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1
Seeley, Marguerite R. "Developmental toxicity of alkylating agents in differentiating cell cultures /." Thesis, Connect to this title online; UW restricted, 1996. http://hdl.handle.net/1773/8464.
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Limoges, Mireille. "Apoptosis in primary cultures of rat Leydig cells." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1998. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp02/NQ36586.pdf.
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Dove, N. S. "Electrophysiological studies on primary cultures of skin cells." Thesis, University of Cambridge, 1997. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.598614.
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To model sweat gland fluid secretion in vitro, human apocrine glands were isolated from axillary skin and their secretory coil cells were cultured and mounted in an Ussing chamber. Cultured apocrine coil cells exhibited an amiloride-sensitive inward resting short circuit current, suggesting an absorptive phenotype. However, evidence of a chloride-secretory phenotype was also observed: transient inward currents were recorded in response to autonomic secretagogues and inflammatory mediators, and these were not abolished by amiloride, but they were reduced by half in chloride-free buffer. This may indicate a role for chloride secretion in mediating the stimulated currents. Further, sustained inward currents were recorded in 58% of tissues which were attenuated by frusemide, implying the involvement of the Na+/K+/Cl- transporter. Assuming that secondary active chloride transport is responsible for aqueous fluid secretion in the apocrine sweat gland, these data are compatible with some maintenance of an in vivo phenotype in culture. To study the cellular determinants of eccrine sweat gland differentiation, eccrine sweat gland-associated fibroblasts were cultured and compared with derminal fibroblasts from the same subject, demonstrated a different pattern of outgrowth and proliferation. Therefore these cells may represent a novel fibroblast subtype. To improve the differentiation of eccrine sweat gland coil cells in vitro, they were co-cultured with eccrine fibroblasts. Eccrine sweat gland cells grown on a Transwell as a control demonstrated apparent dedifferentiation to a reabsorptive phenotype, indicated by the amiloride-sensitivity of their resting and agonist-stimulated inward short circuit currents. Eccrine cells co-cultured with eccrine fibroblasts, however, demonstrated agonist-stimulated outward currents in the presence of amiloride.
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Montgomery, Sarah Lynn. "Impedance measurement system for embryonic stem cell and embryoid body cultures." Thesis, Atlanta, Ga. : Georgia Institute of Technology, 2008. http://hdl.handle.net/1853/24661.
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Dempsey, Katherine. "Monitoring individual cells within cell cultures using image processing and pattern recognition techniques." Thesis, Keele University, 2017. http://eprints.keele.ac.uk/4179/.
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Cells are the building blocks of the human body which are normally specialised by type in accordance with their function. Human cells interact with each other to form the tissues that make up the body. Consequently, it is important to study the behaviour and interactions of these cells at the microscale level, so that the causes of cellular irregularities can be identified; and, possible treatments can be devised. This project aimed to create algorithms that were capable of tracking a variety of cells types within both single cultures and mixed cultures, and from this generate data that was relevant to current clinical trials. There have been successes in tracking some cells types, most notably articular chondrocytes and spinal disk cells. In terms of data generated there has been successes in a whole variety of different types of clinical trials. The algorithms used here have been able to identify the point of mitosis. They have created a better method of determining neural growth and from this have shown that neurons co-cultured with MCSs can grow in places with neural inhibitors. Through the use of algorithms that can analyse culture in three dimensional structures it has been shown that neurons are more affected by topographical cues than chemical cues in their direction of growth. It has also been shown that vesicles are more likely to appear on smaller back disk cells. In the study of gels, it has been found that the more transparent gels are better for imaging. Finally, it has been shown that MSCs and chondrocytes behave differently when in single and co-cultures. These discoveries would not have been possible without the use of the algorithms that allowed for the study of individual cells within a larger culture.
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Szemiel, Agnieszka M. "Replication of Bunyamwera virus in mosquito cells." Thesis, University of St Andrews, 2011. http://hdl.handle.net/10023/2570.
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The Bunyaviridae family is one of the largest among RNA viruses, comprising more than 350 serologically distinct viruses. The family is classified into five genera, Orthobunyavirus, Hantavirus, Nairovirus, Phlebovirus, and Tospovirus. Orthobunyaviruses, nairoviruses and phleboviruses are maintained in nature by a propagative cycle involving blood-feeding arthropods and susceptible vertebrate hosts. Like most arthropod-borne viruses, bunyavirus replication causes little damage to the vector, whereas infection of the mammalian host may lead to death. This situation is mimicked in the laboratory: in cultured mosquito cells no cytopathology is observed and a persistent infection is established, whereas in cultured mammalian cells orthobunyavirus infection is lytic and leads to cell death. Bunyaviruses encode four common structural proteins: an RNA-dependent RNA polymerase, two glycoproteins (Gc and Gn), and a nucleoprotein N. Some viruses also code for nonstructural proteins called NSm and NSs. The NSs protein of the prototype bunyavirus, Bunyamwera virus, seems to be one of the factors responsible for the different outcomes of infection in mammalian and mosquito cell lines. However, only limited information is available on the growth of bunyaviruses in cultured mosquito cell lines other than Aedes albopictus C6/36 cells. Here, I compared the replication of Bunyamwera virus in two additional Aedes albopictus cell clones, C7-10 and U4.4, and two Aedes aegypti cell clones, Ae and A20, and investigated the impact of virus replication on cell function. In addition, whereas the vertebrate innate immune response to arbovirus infection is well studied, relatively little is known about mosquitoes’ reaction to these infections. I investigated the immune responses of the different mosquito cells to Bunyamwera virus infection, in particular antimicrobial signaling pathways (Toll and IMD) and RNA interference (RNAi). The data obtained in U4.4 cells suggest that NSs plays an important role in the infection of mosquitoes. Moreover infection of U4.4 cells more closely resembles infection in Ae and A20 cells and live Aedes aegypti mosquitoes. My data showed that the investigated cell lines have various properties, and therefore they can be used to study different aspects of mosquito-virus interactions.
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Bell, Cindy Lea. "Transport studies in primary cultures of mouse renal epithelial cells." Thesis, McGill University, 1986. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=75363.
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The Hyp (hypophosphatemic) mouse, a murine homologue of X-linked hypophosphatemia (XLH) in man, is a Mendelian disorder of phosphate (Pi) homeostasis. The mutant genotype is characterized by abnormal Pi transport at the brush border membrane (BBM) of the proximal tubule and a defect in renal metabolism of vitamin D$ sb3$. The exact nature of these defects has not been elucidated.In order to determine if the defect is intrinsic to the renal cell or dependent upon an extrinsic humoral factor, I established primary cultures of renal epithelial cells from normal and Hyp mouse kidney. The cultures demonstrated several differentiated properties of epithelial cells of the renal proximal tubule, the site of the Pi transport defect in the Hyp mouse.
Primary cultures initiated from Hyp mice had decreased Pi transport (expressed as an uptake ratio, Pi/$ alpha$-MG), and increased production of 24,25 dihydroxyvitamin D$ sb3$. These results provide evidence for the intrinsic nature of the primary defect in the Hyp mouse.
This appears to be the first time that expression of a mutant transport gene has been demonstrated in cultured renal cells.
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Zhang, Fan. "Electric and electrochemical responses of adherent cells : application of microfabrication technologies." Paris 6, 2011. http://www.theses.fr/2011PA066194.
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La connaissance des comportements cellulaires tels que l'adhésion, migration et prolifération cellulaire est importante pour l'ingénierie tissulaire et de l'implantologie. Ce travail de thèse a été développé pour obtenir une vision plus claire sur les activités cellulaires in-vitro en utilisant des substrats micro/nanofabriqués et des méthodes d’analyses électriques/électrochimiques. Des substrats avec des motifs divers et variés, incluant des nanoélectrodes de haute densité et des microstructures à trois dimensions, ont été obtenu pour culture cellulaire et analyse par méthodes électriques ou électrochimiques. L’intégration de ces structures dans un dispositif microfluidique a été également démontrée. L'adhésion, migration et prolifération cellulaire ainsi que l'activité métabolique des cellules ont été étudié par mesure de voltamétrie cyclique et d'impédance électrique. En combinaison avec les techniques optiques pour l'observation de la morphologie cellulaire et la densité de cellules, les mesures électriques ou électrochimiques nous ont permis d’étudier des nouveaux effets de substrats ou électrodes micro et nano-structurés sur la culture cellulaire et les activités métaboliques de cellules en culture
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Pucihar, Gorazd. "Induced transmembrane voltage and electropermeabilization of cells in cultures in vitro." Toulouse 3, 2006. http://thesesups.ups-tlse.fr/1038/.
