ARS (Attitude Reference System), dove sostanzialmente le misure fornite da giroscopi ed accelerometri, quindi accelerazioni e velocità angolari, vengono elaborate da un filtro osservatore dello stato che permette di ricavare la stima dell'angolo di elevazione Q e dell'angolo di inclinazione f, non misurabili direttamente, e quindi dell'orientamento del sistema rispetto al piano orizzontale.
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Ciaroni, Riccardo. "Stima dello stato nelle reti di distribuzione." Master's thesis, Alma Mater Studiorum - Università di Bologna, 2018. http://amslaurea.unibo.it/17249/.
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La stima dello stato di un sistema elettrico rappresenta un processo di stima di un gruppo di variabili di stato, in un preciso istante di tempo, che lo descrivano in modo univoco partendo da un certo numero di misure tenendo conto delle incertezze ad esse correlate. Conoscere lo stato di un sistema è fondamentale per una gestione efficiente, in sicurezza e nel rispetto dei vincoli tecnici, delle reti elettriche.
La tesi ha come obiettivo la procedura di stima dello stato mediante il metodo dei minimi quadrati ed in particolare alla sua applicazione a reti di distribuzione dell’energia elettrica.
La stima dello stato di reti di distribuzione pone problemi particolari essenzialmente legati al gran numero di carichi, e quindi di nodi, in genere non monitorati in tempo reale, e alla lunghezza limitata delle linee, specie in reti urbane. Il gran numero di nodi e l’assenza di misure in numero sufficiente a rendere la rete osservabile e
La tesi analizza tali aspetti mediante applicazione di un algoritmo di stima dello stato, disponibile nella libreria Matpower di Matlab, a due reti test di diverse caratteristiche. La prima è una rete test IEEE disponibile in letteratura, caratterizzata da lunghi feeder rurali a tensione nominale 4,16 kV, ed una porzione della rete di distribuzione AMAIE di Sanremo, caratterizzata da feeder urbani in cavo, a tensione nominale 15 kV.
Le principali difficoltà riscontrate nell’esecuzione di calcoli di lod flow e stima dello stato della rete reale sono risultati legati alla complessità della topologia della rete, al gran numero di nodi utenti finali e alla presenza di misure distribuite solo in un determinato porzione della rete, dalla presenza di un gran numero di misure in bassa tensione ed alla necessità di identificare la rete a partire dalla sua descrizione contenuta in un file avente un formato particolare (hdc).
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Venieri, Laura. "Stime subellittiche per operatori somma di quadrati." Bachelor's thesis, Alma Mater Studiorum - Università di Bologna, 2011. http://amslaurea.unibo.it/2345/.
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Linguerri, Matteo. "Stima della portata cardiaca dalla pressione aortica." Bachelor's thesis, Alma Mater Studiorum - Università di Bologna, 2013. http://amslaurea.unibo.it/6386/.
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Questo elaborato illustra il problema della determinazione di una tecnica per rendere la misura della cardiac output il più possibile accurata, economica e poco invasiva.
A tale scopo è preso in esame un nuovo metodo basato sul modello WindKessel per la stima continua battito a battito di CO e TPR, partendo dall’analisi delle forme d’onda della pressione arteriosa periferica. Tale metodo ideato nel 2007 da T.A. Parlikar considera informazioni pressorie intrabattito e interbattito, in modo da ottenere stime soddisfacenti, che migliorano ulteriormente assumendo una complianza pressione-dipendente.
Applicando il metodo a un data set di animali suini, contenente misurazioni della CO di riferimento su base battito-battito, si riscontra un errore di stima complessivo pari a un RMNSE medio variabile tra l’11% ed il 13%, inferiore alla soglia del 15% ritenuta accettabile in letteratura per scopi clinici. Confrontando questi risultati con quelli ottenuti attraverso l’applicazione di altri metodi riportati in letteratura allo stesso set di dati, è stato dimostrato che il metodo risulta tra i migliori. Le CO e TPR stimate, dopo le infusioni farmacologiche endovenose effettuate sugli animali, corrispondono abbastanza fedelmente alle risposte emodinamiche attese.
Successivamente viene considerato l’obbiettivo di verificare l’applicabilità della procedura matematica sulla quale si fonda il metodo. Si implementa il procedimento e si applica il metodo su un data set simulato su base battito-battito, comprendente dati relativi a varie condizioni di funzionamento fisiologiche. Le stime di CO e TPR ottenute in questa fase inseguono discretamente le variazioni delle variabili emodinamiche simulate, dimostrandosi migliori nel caso di calibrazione con complianza pressione-dipendente.