Thèses : « Modulation of cell size »

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GOTTI, LAURA. "Nutritional modulation of cell size at s phase initiation in the buddine yeast saccharomyces cerevisiae: a role for glucose sensing and the cyclin dependent kinase inhibitor." Doctoral thesis, Università degli Studi di Milano-Bicocca, 2011. http://hdl.handle.net/10281/19573.

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The budding yeast Saccharomyces cerevisiae is a model organism for studies on cell cycle. For the survival of this cells a tight coordination of cell growth and division occurs at Start, a regulatory area of the cell cycle positioned immediately before beginning of S phase, at the G1-S boundary. Start is the event, or set of events, that commits a cell to a round of division. This mechanism is based on achieving of a critical cell size (protein content per cell at the onset of DNA replication, Ps) to enter into S phase. Ps increases in proportion with ploidy and is modulated by nutrients. In fact, in bach cultures, the average cell size remains at low levels during growth on non-fermentable substrates, while the average size of cells increases in a linear way with the specific growth rate only during growth on fermentable substrates. A genome-wide genetic analysis has suggested that cell size control could be due to ribosome biogenesis rate, one of the most energetic demanding processes in a cell and it is modulated according to nutrient availability. Indeed, a large cluster of genes involved in ribosome biogenesis, have been identified in a screen for small size (whi) mutants. This includes SFP1 and SCH9 genes. The first encode a zinc finger protein, promoting the transcription of a large cluster of genes involved in ribosome biogenesis, where the latter is serine threonine protein kinase involved in stress response and nutrient-sensing signaling pathway. Recent work from our laboratory allowed to identify that Far1, a cyclin kinase dependent inhibitor, and Cln3, a G1 phase cyclin, may form a nutritional modulated threshold controlling the entrance into S phase. Two parallel pathways downstream from the TORC1 complex regulate expression of genes encoding ribosomal proteins (RP) and the so-called RiBi regulon, composed by genes involved in ribosome biogenesis. The two pathways involve Sfp1, and Sch9. Therefore it was of interest to see whether the increase in size (RNA and protein) brought about by FAR1 overexpression was mediated by Sfp1 and Sch9. The effect of FAR1 overexpression on cell size parameters in the wild type BY4741 strain (isogenic to the sch9Δ and sfp1Δ mutants), grown in synthetic complete media supplemented with either ethanol or glucose as a carbon source, was similar to that reported in the W303 background. sfp1Δ and sch9Δ mutants were much smaller than wild type both in glucose - confirming previous data (Jorgensen et al., 2002; Jorgensen et al., 2004) - and ethanol-supplemented media. As observed in wild type cells, in both mutant strains FAR1 overexpression had only minor effects on cell cycle and cell size related parameters on glucose-grown cells. FAR1 overexpression did not affect duplication time in ethanol-grown sch9Δ cells, while sfp1Δ mutants overexpressing the FAR1 gene product were quite unhealthy with a large increase in duplication time. Overall increase in cell size was dramatic in ethanol-grown cells: however, while in wild type cells and sch9Δ mutants the increase in cell size derived from a balanced increase in RNA and protein content, in the sfp1Δ mutant the increase in protein content was not accompanied by an increase in RNA content, as shown by both FACS and chemical analysis, indicating that the Sfp1 is required to maintain proper coupling of RNA and protein syntheses when the Far1 protein is overexpressed in ethanol-grown-cells. The observation that FAR1 overexpression has different effects in sfp1Δ cells grown in ethanol and glucose media was not entirely unexpected. First, in untransformed cells, the Far1 level of glucose-grown cells is larger than in ethanol-grown cells, while ectopically expressed Far1 accumulates to a similar level regardless of the carbon source: as a result, Far1 overexpression is more dramatic in ethanol-grown cells than in glucose-grown cells (Alberghina et al., 2004). Accordingly, the effect of Far1 overexpression on cell size are minor on cells grown in glucose-supplemented media and much more dramatic in ethanol-grown cells. In the second part of this study we try to determine whether (and possibly, to which extent) the regulatory function of glucose can be separated from its nutrient function. To this aim, we characterized yeast strains in which glucose sensing is strongly reduced. An essential requisite for the survival of free living microorganism like the budding yeast Saccharomyces cerevisiae is the capacity to regulate growth and cell cycle progression according to the frequent changes in the nutrient status, so that proliferation is rapid when large supplies of nutrients are available and ceases when these becomes exhausted. Nutrients like glucose must therefore generate signals that are somehow received and elaborated by the complex machinery governing growth and cell cycle progression. Besides being the favorite carbon and energy source for S. cerevisiae, glucose can act as a signaling molecule (“hormone”) to regulate multiple aspects of yeast physiology: addition of glucose to quiescent or ethanol growing cells triggers a fast and massive reconfiguration of the transcriptional program, which enables the switch to fermentative metabolism and promotes an outstanding increase of the cell biosynthetic capacity. Yeast cells evolved several mechanisms for monitoring glucose level in their habitat: the cAMP-PKA pathways (with its two branches comprising Ras and the Gpr1-Gpa2 module), the Rgt2/Snf3-Rgt1 pathway and the main repression pathway involving the kinase Snf1. In order to investigate whether the glucose effect on cell size was due to its function as nutrient, that require metabolism of the sugar, or to sensing of extracellular glucose levels, yeast strains in which one or more of the glucose sensing pathway was impaired, due to gene deletion of glucose receptors (GPR1, SNF3, RGT2), were analyzed. These mutants show only a partial nutritional modulation of cell size and/or of duplication time. The gpa2Δ gpr1Δ strain does not show substantial changes in duplication time compared to its isogenic wild type grown in the same conditions, while its protein content is consistently lower. In the snf3Δ rgt2Δ and in the snf3Δ rgt2Δ gpa2Δ gpr1Δ strains a lower protein content is accompanied by an increase in duplication time, when compared to wild type strain. Furthermore, in all glucose sensing mutants the variation of protein content, as function of glucose levels, is less than the wild type. In the presence of ethanol, the kinetic parameters of mutants strain analyzed are comparable to wild type: there is only a strong increase in the duplication time, while there isn’t a further decrease in protein content compared to 0.05% glucose concentration. Data obtained show that the Gpa2-Gpr1 pathway specifically modulates Ps setting, while the Snf3-Rgt2 pathway plays an important role in Ps and growth rate setting of the cells. In conclusion, this work highlighted that the elements involved in the cell size determination are multiple and interconnected. A strong alteration in cell size and protein content could originate not only from alteration in the dosage of genes involved in the molecular mechanism of the threshold which controls Ps, but also from environmental conditions. Of particular relevance seems to be that glucose effect is largely acting as a signaling molecule, rather than as an energy source. Further studies are necessary in order to clarify the molecular mechanism that link the glucose sensing to the molecular machinery responsible of G1-S transition.

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Hubatsch, Lars. "Interplay between cell size and cell polarity." Thesis, University College London (University of London), 2018. http://discovery.ucl.ac.uk/10055636/.

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Cell polarity is a fundamental phenomenon underlying processes such as asymmetric cell division, tissue homeostasis and directed cell migration. In metazoans, the conserved PAR (-titioning defective) protein network polarizes cells with different shapes and sizes. Here, I investigate whether cell size influences the polarity pattern set up by the PARs. PAR polarity is typically achieved by localizing different sets of antagonizing proteins to opposing membrane domains. Antagonism ensures that mixing of the two species is restricted to a region between the two domains. Theoretically, this can be described using reaction-diffusion models, in which abstract biochemical agents are able to exchange between membrane and cytosol, and are subjected to cytosolic and membrane diffusion. Under suitable conditions, their interactions give rise to a stable pattern. In such a system, the interplay between diffusion and reaction rates determines the pattern by setting key length scales, for example, the extent of the boundary region between the two opposing domains. Using computer simulations, I first show that current reaction-diffusion models fail to adapt such pattern length scales to cell size. Second, this results in pattern breakdown below a certain minimum size, producing completely uniform protein distributions across the membrane. We term this size critical polarizable system size (CPSS). To test the first prediction - failure to adapt to cell size - I measured kinetic parameters and the resulting pattern length scales in differently sized cells, using the early C. elegans embryo as a model. The results suggest a moderate, if any, adaptation, prompting me to examine the second prediction - failure to maintain polarity at small sizes. By combining novel computational methods for 3D membrane reconstruction with cell size mutants, I then show that small cells are indeed unable to maintain polarity, thus establishing a lower size limit for polarity in vivo.

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Maguire, Sharon Marie. "Germ cell modulation of Sertoli cell function." Thesis, University of Edinburgh, 1994. http://hdl.handle.net/1842/20662.

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The aim of this work was to assess the influence of germ cells on the expression of selected Sertoli cell mRNAs. To this end, adult rats were treated with 650mg/kg methoxyacetic acid (MAA) to induce the specific depletion of >80% of pachytene and later spermatocytes from most tubules, and expression of selected Sertoli cell mRNAs was then assessed at various times after treatment when particular germ cell types were depleted selectively (see Bartlett et al. 1988; Allenby et al., 1991). Studies on the expression of cyclic protein 2(CP-2) mRNA supported the hypothesis that germ cells can influence the cyclic function of Sertoli cells. Expression of CP-2 mRNA was shown by Northern blot analysis to decrease significantly 21 days after MAA treatment. In situ hybridisation showed that CP-2 mRNA expression was decreased or absent from tubules at stages at which CP-2 mRNA is normally expressed (stages IV-VII) when elongate spermatids were depleted selectively from these tubules. This decrease was reflected in loss of CP-2 protein production. These observations lead us to hypothesise that elongate spermatids positively modulate CP-2 expression in the Sertoli cell, with this modulation occurring at the level of transcription. In conclusion, this study demonstrated that germ cells do appear to influence Sertoli cell gene expression. This can occur at the level of transcription as was demonstrated by the effect of elongate spermatids on CP-2 mRNA expression, or may occur post-transcriptionally, as would appear to be the case with ABP.

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Cadart, Clotilde. "Cell size homeostasis in animal cells." Thesis, Université Paris-Saclay (ComUE), 2017. http://www.theses.fr/2017SACLS103/document.

