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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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Walls, G. A. "A study of cellular heterogeneity and therapeutic resistance in cultures of human lung cancer." Thesis, University of Cambridge, 1988. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.380720.
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Kalluri, Usha. "Gas chromatographic-mass spectrometry analysis of volatile organic compounds from cancer cell cultures - The effect of hypoxia." Thesis, Federation University Australia, 2018. http://researchonline.federation.edu.au/vital/access/HandleResolver/1959.17/166534.
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Early diagnosis of lung cancer improves patient outcomes which has led to a search for non-invasive diagnostic tests suitable for population screening. Volatile organic compounds (VOCs) in exhaled breath have shown potential, however, confirmation of the metabolic origins and disease specificity of candidate markers is required. Cell culture metabolomics can identify disease biomarkers and their origins. To date VOC profiles from in vitro cultured cancer cells have little similarity to cancer breath VOC profiles. In vivo, cancer cells experience hypoxia whereas in vitro cells are cultured under normoxic conditions. Since hypoxia influences cell metabolism, we hypothesize that cancer cells cultured under hypoxic conditions will have altered cell metabolism and produce VOC profiles more typical of cancer breathe. This study investigates the effect of hypoxia on metabolic reprogramming in A549 lung cancer cells cultured under standard normoxic (atmospheric oxygen) or hypoxic (2% oxygen) conditions. Results from quantitative RT-PCR demonstrated a significant upregulation in hypoxia of the glucose transporter (GLUT1) and the key TCA regulatory gene PDHK1, demonstrating that hypoxia plays a pivotal role in regulating metabolism in A549 cells. A ratio-metric assessment of Lipid Peroxidation (LPO) and the production of reactive oxygen species (ROS) showed an increase in LPO and a slight decrease in the production of ROS in hypoxic cultures, the combined effect of which may serve to equip the cells to adapt to and proliferate under low oxygen. Finally, the comparison of endogenous VOCs produced by A549 cells under hypoxic and normoxic conditions identified twelve VOCs unique to cells grown under hypoxic conditions including n-pentane, a marker of LPO and cancer, and 3-methyl hexane, which has been reported as a biomarker of cancer. This data is consistent with the hypothesis that a hypoxic tumour microenvironment may influence cell metabolism leading to a unique and diagnostic cancer VOC profile.Doctor of Philosophy
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Roberts, Simon A. "Studies on the toxicity and metabolism of T-2 toxin in keratinocyte cultures : evaluation of a keratinocyte cell line and primary cultures as model systems for toxicity testing." Thesis, University of Surrey, 1989. http://epubs.surrey.ac.uk/847950/.
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With a view to establishing a model system for examining toxicity in skin, primary lingual keratinocyte cultures , a keratinocyte cell line and freshly isolated keratinocytes all derived from rat sub-lingual epithelium, were partially characterized both morphologically and enzymically and the toxicity and metabolism of the mycotoxin T-2, studied therein. A number of techniques for obtaining pure suspensions of sublingual keratinocytes (NCK) were examined and dispase is recommended for completely separating the epithelium from its dermis prior to trypsinization. Existing methods for the culture of PLK cells were improved to reduce the number of rats used and minimize the fibroblast contamination and a technique for culturing the epithelium associated with human hair follicles was also examined. The follicle technique was found to produce primary cultures which were 100% epithelial but considerable time and resources were required to generate relatively few cells. The keratinocyte cultures, PLK and RTE5, were shown to produce keratin and undergo stratification like the epithelium in vivo, Using a series of specific enzyme inhibitors, both the cultured and non-cultured epithelial cells were found to possess the same characteristic forms of acetylcholinesterase, butyrylcholinesterase and carboxylesterase. The effect of T-2 on protein synthesis in the keratinocyte cells was examined and a dose-related inhibition was evident. The primary and freshly isolated cells appeared to be the most resistant to synthesis inhibition. The rate of recovery from this inhibition was greatest in the cell line which also lost T-2 at the fastest rate. The uptake into and loss of T-2 from the keratinocyte cells was chiefly by simple diffusion, but studies using rotenone demonstrated that an active process may have been involved in reducing the rates of uptake and loss. The total amount of T-2 absorbed was likely to be dependent on the number of binding sites while the overall rate of loss was dependent on the strength of that binding. The possible nature of these binding sites is discussed. T-2 was metabolized in all the keratinocyte preparations to the same products found in mammalian skin, in vivo, and studies with specific enzyme inhibitors showed that a carboxylesterase was most likely to be involved in its hydrolysis. The greater metabolism of T-2 in the primary cells, which contained the most carboxylesterase, is likely to have been one of the factors involved in reducing the levels of protein synthesis inhibition in these cells. Rat sublingual epithelium and the cultures derived from it have a similar morphology and esterolytic capability to that of skin in viva. The PLK cells were most similar to NCK cells in terms of protein synthesis and esterolytic capability, so if it is assumed that NCK cells are representative of the sublingual epithelial cells in viva and hence skin, then the primary cultures might prove to be the best model culture system for studying toxicity in skin. Nevertheless, the RTE5 cell line was most similar to the NCK cells in respect of T-2 metabolism, and if it shown to have other characteristic skin metabolism systems then it too might prove useful for studying skin toxicity. In addition, the cell line would probably prove to be a more convenient, simpler, reproducible and cheaper means of examining skin toxicity on a large scale.
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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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Bates, Amber Marie. "Matrix metalloproteinase and cytokine profiles from cell co-cultures and their role in oral inflammation and head and neck cancer." Diss., University of Iowa, 2018. https://ir.uiowa.edu/etd/6051.
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Cytokines, chemokines, and MMPs play a major role in both the inflammatory and tumor microenvironments. However, there has been very little research focused on characterizing the MMP and cytokine responses of co-culture models constructed with cells from the oral cavity to study disease. Cells grown in single-cell cultures do not receive signals from other cells as they do in the natural host environment, and the results from single-cell culture studies are often not representative of the host response. Accordingly, studies involving cell cultures with multiple cell types are needed to better represent the host response and responses seen in clinical situation. Therefore, we have developed multiple 3D transwell co-culture systems to study mechanisms involved in periodontal disease and head and neck cancer. Using a unique 3D transwell co-culture, we are able to eliminate cell-cell physical interactions and focus on the effects of cell signals and cell reactions to these signals caused by the presence other cells in the co-culture. Studying the MMP and immunosuppressive biomarker response in this type of model will provide new insights on the tumor microenvironment created by head and neck cancer cells and the pro-inflammatory environment in periodontal disease.
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Kashtl, Ghasaq J. "Differential membrane-type matrix metalloproteinase expression in phenotypically defined breast cancer cell lines: Comparison of MT-MMP expression in environmentally-challenged 2D monolayer cultures and 3D multicellular tumour spheroids." Thesis, University of Bradford, 2018. http://hdl.handle.net/10454/17346.
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Matrix metalloproteinases (MMPs) are a family of zinc endopeptidases capable of digesting the extracellular matrix (ECM), which is essential for tissue structure and transmitting messages between cells. MMPs play an important role in cancer, controlling cell migration, proliferation, apoptosis, regulation of tumour expansion, angiogenesis and invasion. Previous research has indicated high expression of MT1-MMP in breast cancers suggesting a potential role in tumour progression. Our results confirm that 3D multicellular tumour spheroids (MCTS) using phenotype-specific breast cancer cell lines are a valuable experimental model of the tumour microenvironment.
Optimisation of MCTS culture growth conditions using different breast cancer cell lines (MCF-7, T47D, MDA-MB-468 and MDA-MB-231) was performed. Unexpected detection of MT1-MMP in MCF-7 MCTS warranted further investigation. MT1-MMP expression in different micro-environmental conditions, including hypoxia and nutrient deprivation (serum-free induced autophagy) were measured in MCF-7 monolayer cultures and MCTS models using immunofluorescence (IF), immunohistochemistry (IHC) and western blot (WB).
MT1-MMP expression was rapidly and irreversibly up-regulated in MCF-7 breast cancer cells under conditions of stress (hypoxia and autophagy) compared to normal conditions suggesting an important role of the culture environment on cells behaviour and protein expression.
We employed isobaric tags for relative and absolute quantitation (iTRAQ) technology to correlate MT1-MMP increase with proteomic profiles in MCF-7 breast cancer cell grown under hypoxic, serum-free and 3D MCTS conditions. More than 3500 proteins were identified, which were clustered into groups based on response to unique or shared microenvironment changes. Hypoxic monolayer and spheroid cells exhibited changes in anaerobic metabolism and lipid synthesis, respectively, whereas autophagy resulted in up-regulation of cellular component disassembly. The result indicated multiple drivers of MT1-MMP expression in MCF-7 cells.Al-Mstansiriya University, Iraq
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Trakarnpaibul-Kunakornsawat, Sunee. "Effects of 1α,25-Dihydroxyvitamin D₃ and its analogs in Cancer-associated Hypercalcemia in vivo and in vitro, and in normal Prostate Epithelial and Stromal Primary Cell Cultures /." The Ohio State University, 2001. http://rave.ohiolink.edu/etdc/view?acc_num=osu1486572165276222.
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Dolega, Monika Elzbieta. "Developement of microtechnologies for 3D cell culture to study prostate acini formation and carcinogenesis." Thesis, Grenoble, 2014. http://www.theses.fr/2014GRENS022/document.
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Tout épithélium glandulaire sécrétoire est constitué d'une unité structurale et fonctionnelle commune, l'acinus. C'est une architecture sphérique pluricellulaire parfaitement différentiée et polarisée qui, reconstruite en culture 3D, mime l'organisation réelle du tissu. Etudier les déterminants environnementaux et génétiques qui gouvernent la transformation d'un acinus en sphéroïde s'apparentant à une tumeur est l'un des enjeux majeurs des modèles in vitro. Un des défis actuels est d'adapter ces modèles in vitro à des conditions de culture 3D qui soient compatibles avec la réalisation de cribles génétiques en 3D, basés par exemple sur l'ARN interférence (RNAi). Cependant, les formats standards de culture 3D et les méthodes analytiques ne sont pas compatibles aux cribles haut-débit. Ils ne permettent pas de contrôler la taille et la distribution des acini, sont dépendants d'immuno-marquages et les acquisitions sont longues. Par ailleurs, la microscopie confocale et vidéomicroscopie offrent un champ d'observation restreint qui ne permet pas d'observer un grand nombre de structures 3D en même temps, pour permettre une analyse statistique. Ainsi, dans le but i) de développer des modèles cellulaires appropriés en 3D, ii) d'adresser des questions fondamentales relatives au cancer de la prostate et iii) de réaliser des cribles RNAi dans un contexte plus pertinent que la culture 2D, j'ai développé des outils innovants au format microsystèmes adaptés à l'analyse haut-débit d'un grand nombre d'objets 3D. En optimisant les conditions de culture cellulaire 3D sur le modèle de la lignée cellulaire RWPE1, j'ai pu récapituler les étapes de formation des acini prostatiques et montrer que la formation du lumen est indépendante de la polarité et est gouvernée par deux mécanismes, « hollowing » et cavitationIn all secretory epithelia from glandular tissues, there is a common structural and functional unit, the acinus. It is a well polarized and organized pluricellular structure that is spontaneously reconstructed in 3D culture, therefore closely mimics the real structure we find in vivo. For my purpose, acini are used as models for tumor initiation and cancer development. One of the objectives of Biomics laboratory is to identify the genetic and microenvironmental determinants of prostate acini morphogenesis and polarity. The strategy is based on High-Throughput (HT) RNA interference (RNAi)-based screening. To meet this objective, my project was to develop appropriate 3D cell models which closely mimic the cyst-like and duct-like structure of prostate. By optimizing conventional 3D culture in Matrigel, I could recapitulate prostate acini morphogenesis and showed that lumen formation is independent to the polarity, which appears later. However, the conventional 3D cell culture formats and analytical tools are not suited for HT Screening (HTS). They lack control over acini size, are label-dependant and therefore time-consuming and labor intensive. Also, classical microscopy offers a very limited field of view and hence does not allow observing a large amount of 3D structures for statistical analysis
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Huynh, Minh Diem. "Reduction in apparent stromal cell culture density through transient fusions with osteosarcoma cells." University of Sydney, 2007. http://hdl.handle.net/2123/4465.