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Quand une cellule est exposée à un champ électrique externe, la tension électrique transmembranaire (ITV) est modifiée. Pendant l'exposition, l' ITV se superimpose au potentiel de repos (RTV) et quand la somme des deux tensions excède une valeur critique, la perméabilité de la membrane cellulaire augmente transitoirement localement. Ce phénomène est désigné comme electropermeabilisation. Dans beaucoup d'applications de l'electropermeabilisation une permeabilisation efficace et en même temps réversible est essentielle. Ainsi, une prédiction de l'expérience, qui implique l'évaluation de l'amplitude de l'ITV pour déclencher la permeabilisation, est exigée. Le problème est critique dans des tissus, où la géométrie cellulaire est plus compliquée, les cellules sont assez proches pour affecter le champ électrique autour d'elles et elles sont souvent connectées entre elles. Dans tous ces cas, une description analytique de l'ITV n'est en général pas accessible et des méthodes numériques sont ainsi souvent la seule approche envisageable. En raison de la complexité de la structure tissulaire, les modèles sont macroscopiques, et on ne considère pas la structure cellulaire détaillée, ou en cas des modèles microscopiques, les modèles sont construits utilisant des formes géométriques simples (des semi-sphères, des cubes). Pour mieux comprendre comment le champ électrique interagit avec des tissus, nous avons construit les modèles microscopiques réalistes de cellules irrégulièrement formées, des groupes de telles cellules et des suspensions denses. Le travail a alors été développé sur le plan expérimental au niveau de la cellule isolée. Les mesures de cinétique de transport de membrane ont montré que l'electropermeabilisation avec des amplitudes d'impulsion ou des durées d'impulsion progressivement croissantes conduit à des transports accrus dans des cellules. Une large augmentation a été observée dans les milisecondes après le début d'une impulsion, suivie par une augmentation de fluorescence progressive. Les résultats mesurés sur un intervalle de temps de 400 µ S ont révélé que le transport à travers la membrane permeabilisée ne peut être détecté que 100 µ S après le début de l'impulsion. En plus, une dynamique différente d'augmentation de fluorescence pendant et après l'impulsion a été observéeWhen a biological cell is exposed to an external electric field, induced transmembrane voltage (ITV) forms on its membrane. During the exposure, ITV superimposes to the native or resting transmembrane voltage (RTV) and when the sum of both voltages exceeds some threshold value, the permeability of the cell membrane in these regions transiently increases. This phenomenon is termed electropermeabilization. In many applications of electropermeabilization an efficient and at the same time reversible permeabilization is essential (e. G. DNA electrotransfer). Thus, a careful planning of the experiment, which involves the estimation of the amplitude of ITV leading to cell permeabilization, is required. The problem arises in case of tissues, where cell geometry is more complicated, cells are close enough to affect the electric field around each other, and they are often connected with pathways between them. In all these cases, an analytical description of ITV is in general not attainable and numerical methods are often the only feasible approach. Due to the complexity of tissue structure, numerical models are either macroscopic, where detailed cell structure is notconsidered, or in case of microscopic models, the models are constructed using simple geometrical shapes (semi-spheres, cubes). To better understand how the electric field interacts with tissues on a microscopic (single cell) level, which in turn determines the macroscopic behavior of the tissue, we constructed realistic microscopic models of irregularly shaped cells, clusters of such cells, and dense suspensions. Regarding the shape, density and connections between cells, these cell assemblies are in their complexity close to tissues. First, the amplitude of resting transmembrane voltage of cells used in the study was determined. Next, calculations of ITV were performed on models of single spherical, single attached cells, and cell clusters and they were compared to measurements of ITV on the same cells, from which the models were constructed. The course of electropermeabilization of these cells was then monitored and the results were compared with measurements and calculations of ITV. In a separate experiment, a detailed investigation of kinetics of molecular transport into cells after permeabilization was performed. Similarly, for dense cell suspensions, the ITV calculated on a model of suspension was compared with the fraction of permeabilized cells measured in suspensions with increasing cell densities. Measurements of resting transmembrane voltage (RTV) were performed by means of a slow potentiometric fluorescent dye TMRM on different cell lines in culture media and media with progressively decreasing conductivities. ITV was measured on single spherical cells, single irregularly shaped cells, and cell clusters with a fast potentiometric fluorescent dye di-8- ANEPPS. The cross-section fluorescence images of the same cells on which the measurements of ITV were performed, were used to construct realistic numerical models of cells and the ITV on these models was then calculated with finite elements method. Finitethickness, nonzero conductivity cell membrane in the model was replaced by a boundary condition in which a specific surface conductivity was assigned to the interface between the cell interior and the exterior. Electropermeabilization of cells was followed by monitoring thechanges in intracellular fluorescence of membrane-impermeant fluorescent dye Propidium Iodide. Measurements of RTV showed that in physiological conditions (cells in culture medium) and in the presence of pulsing buffer, RTV on investigated cell lines is low (between -4 and -35 mV for suspended cells and between -18 and -27 mV for attached cells). Therefore, in experiments involving electropermeabilization ITV can be used as a rough approximate of the total voltage on the membrane, while RTV can be neglected. RTV in cells in media with decreasing conductivities gradually decreased, but less than expected from theoretical calculations. This was partly attributed to overestimated intracellular concentration of potassium. However, it is also possible that the method for measuring RTV, although reported as efficient, was not suitable for these experiments. Measurements of ITV on single spherical cells, single attached cells, and cell clusters were in qualitative agreement with results of numerical calculations, while in some cases discrepancies in measured and calculated amplitudes could be observed. This was attributed to variations of the slope of calibration curve, the differences between the actual and implemented parameters of the model, physiological state of cells, and experimental setup. In addition, we observed that at pulse parameters used in measurements of ITV, cells in clusters behaved as electrically connected, i. E. A cluster acted as one giant cell. Numerical calculations on models of cells where cell membrane was replaced with a boundary condition resulted in considerably lower number of mesh elements and consequently shorter time needed to solve the problem. We also demonstrated that calculations of ITV on simplified models of irregularly shaped cells can lead to considerable deviations from ITV calculated on a realistic model. Electric field orientation affects the amplitude and distribution of calculated ITV and consequently permeabilization. Namely, cells oriented with their longer axis parallel to the field are more likely to get permeabilized than the same cells oriented perpendicularly to the field. Comparison of measured and calculated ITVs with observations of electropermeabilization on single spherical and single attached cells confirmed that permeabilization occurs in those regions of the membrane, where the absolute value of ITV is the highest (the regions facing the electrodes). Additional experiments performed on single spherical cells showed that during and immediately after the pulse, the fluorescence from cells increases asymmetrically if unipolar pulses were delivered, while symmetrical fluorescence was observed for bipolar pulses. These observations were attributed to electrophoretical effect of the pulse. On a longer time scale, asymmetry in fluorescence was still observed, even for bipolar pulses, and we did not find any reasonable explanation for that. Critical value of ITV, at which permeabilization occurs, was calculated from the polar angle of permeabilization measured immediately after the pulse and was found to be approximately 450 mV, in agreement with reported critical thresholds. Permeabilization results obtained on cell clusters showed that cells in clusters, atpulse parameters used in these experiments, behaved as electrically insulated and were permeabilized individually. This is in contradiction to what we observed during measurements of ITV (i. E. With longer, low voltage pulses), where cells in clusters behaved as electrically connected, and was assumed to be the result of opening and closing of gap junctions at different pulse parameters. Measurements of kinetics of membrane transport showed that electropermeabilization with progressively increasing pulse amplitudes or pulse durations results in increased dye transport into cells. A sharp increase was observed miliseconds after the onset of a pulse, followed by a moderate additional fluorescence increase. Results measured on a time interval of 400 µs revealed that the transport across the permeabilized membrane can be detected within 100 µs after the onset of the pulse. Besides, different dynamics of fluorescence increase was observed during and immediately after the pulse. Experiments carried out on dense cell suspensions showed that with increasing cell density (from 10×106 cells/ml to 400×106 cells/ml) the fraction of permeabilized cells decreased by approximately 50%. We attributed this to the changes in the local electric field, which lead to a decrease in the amplitude of ITV. The uptake of Propidium Iodide also decreased with cell density, but by a larger amount than expected from permeabilization results. We supposed that the additional decrease in fluorescence was mainly due to cell swelling after permeabilization, which reduced extracellular dye availability to the permeabilized membrane and hindered the dye diffusion into the cells. Resealing of cells appeared to be slower in dense suspensions, which can also be attributed to cell swelling resulting from electropermeabilization
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YOSHITO, DANIELE. "Cultivo e irradiação de fibroblastos humanos em meio enriquecido com lisado de plaquetas para obtenção de camada de sustentação em cultura de células da epiderme." reponame:Repositório Institucional do IPEN, 2011. http://repositorio.ipen.br:8080/xmlui/handle/123456789/9959.