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Le mécanisme d’homéostasie de taille chez les cellules animales est très peu compris actuellement. Cette question est pourtant d’un intérêt majeur car le maintien de l’homéostasie de taille dans une population de cellules prolifératives doit se faire par une coordination entre la croissance et la division. Chez la levure S. pombe, il a ainsi été montré que la taille est une information cruciale pour déclencher l’entrée en mitose (Fantes, 1977). Chez plusieurs bactéries et les cellules filles de la levure S. cerevisiae au contraire, de récentes études ont au contraire montré que l’homéostasie de taille était le résultat d’une addition constante de volume, indépendamment de la taille initiale des cellules (Campos et al., 2014; Soifer et al., 2016; Taheri-Araghi et al., 2015). Ce mécanisme est appelé « adder » et génère une régression des tailles à la moyenne, génération après génération. Ces résultats ont été possibles grâce au développement de techniques permettant la mesure dynamique du volume à l’échelle de la cellule unique et sur plusieurs générations. Une telle mesure est cependant très difficile chez les cellules de mammifère dont le volume fluctue constamment et qui cyclent sur des temps plus longs (environ 20 heures). Pour cette raison, la plupart des approches proposées sont indirectes (Kafri et al., 2013; Sung et al., 2013; Tzur et al., 2009) ou reposent sur une mesure de la masse plutôt que du volume (Mir et al. 2014; Son et al., 2012). Ensemble, ces études ont montré que les cellules de mammifère croissaient de manière exponentielle. Elles ont aussi remis en cause le modèle traditionnel qui proposait que l’homéostasie de taille reposait sur l’adaptation de la durée du cycle et mis en avant un rôle de la régulation de la vitesse de croissance. Cependant, aucun modèle n’a réellement été proposé ou démontré. La nature et l’existence même d’un mécanisme maintenant l’homéostasie de taille des cellules de mammifère est en fait discutée (Lloyd, 2013).Pour caractériser l’homéostasie de taille des cellules de mammifères, nous avons développé une technique permettant pour la première fois la mesure du volume de ces cellules sur des cycles complets (Cadart et al., 2017; Zlotek-Zlotkiewicz et al. 2015). Nous montrons que plusieurs types cellulaires (HT29, MDCK et HeLa) se comportent d’une manière similaire à celle d’un « adder ». Pour tester davantage cette observation, nous induisons artificiellement des divisions asymétriques en confinant les cellules dans des micro-canaux. Nous observons que les asymétries de tailles sont réduites mais pas complètement corrigées au cours du cycle suivant, à la manière d’un « adder ». Pour comprendre comment la croissance et la progression dans le cycle sont coordonnées et génère cet « adder », nous combinons notre méthode de mesure de volume avec un suivi de la progression dans les différentes phases du cycle. Nous montrons que la durée de la phase G1 est inversement corrélée au volume initial des cellules. Cependant, cette corrélation semble contrainte par une durée minimale de G1 mise en évidence lors de l’étude de cellules artificiellement poussées à atteindre de grandes tailles. Néanmoins, même dans cette condition où la modulation de la durée du cycle est perdue, l’observation du « adder » est maintenue. Ceci suggère un rôle complémentaire de la régulation de la vitesse de croissance des cellules. Nous proposons donc une méthode pour estimer théoriquement la contribution relative de l’adaptation de la vitesse de croissance et de la durée du cycle dans le contrôle de la taille. Nous utilisons cette méthode pour proposer un cadre général où comparer le processus homéostatique des bactéries et de nos cellules. En conclusion, notre travail apporte pour la première fois la démonstration que les cellules de mammifères maintiennent l’homéostasie grâce à un mécanisme similaire au « adder ». Ce mécanisme semble impliquer à la fois une modulation de la durée du cycle et du taux de croissance The way proliferating mammalian cells maintain a constant size through generations is still unknown. This question is however central because size homeostasis is thought to occur through the coordination of growth and cell cycle progression. In the yeast S. pombe for example, the trigger for cell division is the reach of a target size (Fantes, 1977). This mechanism is referred to as ‘sizer’. The homeostatic behavior of bacteria and daughter cells of the yeast S. cerevisiae on the contrary was recently characterized as an ‘adder’ where all cells grow by the same absolute amount of volume at each cell cycle. This leads to a passive regression towards the mean generation after generation (Campos et al., 2014; Soifer et al., 2016; Taheri-Araghi et al., 2015). These findings were made possible by the development of new technologies enabling direct and dynamic measurement of volume over full cell cycle trajectories. Such measurement is extremely challenging in mammalian cells whose shape constantly fluctuate over time and cycle over 20 hours long periods. Studies therefore privileged indirect approaches (Kafri et al., 2013; Sung et al., 2013; Tzur et al., 2009) or indirect measurement of cell mass rather than cell volume (Mir et al. 2014; Son et al., 2012). These studies showed that cells overall grew exponentially and challenged the classical view that cell cycle duration was adapted to size and instead proposed a role for growth rate regulation. To date however, no clear model was reached. In fact, the nature and even the existence of the size homeostasis behavior of mammalian cells is still debated (Lloyd, 2013).In order to characterize the homeostatic process of mammalian cells, we developed a technique that enable measuring, for the first time, single cell volume over full cell cycle trajectories (Cadart et al., 2017; Zlotek-Zlotkiewicz et al. 2015). We found that several cell types, HT29, HeLa and MDCK cells behaved in an adder-like manner. To further test the existence of homeostasis, we artificially induced asymmetrical divisions through confinement in micro-channels. We observed that asymmetries of sizes were reduced within the following cell cycle through an ‘adder’-like behavior. To then understand how growth and cell cycle progression were coordinated in way that generates the ‘adder’, we combined our volume measurement method with cell cycle tracking. We showed that G1 phase duration is negatively correlated with initial size. This adaptation is however limited by a minimum duration of G1, unraveled by the study of artificially-induced very large cells. Nevertheless, the adder behavior is maintained even in the absence of time modulation, thus suggesting a complementary growth regulatory mechanism. Finally, we propose a method to estimate theoretically the relative contribution of growth and timing modulation in the overall size control and use this framework to compare our results with that of bacteria. Overall, our work provides the first evidence that proliferating mammalian cells behave in an adder-like manner and suggests that both growth and cell cycle duration are involved in size control

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Ricolo, Delia. "Cytoskeletal modulation of single-cell branching." Doctoral thesis, Universitat de Barcelona, 2017. http://hdl.handle.net/10803/404782.

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The embryonic tracheal system of Drosophila melanogaster consists of a network of interconnected epithelial tubes of different size and architecture characterized by different cellular mechanisms of tube formation. The main branches of the Drosophila tracheal system have an extracellular lumen because their cells fold to form a tube. However, terminal cells (TCs), specialized cells designed to connect the tracheal system to target tissues, form unicellular branches by generating of a subcellular lumen. This topology of unicellular tubes is a good model to clarify the mechanisms that orchestrate single-cell branching, a process parallel to capillary sprouting in blood vessels. During tracheal embryonic development, TCs produce seamless tubes, generating a cytoplasmic extension, by cell elongation, and a concurrent intracellular luminal space surrounded by an apical membrane. Cell elongation and subcellular lumen formation are very much dependent on cytoskeleton reorganization. The main aim of this thesis was to understand new aspects of cytoskeletal modulation that orchestrate subcellular lumen formation. In particular, we have addressed this aim analysing mutants displaying an increase in subcellular lumen branching and mutants characterized by the absence of the subcellular lumen. We found that mutations in Regulator of Cyclin A (Rca1) and Cyclin A (CycA) affect subcellular branching, causing TCs to form more than one subcellular lumen. The effect of Rca1 is post-mitotic in the tracheal system, and depends on an amplification of centrosome number. Other mutant conditions, characterized by the increase of centrosome number, such as Slimb (slmb) and the overexpression of SAK also show excess of subcellular lumen branching. Furthermore, we showed that de novo lumen formation is impaired in mutant embryos with low centrosome numbers such as sas4 and is restored in the double mutant Rca1; sas4. The data presented here define a requirement for the centrosome as a microtubule organizing center (MTOC) for the initiation of subcellular lumen formation. We propose that in wt condition two centrosomes are needed to arrange the specific intracellular TC organization necessary to generate a subcellular lumen, and that an excess of centrosome numbers allows for an increase in single- cell branching. We also analysed the involvement of the spectraplakin Short-stop (Shot) in the cytoskeletal organization of the TCs. Shot is a multifunctional protein involved in many aspects of cytoskeletal organization in different tissues, which can operate as a single cytoskeleton component as well as coordinating cytoskeletal elements between them. This functional versatility of Shot is probably reflected by the abundant generation of isoforms and by the modulation of its numerous domains. We found that the overexpression of shot in the tracheal system induces extra-branching of the subcellular lumen and this effect depends mainly on the C-tail domain at the C- terminus and its involvement in the stabilization/polimerization of MTs. On the other hand, by examining loss of function alleles, analysing its structural function and visualizing Shot accumulation, we suggest that Shot is not just involved in MT organization in the TC but it also acts as a crosslinker between MTs and the actin network. The first calponin domain (CH1) of the acting binding domain (ABD) at the N-terminal is involved in this cross-linking activity. Finally, we provide some data indicating a functional overlap between the spectraplakin and the microtubule associated protein (MAP) Tau during subcellular lumen formation. Las células terminales (TCs) de la tráquea del embrión de Drosophila melanogaster son capaces de generar un lumen subcelular y son utilizadas como modelo para la formación de tubos unicelulares de tipo “seamless”. La generación de dicho lumen depende estrictamente de una especifica organización del citoesqueleto que permite la formación de una nueva membrana apical en el interior de la TC. El objetivo del trabajo aquí presentado ha sido lo de aclarar nuevos aspectos de la modulación del citoesqueleto en el contexto de la formación del lumen sub-celular. Los mutantes de Regulator of Cyclin A (Rca1) y CycA (Cyclin A) están caracterizados por TC con mas de un lumen subcelular. El efecto de Rca1 es post-mitótico y esta causado por un aumento del numero de centrosomas. Reportamos, atraves el estudio de Rca1 y otros mutantes afectados en el numero de centrosomas, una estricta asociación entre centrosomas y formación del lumen sub-celular. Nuestros datos revelan, por primer vez, la función de los centrosomas como centros de organización de microtubulos (MTOC) en la TC y que un exceso de centrosomas puede causar un aumento en la capacidad de ramificación del lumen. En este trabajo también hemos analizado la función de la spectraplakina Short-stop (Shot). A través de experimentos de sobre-expresión y falta de función de shot, integrados con estudios estructura-función y de localización de sus productos proteicos hemos concluido que la spectraplakina actúa el la TC acudiendo a diferentes grados de organización citosqueletrica; en nuestro modelo Shot es capaz de promover la estabilización/polymerizacion de microtubulos, y un exceso de esta función puede causar extra ramificación en la TC. Por otro lado, Shot esta implicado en la correcta conexión entre la red de microtubulos y la actina y su falta influye negativamente la formación del lumen sub-celular. También reportamos datos preliminares que indican una superposición funcional entre Shot y la proteína asociada a microtulos (MAP) Tau durante el desarrollo del la TC.

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Forsythe, Paul. "The modulation of mast cell activity." Thesis, Queen's University Belfast, 1997. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.388087.

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Watt, A. P. "The modulation of mast cell activity." Thesis, Queen's University Belfast, 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.368527.

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Luizet, Jean-Baptiste. "Host Cell modulation by Brucella effectors." Thesis, Lyon, 2019. http://www.theses.fr/2019LYSE1157.