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Doctor of PhilosophyBenign tumours grow by expanding and displacing the surrounding tissues, while malignant tumours replace and destroy the surrounding tissues by invasion. Although there is extensive literature on mechanisms of tumour invasion and metastasis, with an emphasis on angiogenesis, adhesion, degradation of the extracellular matrix and migration, an important question not clearly addressed by the literature, but nonetheless approached in this thesis, is that of the fate of normal cells during tissue replacement by migrating invasive malignant cells. Earlier work in the laboratory where this PhD candidature was carried out, investigated the effect of osteosarcoma cells on endothelium. In contrast to the expected angiogenic effect of malignant cells for endothelium, it was found that the human osteosarcoma cell line (SAOS-2) induced apoptosis in human umbilical vein endothelial cells (HUVEC) in contact dependent manner (McEwen et al., 2003). It was suggested that apoptosis of endothelium by malignant tumour cells may facilitate tumour invasion and metastasis (McEwen et al., 2003), and one of the aims of the current study was to extend these findings to include human gingival fibroblasts (HGF) and human umbilical artery smooth muscle cells (HUASMC). The major finding of this thesis was that SAOS-2 induced a reduction in the apparent cell culture density of HGF and HUASMC in a contact-dependent manner. The SW480 colorectal carcinoma cell line did not have any clear effect upon the apparent stromal cell culture density of either HGF or HUASMC, suggesting that the effect under investigation was tumour cell line specific. Surprisingly and in contrast to the similar effect reported for endothelium (Chen et al., 2005; McEwen et al., 2003), the effect of SAOS-2 upon HGF and HUASMC was not due to stromal cell apoptosis. Apoptosis was ruled out as a possible mechanism for the reduced apparent culture density under study, by using widely accepted methods which are dependent upon intermucleosomal fragmentation of DNA, the permeability of plasma membranes to dyes in advanced apoptosis and necrosis, phosphatidylserine translocation as well as inhibitor studies blocking both caspase dependent and independent pathways. While apoptosis was not demonstrated, the possibility emerged that reduced apparent stromal cell culture density reflected fusion events rather than the simple removal of cells as had been earlier reported for HUVEC (McEwen et al., 2003). This idea was supported by reduced SAOS-2 circularity in co-culture. Confocal microscopy of cells pre-labelled with fluorescent dyes further supported this idea, with dual-labelling as evidence of cell fusion. Although occasional homotypic fusion of stromal cells was seen, heterotypic fusion of stromal cells with SAOS-2 was much more prevalent. Time lapse microscopy was performed to further characteristic cell fusion in co-cultures, and revealed multiple transient fusions between SAOS-2 and HGF. To work towards determining the biological relevance of the key observation, two stable SAOS-2 GFP clones were generated for future planned studies using human gingival explants and nude mice. Importantly, the clones were similar to native SAOS-2 with regard to alkaline phosphatise expression and reducing apparent stromal cell culture density. Transient fusions between HGF and SAOS-2, may be a mechanism for cooption of stromal cells into the malignant process, facilitating tumour invasion. Additionally, heterocellular fusion of SAOS-2 with stromal cells may facilitate immune evasion, while it seems likely that despite the absence of an identical activity in SW480 cells, other malignant tumour cells may also express similar activity.
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Kapeleris, Joanna C. "Circulating tumour cells in non-small cell lung cancer." Thesis, Queensland University of Technology, 2022. https://eprints.qut.edu.au/228607/1/Joanna_Kapeleris_Thesis.pdf.
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Circulating tumour cells (CTCs) have the potential to transform the management of patients with non-small cell lung cancer (NSCLC). The applications of CTCs can identify clinically actionable targets to predict treatment response and to better understand metastasis. CTCs isolated using microfluidics can be used as prognostic indicators of NSCLC as well as characterizing for markers of immunotherapy (PD-L1), molecular targets (ALK, EGFR). Short term cultures were successfully expanded in 9/70 NSCLC patients and cultured for up to 3 months. Optimization of this novel CTC culture model provides opportunity to identify new therapeutics for NSCLC patients in a precision medicine approach.
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凌明達 and Ming-tat Patrick Ling. "A study of molecular and cell biology of prostate tumorigenesis in cell culture." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2000. http://hub.hku.hk/bib/B31223102.
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Timmins, Nicholas E. "Extending the third dimension : novel methods and applications for 3D multicellular spheroids /." [St. Lucia, Qld.], 2004. http://www.library.uq.edu.au/pdfserve.php?image=thesisabs/absthe18289.pdf.
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Faustino, Vera. "The study of cell behaviour using biomedical microdevices." Master's thesis, Instituto Politécnico de Bragança, Escola Superior de Tecnologia e Gestão, 2012. http://hdl.handle.net/10198/8216.
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The cell culture has become over the years an important discipline for the thorough knowledge of cells behaviour and migration, as also cell-cell interaction. A few years ago it was only possible to create culture systems in vitro systems, however, with the advance of new technologies it has become possible to create culture systems in vivo closer to reality.
The microfabrication processes such as photolithography were used in this work for the design of a microdevice which is aimed to study the migration of cells into nanofibers.
Beyond the cell studies, microdevices were also used for rheological studies. The behaviour of blood cells is very important, regarding the transport of oxygen in our body as well as nutrients. The knowledge about the factors that influence changes in mechanical properties of erythrocytes is of great importance because it is their deformability that allows them to carry oxygen and pass through smaller arteries and veins. Therefore, another microdevice was used for the purpose of observing the deformation of the erythrocytes after contact with human tumour cell lines (NCI-H460- non-small cell lung cancer and HCT-15 - colon carcinoma) up to two consecutive days. With the flow rate of 500 nl/min, the flowing erythrocytes through the microchannel were captured by a high speed camera and their deformability was examined. The erythrocytes that contacted tumour cells show less deformability than ―healthy‖ erythrocytes.
It was found that the tests were insufficient to conclude clearly the effect of the tumour cells in the deformation of erythrocytes. We have found evidence that erythrocytes also influence the growth of tumour cells, however, further testing would be needed to clarify this observation. A cultura de células tornou-se ao longo dos anos uma importante disciplina para o conhecimento profundo do seu comportamento, migração e interação entre células. Com o avanço de novas tecnologias tornou-se possível criar sistemas de cultura in vivo mais próximos da realidade, quando anteriormente apenas existiam sistemas in vitro.
Neste trabalho, foram aplicados processos de microfabricação, nomeadamente a fotolitografia, para a conceção de um microdispositivo que teve como objetivo principal o estudo da migração de células em nanofibras.
Para além de estudos celulares, os microdispositivos também foram usados em estudos reológicos. O comportamento do sangue é muito importante, pois é ele que transporta o oxigénio no nosso organismo assim como nutrientes. Perceber o que influencia a alteração das características mecânicas dos eritrócitos é de grande importância pois é a sua deformabilidade que lhes permite transportar o oxigénio e passar nas artérias e veias mais pequenas. Assim, utilizou-se um outro microdispositivo, com o propósito de observar a deformação dos eritrócitos após contacto com linhas celulares tumorais humanas (NCI-H460 – cancro do pulmão e HCT-15 – carcinoma do cólon) até um máximo de dois dias consecutivos. Os eritrócitos fluíram pelo microcanal com uma taxa de fluxo de 500 nl/min e foram capturados por uma câmara de alta velocidade para ver a sua capacidade de deformação. Os eritrócitos que estiveram em contacto com as células tumorais revelam menos deformação que os eritrócitos ―saudáveis‖.
Verificou-se que os ensaios realizados foram insuficientes para concluir com clareza o efeito das células tumorais na deformação dos eritrócitos. Encontraram-se também indícios de os eritrócitos influenciarem o crescimento das células tumorais, no entanto, mais ensaios também seriam necessários para clarificar essas observações.This Final Project Report was a collaboration of the University of Navarra and research centre, CEIT
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Marzioch, Julia [Verfasser], and Gerald A. [Akademischer Betreuer] Urban. "Microsensor system for the metabolic monitoring in cancer cell culture." Freiburg : Universität, 2019. http://d-nb.info/118713337X/34.
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Böpple, Kathrin [Verfasser], and Roland [Akademischer Betreuer] Kontermann. "Characterization of persister-cell derived ovarian cancer cells and methods for advanced 3D cell and tissue culture / Kathrin Böpple ; Betreuer: Roland Kontermann." Stuttgart : Universitätsbibliothek der Universität Stuttgart, 2021. http://d-nb.info/1233287818/34.
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Atefi, Ehsan. "Aqueous Biphasic 3D Cell Culture Micro-Technology." University of Akron / OhioLINK, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=akron1443112692.
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Morales, Delphine. "Modèles 3D de mélanome métastatique pour l’évaluation in vitro de l’efficacité de molécules de thérapies ciblées." Thesis, Compiègne, 2019. http://www.theses.fr/2019COMP2498.