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Dissertacao (Mestrado)
IPEN/D
Instituto de Pesquisas Energeticas e Nucleares - IPEN-CNEN/SP
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Freyer, Nora [Verfasser]. "Optimizing culture conditions for hepatic differentiation of human induced pluripotent stem cells : from 3D culture systems to co-cultures / Nora Freyer." Berlin : Medizinische Fakultät Charité - Universitätsmedizin Berlin, 2018. http://d-nb.info/1160514968/34.
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Patel, Leena Suman. "Functional changes in pyramidal neurons surviving an excitotoxic challenge in mouse organotypic slice cultures /." Thesis, Connect to this title online; UW restricted, 2005. http://hdl.handle.net/1773/10658.
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KONDO, TATSUHEI, HIDEO KAMEI, FUMIHIRO KOBAYASHI, TAKAO YUKAWA, KEISUKE TERABE, YASUHISA HASEGAWA, YOHICHI ITOH, and TAKASHI KOJIMA. "Colony-Stimulating Activity in Cultures of Human Spleen and Bone Marrow Cells." Nagoya University School of Medicine, 1986. http://hdl.handle.net/2237/17486.
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Liu, Ning. "Expansion and Neural Differentiation of Embryonic Stem Cells in Three-Dimensional Cultures." The Ohio State University, 2010. http://rave.ohiolink.edu/etdc/view?acc_num=osu1262281522.
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McGoldrick, Trevor A. "C-S lyase-mediated toxicity in primary cultures of proximal tubular cells." Thesis, University of Aberdeen, 2000. http://digitool.abdn.ac.uk/R?func=search-advanced-go&find_code1=WSN&request1=AAIU602010.
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Halogenated alkenes are a group of commercially important chemicals. For example tetrafluoroethylene is the monomer used for the production of poly- tetrafluoroethylene, hexachloro-1:3-butadiene is a by-product from the manufacture of chlorinated solvents and perchloroethylene is widely used as a dry cleaning agent. Due to possible exposure to haloalkenes and the nephrotoxicity observed in animal studies, concern has been expressed for the potential of these compounds to cause toxicity to man. Animal studies have shown that these compounds undergo inter-organ metabolism and are bioactivated by enzymes of glutathione processing. The metabolites are delivered to the kidney where they cause proximal tubular cell necrosis. This site-specific toxicity is due to accumulation of the metabolites via specific transport mechanisms and bioactivation via the enzyme C-S lyase present in high amounts in the proximal tubules. The aim of this research was to investigate the mechanisms of toxicity of haloalkene S'-conjugates in vitro using cultures of rat and human proximal tubular cells. This study demonstrates that human proximal tubular cells are sensitive to haloalkene. -conjugate toxicity, particularly DC VC. Human exposuredata has shown that workers exposed to trichloroethylene (Bimer et al, 1993) and perchloroethylene (Mutti et al, 1992) excrete nephrotoxic metabolites and markers of renal damage respectively. In the light of these findings and the toxicity of DCVC in HPT cells, exposure to halogenated alkenes should be controlled and those exposed monitored.
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Cullen, Daniel Kacy. "Traumatically-induced degeneration and reactive astrogliosis in three-dimensional neural co-cultures." Available online, Georgia Institute of Technology, 2005, 2005. http://etd.gatech.edu/theses/available/etd-11282005-210117/.
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Thesis (Ph. D.)--Biomedical Engineering, Georgia Institute of Technology, 2006.Robert McKeon, Committee Member ; Robert Lee, Committee Member ; Robert Guldberg, Committee Member ; Ravi Bellamkonda, Committee Member ; Michelle LaPlaca, Committee Chair. Vita.
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Park, Deric M., Jinkyu Jung, Jimmy Masjkur, Stylianos Makrogkikas, Doreen Ebermann, Sarama Saha, Roberta Rogliano, et al. "Hes3 regulates cell number in cultures from glioblastoma multiforme with stem cell characteristics." Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2013. http://nbn-resolving.de/urn:nbn:de:bsz:14-qucosa-127014.
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Tumors exhibit complex organization and contain a variety of cell populations. The realization that the regenerative properties of a tumor may be largely confined to a cell subpopulation (cancer stem cell) is driving a new era of anti-cancer research. Cancer stem cells from Glioblastoma Multiforme tumors express markers that are also expressed in non-cancerous neural stem cells, including nestin and Sox2. We previously showed that the transcription factor Hes3 is a marker of neural stem cells, and that its expression is inhibited by JAK activity. Here we show that Hes3 is also expressed in cultures from glioblastoma multiforme which express neural stem cell markers, can differentiate into neurons and glia, and can recapitulate the tumor of origin when transplanted into immunocompromised mice. Similar to observations in neural stem cells, JAK inhibits Hes3 expression. Hes3 RNA interference reduces the number of cultured glioblastoma cells suggesting a novel therapeutic strategy.
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McMenamin, C. C. "Generation and charecterisation of mucosal mast cells in normal rat bone marrow cultures." Thesis, University of Glasgow, 1986. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.381463.
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Khetani, Salman R. "Micropatterned co-cultures of hepatocytes and nonparenchymal cells mechanisms of differentiation, dynamics and applications /." Connect to a 24 p. preview or request complete full text in PDF format. Access restricted to UC campuses, 2006. http://wwwlib.umi.com/cr/ucsd/fullcit?p3204579.
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Thesis (Ph. D.)--University of California, San Diego, 2006.Title from first page of PDF file (viewed Apr. 4, 2006). Available via ProQuest Digital Dissertations. Vita. Includes bibliographical references (p. 242-264).
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PERONI, CIBELE N. "Altos niveis de expressao de tireotrofina humana em celulas de ovario de hamster chines mediante a utilizacao de vetores dicistronicos." reponame:Repositório Institucional do IPEN, 1999. http://repositorio.ipen.br:8080/xmlui/handle/123456789/10758.
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Tese (Doutoramento)
IPEN/T
Instituto de Pesquisas Energeticas e Nucleares - IPEN/CNEN-SP
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Knowles, Christopher. "Generation of mammalian cell culture systems for the rapid and efficient production of recombinant proteins." Thesis, University of Aberdeen, 2016. http://digitool.abdn.ac.uk:80/webclient/DeliveryManager?pid=231905.