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Brucella abortus est une bactérie pathogène responsable d’une zoonose ré-émergente causant plus de 500 000 décès par an. Brucella est une bactérie à Gram négative facultative intracellulaire capable d’infecter un grand spectre de cellules différents en entrant par phagocytose ou macropinocytose. Une fois à l’intérieur de la cellule, Brucella est retrouvée dans une vacuole doublement membranée appelée Brucella-containing vacuole (BCV). Brucella va alors interagir partiellement avec les différents compartiments endosomaux et lysosomaux afin d’établir sa niche réplicative au sein du réticulum endoplasmique est un aspect clé de sa virulence. Pour cela il a été démontré que Brucella abortus possède le système de sécrétion de type IV (SST4) démontré pour être un facteur de virulence majeur très répandu chez de nombreuses bactéries pathogènes. Mon projet de thèse consiste à caractériser un effecteur identifié pour être secrété par ce SST4. Nous avons appelé cet effecteur Brucella secreted protein L (BspL). Nous avons pu par différentes approches biologiques déterminer son mode d’action chez les cellules hôtes infectées ainsi que son importance pour le cycle intracellulaire de la bactérie. Nous avons en effet vu que cet effecteur localisait au sein du réticulum endoplasmique, organelle majeure de la cellule eucaryote, et induisait du stress. Ce stress est ressenti par la cellule grâce à différents senseurs cellulaires qui vont converger vers l’activation d’une réponse cellulaire appelée «UnfoldedProtein Response». Cette voie cellulaire va par différents mécanismes moléculaires tenter de contrer ce désordre induit au sein du réticulum afin de restaurer une homéostasie cellulaire. Cependant en recherchant de potentielles de notre effecteur BspL nous avons identifié Homocysteine-responsive endoplasmic reticulum-resident ubiquitin-like domain member 1 (Herp) une protéine impliquée dans de nombreux processus de régulations physiologiques du réticulum en cas de stress comme la dégradation associée au réticulum (ERAD). Nous avons donc par la suite démontré que l’action de BspL n’était pas uniquement d’induire du stress mais d’augmenter la capacité d’action de l’ERAD au sein de la cellule. Par ailleurs nous avons vu que BspL était important pour Brucella dans le contrôle de la cinétique de formations de ses vésicules de sortie de la cellule afin de pouvoir faire son cycle sans sortir prématurément. Ainsi l’identification de cet effecteur a permis de mettre en évidence l’importance de l’ERAD pour la première fois dans la pathogénie de Brucella. De plus cet effecteur est le premier à être identifié pour participer dans la régulation des vacuoles de sortie de Brucella. En parallèle nous avons vu que BspL impactait d’autres organites qui sont les mitochondries en fragmentant la connectivité de ce réseau mitochondrial. Aujourd’hui nous savons que les mitochondries et le réticulum sont étroitement connectés notamment avec l’implication du réticulum dans le processus de fission mitochondriale. Par ailleurs il est également connu que l’induction de stress par des agents physiques ou chimiques également fragmente le réseau mitochondrial. Nous avons néanmoins vu que le processus de fragmentation mitochondriale des mitochondries étaient indépendants du stress induit par BspL. Ces résultats ont été également confirmés en infection avec la comparaison d’une souche sauvage et mutante pour le gène codant pour BspL. En conclusion nous avons identifié un effecteur de Brucella associé au détournement de diverses fonctions au sein de la cellule. Il a permis de mettre en évidence l’importance du contrôle de l’ERAD pour contrôler la cinétique de réplication de la bactérie. Par ailleurs le fait que BspL soit également associé aux mitochondries semblent indiquer que l’effecteur pourrait avoir un rôle différentiel au cours du temps pendant l’infection This work provides new insights not only into Brucella pathogenesis, but also places effector targeting of the endoplasmic reticulum quality control machinery at the center of bacterial intracellular trafficking, a completely novel research topic that could be of relevance for other bacterial pathogens. The endoplasmic reticulum-associated degradation (ERAD) pathway plays a vital role in this quality control process, co-regulated with protein folding, stress responses and degradation pathways. Dysfunction of these processes can lead to severe diseases due to either the accumulation of misfolded proteins, namely neurodegenerative diseases; or due to the destruction of the entire pool of a given protein, such as the case of cystic fibrosis transmembrane conductance regulator (CFTR). Certain viruses have been shown in the past to directly hijack ERAD components to control immune responses and viral assembly. However, little is known on the control of ERAD during bacterial infections. In addition, insights into ERAD modulation could have an important impact in clinical research, for example, by providing new clues on how to boost the cell’s capacity to handle misfolded proteins. Brucella is an excellent model intracellular pathogen, with a well characterized cellular cycle that relies on a type IV secretion system to establish a replicative niche derived from the endoplasmic reticulum. It is then able to modify this vacuole by a currently unknown mechanism, induce autophagy and form vacuoles that will enable bacterial escape from the cell (Starr et al Cell Host and Microbe 2011). The effector proteins that mediate the formation of these Brucella replicative and autophagy vacuoles remain unknown. Here we identify a new type IV secretion effector that interacts with Herp to facilitate ERAD and delay the formation of autophagic Brucella-containing vacuoles to prevent premature bacterial dissemination. Intriguingly, we also show that ERAD is fine-tuned during infection as at early stages blocking of ERAD is beneficial whereas at late stages of the infection it results in premature egress. The work is obviously of interest for the Brucella community with the characterization of a novel type IV secretion system effector and, in addition, the identification of the first effector implicated in control of the formation of autophagic Brucella-containing vacuoles. However, our study will also appeal to a broader audience as it identifies an eukaryotic target not previously implicated in bacterial pathogenesis, Herp a key modulator of ERAD highlighting a novel mechanism of bacterial regulation of the endoplasmic reticulum quality control machinery for intracellular trafficking

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Brown, Marena Dessette. "Sickle cell-endothelial interactions : modulation of cell adhesion molecule expression." Diss., Georgia Institute of Technology, 1995. http://hdl.handle.net/1853/11306.

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Wu, Chia-Yung. "Control of gene expression by cell size." Thesis, Massachusetts Institute of Technology, 2010. http://hdl.handle.net/1721.1/57564.

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Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Biology, 2010. This electronic version was submitted by the student author. The certified thesis is available in the Institute Archives and Special Collections. Cataloged from student submitted PDF version of thesis. Includes bibliographical references. Polyploidy, increased copy number of whole chromosome sets in the genome, is a common cellular state in evolution, development and disease. Polyploidy enlarges cell size and alters gene expression, producing novel phenotypes and functions. Although many polyploid cell types have been discovered, it is not clear how polyploidy changes physiology. Specifically, whether the enlarged cell size of polyploids causes differential gene regulation has not been investigated. In this thesis, I present the evidence for a size-sensing mechanism that alters gene expression in yeast. My results indicate a causal relationship between cell size and gene expression. Ploidy-associated changes in the transcriptome therefore reflect transcriptional adjustment to a larger cell size. The causal and regulatory connection between cell size and transcription suggests that the physical features of a cell (such as size and shape) are a systematic factor in gene regulation. In addition, cell size homeostasis may have a critical function - maintenance of transcriptional homeostasis. by Chia-Yung Wu. Ph.D.

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Crotti, Pablo. "Phenotypic variability, cell size and population fitness." Thesis, Imperial College London, 2015. http://hdl.handle.net/10044/1/43964.

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In all biological systems, phenotypes are quantitative measures of an organism's traits. To understand the relation between a genotype and phenotypes, the influence of genetic perturbations, such as gene deletions, on phenotypes is studied. Although it is common practice to look at the mean value of phenotypes to detect changes, it is less common to model such modifications by considering phenotypic variability. However, phenotypic variability could also contain important information about the phenotype-genotype map. Also, phenotypic variability is a determinant of an organism's robustness and fitness. This thesis is a collection of five studies devised to identify and analyse phenotypic variability in S. pombe, S. cerevisiae and E. coli. The first chapter determines the probabilistic nature of the dynamic transition from quiescence to proliferation in S. pombe. Based on the viability and the random nature of the quiescence of knockout genes, we demonstrate that two competing stochastic models explain the data equally well. The second and third chapters provide novel approaches to identifying mutant genes that increase/decrease phenotypic variability throughout the cell cycle of S. cerevisiae. Using machine learning algorithms, we observe that mutants divide in four categories acting positively and/or negatively on phenotypic variability. Additionally, we show that phenotypic robustness is inversely related to cell fitness. The fourth chapter builds morphological interaction networks in S. cerevisiae. By employing a Bayesian network framework, we show that cell size dictates nuclear size throughout most of the cell cycle. The fifth chapter investigates the growth rate of bacterial populations subject to fluctuations in the variance of their cell size distributions. By employing a population balance equation combined with an agent-based model, our results indicate that the cell size distribution affects the growth rate. Moreover, we show that cells possess a cell size regulator within their cycle.

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Burgoyne, Celia Helen. "Glucose modulation of endothelial cell in vitro." Thesis, University of Hull, 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.402798.

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Martin, Ashley Diane. "Modulation of endothelial cell characteristics by pericytes." Thesis, University of Ulster, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.287133.

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Iqbal, Z. "Electrochemical modulation of sickle cell haemoglobin polymerisation." Thesis, University College London (University of London), 2008. http://discovery.ucl.ac.uk/1444279/.

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Sickle cell haemoglobin differs from normal haemoglobin by a single amino acid in its chain. This amino acid replacement, from glutamic acid to valine, causes polymerisation of proteins into defined long insoluble fibres with a typical diameter of 21.5 nm. The polymerisation is triggered by the formation of deox haemoglobin from oxyhaemoglobin in low oxygen partial pressures, which results in a conformational change in the secondary structure of the protein. Pathogenesis in sickle cell disease depends on the polymerisation and gelation of deoxygenated HbS molecules. In this work, an electrochemical method has been described to modulate the oxygen concentration in an optically transparent thin layer cell to produce deoxyhaemoglobin whilst monitoring the extent of polymerisation using turbidity measurements. The oxygen was depleted in the vicinity of the electrode and triggered the polymerisation. The dependence of protein concentration, temperature, pH and ionic strength on the nucleation and elongation of HbS polymerisation was characterised at the electrode surface and the kinetics of polymerisation was investigated using a model for fibrillogenesis describing a two-step process of nucleation followed by elongation. The rate constants, determined for a number of conditions, showed that nucleation is far slower than the growth whilst polymerisation at the surface was demonstrated to occur in three stages with an initial time delay when no structures were observed followed b growth of fibrous hair-like strands and finally gel-like aggregation. An understanding of the factors which affect polymerisation at a surface and an insight into the dynamics and mechanism of polymer aggregation and the pathophysiology of sickle cell disease has been provided. A screening method for substances that effect the fibre nucleation and/or growth that could be valuable to the pharmaceutical industry for treating sickle cell disease is also presented.

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Mangani, Christian. "Peptides and polymers for stem cell modulation." Thesis, University of Edinburgh, 2015. http://hdl.handle.net/1842/17909.