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La sensibilité des cellules de mélanomes aux molécules de thérapies ciblées dépend du microenvironnement tumoral (interactions cellule-cellule et cellule-matrice extracellulaire). Les systèmes tridimensionnels (3D) de culture in vitro reflètent mieux l’architecture structurelle native des tissus et sont attrayants pour l’étude des interactions cellulaires. Nous avons développé et comparé plusieurs modèles de mélanome métastatique : les cellules de mélanomes (SK-MEL-28 et SK-MEL-3, mutées BRAF V600E et SK-MEL-2, BRAF sauvages) cultivées en monocouche (2D) et co-cultivées en 3D sur des équivalents de derme avec des fibroblastes, afin de mieux comprendre les facteurs modulant la sensibilité cellulaire à un inhibiteur de BRAF (BRAFi, Vémurafenib) et au Vémurafenib associé à un inhibiteur de MEK (MEKi, Cobimetinib). La sensibilité cellulaire aux traitements a été évaluée sous différents aspects : prolifération cellulaire (numération cellulaire, incorporation d'EdU, test MTS), analyse des voies de signalisation MAPK et PKB / Akt (Western-blot), apoptose (TUNEL), libération de cytokines et de facteurs de croissance (ELISA) et histologie (modèles 3D). Un effet cytostatique de BRAFi a été observé sur les cellules SK-MEL-28 et SK-MEL-3 cultivées dans les modèles 2D et 3D. La lignée cellulaire SK-MEL-2 était résistante au BRAFi lorsqu'elle a été cultivée en monocouche, mais sensible lorsqu'elle a été co-cultivée avec des fibroblastes incorporés dans une matrice de collagène de type I. Les milieux conditionnés par les fibroblastes 3D (équivalents de derme) ont sensibilisé les cellules SK-MEL-2 (2D) au BRAFi. L'analyse des surnageants de culture cellulaire a révélé que les équivalents de derme libéraient certains facteurs solubles (IL-6, IL-8, HGF, TGF-β) : ces sécrétions ont été modifiées au cours du traitement par Vémurafenib. La combinaison du traitement avec MEKi a renforcé l'action du Vémurafenib sur les cellules de mélanomes métastatiques tout en diminuant la capacité de prolifération des fibroblastes. Des populations de cellules contenant des cellules de mélanomes ou des fibroblastes associés au cancer (CAFs) ont été isolées à partir d'une biopsie de métastase cutanée provenant d'une patiente atteinte d'un mélanome métastatique. Ces cellules ont permis de réaliser des modèles de mélanome métastatique patient-spécifique afin d’étudier in vitro la sensibilité des cellules de la patiente aux traitements dans un microenvironnement tumoral (sécrétion paracrine de cellules stromales et matrice de collagène). Ces modèles prédictifs 3D patient-spécifique pourront être utilisés pour déterminer des stratégies de thérapies personnalisées, ainsi que pour comprendre les phénomènes de résistance des cellules de mélanomes aux traitementsMelanoma cell sensitivity to targeted therapy molecules is dependent on the tumor microenvironment (cell-cell and cell-extracellular matrix interactions). Three dimensional (3D) in vitro cell culture systems better reflect the native structural architecture of tissues and are attractive to investigate cellular interactions. We have developed and compared several metastatic melanoma models: melanoma cells (SK-MEL-28 and SK-MEL-3, BRAF V600E mutant and SK-MEL-2 BRAF wt) cultured as a monolayer (2D) and co-cultured on 3D dermal equivalents with fibroblasts to better unravel factors modulating cell sensitivity to a BRAF inhibitor (BRAFi, Vemurafenib) and a BRAFi combined with a MEK inhibitor (MEKi, Cobimetinib). Cell sensitivity to treatments was evaluated under various aspects: cell proliferation (cell counting, EdU incorporation, MTS assay), MAPK and PKB/Akt signaling pathway analysis (Western-blotting), apoptosis (TUNEL), cytokine and growth factor release (ELISA) and histology (3D models). A cytostatic effect of BRAFi was observed on SK-MEL-28 and SK-MEL-3 cells in both models. SK-MEL-2 cell line was clearly resistant to BRAFi when cultured as a monolayer but not when co-cultured with 3D fibroblasts embedded in a type I collagen matrix. Conditioned media provided by 3D fibroblasts (dermal equivalents) underlined 2D SK-MEL-2 sensitivity to BRAFi. Cell culture supernatant analysis revealed that dermal equivalents released some soluble factors (IL-6, IL-8, HGF, TGF-β): these secretions were modified during vemurafenib treatment. The combination of treatment with MEKi enhances the action of Vemurafenib on metastatic melanoma cells while decreasing the proliferation capacity of fibroblasts. Cell populations containing melanoma cells or fibroblasts associated with cancer (CAFs) were isolated from a cutaneous metastasis biopsy of a patient with metastatic melanoma. These cells allowed the realization of patient-specific models of metastatic melanoma in order to study in vitro the sensitivity of the patient’s melanoma cells to treatments in a tumor microenvironment (paracrine secretion of stromal cells and collagen matrix). These 3D predictive patient-specific models could be used to determine personalized therapy strategies, as well as to understand the resistance phenomena of melanoma cells to treatments
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Gaedtke, Lars. "Cell culture models and novel gene therapeutic strategies for colorectal cancer." Diss., lmu, 2006. http://nbn-resolving.de/urn:nbn:de:bvb:19-51714.
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Chinembiri, Tawona Nyasha. "Optimised topical delivery of 5-fluorouracil." Thesis, North-West University, 2012. http://hdl.handle.net/10394/9000.
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Skin cancer is the most widely diagnosed form of cancer and it is split in to non-melanoma skin cancer (NMSC) and cutaneous malignant melanoma (CMM). Cutaneous melanoma has a high propensity for malignancy and it has the highest mortality rate of all skin cancers (de Gruijl, 1999:2004). The first line of treatment for most skin cancers is surgical excision but instances do arise in which surgery is not feasible due to the health of the patient or the location of the lesion. Therefore, viable alternatives are necessary in cases where surgery is not possible (Telfer et al., 2008:36). The skin is readily available for delivery of cytotoxic drugs to treat carcinomas and melanomas so the topical delivery of 5-fluorouracil was investigated in this study.
5-Fluorouracil is a pyrimidine anti-metabolite which interferes with deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) synthesis by inhibiting the nucleotide synthetic enzyme thymidylate synthase (TS) and by becoming misincorporated into RNA and DNA. Thymidylate is essential for replication as well as repair of DNA, in the event of TS inhibition thymidylate is not formed and “thymineless deaths” of cells occur (Chu & Sartorelli, 2009:935; Longley et al., 2003:330). This active pharmaceutical ingredient (API) causes death of atypical and rapidly dividing cells (Tsuji & Karasek, 1986:474). The intravenous and topical routes are approved for 5-fluorouracil and in the case of skin cancer the obvious choice would be topical application (Chu & Sartorelli, 2009:935). Topical application of 5-fluorouracil results in the occurrence of terrible side effects such as severe inflammation, stomatitis, photosensitivity and dermatitis. A reduction in side effects would reduce the stigma associated with topical 5-fluorouracil and in turn increase patient compliance.
Topical drug delivery entails the delivery of an API onto or into the various layers of the skin (Flynn & Weiner, 1993:33) in order to treat conditions on or within the skin. Topical application of APIs is non-invasive, painless and simple plus the target site is readily accessible for topical therapy, thus the API is delivered directly to the site of action (Naik et al., 2000:318). In the case of skin cancer, 5-fluorouracil should be able to reach the epidermis because NMSC originates from the keratinocytes (Marks & Hanson, 2010:305) and CMM from melanocytes (de Gruijl, 1999:2004) which are both found in the epidermis. The barrier function of the skin limits the penetration of molecules into the skin and the rate-limiting step is usually penetration into the stratum corneum (Foldvari, 2000:418).
The aim of this study was to investigate the diffusion of 5-fluorouracil from formulations into and through the skin. Two physico-chemical properties of 5-fluorouracil that influence skin permeation were determined (aqueous solubility and n-octanol-buffer partition coefficient (log D)). The Pheroid™ drug delivery system was used to enhance the delivery of 5-fluorouracil (Grobler et al., 2008:284). Pheroid™ is a novel technology that is used in the delivery of APIs in pharmaceutical products. It enhances the efficacy of delivered compounds while allowing for the reduction of unwanted adverse effects (Grobler et al., 2008:284). Franz cell skin diffusion studies and tape-stripping were conducted with Pheroid™ and non-Pheroid™ formulations to allow for comparison and determination of the effect of Pheroid™. The in vitro efficacy of 5-fluorouracil in inducing apoptosis of human melanoma cells was investigated using a flow cytometric apoptosis assay. Different concentrations of 5-fluorouracil in formulation were utilised in the experiments so as to observe the cytotoxic effect of 5-fluorouracil. The effect of the drug delivery vehicle on the efficacy of 5-fluorouracil was investigated by utilising API solutions in addition to Pheroid™ and non-Pheroid™ formulations in the experiments.
Relatively high concentrations of 5-fluorouracil diffused into and through the skin with Pheroid™ formulations resulting in a greatly enhanced in vitro skin permeation of 5-fluorouracil. The tape-stripping revealed that the Pheroid™ lotions resulted in higher concentrations of 5-fluorouracil in the epidermis and dermis after 12 h as compared to the lotions. There was no deducible trend with respect to the distribution of 5-fluorouracil between the epidermis and dermis. Subsequent to the apoptosis assay it was found that 5-fluorouracil was able to induce apoptosis in A375 cells after a 24 h incubation period. The Pheroid™ treatment of cells resulted in a greater response (mean fluorescence intensity) as compared to treatments with the other drug delivery vehicles at three of the four concentrations. This showed that the drug delivery vehicle played a role in the in vitro efficacy of 5-fluorouracil.
Further research must be done in order to combine these results. Optimum and highly effective topical formulations with low doses of 5-fluorouracil must be formulated for the purpose of treating cutaneous cancers with a reduced incidence of side effects.Thesis (MSc (Pharmaceutics))--North-West University, Potchefstroom Campus, 2013.
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Canbay, Emel. "Responsiveness of cultured human breast cancer cells to prolactin." Thesis, King's College London (University of London), 1996. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.244652.
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Ferreira, Luís Pedro Correia Pinto. "Development of multicelular 3D cancer testing platforms for evaluation of new anti-cancer therapies." Master's thesis, Universidade de Aveiro, 2017. http://hdl.handle.net/10773/22713.
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Mestrado em Bioquímica ClínicaO cancro do pulmão (CP) é um dos cancros mais diagnosticados a nível mundial e também um dos mais mortíferos. Atualmente, as terapias administradas a nível clínico para o tratamento do CP são ainda extremamente ineficazes e limitadas no que diz respeito ao aumento da taxa de sobrevivência dos pacientes oncológicos. Esta realidade demonstra a necessidade de investigar ativamente novas terapias para o tratamento desta neoplasia. No entanto a validação pré-clínica de terapias inovadoras para o CP tem-se revelado extremamente difícil devido à inexistência de plataformas que sejam adequadas para testes a nível laboratorial, uma vez que as culturas celulares in vitro bidimensionais (2D), recomendadas pelas agências regulatórias são incapazes de mimetizar as caraterísticas principais dos tumores humanos. Estas limitações têm originado uma fraca correlação entre a performance das terapias nos estudos in vitro e a obtida em ensaios clínicos controlados. Neste contexto, os modelos de tumores tridimensionais (3D) in vitro têm vindo a ser reconhecidos como uma solução para este problema, pois podem recapitular várias componentes do microambiente tumoral. Das várias plataformas 3D in vitro de CP investigadas atualmente muito poucas avaliaram o papel da inclusão de células estaminais mesenquimais (MSCs). Para colmatar esta lacuna, o trabalho de investigação desenvolvido no âmbito desta dissertação descreve a produção e otimização de novos modelos hétero-celulares 3D in vitro. Estas plataformas são compostas por células tumorais do CP (A549) e do seu estroma, nomeadamente fibroblastos da pele e células estaminais mesenquimais derivadas da medula óssea (BM-MSCs). Estes três tipos de células foram co-cultivadas em micropartículas poliméricas de policaprolactona revestidas por ácido hialurónico, com o objetivo de incluir este componente da matriz extracelular que se encontra presente no microambiente do CP. Esta abordagem permitiu formar a nível laboratorial microtecidos multicelulares 3D híbridos que melhor mimetizam a heterogeneidade celular das neoplasias pulmonares. Os resultados obtidos demonstraram que os microtumores formados através da técnica de sobreposição-líquida são reprodutíveis em termos de morfologia e tamanho, apresentaram núcleos necróticos, organização celular 3D e produziram proteínas do microambiente tumoral. Além destas caraterísticas, os dados obtidos através de microscopia de fluorescência revelaram que as BM-MSCs migram para o interior dos microtumores ao longo do tempo. A avaliação da citotoxicidade da Doxorubicina, um fármaco anti-tumoral rotineiramente utilizado a nível clínico, demonstrou que a inclusão de micropartículas aumenta a resistência das células tumorais em modelos homotípicos. Nos modelos tri-cultura heterotípicos a citotoxicidade foi comparável à obtida em microtumores sem micropartículas. Estes resultados evidenciam assim o papel importante dos fibroblastos e das BM-MSCs na resposta dos microtumores. Numa visão global, os modelos 3D formados recapitulam com mais exatidão o microambiente do cancro do pulmão e poderão servir no futuro como plataformas de teste para descobrir ou aperfeiçoar novas terapias, ou combinações de terapêuticas, para este tipo de neoplasia.