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The overarching objective of this thesis was the development of an improved expression cell line for recombinant proteins, in which a transgene of interest can be inserted into a highly active gene locus using recombinase mediated cassette exchange. Random integration of transgenic DNA is a common route to achieve transgene expression. However, randomly integrated transgenes are susceptible to gene silencing over time, and do not show stable expression for extended periods in culture. Furthermore, every new cell clone generated requires regulatory approval. The improvement of expression strategies may significantly reduce the time required to bring novel recombinant protein products to the market. In order to identify a suitable expression locus for the integration of transgene expression cassettes, total protein samples were derived from the production cell lines HEK293 and CHO. Highly expressed proteins were isolated via 2D PAGE, and identified via peptide mass fingerprinting. Their promoter regions were then validated to express a recombinant transgene in HEK293 cells. The long term stability of these promoter regions was also assessed. Direct gene targeting of the highly active gene loci may or may not be possible in typical producer cell lines. Targeting of a murine homologue to these highly expressed CHO/HEK293 loci may be more efficient in a murine stem cell line. The transfer of a modified allele from HM1 murine embryonic stem cells, into a somatic cell line (HC11) was demonstrated in this thesis. These validated methods were then explored for the generation of viable HM1-HEK293 and HM1-CHO fusion hybrids. For these experiments, a fluorescence based fusion assay was generated, validated and used for in-situ monitoring of the cell fusion process. The random integration of transgenic DNA into mammalian genomes typically results in a highly unpredictable integration architecture. RMCE at such loci would be inefficient. However, a highly efficient RMCE reaction at (rare) single copy transgene integrations, may be possible under the correct conditions. RMCE at randomly integrated loci could therefore be more beneficial (for transgene expression) than random integration alone. This thesis explores this concept with the use of a randomly integrated RMCE platform, and subsequent selection of cell lines post RMCE attempts at these loci CRISPR/Cas9 technology was also applied to a highly expressed locus in HEK293 cells. A framework for successful direction of double strand breaks to a defined locus is demonstrated in this work. The methods used to achieve this can therefore be built upon for the homologous recombination of a transgenic cassette, into a highly expressed locus in HEK293 cells. Monoclonal antibodies have dominated the biologics market for over two decades, and mammalian expression systems are well suited to their production. The work in this thesis attempts to raise and verify antibody molecules against a potential tumour marker using hybridoma and phage display technologies.
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Postigo, Milagros. "Molecular and antigenic characterisation of Ehrlichia ruminantium in Amblyomma variegatum ticks and in vitro cultures." Thesis, University of Edinburgh, 2007. http://hdl.handle.net/1842/6576.
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The rickettsial pathogen Ehrlichia ruminantium, transmitted by ticks of the genus Amblyomma, causes heartwater, an economically important, often fatal disease of domestic and wild ruminants in sub-Saharan Africa and in the Caribbean. The studies described in this thesis have contributed to understanding several aspects of heartwater. First, a real-time PCR method was developed in order to study the kinetics of infection with E. ruminantium in the mammalian host. The assay was validated for specificity and sensitivity and was used to estimate numbers of the organisms in the blood of infected sheep. However, organisms were only detected during the clinical phase of infection, indicating that the way in which it was applied did not provide sufficient sensitivity to follow the early stages of infection. This PCR assay was then used, together with transcription and proteomic analyses, to investigate differential gene expression of E. ruminantium in the arthropod and mammalian hosts, in order to identify genes that may allow the organisms to successfully adapt to different environments. These studies used in vitro tick and mammalian cell culture systems, as well as tissues from infected A. variegatum ticks, and initially focused on the map1 multigene family. Although transcripts for most of the map1 paralogs were detected in organisms grown in vitro, in both mammalian and tick cells, only transcripts from map1 and map1-1 were detected in infected ticks. Moreover, map1-1 transcripts were more abundant in midguts than in salivary glands whereas map1 transcripts were most abundant in salivary glands and were expressed at higher levels following several days of tick feeding on a mammalian host. Because of the quantities of material required, proteomic analysis was only possible using in vitro-cultured organisms. Comparison of proteins encoded by the map1 cluster in E. ruminantium grown in tick or bovine endothelial cell cultures, using 2D gels and MALDI-TOF analysis, revealed that different proteins predominated in the corresponding spots in 2D gels from the different cultures; products of the map1-1 gene were abundant in tick cells, while products of map1 were abundant in endothelial cells. The detection of higher levels of map1 transcripts in salivary glands than in midguts of infected ticks, together with the presence of abundant MAP1 protein in organisms grown in mammalian but not in tick cell lines, suggest that expression of this protein may be associated with infectivity for mammalian cells. In contrast, map1-1 transcripts were abundant both in midguts of infected ticks and in tick cell lines, and the protein was expressed at high levels in infected tick cell cultures. Since both of these stages have low infectivity for sheep, these results suggest that the MAP1-1 protein may play an important role within the vector, possibly associated with colonisation and replication of E. ruminantium in the tick midgut. Collectively these findings suggest that this multigene family is involved in functions of biological relevance in different stages of the life cycle of E. ruminantium. Lastly the suppression subtractive hybridisation (SSH) technique was applied to RNA extracted from E. ruminantium-infected endothelial and tick cell cultures in an attempt to sample a large portion of the E. ruminantium genome for differentially expressed genes; although not resulting in identification of any differentially transcribed genes in the present study, this method was shown to work in principle.
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Mozzetti, Valeria. "Novel technological approaches to enhance stress tolerance of Bifidobacterium longum NCC2705 cells using continuous cultures /." [S.l.] : [s.n.], 2009. http://e-collection.ethbib.ethz.ch/show?type=diss&nr=18419.
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Cēbere, Aleta. "Effects of ethanol on NMDA receptor-mediated functions in primary cultures of cerebellar granule cells /." Stockholm, 2001. http://diss.kib.ki.se/2001/91-628-5052-0.
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Lipscomb, Matthew L. "Perfusion cultures of recombinant CHO cells: Effects on specific productivity, production stability, and protein glycosylation." Diss., Connect to online resource, 2005. http://wwwlib.umi.com/dissertations/fullcit/3165829.
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Osborn, Kay Elizabeth. "Radiation effects on human keratinocyte cultures in relation to growth factors, differentiation and stem cells." Thesis, Queen Mary, University of London, 2006. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.429519.
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Laryea, D., A. Isaksson, Colin W. Wright, R. Larsson, and P. Nygren. "Characterization of the cytotoxic activity of the indoloquinoline alkaloid cryptolepine in human tumour cell lines and primary cultures of tumour cells from patients." Springer, 2009. http://hdl.handle.net/10454/4533.
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noThe plant derived indoloquinoline alkaloid cryptolepine was investigated for its cytotoxic properties in 12 human tumour cell lines and in primary cultures of tumour cells from patients. The fluorometric microculture cytotoxicity assay was used to assess cytotoxicity and DNA mocro-array analysis to evaluate gene expression. Cryptolepine mean IC50 in the cell line panel was 0.9 microM compared with 1.0 and 2.8 microM in haemaotological and solid tumour malignancies respectively. Among patient solid tumour samples, those from breast cancer were the most sensitive and essentially as sensitive as haematological malignancies, respectively. Among patient solid tumour samples, those from breast cancer were the most sensitive and essentially as senstive as haematological malignancies. Cryptolepine activity showed highest correlations to topoisomerase II and microtubule targetting drugs. In the cell lines cryptolepine activity was essentially unaffected by established mechanisms of drug resistance. A number of genes were identified as associated with cryptolepine activity. In conclusion, cryptolepine shows interesting in vitro cytotoxic properties and its further evaluation as an anticancer drug seems warranted.
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Tomasi, Raphaël. "Multiscale cytometry of 3D cell cultures in microfluidic hydrogel arrays." Thesis, Université Paris-Saclay (ComUE), 2016. http://www.theses.fr/2016SACLX114/document.