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One of the requisites for a growth factor and a biomaterial in tissue engineering, cell therapy and regenerative medicine is the ability to control cell fate. Cells exist in a complex micro-environment consisting of extra-cellular matrix, growth factors, together with adjacent cells. Stem cell culture and modulation remains a challenge due to insufficient, undefined and costly culture systems. This thesis describes synthetic approaches that can modulate stem cell fate by the identification of new synthetic substrates for the growth of cancer, embryonic stem cells and potential short peptide sequences that can mimic the biological functions of the native cytokine used to culture stem cells. Glioma cancers exist as a heterogeneous population of cancer stem cells and cancer progenies. Scale up and spin coating of a polyurethane and polyacrylate polymers was done on agarose for the enrichment of the cancer stem cell population from glioma cells. A polyurethane, synthesised from poly(tetramethylene glycol) and 1,3-bis(isocyanatomethyl)cyclohexane spin (PU10) coated on an agarose surface, was identified to have a higher affinity for the cancer stem cell population over its progenies. By using this polymer to study the mechanism of the cancer stem cell adhesion, two niche components i.e. galectin, transferrin that are enriched by the polymer that contributed to the growth of the cancer stem cells were identified. A synthetic hydrogel (HG21) was identified as substrate for the culture of mouse embryonic stem cells (mESC) as a replacement for gelatin. mESCs were cultured on the hydrogel under undefined and defined conditions. Under both culture conditions, mESC pluripotency and naïve phenotype markers were verified. Marker profiles by immunostaining (Oct-4, Nanog), flow cytometry (SSEA-1) and qPCR (14 gene markers) of mESC grown on the hydrogel were comparable to gelatin, while enabling thermo-detachment for enzyme free passaging of mESC. To identify alternative substances to the cytokines used in stem cell culture, a microarray system was developed. The microarray system was developed initially with adhesion cellulose peptides printed onto polyacrylamide coated microscope slides. These slides were then screened for interaction with human embryonic stem cells (hESCs). After successful development of the cell based cellulose peptide microarray system, overlapping 25-mer peptides based on of basic fibroblast growth factor were synthesised, printed onto the same type of slide and screened with hESC. The screen identified “hit” peptides, which could potentially mimic the biological effects of the native cytokine on hESCs. These “hit” peptides were scaled up and tested in solution with hESC. In the linear form the peptides were not sufficient to sustain pluripotency and further optimisation is needed.

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Muñoz, Frances M. "Calcium Modulation of PARP-mediated Cell Death." Diss., The University of Arizona, 2015. http://hdl.handle.net/10150/347337.

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Many pathological conditions, including renal disease, are associated with oxidative stress. 2,3,5-tris(Glutathion-S-yl)hydroquinone (TGHQ), a potent nephrotoxic and nephrocarcinogenic metabolite of benzene and hydroquinone, generates reactive oxygen species (ROS), can cause DNA strand breaks, and the subsequent activation of DNA repair proteins, including poly(ADP-ribose) polymerase (PARP)-1. Under robust oxidative damage, PARP-1 is hyper-activated, which causes elevations in intracellular calcium concentrations (iCa²⁺), NAD⁺ and ATP depletion, and ultimately necrotic cell death. The role of Ca²⁺ in PARP-dependent necrotic cell death remains unclear. We therefore sought to determine the relationship between Ca²⁺ and PARP-1 during TGHQ-induced necrotic cell death in human renal proximal tubule epithelial cells (HK-2). Extracellular Ca²⁺ is responsible for coupling PARP-1 activation to increases in iCa²⁺ during TGHQ-induced cell death. Moreover, organelles such as the endoplasmic reticulum and the mitochondria, which contain intracellular Ca²⁺ stores play no role in increases of iCa²⁺. PARP-1 inhibition attenuates increases in iCa²⁺ induced by TGHQ, and treatment with 2-aminoethoxydiphenyl borate (2-APB), a store-operated Ca²⁺ channel (SOC) inhibitor, restored cell viability, levels of NAD⁺, and attenuated PAR protein-ribosylation (PARylation). Concurrent with SOC activation having a direct effect on PARP-1 activity, and PARP-1 inhibition attenuating increases in iCa²⁺, the results suggest that PARP-1 and SOCs are coupled during TGHQ-induced cell death. We also explored the relationship between SOC activation and PARP-1 downstream of PARP-1 activity. Poly(ADP-ribose)glycohydrolase (PARG), which catalyzes the degradation of PARs to yield free ADP-ribose (ADPR), is known to activate SOCs. Interestingly, siRNA knockdown of PARG modestly increased PAR ribosylation, but did not restore cell viability in the presence of TGHQ, indicating that free ADPR is not responsible for SOC activation in HK-2 cells. Overall, our results suggest that PARP-1 and Ca²⁺ are coupled through SOC entry, and that this relationship may involve alternative PAR-mediated signaling that leads to necrotic cell death. To further elucidate the role of PAR polymers in response to TGHQ, we determined the cellular co-localization of PAR by immunofluorescent staining. PAR polymers originally co-localized in the nucleus, and in the cytosol at later time points. Immunoprecipitation with a pADPr antibody and further analysis via mass spectrometry revealed PARylation of many stress-related proteins and Ca²⁺-related proteins upon TGHQ treatment. We therefore speculate that cytosolic PAR may cause downstream signaling, PARylating proteins that activate store-operated Ca²⁺ entry either directly through Ca²⁺-related proteins or PARylation of stress-related proteins. Thus, PARylation of proteins may contribute to increases in iCa²⁺ concentrations, leading to PARP-1-dependent necrotic cell death. Our studies provide new insight into PARP-mediated necrotic cell death. Ca²⁺ is coupled to PARP-1 hyperactivation through SOCs, where iCa²⁺ increases are independent of PARG activity, demonstrating a novel signaling pathway for PARP-dependent necrotic cell death.

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Bruckner, Markus [Verfasser]. "Modulation of Dendritic Cell Behaviour / Markus Bruckner." Konstanz : Bibliothek der Universität Konstanz, 2011. http://d-nb.info/1041832877/34.

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McSherry, Iain Neil. "Endothelial cell modulation of smooth muscle contraction." Thesis, University of Bath, 2005. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.423481.

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Sharma, Manu. "Host cell death modulation by Chlamydia trachomatis." Doctoral thesis, Humboldt-Universität zu Berlin, Mathematisch-Naturwissenschaftliche Fakultät I, 2010. http://dx.doi.org/10.18452/16253.

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Chlamydien durch die Modulation spezifischer Wirtszellproteine verschiedene Wege der Apoptose verhindern können. Mcl-1 und cIAP-2 erwiesen sich als bedeutende Faktoren, die durch die Infektion hochreguliert und absolut notwendig für die Inhibierung der Apoptose durch Chlamydien waren. Hochregulation der Mcl-1 Expression führte zu einem Block im apoptotischen Weg oberhalb der Mitochondrien. cIAP-2 zusammen mit anderen Inhibitor of Apoptosis Proteins (IAP) verhinderten die Aktivierung von Caspase-3, denfinalen Schritt in der apoptotischen Kaskade. Weiterhin wurde beobachtet, dass die Aktivierung des MAPKinase-Signalweges durch die Infektion wichtig war für die Hochregulierung von Mcl-1 und cIAP-2. Ein Hochdurchsatz-Screen wurde durchgeführt, um andere Wirtszellfaktoren, die für die Apoptoseinhibierung durch die Chlamydien verantwortlich sind, zu identifizieren. Neben Mcl-1 waren die identifizierten Faktoren hauptsächlich Mitglieder des MAPKinase-Signalweges. Dabei wurde deren Rolle für die Apoptoseresistenz bestätigt. Eine weiterführende Analyse der im Screen ermittelten Faktoren identifizierte eine Funktion von HIF-1a bei der Modulation der Expression anti-apoptotischer Faktoren während der Infektion. Es wurde beobachtet, dass HIF-1a stabilisiert und zum Nukleus transloziert wird. Es ist bekannt, dass HIF-1a HIF-1a im Nukleus binden kann, um den funktionalen Transkriptionsfaktor HIF zu bilden. Dieser reguliert die Expression verschiedener Überlebensfaktoren, unter anderem Mcl-1. HIF-1a Knockdown inhibierte die Chlamydien-induzierte Hochregulation von Mcl-1 mRNA-Expression. chlamydial infection blocked the apoptotic pathway at multiple levels by modulation of specific host cell proteins. Mcl-1 and cIAP-2 were two most prominent factors that were up-regulated during the infection, and absolutely required for apoptosis inhibition. Increased expression of Mcl-1 led to a block in the apoptotic pathway upstream of the mitochondria. cIAP-2, together with other inhibitor of apoptosis proteins (IAPs), blocked the activation of caspase-3 at the final step of the apoptosis cascade. Further, it was observed that the activation of the MAPK pathways during infection was needed for the up-regulation of Mcl-1 and cIAP-2. A high throughput RNAi screen was performed to identify other host factors required for the apoptosis resistance during the infection. Besides Mcl-1, the targets from the screen prominently included members of the MAPK pathways, confirming their role in the apoptosis resistance. Pathway analysis of the targets identified the role of HIF-1a in modulating the expression of the anti-apoptotic factors during infection. It was observed that during infection, HIF-1a gets stabilized and translocates to the nucleus. It is known that HIF-1a can bind to HIF-1a in the nucleus to form the functional transcription factor HIF, which can regulate the expression of survival factors like Mcl-1. This was seen to be the case, because knock down of HIF-1a abrogated the infection induced up-regulation of Mcl-1 at the mRNA levels.

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Grangeray-Vilmint, Anais. "Modulation of cerebellar Purkinje cell discharge by subthreshold granule cell inputs." Thesis, Strasbourg, 2016. http://www.theses.fr/2016STRAJ023/document.

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La décharge des cellules de Purkinje (CP), neurone de sortie du cortex cérébelleux, joue un rôle majeur dans le contrôle moteur. Les CP reçoivent des entrées excitatrices provenant des cellules des grains (CG), lesquelles génèrent également une inhibition antérograde sur les CP via l’activation d’interneurones de la couche moléculaire (IN). Lors de ma thèse, j’ai étudié l’influence simultanée de la balance excitation-inhibition (E/I) et des plasticités à court terme aux synapses CG-IN-CP sur la décharge des CP, par des techniques d’électrophysiologie, d’optogénétique et de simulation. Ces travaux démontrent l’existence d’une hétérogénéité d’E/I dans le cortex cérébelleux ainsi qu’une grande diversité de modulation des CP en réponse à la stimulation de CG. Le nombre de stimulation des CG influence fortement la direction et l’intensité de la modulation observée. Enfin, la combinaison de plasticités à court terme et d’E/I génère dans la décharge des CP des motifs de réponses complexes mais reproductibles, ayant sans doute un rôle essentiel dans l’encodage sensoriel Rate and temporal coding in Purkinje cells (PC), the sole output of the cerebellar cortex, play a major role in motor control. PC receives excitatory inputs from granule cells (GC) which also provide feedforward inhibition on PC through the activation of molecular layer interneurons (MLI). In this thesis, I studied the influence of the combined action of excitation/inhibition (E/I) balance and short-term plasticity of GC-MLI-PC synapses on PC discharge, by using electrophysiological recordings, optogenetic stimulation and modelling. This work demonstrates that E/I balances are not equalized in the cerebellar cortex and showed a wide distribution of PC discharge modulation in response to GC inputs, from an increase to a shut down of the discharge. The number of stims in GC bursts strongly controls the strength and sign of PC modulation. Lastly, the interplay between short-term plasticity and E/I balance implements complex but reproducible output patterns of PC responses to GC inputs that should play a key role in stimulus encoding by the cerebellar cortex

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Wang, Wei. "Modulation of immune cell responses by small cell lung cancer cells." Thesis, King's College London (University of London), 2016. https://kclpure.kcl.ac.uk/portal/en/theses/modulation-of-immune-cell-responses-by-small-cell-lung-cancer-cells(7bdc85c2-acd8-4f13-9d2b-e2ce07d1567b).html.