Lung cancer (LC) is one the most commonly diagnosed cancers worldwide, being also one of the deadliest. Currently, clinically administered therapies for treatment of LC are still extremely ineffective and limited in increasing oncologic patients survival rates. This reality evidences the necessity of actively investigating novel therapies for the treatment of LC. However, preclinical validation of novel therapies as revealed itself as an extremely arduous process, due to the lack of suitable laboratory testing platforms since the recommend in vitro bi-dimensional (2D) cell cultures are unable to fully mimic the main hallmarks of human tumors. In this context, in vitro tridimensional (3D) tumor models are being increasingly recognized as a solution due to their ability to correctly recapitulate several characteristics of the tumor microenvironment (TME). Amongst currently developed 3D in vitro platforms for the study of LC, few have included or studied the role of mesenchymal stem cells (MSCs). To provide further insights into this hypothesis, the research work developed in this thesis describes the production and optimization of novel heterotypic in vitro 3D models, comprised by non-small-cell lung cancer cells (A549) and stromal cells, namely skin fibroblasts (HFs), and bone-marrow derived mesenchymal stem cells (BM-MSCs). These three diverse cell populations were co-cultured in hyaluronic acid coated polymeric polycaprolactone microparticles (LbL-MPs) as to include this key extracellular matrix component of LC TME. This approach allowed the formation of 3D multicellular heterotypic microtissues (3D-MCTS) that better recapitulate the cellular heterogeneity of LC TME in the laboratory. The obtained findings demonstrate that these models formed via the liquid-overlay technique were reproducible in terms of morphology and size, presented necrotic core formation, 3D cellular organization, and deposited matrix proteins in a similar manner as in the TME. Besides this, fluorescence microscopy data revealed that BM-MSCs migrated overtime into the microtumors core . Performed doxorubicin in vitro cytotoxicity assays revealed that the inclusion of LbL-MPs lead to an increased resistance of homotypic A549 monoculture models against this anti-cancer drug commonly used in clinical treatments. Alongside, the cytotoxicity obtained in triculture heterotypic models was comparable to that of microtumors without LbL-MPs inclusion, showcasing the role of HFs and BM-MSCs in microtumors response to therapy. Globally, the herein bioengineered 3D models were able to recapitulate with an increased precision the TME of LC, making them suitable test platforms for development or improvement of standalone or combinatorial therapies for this type of neoplasia.
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Kingston, Shaun Thomas. "Genotoxicology of diesel engine exhaust emissions in cultured mammalian cells." Thesis, University of Plymouth, 1994. http://hdl.handle.net/10026.1/2131.
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Diesel exhaust emissions were collected from a 2 litre direct injection diesel engine using the Total Exhaust Solvent Scrubbing Apparatus (TESSA). Emission samples were collected from a series of two minute engine runs, at a variety of engine speeds and loads which covered the full operating range of the engine. The exhaust extracts collected were then tested for their cytotoxicity and mutagenic potential in cultured Chinese hamster cells. Emission samples were found to be extremely toxic, with most causing 100% cytotoxicity at concentrations of less than 100 /µg/ml. Chromosome aberration studies indicated that less than 50% of the emission samples collected induced increases in the numbers of cells with aberrations, and less than 10% induced aberrations in cell cultures exposed to emission samples with a supplementary metabolic activation system. Samples tested in in vitro sister chromatid exchange assays, induced significant increases in the frequencies of exchanges. Linear trend statistics calculated from chromosome aberration data, were used to reflect the relative clastogenic potential of individual emission samples. Linear regression of these trend values against physical components of the exhaust emissions, showed a significant correlation between sample mutagenicity in aberration assays and the emission of oxides of nitrogen (NOx) in the exhaust. Mapping of NOx emissions to engine conditions has shown that the mutagenicity of diesel emissions from the test engine tend to be highest under condition of low speed/high load and high speed with increasing load. Toxicity assays of subfractions of emission samples isolated by column chromatography has shown that the toxicity of the emission samples is associated with the aromatic and polar components of the samples. The dose responses obtained from mutagenicity assays, in which samples only caused increases in aberrations and chromatid exchanges at the same concentrations at which cytotoxic effects were observed, suggests that the emission may have a limited cytogenetic effects in vivo. Mapping of the clastogenicity of emission samples against engine speed and load, has shown that the most clastogenic samples collected, were emitted under engine conditions that might be expected to occur under urban driving conditions. Results of epidemiological studies of health effects of diesel emissions, and recent associations between particulate emissions and health effects are discussed.
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Daukste, Liene. "Mathematical Modelling of Cancer Cell Population Dynamics." Thesis, University of Canterbury. Department of Mathematics and Statistics, 2012. http://hdl.handle.net/10092/10057.
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Mathematical models, that depict the dynamics of a cancer cell population growing out of the human body (in vitro) in unconstrained microenvironment conditions, are considered in this thesis. Cancer cells in vitro grow and divide much faster than cancer cells in the human body, therefore, the effects of various cancer treatments applied to them can be identified much faster. These cell populations, when not exposed to any cancer treatment, exhibit exponential growth that we refer to as the balanced exponential growth (BEG) state. This observation has led to several effective methods of estimating parameters that thereafter are not required to be determined experimentally.
We present derivation of the age-structured model and its theoretical analysis of the existence of the solution. Furthermore, we have obtained the condition for BEG existence using the Perron-Frobenius theorem. A mathematical description of the cell-cycle control is shown for one-compartment and two-compartment populations, where a compartment refers to a cell population consisting of cells that exhibit similar kinetic properties. We have incorporated into our mathematical model the required growing/aging times in each phase of the cell cycle for the biological viability. Moreover, we have derived analytical formulae for vital parameters in cancer research, such as population doubling time, the average cell-cycle age, and the average removal age from all phases, which we argue is the average cell-cycle time of the population. An estimate of the average cell-cycle time is of a particular interest for biologists and clinicians, and for patient survival prognoses as it is considered that short cell-cycle times correlate with poor survival prognoses for patients.
Applications of our mathematical model to experimental data have been shown. First, we have derived algebraic expressions to determine the population doubling time from single experimental observation as an alternative to empirically constructed growth curve. This result is applicable to various types of cancer cell lines. One option to extend this model would be to derive the cell cycle time from a single experimental measurement. Second, we have applied our mathematical model to interpret and derive dynamic-depicting parameters of five melanoma cell lines exposed to radiotherapy. The mathematical result suggests there are shortcomings in the experimental methods and provides an insight into the cancer cell population dynamics during post radiotherapy. Finally, a mathematical model depicting a theoretical cancer cell population that comprises two sub-populations with different kinetic properties is presented to describe the transition of a primary culture to a cell line cell population.
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Pyne, Emily Seton. "The Impact of Stromal Cells on the Metabolism of Ovarian Cancer Cells in 3D Culture." Thesis, Virginia Tech, 2017. http://hdl.handle.net/10919/74931.
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Academic: Ovarian cancer is the leading cause of death among female gynecologic cancers. Current treatments include surgical debulking, and chemotherapy. However, better interventions are needed to reduce the mortality rate of metastatic disease. Ovarian cancer cells have displayed the ability to aggregate and form 3D homogeneous and heterogeneous spheroids, which can function as micrometastases. Ovarian cancer spheroids survive independently prior to adhering to an endothelial tissue. Since aggregation has been shown to provide a survival advantage to the spheroids and increased their aggressive phenotype, this study aimed to investigate how the metabolism of ovarian cancer cells change in 3-dimensional (3D) culture. Examining metabolic pathways and identifying markers of metabolic change could provide the scientific base for new, targeted interventions for this disease. Spheroids of both homogeneous and heterogeneous composition demonstrated overall lower metabolic capacity than their adherent counterparts. Spheroids had a lower basal energetic demand than adherent cells, paralleled by lower maximal respiration capacity, glycolytic capacity, and spare respiratory capacity. We conclude that the lower energetic demand of spheroids may be a mechanism to prolong death by reserving energy and metabolic cellular processes; this may render anti-metabolic drug treatment with AICAR or metformin ineffective against disseminating ovarian cancer aggregates.
General: Ovarian cancer is currently the leading cause of death among female gynecologic cancers. While treatments exist, better interventions are needed to reduce the mortality rate in this form of cancer. Ovarian cancer cells have displayed the ability to aggregate and form 3D homogeneous and heterogeneous spheroids, which can function as micrometastases. Ovarian cancer spheroids survive independently prior to adhering to an endothelial tissue. Since aggregation has been shown to provide a survival advantage to the spheroids and increased their aggressive phenotype, this study aims to investigate how the metabolism of ovarian cancer cells change in 3-dimensional (3D) culture. Examining metabolic pathways and identifying markers of metabolic change could provide the scientific base for new, targeted interventions for this disease.Master of Science
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Liddil, James Duncan 1960. "Mechanisms of the cytotoxic actions of tumor necrosis factor (TNF) in cultured cancer cells." Thesis, The University of Arizona, 1987. http://hdl.handle.net/10150/276602.
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Tumor necrosis factor's (TNF) cytotoxic mechanism of action was examined using cultured cancer cell lines. TNF demonstrated cytolytic and cytostatic effects on L929 fibrosarcoma and MCF-7 adenocarcinoma cells. TNF failed to show any specific effects on RNA, DNA or protein synthesis or ATP content in tumor cells in vitro. It did not cause DNA single strand breaks. Decreased cellular levels of reduced thiols did not predict sensitivity to the cytotoxic effects of TNF. Depletion of cellular glutathione failed to increase the sensitivity of TNF-sensitive or resistant cells. However, various non-specific and specific lysosomotropic agents lead to an inhibition of TNF's cytotoxic action. Differences in enzyme activity, primarily lysosomal, were noted between TNF-sensitive and resistant cells. These changes involved a general halving of lysosomal proteins and enzymes in the TNF-resistant cells. The antitumor activity of TNF does not involve specific inhibition of macromolecular synthesis but may involve alterations in lysosomes.
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Goschorska, Maja. "Investigating the mechanisms of cell competition in mammals using in vitro systems." Thesis, University of Cambridge, 2019. https://www.repository.cam.ac.uk/handle/1810/290214.