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Les conditions du corps humain ne sont pas reproduites fidèlement par la culture cellulaire traditionnelle en 2D. Dans cette thèse, des cultures cellulaires 3D sont réalisées dans une plateforme microfluidique hautement intégrée. Des cellules mammifères adhérentes sont encapsulées dans des gouttes immobilisées dans un tableau de pièges capillaires à haute densité. Dans chaque goutte, les cellules se réorganisent pour former un unique microtissu 3D et fonctionnel appelé sphéroïde. L'utilisation d'un hydrogel permet d'alonger le temps de culture et de perfuser le tableau avec des solutions aqueuses, par exemple pour de l'immuno-cyto-chimie. Un unique sphéroïde, viable, peut aussi être extrait de cette puce microfluidique. Des données quantitatives sont extraites à haut débit au niveau de la population, du sphéroïde (dizaines de miliers de sphéroïdes) et au niveau cellulaire emph{in situ} (centaines de miliers de cellules) grâce à de l'imagerie de fluorescence et au dévelopement d'un code d'analyse d'image. Une première preuve de concept a été obtenue en démontrant la viabilité, la prolifération et la fonctionalité de sphéroïdes d'hépatocytes et en les corrélant à des paramètres morphologiques. Ensuite, des aggrégats de cellules souches mésenchymales ont été produits et les hétérogénéités spatiales dans l'expression de protéines impliquées dans leurs propriétés thérapeutiques ont été étudiées. Enfin, cette technologie a été encore dévelopée pour permettre d'appliquer des conditions biochimiques différentes dans chaque goutte. La production et la culture de sphéroïdes dans cette plateforme microfluidique peut mener à des dévelopements importants dans beaucoup de domaines tels que l'analyse de la toxicité des médicaments, le criblage de médicaments à haut débit, le traitement personnalisé du cancer, l'ingénierie tissulaire ou la modélisation de maladiesConventional 2D cell culture fails to reproduce emph{in vivo} conditions. In this PhD thesis, 3D cell culture is implemented into a highly integrated microfluidic platform. Adherent mammalian cells are encapsulated in droplets immobilized on a high density array of capillary traps called anchors. In each droplet, the cells reorganize into a single functional 3D microtissue called spheroid. The use of an hydrogel allows to extend the culturing time in microdroplets and to perfuse the array with aqueous solutions, for instance for immuno-cyto-chemistry. A single and viable spheroid can also be selectively retrieved from the microfluidic chip. High throughput and quantitative data is extracted at the population, spheroid (tens of thousands of spheroids) and cellular level emph{in situ} (hundreds of thousands of cells) thanks to fluorescent imaging and a custom image analysis software. As a first proof of concept, the viability, proliferation and functionality of hp sh s were demonstrated and correlated with morphological parameters. Drug toxicity experiments were also performed on this liver model. Then, human mesenchymal stem cell aggregates were produced and the spatial heterogeneities of the expression of proteins involved in their therapeutic properties were investigated. Finally, this technology was further developed to enable applying different biochemical conditions in each droplet. The production and culture of spheroids in this microfluidic platform could lead to major advances in many fields such as drug toxicity, high throughput drug screening, personalized cancer treatment, tissue engineering or disease modeling
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Samuels, Peter L. "Development and survival of a postsynaptic specialization in cultures of embryonic xenopus nerve and muscle cells." Thesis, McGill University, 1989. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=59248.
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This study has focussed on the formation and survival of acetylcholine receptor (AChR) clusters at neuromuscular synapses formed in culture between myotomal muscle cells and spinal cord neurons derived from embryos of Xenopus laevis. AChRs were labelled with tetramethylrhodamine-conjugated $ alpha$-bungarotoxin so that the neurite associated receptor patches (NARPs) on the muscle cells could be viewed by fluorescence microscopy. Reduced fluorescence excitation was used in combination with a low light level TV camera and a computer based image calculator to make daily observations on all NARPs. Observations suggest the following conclusions. Neurons retain the capacity to trigger NARP formation as long as they continue to grow. Changes in NARP shape as well as decreases (and increases) in NARP size can occur even in the absence of competitive interactions between neurons and these changes are locally regulated along the contact. The capacity of neurons to maintain NARPs in more proximal portions of their neuritic arbor persists even as growing distal portions continue to induce the formation of new NARPs.
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胡, 興柏. "ACTIVATION OF FIBROBLAST-DERIVED MATRIX METALLOPROTEINASE-2 BY COLON CANCER CELLS IN NON-CONTACT CO-CULTURES." Doctoral thesis, Kyoto University, 2000. http://hdl.handle.net/2433/151419.
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京都大学0048
新制・課程博士
博士(医学)
甲第8550号
医博第2272号
新制||医||747(附属図書館)
UT51-2000-M14
京都大学大学院医学研究科内科系専攻
(主査)教授 鍋島 陽一, 教授 月田 承一郎, 教授 千葉 勉
学位規則第4条第1項該当
Doctor of Medical Science
Kyoto University
DAM
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Rotti, Pavana Gururaj. "3D differentiation enhances the efficiency of differentiation of human induced pluripotent stem cells to insulin producing cells." Thesis, University of Iowa, 2014. https://ir.uiowa.edu/etd/2266.
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Type 1 Diabetes (T1D) is an autoimmune disorder in which the pancreatic β-cells are destroyed by the body's immune system. The reduced number of β-cells leads to inadequate insulin secretion and high glucose levels in the body. The requirement of insulin injection throughout life and lack of donors for islet transplantations has prompted a search for more accessible and available sources of insulin producing cells that can be transplanted in T1D patients. To that end, the discovery of induced pluripotent stem (iPS) cells has provided a potential source of precursors for cell therapy for T1D. iPS cells are reprogrammed somatic cells which can be transplanted back into the patient from whom the somatic cells were initially derived, thus potentially avoiding immune rejection when transplanted. As a potential therapy for T1D, we aim to derive insulin producing cells (IPCs) from human iPS cells. In contrast to the conventional two dimensional (2D) cell culture systems used in many iPS derived IPC studies, the inner cell mass (ICM) from which various organs differentiate during embryogenesis is a cluster of cells that enables signaling crosstalk between cells of different types. Three dimensional (3D) cell culture systems allows cells to form cell clusters that promote cell - cell signaling. Hence, we hypothesized that 3D cell culture systems will yield better efficiency of differentiation to functional IPCs in vitro than 2D cultures.
Initially, the synthetic polymers sodium alginate and matrigel were analyzed for their ability to enable cell clustering to establish 3D cell culture systems. The 3D cell environment established using matrigel was used for the differentiation of human iPS cells to Insulin Producing Cells (IPC). The cells were first converted to endodermal cells. A mixture of growth factors then induced the differentiation of endodermal cells to pancreatic cells. The pancreatic cells were converted to IPCs that resemble pancreatic β-cells. Our 3D differentiated IPCs strongly expressed pancreatic endocrine transcription factors and pancreatic hormones. The IPCs also produced insulin when exposed to a high glucose environment. But the number of IPCs obtained at the end of the differentiation was low.
Hence, our results demonstrate that 3D differentiation generates functional IPCs in vitro unlike 2D differentiation. In the future we aim to improve the percentage of IPCs that we generate from the 3D differentiation. Our expectation is that these cells will be able to cure hyperglycemia in diabetic mice more rapidly compared to the 2D differentiated cells owing to their proven insulin production in the presence of a high glucose environment in vitro.
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Scott, R. "Growth of mesenchymal stromal cells in automated microwell cultures : influence of the engineering environment on cell growth kinetics and non-directed differentiation." Thesis, University College London (University of London), 2009. http://discovery.ucl.ac.uk/18778/.
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Human Mesenchymal Stromal Cells (hMSC) have the potential to differentiate into lineages of mesoderm origin, such as osteogenic, chondrogenic, and adipogenic lineages, presenting a promising potential for autologous regenerative medicine applications. However, one of the major challenges associated with delivering hMSC to the clinic is the propagation of undifferentiated cells in vitro in order to reach the quantity required for therapeutic applications. This thesis investigates the effect of environmental parameters that impact cell growth characteristics of hMSC in microwell plates, for the design of an automated cell expansion process in a robotic platform. As a result of this work, the main parameters that can be used to control the growth rate and differentiation potential of hMSC at all stages of the cell expansion process have been identified. A mathematical model has also been developed to forward predict the cell growth characteristic of hMSC for an individual donor, allowing for a patient specific bioprocess design that will ensure enough cells can be supplied back to the patient in a timely manner while assuring the quality of the final product. Process parameters for the cell expansion process of hMSC in automated microwells have been characterised. Optimum values for inoculation cell density and medium exchange strategies have been proven to reduce the overall time of the cell expansion process for hMSC in an undifferentiated state for their use in regenerative medicine therapies. These parameters were implemented in the cell expansion in an automated platform for the duration of one passage, proving the potential of such technology for the delivery of hMSC to the clinic.