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Small Cell Lung Cancer (SCLC) accounts for 15-20% of all lung cancers and kills at least one person every 2 hours in the UK. There is no effective treatment and overall 2-year survival is less than 5%. Patients with SCLC have poorly understood local and systemic immune defects. Previous studies have shown several important defects in cell-mediated immune responses in patients with SCLC. A better understanding of interactions between SCLC tumour cells and immune cells may lead to the development of novel therapeutic approaches. There is increasing recognition that immunological biomarkers may add to traditional histological analyses and can be exploited in the management of multiple epithelial malignancies. There are currently no such markers used in the management of SCLC. In my PhD project, I have shown that cell lines from different SCLC patients have differential immunosuppressive capabilities. These properties are mediated by the secretion of differing levels of soluble molecules that can suppress the mixed leukocyte reaction (MLR) and CD4+ T cell proliferation, induce IL-10 secretion and differentiation of functional CD4+CD25+CD127+FoxP3+Helios- regulatory T cells (Tregs) from naïve CD4+ T cells. IL-15 is secreted by SCLC cells in culture in proportion to their immunosuppressive capability. Its in vivo relevance is supported by its presence in tumour biopsy samples. The suppressive effect on CD4+ T cell proliferation and the induction of Treg cell population was not affected by blocking IL-10 or TGF-β signalling but was partially reversed by blocking IL-15 activity. Therefore, IL-15 is one, though not the only, soluble molecule produced by SCLC cells to mediate immune suppression by inducing increased population of Treg cells. This may represent a mechanism by which SCLC cells can suppress the immune response. In addition, SCLC cells supressed TNF-α release from monocytes in response to LPS stimulation, down-regulated expression of CD16 and CD86 and upregulated expression of CD163 and CD206 on monocyte-derived macrophages (MDMs) upon activation. This M2-like phenotype poralization was associated with decreased TNF-α and IL-6 production and increased IL-10 secretion. These effects were abrogated by blocking the signalling of bombesin-like peptides (BLPs) that are neuropeptides produced by SCLC cells using a GRP receptor (GRP-R) antagonist. Therefore, the polarization of macrophages to an M2-like phenotype by SCLC cell-derived BLPs may represent another mechanism by which SCLC tumours suppress the immune response. Finally, SCLC tumour biopsies were shown to be infiltrated with various mononuclear immune cells and Treg cells. CD45 and FoxP3 were used as paninflammatory cell and Treg cell markers respectively. An elevated CD45+ infiltrate was predictive of prolonged survival in SCLC independent of age, sex, stage or treatment strategy. An elevated FoxP3+/CD45+ ratio was predictive of a significantly worse prognosis. This study identifies potential mechanisms by which SCLC tumour cells may downregulate local and systemic immune response, and also identifies an independent prognostic marker to predict patient survival in SCLC. Further, IL- 15 and BLPs are potential novel therapeutic targets in SCLC.

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Matsiaka, Oleksii. "New mathematical models for cell biology assays incorporating realistic cell size dynamics." Thesis, Queensland University of Technology, 2020. https://eprints.qut.edu.au/198192/1/Oleksii_Matsiaka_Thesis.pdf.

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This thesis provides novel insights into several contemporary problems in the mathematical biology involving migration of living cells. Primarily, we focus on cell motility and how dynamic changes in cell size affect collective cell migration. Additionally, this thesis investigates the importance of cellular heterogeneity and how it might affect the choice of modelling techniques we use to describe in vitro cell cultures.

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Martínez, Segura Amalia. "Coordination between cell size and global gene expression." Thesis, Imperial College London, 2017. http://hdl.handle.net/10044/1/62906.

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All cells change size during the cell cycle, as they have to double their mass in order to produce two equally sized daughter cells. For biochemical reactions to work as intended in a changing volume, the reactants’ concentration should be stable. Data shows that the total mass of RNA and protein per cell are proportional to cell size. Expression data also shows a coordinated increase of the majority of the transcripts when there is an increase in average cell length \citep{Zhurinsky2010}, the mechanism behind it is still poorly understood. In this study I took advantage of a previously described genetic model in fission yeast to investigate this question. This particular strain has a mutation in the \textit{cdc2} gene that makes the gene product sensitive to a nucleotide analogue. When the drug is added, cells arrest in G2 and start growing in size without replicating their genome or dividing. Using transcriptomics and proteomics, I characterised how all transcripts and the majority of proteins respond to an increase in cell size. As previously described, most transcripts and proteins concentration is proportional to cell size. However, there is a subset of molecules that scape the global regulation that the rest of the genome is subjected to. Examining the features that makes this molecule circumvent the coordination of size and gene expression could reveal what is the mechanism behind it. I also applied a mathematical model to study how the cell allocates its limited resources at a maximal size.

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Ginzberg, Miriam Bracha. "Size Control and Uniformity in Animal Cells." Thesis, Harvard University, 2015. http://nrs.harvard.edu/urn-3:HUL.InstRepos:17463956.

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The homogeneity in cell size observed in many normal proliferating tissues, and the contrasting size disparities characteristic of several cancers, suggest that control mechanisms coordinate growth and cell cycle progression, but the existence of such mechanisms has not been firmly established. To address this problem, we used quantitative fluorescence microscopy to measure cell cycle position, total protein content, and the level of cell growth regulators such as phosphorylated ribosomal protein S6, in tandem in single cells. Measurements were made on large numbers of cells drawn from proliferating populations of both non-transformed and cancerous cells. Analysis of the joint distribution of cell size and cell cycle position revealed a control mechanism that restricts cells to a specified size range at several points in the cell cycle. Combining our measurements with live-cell imaging showed that this restriction is the result of a negative correlation between cell size and subsequent growth rate, indicating that cells can sense their own size and modulate their growth accordingly. We also observed cell-size-dependent adjustments of cell cycle length, which further reduced size variability. We then identified drugs that change the mean cell size without disrupting the cell-autonomous control mechanism, as well as drugs that weaken the size-dependence of either growth rate or cell cycle progression and increase cell-to-cell size variability. In particular, long- and short-term drug treatments revealed that mTORC1 inhibition decouples the rate of cell growth from cell size, impairing the efficiency of cell size specification and increasing size variability. Our measurements of mTORC1 activity as a function of size and cell cycle position suggest that mTORC1 assumes its role in growth control following S-phase entry. Although mTORC1 activity, assayed by levels of mTORC1-target phosphorylation, increases abruptly during G1, it becomes correlated with cell size only upon G1 exit. The mTORC1 inhibitor rapamycin preferentially inhibits growth in S-phase cells. Taken together, these results indicate that the mTOR pathway maintains size homogeneity by stimulating growth in a cell-size-dependent manner after G1 exit. The screening and analysis methods developed here will be used to further elucidate the cellular size-control mechanism. Chemical Biology

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Pinent, Armengol Montserrat. "Adipose cell metabolism modulation by red wine procyanidins." Doctoral thesis, Universitat Rovira i Virgili, 2005. http://hdl.handle.net/10803/8650.

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Flavonoids, and more specifically, red wine procyanidins, have many beneficial effects against pathologies such as cardiovascular heart disease and related illnesses. Although adipose tissue has a central role in some of these pathologies, including obesity and diabetes, there is a lack of information about the effects of procyanidins on this tissue. This thesis addresses this question. The effects of a grape seed procyanidin extract (GSPE) on the lipid and glucose metabolism of adipocytes were evaluated by taking the 3T3-L1 cell line as a model of study. Results show that the GSPE has insulinomimetic effects, stimulating glucose uptake, glycogen synthesis and trigliceride synthesis. To achieve this, the GSPE shares some of the mechanisms and intracellular mediators of the insulin-signalling pathways (such as GLUT-4 translocation, PI3K and p38 MAPK) but it must also use other, complementary, mechanisms. These results suggest that procyanidins have beneficial effects on diabetes and/or insulin resistance. This is partially proven by in vivo studies that show that GSPE has antihyperglycemic properties on streptozotozin-induced diabetic rats. Also analyzed in this thesis are the molecular mechanisms used by GSPE to explain the already described lipolytic effects. Protein kinase A and PPARã are shown to be involved in these effects. Some of these results opened up another line of study into the effects of GSPE on the differentiation process of the 3T3-L1. These studies showed that procyanidins alter the differentiation of preadipocytes when added at the induction of differentiation. Since an increase in the number of adipocytes has a negative effect on obesity, this is a promising characteristic of GSPE that should be taken into account when its possible antiobesity properties are studied. Als flavonoides, i més concretament a les procianidines del vi negre, se'ls han atribuït moltes propietats beneficioses contra diverses patologies, com les malalties cardiovasculars i altres patologies relacionades. Tot i que el teixit adipós juga un paper important en algunes d'aquestes patologies, com la obesitat i la diabetis, la informació referent l'acció de les procianidines en aquest teixit és escassa. Aquesta tesis estudia els efectes de les procianidines derivades de pinyol de raïm (GSPE) en l'adipòcit, i per a dur-ho a terme es pren com a model d'estudi la línia cel.lular 3T3-L1. Per una banda es descriuen els efectes del GSPE en el metabolisme de lípids i glúcids de la cèl.lula adiposa. El GSPE fa un paper insulinomimètic: estimula la captació de glucosa, la síntesi de glicògen i la síntesi de triacil glicerols. L'anàlisi dels mecanismes moleculars per exercir aquests efectes mostra que GSPE en part comparteix mecanismes i vies de senyalització propis de la insulina (translocació de GLUT-4, PI3K, p38 MAPK); tanmateix, s'observa que GSPE ha d'usar també altres mecanismes complementaris. Aquests resultats suggereixen que GSPE pot tenir efectes positius en situacions de diabetis i/o resistència a insulina, donat que a més a més, els estudis in vivo mostren que GSPE és antihiperglicèmic en condicions de diabetis induïda per estreptozotocina. En aquesta tesis també s'analitzen els mecanismes moleculars que explicarien els efectes lipolítics de les procianidines descrits en estudis previs, i s'ha trobat que la proteina kinasa A i PPARã hi estan involucrats. Part d'aquests resultats han obert una altra via d'estudi sobre els efectes de la GSPE en el procés de diferenciació de la cèl.lula adiposa on s'ha observat que el tractament amb procianidines a l'inici de la diferenciació dificulta aquesta transformació. Donat que l'augment del nombre d'adipòcits afecta negativament la obesitat, aquest efecte de les procianidines és una característica prometedora que caldrà tenir en compte en l'estudi del seu possible paper antiobesitat.

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Ehigiator, Humphrey Nosayaba. "Immune modulation of B cell function by nematodes." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1999. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape4/PQDD_0018/NQ57365.pdf.

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Saleh, A. W. "Modulation of fetal hemoglobin in sickle cell anemia." Maastricht : Maastricht : Universiteit Maastricht ; University Library, Maastricht University [Host], 1998. http://arno.unimaas.nl/show.cgi?fid=8498.

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Bundy, Ruth Eldeca. "Redox modulation of vascular cell injury and adaptation." Thesis, Imperial College London, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.414511.