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Cell competition leads to elimination of a viable cell population, by fitter cells. Despite over forty years of research, the molecular mechanisms of competition in mammals are poorly understood. During my PhD I have investigated the mechanisms of competition by exploring an established mammalian cell culture system, in which wild-type MDCK cells eliminate scribble-deficient cells, and I have also developed a novel cell culture system to model mammalian competition. My work contributed to the discovery that scribble-deficient cells are eliminated not by biochemical exchange among cells, but by mechanical compaction. We termed this phenomenon mechanical competition. I employed transcriptional profiling to determine the molecular signature of mechanical losers, and identified activation of p53 signalling as their hallmark. My colleagues and I then demonstrated that elevation of p53 is both necessary and sufficient to trigger mechanical competition. In further investigating the mechanisms of mechanical competition, I found that compaction activates ROCK in scribble-deficient cells, and that this is required for their elimination. Inhibition of Src signalling in mechanical losers also protected them form out-competition, and integrin signalling is another pathway likely involved in mechanical competition. While investigating p53 competition, we observed that p53-high and p53-low cells engage in directional migration, with p53-high cells always at the migrating front. As a side-project, I investigated the role of p53 in directional migration, by exploring an established model with a single leader cell and multiple followers. We established a method to generate multinucleated leaders on demand. By creating leaders from p53-deficient cells, I established that p53 signalling is required for some, but not all multinucleated cells to trigger collective migration, thus implicating p53 signalling in a type of migration involved in wound healing. Finally, I successfully modelled p53-driven mechanical competition in a differentiated primary tracheal epithelial cell culture, thereby establishing a novel system to study mammalian competition, and also proving that p53 competition is conserved between different mammalian epithelia. Considering the involvement of p53, mechanical competition may play a major role in cancer.
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Gilbert, Robin Leon. "Mechanism of growth inhibition of cAMP analogues on cultured cancer cells." Thesis, University of Edinburgh, 1991. http://hdl.handle.net/1842/19789.
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Adenosine 3',5'-cyclic monophosphate (cAMP) analogues have antiproliferative effects on a range of cancer cells including breast, colon and lung. The most potent analogue, described to date, is 8-chloro cAMP but its mechanism of inhibition is poorly understood. The aim of this thesis was to investigate the mechanism of action of cAMP analogues on human cancer cell lines, in particular the hormone-responsive breast cancer cell line MCF-7. Growth inhibitory effects of 8-chloro cAMP was confirmed on MCF-7 cells. However, this effect was not present in serum-free conditions and the degree of inhibition was dependent on the concentration of foetal calf serum in the medium. This effect could also be demonstrated in the presence and absence of added oestradiol suggesting that 8-chloro cAMP does not antagonise the oestradiol stimulation of MCF-7 cells. Effects of 8-chloro cAMP were also tested on other cancer cell lines derived from breast, ovary and colon. Inhibition of cellular proliferation was again dependent on the addition of serum in the culture medium. Partial characterisation and purification of serum was attempted in order to identify the component(s) responsible for this serum-dependent 8-chloro cAMP inhibition. The component(s) was heat labile and fractions separated by gel filtration chromatography containing inhibitory activity were eluted before the major albumin peak. When medium containing 8-chloro cAMP and serum was pre-incubated, antiproliferative activity remained after removal of most of the serum components by ultrafiltration. In serum-free conditions, phosphodiesterases conferred the ability to 8-chloro cAMP to inhibit growth of MCF-7 cells; addition of a phosphodiesterase inhibitor appeared to protect the cells from the antiproliferative effect in serum containing medium. High pressure liquid chromatography of cAMP analogues incubated with serum or with gel filtration fractions detected a peak that had identical UV spectrum and a column retention time as an 8-chloro adenosine standard.
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Machado, Camila Maria Longo. "Caracterização morfologica, imunofenotipica, citogenetica e de sensibilidade ao quimioterapico cisplatina da linhagem NG97." [s.n.], 2008. http://repositorio.unicamp.br/jspui/handle/REPOSIP/316951.
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Orientador: Liana Maria Cardoso VerinaudTese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia
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Resumo: Gliomas malignos são tumores de Sistema Nervoso Central extremamente freqüentes, correspondendo a 46% dos casos diagnosticados de Neoplasias Cerebrais. Segundo a Organização Mundial de saúde o glioblastoma multiforme representa aproximadamente 3,9% dos óbitos a cada 100.000 habitantes nos Estados Unidos da América e Europa. Este tipo tumoral é extremamente agressivo e os pacientes acometidos apresentam média de vida de apenas um a dois anos após ressecção cirúrgica. Os tratamentos posteriores à retirada da massa tumoral são a quimioterapia e radioterapia. Assim, há um grande interesse na busca por novas estratégias terapêuticas que se mostrem eficazes no tratamento de pacientes acometidos por glioblastomas. Neste sentido, a utilização de linhagens celulares, estabelecidas à partir de tumores brutos extraídos de pacientes, tem-se mostrado uma ferramenta útil tanto ao avanço nos conhecimentos biológicos sobre tumores primários; quanto ao desenvolvimento de novos agentes terapêuticos. Foi estabelecida em nosso laboratório uma nova linhagem derivada de glioma humano, denominada NG97. Estudos preliminares confirmaram a origem astrocitária das células e a preservação de malignização, demonstrada pelo desenvolvimento de massa tumoral a partir do xenotransplante de células cultivadas in vitro em camundongos atímicos. O presente trabalho teve como objetivo ampliar os conhecimentos biológicos acerca desta linhagem tumoral. Assim foram atribuídas as células da linhagem ultraestruturas típicas de células indiferenciadas e tumores astrocíticos. Também, elucidaram-se características mofológicas atribuídas a linhagens tumorais astrocíticas como a expressão de proteínas GFAP, vimentina, integrina a5ß1, fibronectina, laminina e CD44. Combinada a expressão destas moléculas responsáveis pela adesão célula ao substrato, foi demonstrado que as células uma vez desprovidas do contato adesivo desenvolvem uma estrutura tridimensional organotipica. Quanto a expressao de moléculas envolvidas no processo de resposta imune das células in vivo, não foram detectadas nas células NG97: HLA I,HLAII, B7 (CD80), TGF-ß e Fas (CD95). Em cultura, as células NG97 apresentam variadas aberrações numéricas com cariótipo hiperplóide e inumeros cromossomos aneuploides classificados como um de lote cromossomos murinos associados. Este ultimo, pode ser representado por um lote próximo ao triploide murino (neartriploid, i.e.; 30 cromossomos murinos mais ou menos 11 cromossomos murinos). Finalmente demonstrou-se a sensibilidade das células NG97 ao quimioterapico Cisplatina tanto in vitro quanto in vivo. Portanto a caracterização da linhagem NG97 aqui realizada fundamenta a sua utilização como modelo biológico aos mais diversos estudos.
Abstract: Astrocytomas are extremely aggressive malignancies of the Central Nervous System (CNS) and account for 46% of all primary malignant brain tumors. The perspective for patients with malignant gliomas is poor and the average survival expectancy is less than a year. Progression to a worsened prognosis occurs in 85% of the cases due to changes in cell tumor microenvironment and through biological pathways that are still unclear. These neoplasias grow rapidly and contain high cellularity with marked hyperchromatism and pleomorphism. The hystophatological identity of glioblastoma is the prominent vascularity as well as the area of necrosis surrounded by neoplastic pseudopalisading cells, as a result of microenvironment hypoxic conditions. In order to clarify those issues, cultured cell lines are valuable tools to be employed on tumor related assays as long as they are established and well characterized. Our laboratory set up the NG97 cell line derived from a human astrocytoma grade III, which was injected in the athymic nude mice flank and turned to develop and express phenotypic characteristics of an astrocytoma grade IV. This study aimed to expand the biological knowledge about this tumor cell line. In this way, clarified morphological, immunophenotypic, and cytogenetic characteristics as well as confirmed sensitivity to cisplatin chemotherapeutic agent in cells NG97. These features attributed to NG97 cell line are unique qualities that make them an important model for the study of biology and therapeutic strategies in astrocytomas.
Doutorado
Imunologia
Doutor em Genetica e Biologia Molecular
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Clarke, Catherine Louise. "High density culture of purified populations of human mammary luminal and myoepithelial cells." Thesis, Institute of Cancer Research (University Of London), 1997. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.265842.
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Mackarel, A. Jill. "A study of polyamine efflux from human cancer cells in culture." Thesis, University of Aberdeen, 1995. http://digitool.abdn.ac.uk/R?func=search-advanced-go&find_code1=WSN&request1=AAIU539518.
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The aim of this study was to investigate the relationship between cellular growth status and polyamine efflux, to examine the functional link between acetylation of polyamines and their release from the cell, and to analyse the membrane transport mechanism involved in the polyamine efflux process. The study was based on a human colonic cancer cell line, HT115. Efflux of polyamines was monitored by radiolabelling cellular polyamines and then tracing their appearance in the culture medium. Although the predominant cellular polyamine was spermine, the polyamine almost exclusively effluxed from HT115 cells under normal conditions was N1-acetylspermidine. Actively growing cells released a small but constant amount of N1-acetylspermidine. When cell growth was retarded by serum-deprivation or inhibited by treatment with 5-FU, polyamine efflux increased. The increase in efflux corresponded to the degree of growth inhibition, with maximum release occurring from cells treated with toxic doses (10 M) of 5-FU. Efflux was predominantly of N1-acetylspermidine. Growth inhibition was reversed by re-addition of serum to the culture medium or removal of 5-FU. In response to the re-initiation of cell growth, polyamine efflux decreased. Treatment with 10 M 5-FU, in addition to causing maximum inhibition of cell growth, induced SAT, the enzyme responsible for the formation of N1-acetylspermidine. Upregulated polyamine backconversion coupled with increased efflux of the resulting N1-acetylspermidine was accompanied by a decrease in the total cellular polyamine content. Serum deprivation or doses of 5-FU that were less growth inhibitory did not induce SAT and did not decrease cellular polyamine levels.
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Yamaura, Tadayoshi. "Accurate diagnosis of mismatch repair deficiency in colorectal cancer using high-quality DNA samples from cultured stem cells." Kyoto University, 2019. http://hdl.handle.net/2433/242418.
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Thomas, Mark Peter. "Differential tolerance of a cancer and a non-cancer cell line to amino acid deprivation : mechanistic insight and clinical potential." Thesis, Stellenbosch : Stellenbosch University, 2012. http://hdl.handle.net/10019.1/19912.