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Pierce, Paul Randall 1951. "Control of fibroblast contamination in primary rat skeletal muscle cell cultures: Effects of an epidermal growth factor linked cytotoxin." Thesis, The University of Arizona, 1988. http://hdl.handle.net/10150/276812.
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The in vitro study of muscle cell growth is hampered by the presence of non-muscle cells, particularly fibroblasts. The heterobifunctional cross-linking agent, N-succinimidyl 3-(2-pyridyldithio) propionate (SPDP) has been used to create a novel "toxic growth factor" to address the problem. Epidermal growth factor (EGF), which stimulates fibroblast but not satellite cell proliferation, was conjugated via SPDP to a potent ribosome inhibitor, pokeweed antiviral protein (PAP). By preferentially binding to fibroblasts, it was hoped that EGF-PAP could cytotoxically eliminate fibroblasts from primary cultures of rat skeletal muscle satellite cells. While EGF-PAP did prove to be a fibroblast cytotoxin, it could not completely eliminate them from cell cultures. Low dose-time exposures improved the ratio of multinucleated cells to mononucleated cells (percent fusion) by up to 66% over controls, but increased concentrations, or durations of EGF-PAP treatment, proved detrimental to satellite cell growth and/or differentiation.
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ROSAURO, CRISTIANE W. "Altos niveis de expressao de hormonio de crescimento de camundongo em queratinocitos humanos visando a obtencao de um modelo animal de terapia genica." reponame:Repositório Institucional do IPEN, 2003. http://repositorio.ipen.br:8080/xmlui/handle/123456789/11151.
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Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
Dissertacao (Mestrado)
IPEN/D
Instituto de Pesquisas Energeticas e Nucleares - IPEN/CNEN-SP
FAPESP:00/08457-0
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MENDONCA, FERNANDA de. "Expressao, purificacao e caracterizacao de tireotrofina recombinante humana (rec-hTSH) em celulas de ovario de hamster chines (CHO)." reponame:Repositório Institucional do IPEN, 2003. http://repositorio.ipen.br:8080/xmlui/handle/123456789/11150.
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Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
Dissertacao [Mestrado]
IPEN/D
Instituto de Pesquisas Energeticas e Nucleares - IPEN/CNEN-SP
FAPESP:00/09008-4
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DAMIANI, RENATA. "Expressao estavel de tireotrofina humana (r-hTSH) em celulas de mamifero (CHO) que expressam 'alfa'2,6-sialiltransferase." reponame:Repositório Institucional do IPEN, 2009. http://repositorio.ipen.br:8080/xmlui/handle/123456789/9436.
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Dissertacao (Mestrado)
IPEN/D
Instituto de Pesquisas Energeticas e Nucleares - IPEN-CNEN/SP
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Norcio, Lawrence P. "Effects of microcarrier concentration, agitation rate, and serum concentration on the specific growth rate of mouse L cells in batch cultures." Ohio : Ohio University, 1995. http://www.ohiolink.edu/etd/view.cgi?ohiou1179949129.
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Shah-Derler, Brigitte Anita. "Flat-sedimented and reaggregate cultures of chicken embryonic neuronal retinal cells : developmental patterns and effects of taurine /." [S.l.] : [s.n.], 1994. http://e-collection.ethbib.ethz.ch/show?type=diss&nr=10523.
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Lightly, Eric R. T. "Adrenergic and cholinergic stimulation of cortisol production in primary cultures of bovine adrenocortical zona fasciculata/reticularis cells." Thesis, University of Edinburgh, 1991. http://hdl.handle.net/1842/28430.
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The classical secretagogues, adrenocorticotrophic hormone (ACTH) and angiotensin II (AII), are known to stimulate cortisol production from bovine adrenocortical zona fasciculata/reticulariscells. This is seen in both freshly isolated, enzymically dispersed, cells and in primary cultures prepared from these cells. Previous published work has also shown that adrenergic agonists stimulate steroidogenesis in primary cultures of bovine adrenocortical cells, but not in freshly isolated cells. In addition, cholinergic agonists have been shown to stimulate steroidogenesis in both freshly isolated cells, and in cells in primary culture, and that this process is linked to increased turnover of inositol containing phospholipids. Initial characterisation of primary cultures of bovine adrenocortical zona fasciculata/reticularis cells, prepared by a collagenase digestion procedure, showed that both ACTH and AII stimulated cortisol production from both freshly isolated and cultured cells. Adrenergic agonists stimulated cortisol production in cultured cells, but failed to stimulate cortisol production from freshly isolated cells. Cholinergic agonists stimulated cortisol production from both freshly isolated and cultured cells. Using specific alpha and beta adrenergic agonists and antagonists it was shown that adrenergic stimulation of cortisol production from cultured cells was mediated by beta-adrenergic receptors. Schild analysis, using specific beta-adrenergic antagonists, showed that beta1- adrenoceptors were responsible for this. Adrenergic agonists were also shown to increase intracellular cyclic AMP levels in cultured cells which was directly responsible for stimulating steroidogenesis. These agonists had no effect on turnover of cellular phosphoinositides. Using specific cholinergic agonists and antagonists, it was shown that M3-cholinergic receptors were responsible for mediating stimulation of cortisol production from cultured cells produced by cholinergic agonists. It was also shown that cholinergic agonists activated a phospholipase-C-dependent breakdown of phosphoinositides, and that this was responsible for initiating steroidogenesis. Cholinergic agonists had no effect on cellular cyclic AMP levels.
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Hayes, Marianne Kay. "Bovine testicular cells in vitro: establishment of primary cultures and investigations of secretory functions : a thesis presented for the degree of Doctor of Philosophy in the University of Adelaide." Title page, contents and summary only, 1986. http://web4.library.adelaide.edu.au/theses/09PH/09phh4178.pdf.
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Includes bibliographical references (leaves 98-128). Investigates protein secretion by bovine Sertoli cells in culture. Cultures were obtained from bulls at all stages of post natal development and from sexually mature animals.
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Chan, Po-tong Timothy. "Adrenergic control and its mechanism of stimulation of electrogenic anion secretion in primary cultures of rat epididymal eipthelial cells /." [Hong Kong : University of Hong Kong], 1990. http://sunzi.lib.hku.hk/hkuto/record.jsp?B13183308.
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Calles, Karin. "Regulation of productivity in Trichoplusia ni and Spodoptera frugiperda Sf9 serum-free cultures." Licentiate thesis, Stockholm, 2005. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-243.
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Estévez, Priego Estefanía. "Dynamics and Effective Connectivity in Bi- and Three–dimensional Neuronal Cultures: from Self–organization to Engineering." Doctoral thesis, Universitat de Barcelona, 2019. http://hdl.handle.net/10803/668218.
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This thesis was part of the European consortium MESOBRAIN, a team of 5 organizations that joined efforts in nanofabrication, cell culturing, imaging and data analysis to build tailored human 3D networks. The thesis timing was limited to 3 years, and several of the resources needed for its development were built from scratch.
The main objective of this Ph.D. thesis was to explore complex characteristics of cortical neuronal cultures in terms of effective connectivity and exhaustive network analyses. This objective comprised four research lines: (i) The evaluation of neuronal network resilience and emerging plasticity mechanisms, (ii) the characterization of functional development to underline crucial timepoints in healthy neuronal networks, (iii) the study of 3D network interactions of neurons embedded inside an ECM--like environment, and (iv) the design, construction and viability inspection of neurons seeded on tiny 3D nanoprinted solid scaffold structures as a first step towards recreating cortical columns in vitro.