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Strong, Victoria. "Modulation of dendritic cell function by interleukin-10." Thesis, University of Oxford, 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.393790.

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Longhi, Maria Paula. "Modulation of CD4+ T cell responses by CD59a." Thesis, Cardiff University, 2006. http://orca.cf.ac.uk/56047/.

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CD59 is a GPI-anchored protein found in membrane microdomains known as lipid rafts. It is a complement regulator protein, which blocks the formation of the membrane attack complex by inhibiting binding of C9 to the C5-8 complex. Human CD59 has also been described as a co-stimulator of T cell activation. The aim of this project was to analyze the role of CD59 on T cell activation in vivo. For this purpose, anti-viral CD4+ T cell responses were analyzed in mice deficient in the mouse analogue of CD59 CD59a. Infection with recombinant vaccinia virus (rVV) and influenza virus, resulted in stronger virus-specific CD4+ T cell responses in Cd59a -/- mice compared to WT mice. This effect, which indicates that CD59a downmodulates antigen-specific T cell activity, was found to be complement independent. Experiments were performed to investigate the effect of CD59 expression on human CD4+ T cells. Blocking CD59 increased proliferation of the cells in vitro indicating that CD59 might similarly downmodulate human CD4+ T cell activity. Using mouse T cells, mechanisms underlying the effect of CD59a on CD4+ T cell activity were investigated. Results of these studies indicated that downmodulation of T cell activity through CD59a requires engagement of CD59a with a ligand expressed on APCs. To assess the biological consequences of CD59a deficiency, the extent of immunopathology induced following infection with influenza virus was compared in WT and Cd59a-/- mice. Immunopathology was exacerbated in Cd59a-/- mice, correlating with increased numbers of neutrophils and CD4+ T cells in infected lungs. When complement was inhibited, lung-infiltrating neutrophils in Cd59a-/- mice were much reduced while numbers of infiltrating-CD4+ T cell remained unchanged. These results demonstrate that CD59a is more than a regulator of complement but can in fact, alter both the innate and adaptive immune responses using both complement dependent and independent mechanisms.

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Shepherd, Ruth Juliet Basten. "Modulation of endothelial cell properties in systemic sclerosis." Thesis, King's College London (University of London), 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.404587.

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Chen, Naiyan. "Cell-type specific cholinergic modulation of the cortex." Thesis, Massachusetts Institute of Technology, 2013. http://hdl.handle.net/1721.1/84383.

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Thesis (Ph. D.)--Massachusetts Institute of Technology, Computational and Systems Biology Program, 2013. Cataloged from PDF version of thesis. "September 2013." Page 126 blank. Includes bibliographical references. The cholinergic innervation of the neocortex by afferent fibers originating in the nucleus basalis (NB) of the basal forebrain is implicated in modulating diverse neocortical functions including information processing, cortical plasticity, arousal and attention. To understand the physiological basis of these brain functions during cholinergic modulation, it is critical to identify the cortical circuit elements involved and define how their interactions contribute to cortical network dynamics. In this thesis, I present evidence showing how specific neuronal and glial cell types can be differentially modulated by acetylcholine (Ach), resulting in dynamic functional interactions during ACh-modulated information processing and cortical plasticity. Chapter 2 identifies somatostatin-expressing neurons as a dominant player in driving decorrelation and information processing through its intimate interactions with parvalbumin-expressing and pyramidal neurons. Chapter 3 highlights astrocytes and their interactions with pyramidal neurons as important drives for stimulus-specific cortical plasticity during cholinergic modulation. This is one of the earliest works that has mapped the functional role of distinct cell-types and their interactions to specific brain functions modulated by ACh, thereby setting the foundation for future studies to manipulate these specific functional interactions in both normal and diseased brains. by Naiyan Chen. Ph.D.

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Kovac, S. "Mechanisms and modulation of seizure induced cell death." Thesis, University College London (University of London), 2013. http://discovery.ucl.ac.uk/1383791/.

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Epilepsy, one of the most prevalent neurological diseases, is characterized by recurrent seizures, which are often poorly controlled by present treatments. Neuronal cell death, one of the complications of uncontrolled seizures, contributes substantially to the disease-burden through cognitive decline. This thesis explores mechanisms underlying neuronal death following seizure activity with a focus on mitochondria and ROS as major culprits of seizure induced neuronal cell death. By applying fluorescent dye imaging to glio-neuronal cell cultures, the first study examines mitochondrial mechanisms of seizure induced cell death and how such cell death can be reversed. The second study, using both in vitro and in vivo epilepsy models, explores sources of ROS production during seizures and in chronic epilepsy and how these can be reversed by blocking ROS producing enzymes. Experiments performed in the first part demonstrate that seizure activity leads to a sustained cyclosporine-A-sensitive depolarization of the mitochondrial membrane potential indicating mitochondrial permeability transition pore opening. This work proposes that neuronal ATP levels decrease, and correlate with the frequency of the oscillatory Ca2+ signal, indicative of activity-dependent ATP consumption. Cellular ATP levels during seizure like activity are dependent on the functioning of the mitochondrial complexes and seizure induced neuronal death is reduced when mitochondrial complex I substrate pyruvate is supplemented. In the second part I show that seizure activity induced increases in ROS production were seen both in vitro and in vivo. ROS production in vivo is elevated during periods of spontaneous brief seizures leading to a brain-region specific decrease in the major antioxidant system glutathione. Seizure-induced ROS were not dependent upon mitochondrial dysfunction but were generated in a Ca2+ -independent fashion through initially NADPH oxidase and later xanthine oxidase activity. Inhibition of either enzyme, consequently, reduced seizure-induced neuronal cell death. This works outlines strategies to treat seizure induced cell death.

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Moore, Anthony Norman. "Selenium modulation of gut epithelial cell stress responses." Thesis, University of Newcastle upon Tyne, 2017. http://hdl.handle.net/10443/3679.

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Selenium (Se) is an essential micronutrient necessary for human health. In humans, Se de ciency has been associated with in ammatory bowel disease (IBD) and increased risk of certain cancers, including colorectal cancer. Se has well established antioxidant and anti-in ammatory properties which are medi- ated, in part, though the actions of the selenoproteins, in which Se is present in the form of the amino acid selenocysteine (Sec). The cells of the gastrointestinal tract are exposed to stresses from pro-oxidative and hypoxic conditions, which have been suggested to be involved in the pathogenesis and pathology of IBD. Further characteristics of IBD are inappropriate immune responses of the gut epithelial cells to the gut microbiota. Thus, to help explain the roles of Se in IBD, it is important to understand the modulatory e ects of Se on the cell innate immune responses following challenge of intestinal epithelial cells with pathogen-associated molecular patterns (PAMPs), as well as oxidative and hypoxic stresses. The present work aimed to assess the roles of Se and the selenoproteins, SelH and TR1, in the responses of Caco-2 cell, modelling the gut epithelium, to hypoxia and infection, the latter replicated by challenge with S. typhimurium agellin. To investigate the responses of gut cells to low Se and PAMPs, undi erentiated Caco-2 cells with either supplemented with Se (40 nM selenite) or depleted of Se for 72 h before challenging with agellin (F) (500 ng/mL). The gene expression of the pro-in ammatory cytokines IL-8 and TNF- were measured in addition to the genes encoding the antimicrobial peptides (AMPs) hBD1 and hBD2. Data showed that Se depletion signi cantly a ected hBD1 expression (0.88-fold increase, P < 0.05), but that Se depletion plus F signi cantly increased the induced expression of all genes (IL-8: 1.68-fold, P < 0.001; TNF- : 0.71-fold, P < 0.001; hBD2: 1.74-fold, P < 0.001) compared with the Se supplemented cells. F and Se depletion were also associated with a signi cant increase in expression of TR1 (F: 1.68-fold, P < 0.001; Se depletion: 0.33-fold, P < 0.01) and GPX2 (F: 3-fold, P < 0.001; Se depletion: 11-fold, P < 0.001), but a signi cant decrease due to Se depletion in SelH (62 %, P < 0.001) and GPX1 (47 %, P < 0.001). The selenoprotein TR1 is an antioxidant enzyme and the primary regulator of the thioredoxin system (TXN), which has previously been shown to regulate immune responses. Knockdown of TR1 expression resulted in the reduced agellin-induced expression of IL-8 (40 %, P < 0.001), TNF-a (45 %, P < 0.01), hBD1 (40 %, P < 0.01) and hBD2 (45 %, P < 0.001). These data suggested that Se, through TR1, is involved in regulating the expression of agellin-induced immune e ectors. The selenoprotein SelH has also been suggested to have antioxidant functions. Knockdown of SelH was associated with the increased expression of the oxidative stress-associated genes NQO1 (0.41-fold, P < 0.001), and HMOX1 (1.78-fold, P < 0.001), supporting a role for SelH in the expression of oxidative stress-associated genes. The role of Se, through SelH and oxidative stress, in regulating the gut responses to agellin, has been discussed. The Caco-2 cell model is more representative of intestinal epithelial cells in vivo, when the cells are di erentiated and placed in a gaseous environment re ecting the oxygen gradient of the gut. Thus the F challenge experiments using di erentiated Caco-2 cells were repeated using a dualoxic environment. Interestingly, no potentiation of gene expression relating to the pro-in ammatory agents IL-8 and TNF- , and the defensins hBD1 and hBD2 was observed. These data suggested that the dualoxic environment completely diminished the e ects of Se depletion on the expression of immune e ectors IL-8, TNF- , hBD2 and hBD1, following agellin challenge. These data suggested the e ects of Se in more physiologically relevant intestinal epithelial cell models, more representative of the in vivo state, are required.

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Kim, Sun Wook. "Modulation of Stem Cell Fate by Electrical Stimulation." University of Cincinnati / OhioLINK, 2013. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1383812480.

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Dobbs, Cathleen M. "Neuroendocrine modulation of anti-viral cell-mediated immunity /." The Ohio State University, 1996. http://rave.ohiolink.edu/etdc/view?acc_num=osu1487941504294026.

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Santiago, Joana Filipa Marques. "Modulation of sperm motility using cell-penetrating peptides." Master's thesis, Universidade de Aveiro, 2017. http://hdl.handle.net/10773/22325.