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Thesis (PhD)--Stellenbosch University, 2012.ENGLISH ABSTRACT: Introduction – Due to spatial separation from the native vascular bed, solid tumours develop regions with limited access to nutrients essential for growth and survival. The promotion of a process known as macroautophagy may facilitate in the maintenance of intracellular amino acid levels, through breakdown of cytoplasmic proteins, so that they remain available for macromolecular biosynthesis and ATP production. Several studies point to the potential ability of some cancers to temporarily increase autophagy and thereby prolong cell survival during metabolic stress. The validity of these claims is assessed when a commonly used breast cancer cell line and an epithelial breast cell line are starved of amino acids in this study. Furthermore, we go on to hypothesize that acute amino acid deprivation during treatment will result in an elevated sensitivity of MDAMB231 cells to doxorubicin toxicity but limit its cytotoxic side-effects in MCF12A cells. Methods and study design- Human breast cancer cells (MDAMB231) and breast epithelial cells (MCF12A) cultured in complete growth medium were compared to those incubated in medium containing no amino acids. Steady state autophagy levels were monitored using classical protein markers of autophagy (LC3-II and beclin-1) and the acidic compartmentalization in cells (Lysotracker™ red dye) in conjunction with autophagy inhibition (bafilomycin A1 and ATG5 siRNA). Cell viability was monitored using several techniques, including caspase 3/7 activity. ATP levels were assessed using a bioluminescent assay, while mass spectrometry based proteomics was used to quantify cellular amino acid levels. Similar techniques were used to monitor autophagy during doxorubicin treatment, while cellular doxorubicin localization was monitored using immunofluorescence microscopy. Finally, a completely novel GFP-LC3 mouse tumour model was designed to assess autophagy and caspase activity within tumours in vivo, during protein limitation and doxorubicin treatment. Results - Amino acid deprivation resulted in a transient increase in autophagy at approximately 6 hours of amino acid starvation in MDAMB231 cells. The amino acid content was preserved within these cells in an autophagy-dependent manner, a phenomenon that correlated with the maintenance of ATP levels. Inhibition of autophagy during these conditions resulted in decreased amino acid and ATP levels and increased signs of cell death. MCF12A cells displayed a greater tolerance to amino acid starvation during 24 hours of amino acid starvation. Evidence indicated that autophagy was important for the maintenance of amino acid and ATP levels in these cells and helped prevent starvation-induced cell death. Furthermore, data showed that concomitant amino acid withdrawal resulted in decreased cellular acidity in MDAMB231 cells, and increased acidity in MCF12A cells, during doxorubicin treatment. These changes correlated with evidence of increased cell death in MDAMB231 cells, but a relative protection in MCF12A cells. A novel model was used to apply these techniques in vivo, and although mice fed on a low protein diet during high dose doxorubicin treatment had increased mean survival and smaller tumour sizes, evidence suggested that autophagy is protecting a population of cells within these tumours. Conclusions - This novel approach to tumour sensitization could have several implications in the context of cancer therapy, and given the delicate relationship that autophagy has with the cancer microenvironment, efforts to determine the mechanisms involved in autophagy and sensitization could lead to new and innovative treatment opportunities for cancer management.
AFRIKAANSE OPSOMMING: Inleiding – As gevolg van hul skeiding van die oorpronklike vaskulêre netwerk, ontwikkel soliede gewasse areas met beperkte toegang tot noodsaaklike voedingstowwe. Die bevordering van 'n proses wat as makro-autofagie bekend staan, kan die handhawing van intrasellulêre aminosuur vlakke fasiliteer. Voorafgenoemde proses word waarskynlik deur die afbreek van sitoplasmiese proteïene teweegebring om sodoende vir makro-molekulêre biosintese en ATP produksie beskikbaar te kan wees. Verskeie studies dui daarop dat sommige kankersoorte die vermoë het om autofagie tydelik te verhoog, en daarby sel oorlewing gedurende metaboliese stress te verleng. Die geldigheid van hierdie eise word evalueer wanneer 'n algemeen beskikbare borskanker sellyn, en 'n borsepiteelsellyn in hierdie studie van aminosure verhonger word. Verder, veronderstel ons dat akute aminosuur ontneming gedurende behandeling 'n verhoogde sensitiwiteit van MDAMB231 selle tot doxorubicin toksisiteit tot gevolg sal hê, maar terselfdetyd die middel se sitotoksiese newe-effekte in MCF12A selle sal beperk. Metodes en studie ontwerp – Menslike borskanker- (MDAMB231) en bors epiteel selle (MCF12A) wat in volledige groeimedium gekweek is, is vergelyk met selle wat in aminosuur vrye medium gekweek is. Basislyn autofagie-vlakke is gemonitor deur die gebruik van klassieke autofagie proteïen merkers (LC3-II en beclin-1) en die asidiese kompartementalisering in selle (Lysotracker™ rooi kleurstof) saam met autofagie inhibisie (bafilomycin A1 and ATG5 siRNA). Sellewensvatbaarheid is deur die gebruik van verskeie tegnieke, insluitend caspase 3/7 aktiwiteit, gemonitor. ATP-vlakke is deur die gebruik van 'n bioluminiserende tegniek gemeet, terwyl massa-spektrometrie-gebaseerde “proteomics” gebruik is om sel aminosuur vlakke te kwantifiseer. Soortgelyke tegnieke is gebruik om autofagie gedurende doxorubicin behandeling waar te neem, terwyl sellulêre doxorubicin lokalisasie deur die gebruik van immunofluoresensie mikroskopie gemonitor is. Ten slotte, is 'n unieke GFP-LC3 muismodel in hierdie studie ontwikkel. Hierdie model is gebruik om autofagie en caspase aktiwiteit in gewasse in vivo te bestudeer tydens proteïen beperking en doxorubicin behandeling. Resultate – Aminosuur ontneming het tot 'n tydelike verhoging in autofagie na ongeveer 6 ure van aminosuur verhongering in MDAMB231 selle gelei. Die aminosuur inhoud van hierdie selle het op 'n autofagie-afhanklike manier behoue gebly. Hierdie verskynsel het met die handhawing van ATP-vlakke gekorreleer. Autofagie inhibisie gedurende hierdie kondisies het 'n verlaging in aminosuur en ATP-vlakke teweeggebring, sowel as vermeerderde tekens van seldood tot gevolg gehad. MCF12A selle het 'n groter toleransie tot aminosuur verhongering tydens die 24 uur aminosuur verhongeringsperiode getoon. Getuienis het aangedui dat autofagie belangrik vir die handhawing van aminosuur en ATP-vlakke in hierdie selle was, en gehelp het om verhongerings-geïnduseerde seldood te voorkom. Verder het data gewys dat aminosuur ontrekking tot verminderde sellulêre asiditeit in MDAMB231 selle, en verhoogde asiditeit in MCF12A selle gedurende doxorubicin behandeling gelei het. Hierdie veranderinge stem ooreen met getuienis van toenemende seldood in MDAMB231 selle, maar 'n relatiewe beskerming in MCF12A selle. 'n Unieke model was gebruik om hierdie tegnieke in vivo toe te pas. Alhoewel verhoogde oorlewing en kleiner gewasse in muise op 'n lae proteïen dieet gedurende hoë dosis doxorubicin behandeling opgemerk is, het bewyse voorgestel dat autofagie 'n populasie selle binne die gewasse beskerm. Gevolgtrekkings – Hierdie unieke benadering tot tumor sensitisering kan verskeie implikasies in die konteks van kanker behandeling hê. Gegewe die delikate verhouding van autofagie met die kanker mikro-omgewing, kan pogings om die meganismes betrokke in autofagie en sensitisering te bepaal, tot nuwe en innoverende behandelings vir kanker lei.
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Fuzer, Angelina Maria. "Mecanismos celulares envolvidos na ação antiproliferativa do [10]-gingerol sobre células de tumor de mama." Universidade Federal de São Carlos, 2016. https://repositorio.ufscar.br/handle/ufscar/8761.
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Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
Cancer is a leading cause of death, according to World Health Organization, preceding diseases as diabetes and tuberculosis, conditions as malnutrition and even though interpersonal violence. Many types of cancer are also correlated to major risk factors associated to mortality and morbidity, such as obesity, alcohol abuse and smoking. In 2012 14.1 millions of new cases of cancer arisen with 8.2 million of deaths worldwide. Ginger rhizome (Zingiber officinale Roscoe) is word know use as spice on cooking and widely use as medicinal herb in preparations. Several studies describe its anti-inflammatory, antimicrobial, antioxidant and anti-emetic activities. Much of the ginger bioactivity is due to its phenolic compounds [6], [8], and [10]- gingerol, which have anti-proliferative and antiangiogenic action on tumor cells, as demonstrated by several in vivo and in vitro studies. Several studies have revealed advantages of three-dimensional culture techniques (3D) over traditional two-dimensional monolayer cultures (2D). 3D cultures better mimic the tumor microenvironment found in vivo, as well as the interaction of cells with the extracellular matrix (ECM). Three-dimensional culture has produced different responses compared to those found in 2D cultures, such as increased resistance of tumor cells to various drugs, and increased selective sensitivity to tumor cells in relation to normal cells. On this work, techniques for three-dimensional culture were used to test the effects of [10]-gingerol on breast tumor cells. The aim of this study was to evaluate the effects of [10]-gingerol in different hallmarks of malignancy correlated with metastatic process, such as adhesion, proliferation, migration, invasion and also its effects on apoptosis, both in 2D and 3D. Results demonstrated that [10]-gingerol changes the morphology of MDA-MB-231 malignant cells in lower concentrations and shorter times when compared to nonmalignant MCF-10A cells, suggesting an specific and concentration-dependent action for [10]-gingerol on malignant cells. [10]-gingerol was also able to inhibit migration and invasion of MDA-MB- 231 cells at low concentrations and to induce apoptosis at higher concentrations. We observed that [10]-gingerol presented higher IC50 in proliferation assays with MCF-10A non tumor cells compared to tumor cells. The compound also inhibited migration of non-tumor cell lines at higher concentrations compared to tumor cells. Moreover, [10]-gingerol inhibited MDA-MB- 231 cell adhesion to different ECM components, such as laminin, fibronectin and vitronectin, even at low concentrations. Western blotting and real time quantitative PCR assays suggested that [10]-gingerol was able to act by the intrinsic pathway of apoptosis, increasing Bax/Bcl-2 ratio and caspase-9 and caspase-3 mRNA and protein levels in MDA-MB-231 cells. On 3D assays the results showed selectivity of [10]-gingerol against the malignant T4-2 lineage. The compound was also able to revert the malignant phenotype and to induce apoptosis in this cell line. These results suggest that [10]-gingerol has potential to be a new anticancer drug in the future.