For these multiple lines, we used either primary rat cultures (i,iii,iv) or human--derived neurons (ii). The former group corresponds to cultures with long established protocols that have been thoroughly studied in the field. The latter group corresponds to human neurons derived from iPSCs, a relatively novel model with promising and thrilling applications in regenerative medicine. Despite the increasing use of stem cells in neuroscience, complex systems and medicine, they still lack a thorough exploration in terms of neuronal and circuit formation as well as the properties of the emergent activity patterns.
With either primary or stem cells, we explored the formation of neuronal circuits in 2D and 3D, characterized the effective connectivity and rendered a number of network traits. This Thesis combines experiments of highly difficult implementation with detailed data analysis. It was necessary to develop brand new protocols for culturing 3D neuronal networks and for human-derived neurons, the use of different microscopy setups the programming of object detection and tracking software and advance the analysis toolbox of calcium fluorescence data.
First, resilience experiments on primary clustered neuronal cultures consisted on progressive perturbations through chemical receptor antagonists. This study represents an inspiring numerical--experimental model to comprehend the impact of plasticity mechanisms in the spontaneous activity of neuronal circuits. The results showed that, upon progressive connectivity blockade through chemical receptors' antagonists, only--excitatory neuronal networks displayed a surprising hyper--efficiency (HE) state for early--onset doses. As plasticity mechanisms influence the response of effective connectivity in the presence of perturbations, these compensatory mechanisms, usually disregarded, must be included in biological modeling as accurately as possible. Otherwise, episodes of functional rewiring and synaptic strengthening could mask important phenomena during experiments that alter channel communication. A simple algorithm that hypothesized an effective synaptic scaling was able to capture the hyper--efficiency state seen in experimental data, while percolation models wrongly predicted a progressive decay.
The second research line was a sum of engineering efforts within the MESOBRAIN consortium, the European adventure to build 3D neuronal cultures embedded in hydrogels and with the presence of scaffolds. After several months of biomaterials testing, the candidate D--Clear resulted suitable for the construction of scaffolds, both with primary rat cells and hiPSCs, due to its good optical properties, manageability and biocompatibility. To our knowledge, D--Clear was never used before outside the orthodontic field and could provide a new catalogue of interesting designs for support and guidance of neuronal assemblies. Using this material, we developed a series of designs to offer support and guidance to cortical neurons in a 3D platform.
The third research line focused on the study of neuronal development and cell-to-cell interactions in a semi-synthetic hydrogel that resembles the extracellular matrix of the brain. These hydrogel cultures keep the advantages of in vitro models while achieving an effective connectivity and architecture closer to in vivo.
Finally, the fourth line of research applied cortical neurons from human-derived pluripotent stem cells to study key developmental stages and characterize the healthy maturation of these cells in vitro. As this technology has tremendous potential for regenerative medicine and to model neuronal diseases, it is urgent to consolidate the capacity of these human neuronal networks to reproduce efficient activity patterns of healthy patients, and explore the differences against the results obtained with animal models.La presente tesis doctoral se enmarca en el contexto de la Física de la Materia Condensada, la Biofísica y la Neurociencia. Principalmente, se centra en el estudio de la conectividad funcional en cultivos neuronales bidimensionales (2D) y tridimensionales (3D). El trabajo se ha desarrollado en el Laboratorio del director de tesis Dr. Jordi Soriano, en la Facultad de Física de la Universitat de Barcelona, junto con el codirector Dr. Daniel Tornero, en el Hospital Clínic de Barcelona. Esta tesis forma parte del proyecto europeo MESO-BRAIN, del programa Future and Emergent Technologies (FET) de la Comisión Europea, Horizon2020. El trabajo de investigación combina experimentos con cultivos neuronales (de rata embrionaria o células humanas pluripotentes) y un análisis detallado en el contexto de teoría de redes y sistemas complejos. Los principales núcleos del trabajo realizado son los siguientes: (i) Actividad funcional en cultivos de redes neuronales y los mecanismos homeostáticos que emergen en presencia de perturbaciones; (ii) el desarrollo de herramientas de neuroingeniería para preparar cultivos ad hoc con conectividad dirigida mediante scaffolds; (iii) el análisis exhaustivo de los procesos de formación y madurez de redes funcionales humanas obtenidas de células madre pluripotentes inducidas, una nueva tecnología que promete revolucionar el campo de la medicina regenerativa; y (iv) la caracterización de cultivos neuronales 3D en estructuras que imitan la matriz extracelular natural de su entorno. Entre las diversas técnicas para la realización de cultivos tridimensionales, destacan los hidrogeles semi-sintéticos, constituidos en base a polímeros altamente hidratados con alta biocompatibilidad y cuyas propiedades mecánicas pueden ser manipuladas para obtener la estructura óptima según el tipo de tejido. En conjunto, los resultados de la presente tesis muestran la gran versatilidad de los cultivos neuronales y aportan avances relevantes en el estudio de plasticidad en redes neuronales, madurez y desarrollo tanto en 2D como en 3D, con sus correspondientes diferencias, incluyendo el uso de neuronas humanas derivadas de células madre inducidas. En el futuro, estos estudios nos permitirán incrementar nuestro conocimiento sobre el funcionamiento global del cerebro y avanzar en la investigación de diferentes enfermedades neurodegenerativas.
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陳浦棠 and Po-tong Timothy Chan. "Adrenergic control and its mechanism of stimulation of electrogenic anion secretion in primary cultures of rat epididymal eipthelialcells." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 1990. http://hub.hku.hk/bib/B31210156.
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SOARES, CARLOS R. J. "Sintese de prolactina humana em celulas de ovario de hamster chines (CHO)." reponame:Repositório Institucional do IPEN, 2000. http://repositorio.ipen.br:8080/xmlui/handle/123456789/10770.
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Tese (Doutoramento)
IPEN/T
Instituto de Pesquisas Energeticas e Nucleares - IPEN/CNEN-SP
46
Andersson, Maria. "Chemically Induced DNA Damage in Extended-term Cultures of Human Lymphocytes." Doctoral thesis, Uppsala : Acta Universitatis Upsaliensis : Universitetsbiblioteket [distributör], 2006. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-7179.
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Nassra, Merian. "Effets anti-inflammatoires des stilbènes sur des cultures cellulaires de microglies et mécanismes d'action." Thesis, Bordeaux 2, 2012. http://www.theses.fr/2012BOR22018/document.