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Mestrado em Biologia Molecular e Celular The large number of unintended pregnancies worldwide due to the non-use or failure of contraceptive methods and the fact that male contraceptives are limited to condom and vasectomy, highlight the urgent need for the development of new contraceptive methods. The mechanism of sperm motility acquisition in the epididymis constitutes an ideal target for new pharmacological male contraceptives since only the post-testicular sperm maturation is affected. It is known that protein phosphatase 1 subunit gamma 2 (PPP1CC2), a PPP1 isoform only present in testes and sperm, is essential for sperm motility acquisition. Protein-protein interactions (PPIs) have emerged as a promising class of drug targets and cell-penetrating peptides (CPPs) represents a recognized intracellular delivery system to target PPIs. The main goal of this work is to modulate PPP1CC2 complexes and, consequently, spermatozoa motility using peptides covalently coupled to CPPs. The results showed that both peptides tested could modulate sperm motility with a short incubation period, generally increasing the number of immotile spermatozoa. Additionally, we demonstrated that the peptide sequence that mimics the interaction interface between PPP1CC2 and a sperm-specific interactor – Akinase anchor protein 4 (AKAP4) – disrupted the PPP1CC2-AKAP4 interaction, resulting in arrest of sperm motility. The peptide that mimics the 22 amino-acid C-terminus of PPP1CC2 possible acts by disrupting the interaction between PPP1CC2 and isoform-specific interactors. Fifty putative isoform-specific interactors of PPP1CC2 C-terminus were identified by mass spectrometry and one of them was further validated (GPx4), suggesting new targets for similar contraceptive agents. In conclusion, this work confirmed the potential of CPPs to deliver peptide sequences that target unique PPIs in spermatozoa, clarified the mechanism of action of the peptides testes and identified other potential targets for new male contraceptives. O elevado número de gravidezes indesejadas a nível mundial e o facto de os contracetivos masculinos estarem limitados ao preservativo e à vasectomia refletem a necessidade urgente de desenvolvimento de novos métodos contracetivos. O mecanismo de aquisição de mobilidade dos espermatozoides no epidídimo constitui um alvo perfeito para novos agentes contracetivos dado que apenas a maturação pós-testicular é afetada. Sabe-se que a proteína fosfatase 1 subunidade gama 2 (PPP1CC2), uma isoforma presente apenas nos testículos e espermatozoides, é essencial para a aquisição de mobilidade no epidídimo. As interações proteína-proteína (PPIs) têm surgido como uma promissora classe de alvos terapêuticos e os cell-penetrating peptides (CPPs) representam um reconhecido sistema de entrega intracelular de sequências peptídicas com o potencial de modular PPIs. Assim, o principal objetivo deste trabalho é modular complexos PPP1CC2 específicos de testículo e espermatozoide e, consequentemente, a mobilidade dos espermatozoides recorrendo a sequências peptídicas covalentemente ligadas a CPPs. Os resultados mostram que ambos os péptidos testados são capazes de modular a mobilidade dos espermatozoides, mesmo com curtos períodos de incubação, aumentando o número de espermatozoides imóveis. Adicionalmente, foi demonstrado que o péptido que mimetiza a interface de interação entre PPP1CC2 e uma a A-kinase anchor protein (AKAP4) – um interactor específico no espermatozoide – interfere com a interação PPP1CC2-AKAP4, resultando em espermatozoides imóveis. O péptido que mimetiza os 22 aminoácidos do C-terminal da PPP1CC2 atua disrompendo a interação entre a PPP1CC2 e interatores específicos desta isoforma. Cinquenta interatores específicos do Cterminal da PPP1CC2 foram identificados por espectrometria de massa, sugerindo novos potenciais alvos para futura modulação. Um desses interatores (GPx4) foi posteriormente validado. Concluindo, este trabalho confirmou o potencial dos CPPs na entrega de sequências peptídicas que têm como alvo PPIs únicas do espermatozoide, clarificou o mecanismo de ação dos péptidos testados e identificou potenciais alvos para novos contracetivos masculinos.

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Wang, Kai. "Substrate Nanotopography and Stiffness Modulation of Cell Behavior." Thesis, University of North Texas, 2019. https://digital.library.unt.edu/ark:/67531/metadc1505286/.

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The physical characteristics (i.e., nanostructure and stiffness) of the extracellular matrix where cells reside have been shown to profoundly affect numerous cellular events in vivo and also been employed to modulate cell behavior in vitro, yet how these physical cues regulate cell behavior is still elusive. Therefore, we engineered a variety of nanotopographies with different shapes and dimensions, and investigated how the nanotopographical cue, through focal adhesions-cytoskeleton-nucleus pathway, affected cell phenotype and function. We further designed and fabricated well-defined substrates which had either identical biochemical cue (adhesive ligand presentation) but different nanotopographical cues or identical nanotopography but different biochemical cues, and dissected the roles of these cues in cell modulation. In addition, we revealed that the human mesenchymal stem cells (hMSCs) could obtain nanotopographical memory from the past culture environment, and the nanotopographical memory influenced the future fate decision of the hMSCs. Moreover, we evaluated the effects of substrate nanotopographical and stiffness cues on the fibrogenesis of human lung fibroblasts in response to carbon nanotubes and highlighted the significance of these physical cues in the development of physiologically relevant in vitro models for nanotoxicological study. The mechanistic understanding of the physical regulation of cell behaviors will provide important insight into the advancement of cell culture technologies and the recreation of biomimetic in vitro tissue/organ models.

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Keifenheim, Daniel L. "Cell Size Control in the Fission Yeast Schizosaccharomyces pombe: A Dissertation." eScholarship@UMMS, 2015. http://escholarship.umassmed.edu/gsbs_diss/784.

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The coordination between cell growth and division is a highly regulated process that is intimately linked to the cell cycle. Efforts to identify an independent mechanism that measures cell size have been unsuccessful. Instead, we propose that size control is an intrinsic function of the basic cell cycle machinery. My work shows that in the fission yeast Schizosaccharomyces pombe Cdc25 accumulates in a size dependent manner. This accumulation of Cdc25 occurs over a large range of cell sizes. Additionally, experiments with short pulses of cycloheximide have shown that Cdc25 is an inherently unstable protein that quickly returns to a size dependent equilibrium in the cell suggesting that Cdc25 concentration is dependent on size and not time. Thus, Cdc25 can act as a sizer for the cell. However, cells are still viable when Cdc25 is constitutively expressed suggesting that there is another sizer in the case that Cdc25 expression is compromised. Cdc13 is a likely candidate due to the similar characteristics to Cdc25 and the ability to activate Cdc2. Cdc13 accumulates during the cell cycle in a manner similar to Cdc25. I show that in the absence of Cdc2 tyrosine phosphorylation, the cell size is sensitive to Cdc13 activity showing that Cdc13 accumulation can determine when cells enter mitosis. These results suggest a two sizer model where Cdc25 is the main sizer with Cdc13 acting as a backup sizer in the event of Cdc25 expression is compromised. Additionally, in the absence of Cdc2 phosphorylation by the kinases Wee1 and Mik1, mitotic entry is regulated by the activity of Cdc2. In the absence of Cdc2 phosphorylation, this activity is regulated by binding of cyclins to Cdc2. Under these circumstances, the activity of Cdc13 can regulate mitotic entry provide further evidence that Cdc13 could be a sizer of the cell in the case where Cdc25 expression is compromised. The results I present in this dissertation provide the groundwork for understanding how cells regulate size and how this size regulation affects cell cycle control in S. pombe . The results show how the intrinsic cell cycle machinery can act as a sizer for the G2/M transition in S. pombe . Interestingly, this mitotic commitment pathway is well conserved suggesting a general solution for size control in eukaryotes at the G2/M transition. Understanding the mechanism of how protein concentration is regulated in a size dependent manner will give much needed insight into how cells control size. Elucidating the mechanism for size control will capitalize on decades of research and deepen our understanding of basic cell biology.

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Gerrish, Kevin Edward 1965. "Modulation of Nb2 cell mitogenesis by peripheral benzodiazepine ligands." Thesis, The University of Arizona, 1989. http://hdl.handle.net/10150/277200.

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In this study, we investigated the effects of the peripheral benzodiazepine ligands, Ro5-4864 (putative antagonist) and PK 11195 (putative antagonist) on prolactin stimulated mitogenesis in Nb2 cells. Ro5-4864 and PK 11195 at 10⁻⁹ M maximally enhanced prolactin stimulated mitogenesis. At 10⁻⁶ M Ro5-4864 inhibited prolactin stimulated mitogenesis. Clonazepam, a ligand for the central benzodiazepine receptor had no effect on mitogenesis. Interaction studies were undertaken to determine if Ro5-4864 and PK 11195 act on the same site. The ability of each ligand to enhance the mitogenic action of prolactin was blocked by a 10⁻⁶ M concentration of the other ligand. Finally, simultaneous addition of 10⁻⁹ M of the ligands resulted in no additive effect over each ligand alone. These data show that peripheral benzodiazepine ligands modulate prolactin-stimulated mitogenesis and suggests they interact at the same binding site.

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Mazade, Reece Eric. "Modulation Of Inner Retinal Inhibition With Light Adaptation." Diss., The University of Arizona, 2015. http://hdl.handle.net/10150/565903.

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The retina is able to adjust its signaling over a wide range of light levels. A functional result of this is increased visual acuity at brighter luminance levels, such as during the day, due to changes in the organization of retinal receptive fields. This process is commonly referred to as light adaptation. These organizational changes have been shown to occur at the level of the ganglion cells, the output neurons of the retina, which have shifts in their excitatory center-inhibitory surround receptive fields that increase their sensitivity to small stimuli. Recent work supports the idea that light-adapted changes in ganglion cell spatial sensitivity are due in part to inner retinal signaling changes, possibly including changes to inhibition onto bipolar cells, the interneurons at the center of retinal signal processing. However, it is unknown how inhibition to the bipolar cells changes with light adaptation, how any changes affect the light signal or what mediates the changes to the bipolar cells that have been suggested by previous reports. To determine how light adaptation affects bipolar cell inhibition, the inhibitory inputs to OFF bipolar cells were measured. OFF bipolar cells, which respond to the offset of light, in particular may be involved in retinal adaptation as they bridge dim- and bright-light retinal pathways. Their inputs were compared between dark- and light-adapted conditions to determine how any inhibitory changes affects their output onto downstream ganglion cells. We found that there was a compensatory switch from primarily glycinergic-mediated inhibition to OFF bipolar cells in the dark to primarily GABAergic-mediated inhibition in the light. Since glycinergic and GABAergic inhibition perform very different roles and are mediated by morphologically different cells, it is likely this switch underlies a change in the spatial distribution of inhibition to these cells. We found that the spatial inhibitory input to OFF bipolar cells became significantly smaller and narrower with light adaptation, translating to smaller inhibitory surrounds of the OFF bipolar cell receptive fields. Through a model, our data suggested that the OFF bipolar cell output to downstream ganglion cells was stronger in the light, due to the narrowing and reduction in the spatial input, to small light stimuli. This would effectively be one way the retina could use to increase visual acuity. Additionally, we found that the inhibitory changes to OFF bipolar cells with light-adaptation are partially mediated by dopamine D1 receptor signaling. Dopamine is released in the light and has been shown to be an important modulator of retinal light-adaptation. However, there are likely other factors involved in mediating inhibitory changes to OFF bipolar cells. Through these studies, we show that light adaptation heavily influences inner retina inhibition and likely plays a prominent role in determining and shaping light signals under different ambient light conditions which may ultimately be one mechanism for increasing visual sensitivity and acuity.

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Teixeira, de Matos Cristina. "Modulation of natural killer cell and T-cell functions by CD94/NKG2A receptors /." Stockholm, 2006. http://diss.kib.ki.se/2006/91-7140-846-0/.

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Garrison, Jennifer L. "Small molecule modulation of protein secretion." Diss., Search in ProQuest Dissertations & Theses. UC Only, 2007. http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqdiss&rft_dat=xri:pqdiss:3261264.