O câncer é uma das principais causas de morte no mundo, segundo dados da Organização Mundial da Saúde, estando à frente de doenças como o diabetes e a tuberculose, de condições como a desnutrição e até mesmo da violência interpessoal. Diversos tipos de câncer estão ainda relacionados aos principais fatores de risco associados à mortalidade e morbidade no mundo, tais como a obesidade, o consumo excessivo de álcool e o tabagismo. Em 2012 foram 14,1 milhões de casos novos de câncer com 8,2 milhões de mortes no mundo. O rizoma do gengibre (Zingiber officinale Roscoe) é utilizado no mundo todo como especiaria culinária e como erva medicinal em diversas preparações. Existem muitas pesquisas descrevendo suas propriedades antieméticas, antimicrobianas, anti-inflamatórias, antioxidantes e antitumorais. Os compostos farmacologicamente ativos do gengibre são os compostos fenólicos pungentes [6]-gingerol, [8]- gingerol e [10]-gingerol, os principais componentes do seu extrato bruto. Tais compostos são os responsáveis pela bioatividade do gengibre e por seus efeitos sobre células tumorais, demonstrados em diversos modelos de câncer, em estudos in vivo e in vitro. Muitas pesquisas revelam vantagens das técnicas de cultura tridimensional (3D) sobre as culturas tradicionais em monocamada. Culturas em 3D mimetizam melhor o microambiente tumoral encontrado in vivo, bem como a interação das células com a matriz extracelular (MEC). Em culturas 3D já foram observadas respostas diferentes das encontradas em culturas bidimensionais (2D), como a maior resistência de células tumorais a diversas drogas, além de sensibilidade seletiva aumentada para células tumorais em relação às células normais. Neste trabalho, técnicas de cultura tridimensional foram aplicadas em testes utilizando a molécula de [10]-gingerol em células tumorais de mama. O objetivo deste estudo foi avaliar os efeitos do [10]-gingerol em diferentes marcadores de malignidade, importantes no processo metastático, como adesão, proliferação, migração, invasão, bem como seus efeitos sobre a apoptose, tanto em cultura 2D, quanto 3D. Os resultados mostraram que o [10]-gingerol altera a morfologia de células tumorais da linhagem MDA-MB-231 em concentrações e tempos menores que a de células não tumorais de mama da linhagem MCF-10A, o que sugere uma ação seletiva do [10]-gingerol dependente de concentração e tempo de tratamento. O [10]-gingerol também inibiu a migração e a invasão das células da linhagem MDA-MB-231 em baixas concentrações e induziu apoptose em concentrações mais altas. Observamos que o [10]-gingerol apresentou, em ensaios de proliferação com células não tumorais da linhagem MCF-10A, um IC50 maior comparado com as células tumorais. O composto também inibiu a migração das células não tumorais em concentrações maiores comparado às células tumorais. Além disso, o [10]-gingerol diminuiu a capacidade de adesão das células MDA-MB-231 a diferentes componentes da MEC, como laminina, fibronectina e vitronectina mesmo em baixas concentrações. Ainda, confirmamos sua capacidade de causar danos ao DNA e induzir apoptose. Ensaios de western blotting e qPCR sugerem que o [10]-gingerol atua pela via intrínseca da apoptose, aumentando a razão Bax/Bcl- 2 e ativando as caspases-9 e -3. Com relação aos ensaios em 3D, os resultados demonstraram que o composto [10]-gingerol age seletivamente sobre células tumorais, sendo capaz de reverter o fenótipo maligno e induzir apoptose nas células da linhagem T4-2. Estes resultados sugerem que o [10]-gingerol tem potencial para futuramente se tornar uma nova droga antitumoral.
FAPESP: 2012/18908-6
FAPESP: 2015/08146-0
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Kawai, Takayuki. "Keratin 19, a Cancer Stem Cell Marker in Human Hepatocellular Carcinoma." Kyoto University, 2016. http://hdl.handle.net/2433/215377.
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Coleman, Catherine S. "A study of polyamine acetylation and excretion in cultured human cancer cells." Thesis, University of Aberdeen, 1990. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.315072.
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HT29/219 and HT115 human colon cancer cells were adapted to grow in medium supplemented with serum lacking amine oxidase activity. In HepG2 human hepatoma cells grown in the presence of foetal calf serum, aminoguanidine was added to inhibit extracellular polyamine oxidation. All three cell lines contained spermine as their major polyamine > spermidine > > putrescine. No acetylpolyamines were detected in HT29/219 or HT115 cells although HepG2 cells did contain N1-acetylspermidine. The total intracellular polyamine content of all cells were depleted by treatment with growth inhibitory concentrations of certain antimetabolites including the polyamine inhibitors, DFMO and MGBG. The growth inhibitory effects of DFMO were reversed by putrescine. Pretreatment of HT29/219 cells with DFMO increased the intracellular accumulation of MGBG to levels which were toxic to the cells. MGBG caused structural damage to HT29219 cell mitochondria although the cells were able to recover slowly from its growth inhibitory effects - exogenous spermidine enhanced the rate of recovery. Three spermidine acetyltransferase (SAT) activities were distinguished using an improved assay coupled with identification of products. N8-acetylspermidine was formed by nuclear and cytosolic enzymes. MGBG was the only growth inhibitor treatment to increase significantly a cytosolic spermidine N1-acetyltransferase (N1-SAT). HepG2 cells contained the highest inducible N1-SAT activity > HT115 > HT29/219 cells. Analysis of the substrate specificity of crude cytosolic extracts from HepG2 and HT29/219 cells showed that MGBG treatment caused a specific increase in the acetylation of substrates containing aminopropyl moieties. N1- and N8-acetylspermidine were identified in the extracellular medium of HT29/219 and HT115 cells. These were the major excretory products of HT115 cells but accounted for < 1% of polyamines excreted from HT29/219 cells: putrescine > > spermidine were the main polyamines excreted from HT29/219 cells although no spermine was detected. Growth inhibitory treatments increased polyamine excretion from cells mainly in the form of putrescine and spermidine. In contrast, HepG2 cells excreted spermine > N1-acetylspermidine > spermidine but no putrescine was detected. Putrescine was however excreted from HepG2 cells after treatment with MGBG. These results support the direct excretion of preformed polyamines from growing cells, a process which is increased in response to growth inhibition. While acetylation and excretion may be linked, acetylation may not be a prerequisite to excretion in all cell types.
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Arthur, Christopher Ryan. "The Substituted Pyrrole JB-03-14 Induces Autophagic Cell Death and Growth Arrest in Breast Tumor Cells." VCU Scholars Compass, 2007. http://scholarscompass.vcu.edu/etd/823.
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The use of chemotherapy in the treatment of cancer has stimulated the demand for better chemotherapeutic agents that are more potent at destroying tumor cell populations and more selective for the specific tumor versus normal host tissues. This project is directed at discovering new anti-tumor agents that are effective against breast cancer based on structures derived from marine organisms, specifically brominated pyrroles. We utilized an in vitro breast cancer model to study the effects of pyrroles on tumor proliferation and survival, as well as growth arrest and cell death. Our findings indicate that the substituted pyrrole JG-03-14 induces time dependent cell death in breast tumor cells where the cell death involves apoptosis and autophagy. Residual growth arrest in p53 wild type cells is characteristic of senescence. JG-03-14 also demonstrated substantial anti-proliferative effects in multi-drug resistant cells. These findings indicate JG-03-14 would potentially be developed for the treatment of breast cancer.
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Robinson, Clayt Austin. "Development of an in vitro three-dimensional model for colon cancer study and drug efficacy analysis." Connect to resource, 2005. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=osu1124223577.
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Thesis (Ph. D.)--Ohio State University, 2005.Title from first page of PDF file. Document formatted into pages; contains xvi, 204 p.; also includes graphics (some col.). Includes bibliographical references (p. 196-204). Available online via OhioLINK's ETD Center
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Tayyem, Hasan Mohammad. "Studies on new tumour active compounds with one or more metal centres." Faculty of Health Sciences, 2006. http://hdl.handle.net/2123/1727.
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Doctor of Philosophy(PhD)The present study deals with the synthesis, characterization, determination of anticancer activity of three mononuclear trans-planaraminepalladium(II) complexes code named TH5, TH6 and TH7 and three trinuclear complexes code named TH1, TH8 and TH14. The activity of the compounds against human cancer cell lines: A2780, A2780cisR and A2780ZD0473R, cell uptake, DNA-binding and nature of interaction with pBR322 plasmid DNA have been determined. Whereas cisplatin binds with DNA forming mainly intrastrand GG adduct that causes local bending of a DNA strand, TH5, TH6, TH7, TH1 and TH8 bind with DNA forming mainly interstrand GG adducts that causes more of a global change in DNA conformation. Although TH5, TH6 and TH7 each have two substituted pyridine ligands in a trans-geometry (3-hydroxypyridine in TH5, 2-hydroxypyridine in TH6 and 4-hydroxypyridine in TH7), the compounds differ in their activity against ovarian cancer cell lines, indicating that non-covalent interactions involving the hydroxyl group may be playing a significant role in activity of the compounds. Among trinuclear complexes TH1 is found to be significantly more active than cisplatin. It is actually twice as active as cisplatin against the parent cell line A2780, thirteen times as active as cisplatin against the cisplatin-resistant cell line A2780cisR and 11.5 times as active as cisplatin against the cell line A2780ZD0473R. Whereas the resistance factor for cisplatin as applied to the cell lines A2780 and A2780cisR cell lines is 12.9 that for TH1 is 1.98. The results suggest that TH1 has been able to significantly overcome resistance operating in A2780cisR cell line. The compound is soluble in water so that it may be taken orally. Provided it has favourable toxicity profile, TH1 has the potential to be developed into a highly active anticancer drug with a wider spectrum of activity than cisplatin. Although platinum drugs use a shot-gun approach to kill cancerous cells, widespread use in the clinic and increasing volume of their sale indicate that even in the genomic age, there is still need for shot-gun drugs in the clinic.
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Kulasinghe, Arutha Jeevana. "Circulating tumour cells in head and neck cancers." Thesis, Queensland University of Technology, 2017. https://eprints.qut.edu.au/110534/1/Arutha%20Jeevana_Kulasinghe_Thesis.pdf.
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Metastasis in head and neck cancer patients is responsible for over 50% of deaths. There are currently no tools to identify patients at risk of developing metastasis. Circulating tumour cells (CTC) represent a transient cancer cell population in the blood. In this study, the researcher has developed CTC isolation methodologies and used novel culture formulations to expand patient derived CTCs for therapy testing. Furthermore, the researcher identified biomarkers present on CTCs which could select patients for immunotherapies, a current unmet need. This work sets the foundation for a personalized medicine approach to treating head and neck cancer patients.
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Chan, Christina M. W. "A study of hormone-regulated mRNA in human breast cancer cells in culture." Thesis, University of Liverpool, 1993. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.357364.
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Wright, Muelas Marina. "A systems biology approach to cancer metabolism." Thesis, University of Manchester, 2016. https://www.research.manchester.ac.uk/portal/en/theses/a-systems-biology-approach-to-cancer-metabolism(27286c8a-0281-4256-b749-2ec9bd36370f).html.
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Cancer cells have been known for some time to have very different metabolismas compared to that of normal non proliferating cells. As metabolism is involvedin almost every aspect of cell function, there has been a recent resurgence ofinterest in inhibiting cancer metabolism as a therapeutic strategy. Inhibitors thatspecifically target altered metabolic components in cancer cells are being developedas antiproliferative agents. However, many such inhibitors have not progressedinto the clinic due to limited efficacy either in vitro or in vivo. In this study weexplore the hypothesis that this is often due to the robustness of the metabolicnetwork and the differences between individual cancer cell lines in their metaboliccharacteristics. We take a systems biology approach. We investigate the cellular bioenergetic profiles of a panel of five non-small celllung cancer cell lines before and after treatment with a novel inhibitor of theglutaminase-1 (GLS1) enzyme. Additionally, we explore the effects of this inhibitoron intracellular metabolism of these cell lines as well as on the uptake and secretionof glucose, lactate and amino acids. To be able to do the latter robustly, wehad to modify the experimental assay considerably from procedures that seemto be standard in the literature; using these earlier procedures the metabolicenvironment of the cells was highly variable, leading to misleading results onthe metabolic effects of the inhibitor. We reduced cell density, altered mediumvolume and changed the time-window of the assay. This led to the cells growingexponentially, appearing indifferent to the few remaining changes. In this newassay, the metabolic effects of the glutaminase inhibitor became robust. One of the most significant results of this study is the metabolic heterogeneitydisplayed across the cell line panel under basal conditions. Differences in themetabolic functioning of the cell lines were observed in terms of both theirbioenergetic and metabolic profile. The amount of respiration attributed tooxidative phosphorylation differed between cell lines and respiratory capacity wasattenuated in most cells. However, the rate of glycolysis was similar betweencell lines in this assay. These results suggest that the Warburg effect arisesthrough a greater diversity of mechanisms than traditionally assumed, involvingvarious combinations of changes in the expression of glycolytic and mitochondrialmetabolic enzymes. The effects of GLS1 inhibition on cellular bioenergetics and metabolism alsodiffered between cell lines, even between resistant cell lines, indicating that theremay also be a diversity of resistance mechanisms. The metabolomic response ofcell lines to treatment suggests potential resistance mechanisms through metabolicadaptation or through the prior differences in the metabolic function of resistantcell lines. Part of the metabolome response to GLS1 inhibition was quite specificfor sensitive cells, with high concentrations of IMP as the strongest marker. Our results suggest that the metabolome is a significant player in what determinesthe response of cells to metabolic inhibitors, that its responses differ between cancercells, that responses are not beyond systems understanding, and that thereforethe metabolome should be taken into account in the design of and therapy withanti-cancer drugs.