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Les processus neuro-inflammatoires sont observés dans de nombreux troubles neurodégénératifs. Les cellules microgliales étant les principales cellules immunitaires du système nerveux central, plusieurs recherches ont été menées afin de déterminer des molécules possédant des propriétés anti-inflammatoires au niveau de ces cellules. Une famille de polyphénols, les stilbènes (des dérivés du resvératrol), présentent des activités anti-inflammatoires au niveau périphérique et central. Dans notre étude, nous avons premièrement évalué les propriétés anti-inflammatoires de 25 stilbènes en déterminant leur capacité à inhiber la libération de NO dans un modèle de cellules microgliales BV-2 activées par le LPS. Dix stilbènes inhibent significativement cette production avec des IC50 comprises entre 3,9 ± 0,7 et 23,4 ±1,0 µM. Parmi ces composés, 1 monomère (moracine M) et 2 tétramères du resvératrol (vitisines A et B) diminuent la synthèse d’iNOS, enzyme responsable à la libération de NO, au niveau transcriptionnel et traductionnel. Nous avons ensuite mis en évidence que la moracine M inhibe la phosphorylation d’ERK1/2 et JNK (deux MAPK) et d’Akt (la voie PI3K/Akt) dans des cellules BV-2 activées. Ces enzymes étant impliquées dans la signalisation de la réponse inflammatoire, elles induisent la production de plusieurs médiateurs inflammatoires. La moracine M inhibe significativement la production de certains d'entre eux (NO, TNF-α, IL-1β et PGE2). Ce stilbène attenue également la synthèse de la protéine de mPGEs-1 (enzyme impliquée à la production de PGE2). Ainsi, la moracine M pourrait être un candidat potentiel pour prévenir l’inflammation impliquée dans les maladies neurodégénérativesChronic neuro-inflammatory processes observed in many neurodegenerative disorders. Microglia are the main immune cells of the central nervous system. Many studies have been conducted to find molecules with anti-inflammatory properties in the central nervous system. A family of polyphénols, Stilbenoids (resveratrol derivatives), showed anti-inflammatory effects in peripheral and central levels.In this study, we have first evaluated anti-inflammatory effects of 25 stilbenes for their potential to inhibit NO release by LPS-activated BV-2 microglial cells. Ten stilbenes significantly reduced LPS-induced NO production with IC50 ranging from 3.9 ± 0.7 to 23.4 ±1.0 µM. Among these molecules 1 monomer (moracin M) and two tetramers (vitisins A and B) attenuated the expression of iNOS, a responsible enzyme for NO release on transcriptional and translational levels. Then, we have demonstrated that moracin M inhibits ERK1/2 and JNK phosphorylation of MAPK pathway and Akt phosphorylation of PI3K/Akt pathway, two signaling pathways involved in the inflammatory response in activated BV-2 cells. Indeed, the activation of these pathways leads to the production of several inflammatory mediators. We have shown that moracin M significantly inhibits the production of certain mediators such as NO, TNF-α, IL-1β and PGE2. This stilbene reduces the synthesis of mPGES-1 protein (an enzyme involved in the production of PGE2). In conclusion, we suggest that moracine M could be a potential candidate prevents the inflammation which involved in the progress of neurodegenerative diseases
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Capelli, Irene <1979>. "A step forwards in immunomodulation and immunetolerance knowledge. HLA-G expression in co-cultures of peripheral blood mononuclear cells and stem cells after in vitro NGAL stimulation." Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2013. http://amsdottorato.unibo.it/5795/.
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NGAL (Neutrophil Gelatinase-associated Lipocalin ) is a protein of lipocalin superfamily. Recent literature focused on its biomarkers function in several pathological condition (acute and chronic kidney damage, autoimmune disease, malignancy). NGAL biological role is not well elucidated. Several are the demonstration of its bacteriostatic role. Recent papers have indeed highlight NGAL role in NFkB modulation. The aim of this study is to understand whether NGAL may exert a role in the activation (modulation) of T cell response through the regulation of HLA-G complex, a mediator of tolerance.
From 8 healthy donors we obtained peripheral blood mononuclear cells (PBMCs) and we isolated by centrifugation on a Ficoll gradient. Cells were then treated with four concentrations of NGAL (40-320 ng/ml) with or without iron. We performed flow cytometry analysis and ELISA test. NGAL increased the HLA-G expression on CD4+ T cells, with an increasing corresponding to the dose. Iron effect is not of unique interpretation. NGAL adiction affects regulatory T cells increasing in vitro expansion of CD4+ CD25+ FoxP3+ cells. Neutralizing antibody against NGAL decreased HLA-G expression and reduced significantly CD4+ CD25+ FoxP3+ cells percentage.
In conclusion, we provided in vitro evidence of NGAL involvement in cellular immunity. The potential role of NGAL as an immunomodulatory molecule has been evaluated: it has been shown that NGAL plays a pivotal role in the induction of immune tolerance up regulating HLA-G and T regulatory cells expression in healthy donors. As potential future scenario we highlight the in vivo role of NGAL in immunology and immunomodulation, and its possible relationship with immunosuppressive therapy efficacy, tolerance induction in transplant patients, and/or in other immunological disorders.Ngal (Neutrophil Gelatinase-associated Lipocalin ) è una proteina appartenente alla famiglia delle lipocaline verso cui la recente letteratura ha mostrato una notevole attenzione, soprattutto in quanto biomarcatore in alcune condizioni patologiche (danno renale acuto e cronico, patologie autoimmuni, neoplasie). Il ruolo biologico di NGAL non è però ancora del tutto compreso. Numerose sono le dimostrazioni della sua azione batteriostatica. Recenti lavori hanno inoltre evidenziato un ruolo di NGAL nella modulazione di NFkB. Nessun lavoro ha valutato il ruolo di NGAL nell’immunità umorale. Lo scopo dello studio è quello di capire se NGAL possa esercitare un ruolo di attivazione (modulazione) della risposta T cellulare attraverso la regolazione del complesso HLA-G, un mediatore di tolleranza. Cellule mononucleate da sangue periferico (PBMCs) sono state ottenute da 8 donatori sani dopo consenso informato e isolate tramite centrifugazione (Ficoll). PBMC sono poi state trattate con 4 concentrazioni crescenti di NGAL (da 40 a 320 ng/mL), associate o meno a ferro e analizzate con tecnica fluorimetrica ed elisa.Alle analisi eseguite NGAL stimola l’espressione di HLA-G sulle cellule T CD4+ con un andamento dose dipendente. L’effetto del ferro sull’espressione di HLA-G non è di univoca interpretazione.Inoltre L’aggiunta di NGAL in vitro modifica il pattern di espressione delle cellule T, aumentando la popolazione delle cellule CD4+ CD25+ FoxP3. L’utilizzo di anticorpi anti NGAL limita l’espressione di HLA-G e diminuisce significativamente la percentuale di CD4+ CD25+ FoxP3+ . In conclusione abbiamo mostrato un coinvolgimento di NGAL nell’immunità cellulare. Valutando il ruolo di NGAL come molecola immunomodulatoria, abbiamo mostrato che NGAL gioca un ruolo chiave neell’immunotolleranza aumentando l’espressione di HLA-G e cellule T regolatorie nei donatori sani. Un possibilAs potential future scenario applicativo di tale studio riguarda l’utilizzo in vivo di NGAL nell’immunomodulazione dei pazienti sottoposti a trapianto o affetti da patologie autoimmuni.
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Cullen, Daniel Kacy. "Traumatically-Induced Degeneration and Reactive Astrogliosis in 3-D Neural Co-Cultures: Factors Influencing Neural Stem Cell Survival and Integration." Diss., Georgia Institute of Technology, 2005. http://hdl.handle.net/1853/7584.
Full textAbstract:
Traumatic brain injury (TBI) results from a physical insult to the head and often results in temporary or permanent brain dysfunction. However, the cellular pathology remains poorly understood and there are currently no clinically effective treatments. The overall goal of this work was to develop and characterize a novel three-dimensional (3-D) in vitro paradigm of neural trauma integrating a robust 3-D neural co-culture system and a well-defined biomechanical input representative of clinical TBI. Specifically, a novel 3-D neuronal-astrocytic co-culture system was characterized, establishing parameters resulting in the growth and vitality of mature 3-D networks, potentially providing enhanced physiological relevance and providing an experimental platform for the mechanistic study of neurobiological phenomena. Furthermore, an electromechanical device was developed that is capable of subjecting 3-D cell-containing matrices to a defined mechanical insult, with a predicted strain manifestation at the cellular level. Following independent development and validation, these novel 3-D neural cell and mechanical trauma paradigms were used in combination to develop a mechanically-induced model of neural degeneration and reactive astrogliosis. This in vitro surrogate model of neural degeneration and reactive astrogliosis was then exploited to assess factors influencing neural stem cell (NSC) survival and integration upon delivery to this environment, revealing that specific factors in an injured environment were detrimental to NSC survival. This work has developed enabling technologies for the in vitro study of neurobiological phenomena and responses to injury, and may aid in elucidating the complex biochemical cascades that occur after a traumatic insult. Furthermore, the novel paradigm developed here may provide a powerful experimental framework for improving treatment strategies following neural trauma, and therefore serve as a valid pre-animal test-bed.
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Eriksson, Ulrika. "Impact of autocrine factors on physiology and productivity in Trichoplusia ni serum-free cultures." Licentiate thesis, Stockholm, 2005. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-191.
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