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Miettinen, Teemu P. "On connections between Metazoan cellular metabolism and cell size." Thesis, University of Dundee, 2015. https://discovery.dundee.ac.uk/en/studentTheses/3cdc8663-1167-4a8b-ad5c-698c20695664.

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All animal cells maintain cell size homeostasis, where cell growth (increase in size) is balanced with proliferation (reduction in size via cell division). Yet, different cell types have different sizes and there are physiologically relevant situations where animal cells undergo major cell size changes. So how is cell size regulated? And why is cell size regulated? Are there specific cellular processes that have different functionality in different sized cells? This thesis investigates these questions from the perspective of cellular metabolism. Using a Cyclin dependent kinase 1 knockout mouse model with different degrees of hepatocytes enlargement, gene expression levels were correlated with cell size in vivo. This revealed that the relative expression of mitochondrial and lipid biosynthesis genes are downregulated with increasing cell size. However, mitochondrial content of the liver samples was not decreased, suggesting that cell functions and cell contents scale differently with cell size. To better investigate how mitochondrial functions scale with cell size in non-mutant cells, a novel and high throughput flow cytometry based single-cell analysis method called CoSRA was developed. Using fluorescence mitochondrial probes CoSRA revealed that, while mitochondrial content increases linearly with cell size, mitochondrial membrane potential is decreased in the very smallest and the largest cells. These effects were independent of cell cycle and all animal cell types examined displayed similar effects. Similar nonlinearity was observed in mitochondrial respiration. Furthermore, cell-to-cell variability in mitochondrial membrane potential was minimised in cells which are close to the median cell size of the whole population. The cell size dependence of mitochondrial functions was regulated by mitochondrial dynamics. It was also investigated if mitochondrial functions or lipid biosynthesis are capable of regulating cell size in human cell culture models. Various mitochondrial inhibitions increased cell size by reducing proliferation. Similar results were seen with inhibitions on lipid biosynthesis and especially with inhibitions of mevalonate pathway. Systematic dissection of the mevalonate pathway revealed that protein geranylgeranylation is required for maintaining normal cell size and proliferation ratio. Geranylgeranylation of the recycling endosome regulating protein RAB11 was identified to be at least partially responsible for the cell size regulation by the mevalonate pathway. Furthermore, the link from the mevalonate pathway to RAB11 was found to regulate basal autophagic flux, thus providing a novel connection from lipid biosynthesis to other growth regulating processes. In conclusion, this thesis provides evidence for cell size dependent metabolism, where mitochondrial functions do not increase linearly with cell size. This provides conceptual insights into organelle scaling with cell size and a potential mechanism for maintenance of cell size homeostasis. In addition, mitochondria and lipid synthesis are identified as critical processes for normal cell size homeostasis.

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Silva, Michael Santos. "Flow cytometric of c-FLIPl-mediated regulation of cell cycle and cell population size." Master's thesis, Universidade de Aveiro, 2017. http://hdl.handle.net/10773/22487.

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Mestrado em Bioquímica - Bioquímica Clínica c-FLIP é uma proteína conhecida pela sua capacidade de se ligar ao DISC, onde compete com a procaspase-8 pela interação com FADD. No entanto, existem evidências que a sua isoforma longa consegue regular também o ciclo celular e mecanismos de proliferação. Para além disso, a atividade de c-FLIPL pode ser controlada por fosforilação. Assim, o objetivo deste estudo é perceber como a fosforilação no resíduo de serina 227 nesta proteína afeta a proliferação e ciclo celular. Neste estudo, observamos que a sobre-expressão de c-FLIPL com uma mutação de serina para alanina no resíduo 227 levou a uma diminuição da capacidade proliferativa dessas células. O uso de citometria de fluxo permitiu verificar este decréscimo na capacidade proliferativa, assim como uma acumulação de células na fase G1 do ciclo celular aquando da sobre-expressão de S227A c-FLIPL. Os resultados obtidos sugerem que a sobre-expressão de c-FLIPL controla a população celular através da transição G1/S, através da sua fosforilação no resíduo 227. No entanto, mais estudos são necessários para se perceber a partir de qual mecanismo esta transição é afetada c-FLIP is a protein known for its capacity to bind to the DISC and compete with procaspase-8 for FADD interaction. However, published studies have shown that c-FLIPL can regulate cell cycle and proliferation. Similarly to many other proteins, c-FLIP can be regulated by phosphorylation. Therefore, the aim of this work was to understand how the phosphorylation of S227 residue on c-FLIPL affects cell cycle and cell proliferation. We observed that overexpression of phosphodeficient mutant c-FLIPL lead to a decrease in cell proliferation. Flow cytometric analysis confirmed this decrease, as well as an accumulation of cell at G1 phase of cell cycle, when overexpressing S227A c-FLIPL. Our results suggest that c-FLIPL overexpression controls cell population size by controlling the G1/S transition, via its phosphorylation. Nonetheless, further studies need to be done to understand which mechanism affects this transition.

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Salako, Michael Albert. "Enteroviral modulation of host cell and drug-induced apoptosis." Thesis, University of Surrey, 2005. http://epubs.surrey.ac.uk/843342/.

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Coxsackievirus B4 (CVB4) is a common infection. Most cases are symptom-free but occasionally chronic sequelae emerge such as insulin-dependent diabetes mellitus, and dilated myocardiopathy and virus persistence is thought to contribute to these conditions. Persistent viruses frequently possess mechanisms that promote survival of the infected cell and inhibit apoptosis. Mitochondria play a central role in mediating apoptotic effects and may thus be targeted by viruses. Recently, poliovirus was shown both to inhibit apoptosis and to disrupt mitochondrial function and thus we have sought similar effects in the related virus CVB4. CVB4 infection of HeLa cells did not disrupt mitochondrial function and energy levels were maintained. In contrast to cells entering apoptosis, mitochondrial membrane potential increased after infection; at the same time infected cells developed resistance to drug-induced apoptosis via both death receptor and mitochondrial pathways. Despite this resistance, the apoptotic machinery was fully activated by drug treatment of infected cells and the processing of procaspase-3 to its cleaved form (caspase-3) was normal. However, DEVDase activity was inhibited in drug-treated CVB4 infected cells and extracts from infected cells suppressed the activity of recombinant caspase-3 in vitro. We conclude that this effect was mediated at least in part by virus protein 2BC since immunoprecipitation of caspase-3 from infected cells co-precipitated a 2BC-sized (48kDa protein). Further, protein 2BC expressed in vitro was shown to bind to caspase-3, and expression of 2BC alone in HeLa cells led to the inhibition of caspase-3 activity. Taken together these data imply that CVB4 protein 2BC acts to mitigate the consequences of apoptosis induction by associating with caspase-3 and inhibiting its function. These results suggest that 2BC is acting as a viral IAP and this is the first report of such an ability for an RNA virus.

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Liwski, Robert Stefan. "Modulation of T cell-mediated responses by Nippostrongylus brasiliensis." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1999. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape8/PQDD_0020/NQ49276.pdf.

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Heijstek, Helena Cornelia. "Modulation of human dendritic cell function by therapeutic agents." [S.l. : Amsterdam : s.n.] ; Universiteit van Amsterdam [Host], 2002. http://dare.uva.nl/document/64240.

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Szabat, Marta. "Characterization and modulation of adult pancreatic β-cell maturity". Thesis, University of British Columbia, 2010. http://hdl.handle.net/2429/27295.

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The functional maturation and dedifferentiation of β-cells is central to diabetes pathogenesis and to beta-cell replacement therapy. Despite its importance, the dynamics of adult β-cell maturation remain poorly understood because it has previously been difficult to study the process directly. A novel dual fluorescent reporter lentiviral vector was developed capable of tracking the differentiation status of single β-cells in culture. Using this labeling tool, an immature β-cell state was identified in adult primary human and mouse islets and beta-cell lines. The immature β-cell state was characterized by Pdx1 promoter activity but undetectable insulin promoter activity. Lineage analysis of labeled single adult human, mouse and MIN6 β-cells revealed that a fraction of the immature β-cells underwent maturation over time in culture by robustly activating the insulin promoter. Immature beta-cells also exhibited a significantly downregulated profile of mature β-cell genes. These cells had increased proliferation and a reduced glucose-stimulated insulin secreting function. In order to manipulate the adult β-cell maturation state, a screen for candidate growth/differentiation factors using image-based approaches was performed. Activin A and its antagonist follistatin were found to modulate adult β-cell maturity. Activin A had a strong negative effect on β-cell maturity, reducing insulin promoter activity, insulin secretion and the expression of mature β-cell genes. Follistatin reversed the effects of endogenous activin A and augmented β-cell maturity. These results uncovered a local autocrine/paracrine regulatory mechanism that controls the maturation state of adult β-cells. In addition, gene expression profiling at the whole genome level was used to analyze purified immature and mature β-cells from humans, mice and the MIN6 β-cell line. These analyses revealed that immature β-cells have increased expression of multiple islet hormones and have enriched expression levels of many genes known to be involved in pancreatic development, stem cell plasticity, proliferation and apoptosis. Conversely, mature β-cells are enriched in genes related to maintaining the mature β-cell phenotype. Collectively, these experiments contribute to the understanding of maturation and plasticity of adult pancreatic β-cells. The results have significant implications for islet regeneration and for in vitro generation of functional β-cells to treat diabetes.

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Nsiah, Barbara Akua. "Fluid shear stress modulation of embryonic stem cell differentiation." Diss., Georgia Institute of Technology, 2012. http://hdl.handle.net/1853/47552.

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Vascularization of tissue-engineered substitutes is imperative for successful implantation into sites of injury. Strategies to promote vascularization within tissue-engineered constructs have focused on incorporating endothelial or endothelial progenitor cells within the construct. However, since endothelial and endothelial progenitor cells are adult cell types and limited in number, acquiring quantities needed for regenerative medicine applications is not feasible. Pluriopotent stem cells have been explored as a cell source for tissue-engineered substitutes because of their inherent ability to differentiate into all somatic cell types, including endothelial cells (ECs). Current EC differentiation strategies require laborious and extensive culture periods, utilize large quantities of expensive growth factors and extracellular matrix, and generally yield heterogenous populations for which only a small percentage of the differentiated cells are ECs. In order to recapitulate in vivo embryonic stem cell (ESC) differentiation, 3D stem cell aggregates or embryoid bodies (EBs) have been employed in vitro. In the developing embryo, fluid shear stress, VEGF, and oxygen are instructive cues for endothelial differentiation and vasculogenesis. Thus, the objective of this work was to study the effects of fluid shear stress pre-conditioning of ESCs on EB endothelial differentiation and vasculogensis. The overall hypothesis is that exposing ESCs to fluid shear stress prior to EB differentiation will promote EB endothelial differentiation and vasculogenesis. Pre-conditioning ESCs with fluid shear stress modulated EB differentiation as well as endothelial cell-like cellular organization and EB morphogenesis. To further promote endothelial differentiation, ESCs pre-conditioned with shear were treated with VEGF. Exposing EBs formed from ESCs pre-conditioned with shear to low oxygen resulted in increased production of VEGF and formation of endothelial networks. The results of this work demonstrate the role that physical forces play in modulating stem cell fate and morphogenesis.

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Thèses sur le sujet « Modulation of cell size »

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