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Kaminski, Bettina [Verfasser]. "Chemopreventive and sensitizing effects of phytochemicals in a cell culture model of colorectal cancer / Bettina Kaminski." Gießen : Universitätsbibliothek, 2011. http://d-nb.info/1061195813/34.
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Christakou, Athanasia. "Ultrasound-assisted Interactions of Natural Killer Cells with Cancer Cells and Solid Tumors." Doctoral thesis, KTH, Biomedicinsk fysik och röntgenfysik, 2014. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-158522.
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In this Thesis, we have developed a microtechnology-based method for culturing and visualizing high numbers of individual cells and cell-cell interactions over extended periods of time. The foundation of the device is a silicon-glass multiwell microplate (also referred as microchip) directly compatible with fluorescence microscopy. The initial microchip design involved thousands of square wells of sizes up to 80 µm, for screening large numbers of cell-cell interactions at the single cell level. Biocompatibility and confinement tests proved the feasibility of the idea, and further investigation showed the conservation of immune cellular processes within the wells. Although the system is very reliable for screening, limitations related to synchronization of the interaction events, and the inability to maintain conjugations for long time periods, led to the development of a novel ultrasonic manipulation multiwell microdevice. The main components of the ultrasonic device is a 100-well silicon-glass microchip and an ultrasonic transducer. The transducer is used for ultrasonic actuation on the chip with a frequency causing half-wave resonances in each of the wells (2.0-2.5 MHz for wells with sizes 300-350 µm). Therefore, cells in suspension are directed by acoustic radiation forces towards a pressure node formed in the center of each well. This method allows simultaneous aggregation of cells in all wells and sustains cells confined within a small area for long time periods (even up to several days). The biological target of investigation in this Thesis is the natural killer (NK) cells and their functional properties. NK cells belong to the lymphatic group and they are important factors for host defense and immune regulation. They are characterized by the ability to interact with virus infected cells and cancer cells upon contact, and under suitable conditions they can induce target cell death. We have utilized the ultrasonic microdevice to induce NK-target cell interactions at the single cell level. Our results confirm a heterogeneity within IL-2 activated NK cell populations, with some cells being inactive, while others are capable to kill quickly and in a consecutive manner. Furthermore, we have integrated the ultrasonic microdevice in a temperature regulation system that allows to actuate with high-voltage ultrasound, but still sustain the cell physiological temperature. Using this system we have been able to induce formation of up to 100 solid tumors (HepG2 cells) in parallel without using surface modification or hydrogels. Finally, we used the tumors as targets for investigating NK cells ability to infiltrate and kill solid tumors. To summarize, a method is presented for investigating individual NK cell behavior against target cells and solid tumors. Although we have utilized our technique to investigate NK cells, there is no limitation of the target of investigation. In the future, the device could be used for any type of cells where interactions at the single cell level can reveal critical information, but also to form solid tumors of primary cancer cells for toxicology studies.QC 20150113
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Wan, Xiao. "Development of advanced three-dimensional tumour models for anti-cancer drug testing." Thesis, University of Oxford, 2014. http://ora.ox.ac.uk/objects/uuid:5342fe46-c676-4fe8-8b6e-96d17a18d17d.
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Animal testing is still the common method to test the efficacy of new drugs, but tissue engineered in vitro models are becoming more acceptable for replacing and reducing animal testing in anti-cancer drug screening by developing in vitro three-dimensional (3D) tumour models for anti-cancer drug testing. In this study, three-dimensional (3D) culture methods were developed to mimic the tumour microenvironment. 3D culturing is to seed, maintain and expand cultured cells in three-dimensional space, in contrast to the traditional two-dimensional (2D) method in which the cells attach to the bottom of culture containers as monolayers. To mimic the intercellular interplay for tumour study, cell co-culture was applied. In this thesis, perfusion culture showed a better homeostasis for 3D tumour model growth over 17 days, with a more controllable working platform and a more reliable response-dose correlation for data interpretation. In the Matrigel sandwich system, the co-culture of breast cancer cells and endothelial cells demonstrated the morphology featuring a vascular network and tumour structures, with the thickness of the three-dimensional structure around 100µm and tubule length 200-400 µm, and maintained for 10 days. The comparisons studies between Matrigel sandwich and other methods suggest that though not fully characterised, Matrigel is still a valuable scaffold choice for developing co-culture 3D tumour model. Finally, the combination of perfusion and co-culture showed the potential of applying this model in angiogenesis assay, with a drug response profile combining cell viability and morphology to mimic in vivo tumour physiology.
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Chen, Chao [Verfasser]. "Enrichment of cancer stem cells from head and neck cancer by anchorage independent culture and related research / Chao Chen." Berlin : Medizinische Fakultät Charité - Universitätsmedizin Berlin, 2015. http://d-nb.info/1068208856/34.
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De, Brito Galvao Joao Felipe. "Antitumor effects of combined carboplatin and gemcitabine in canine transitional cell carcinoma." The Ohio State University, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=osu1305257807.
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Jardim-Perassi, Bruna Victorasso. "Avaliação da glutationa e suas enzimas como marcadores prognósticos e preditivos do câncer de mama /." São José do Rio Preto : [s.n.], 2011. http://hdl.handle.net/11449/92529.
Full textAbstract:
Orientador: Débora Aparecida Pires de Campos ZuccariBanca: Dorotéia Rossi Silva
Banca: Maria Tercília Vilela de Azeredo Oliveira
Resumo: O estudo de marcadores prognósticos e preditivos no câncer tem se mostrado efetivo na pesquisa e rotina diagnóstica. A glutationa (GSH) e as enzimas glutationa peroxidase (GPX) e glutationa S transferase pi (GSTpi) exercem papel fundamental na defesa antioxidante das células e na detoxificação de quimioterápicos. Nesse contexto, o objetivo deste estudo foi avaliar a expressão das proteínas GSH, GPX e GSTpi em pacientes com câncer de mama, além de avaliar a expressão gênica dessas proteínas em amostras tumorais in vitro após o tratamento com quimioterápico. As proteínas foram detectadas no tecido tumoral de 63 pacientes por imuno-histoquímica e quantificadas pela técnica de densitometria óptica. A expressão dos genes que sintetizam GSH, glutamato cisteina ligase (GCLC) e glutationa sintetase (GSS) e dos genes codificadores da GPX e GSTpi foi analisada por PCR em tempo real em células cultivadas provenientes de 12 amostras tumorais de mama. As células foram submetidas in vitro ao quimioterápico doxorrubicina, e a expressão gênica foi analisada antes e após o tratamento. A expressão da GSH relacionou-se com tumor receptor de estrógeno (RE) negativo (p<0,05). A expressão da GPX foi maior em tumor receptor de progesterona (RP) negativo e em pacientes que vieram a óbito (p<0,05). Alta expressão da GSTpi relacionou-se com características tumorais de prognóstico desfavorável como positividade para p53, grau histológico III, maior tamanho tumoral e óbito (p<0,05). Além disso, as pacientes foram divididas em subgrupos de acordo com o tratamento recebido. Assim, a alta expressão da GSH relacionou-se com a ocorrência de metástase no grupo de pacientes tratadas apenas com quimioterapia adjuvante (p<0,05). Nas pacientes que receberam quimioterapia e radioterapia adjuvantes, a alta expressão da GPX foi relacionada com óbito e a alta expressão da GSTpi... (Resumo completo, clicar acesso eletrônico abaixo)
Abstract: The study of prognostic and predictive markers in cancer has been proven effective in research and diagnostic routine. Glutathione (GSH) and glutathione peroxidase (GPX) and glutathione S transferase pi (GSTpi) play a crucial role in antioxidant defense of cells and detoxification of chemotherapeutic agents. In this context, the objective of this study was to evaluate the protein expression of GSH, GPX and GSTpi in patients with invasive ductal carcinoma, and to evaluate the expression of genes encoding these proteins in tumor samples in vitro before and after treatment with chemotherapy. The proteins were detected in tumor tissue of 63 patients by immunohistochemistry and quantified by optical densitometry technique. The expression of genes that synthesize GSH, glutamate cysteine ligase (GCLC) and glutathione synthetase (GSS) and the genes encoding GPX and GSTpi were analyzed by real time PCR in cultured cells from 12 tumor samples from patients with breast cancer. The cells were treated in vitro to doxorubicin chemotherapy, and gene expression was analyzed before and after treatment. The expression of GSH was related to tumor estrogen receptor (ER) negative (p <0.05). The expression of GPX was higher in tumor progesterone receptor (PR) negative and patients who died (p <0.05). High expression of GSTpi was related to tumor characteristics of poor prognosis such as p53 positivity, histologic grade III, larger tumor size and death (p <0.05). In addition, patients were divided into subgroups according to treatment received. Thus, high expression of GSH was related to the occurrence of metastasis in patients treated only with adjuvant chemotherapy (p <0.05). In patients who received adjuvant chemotherapy and radiotherapy, high expression of GPX was associated with death and high expression of GSTpi was correlated to local tumor recurrence, metastasis and death (p <0.05)... (Complete abstract click electronic access below)
Mestre
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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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Dean, Zachary S. "Collective Migration Models: Dynamic Monitoring of Leader Cells in Migratory/Invasive Disease Processes." Diss., The University of Arizona, 2015. http://hdl.handle.net/10150/560817.
Full textAbstract:
Leader cells are a fundamental biological process that have only been investigated since the early 2000s. These cells have often been observed emerging at the edge of an artificial wound in 2D epithelial cell collective invasion, created with either a mechanical scrape from a pipette tip or from the removal of a plastic, physical blocker. During migration, the moving cells maintain cell-cell contacts, an important quality of collective migration; the leader cells originate from either the first or the second row, they increase in size compared to other cells, and they establish ruffled lamellipodia. Recent studies in 3D have also shown that cells emerging from an invading collective group that also exhibit leader-like properties. Exactly how leader cells influence and interact with follower cells as well as other cells types during collective migration, however, is another matter, and is a subject of intense investigation between many different labs and researchers. The majority of leader cell research to date has involved epithelial cells, but as collective migration is implicated in many different pathogenic diseases, such as cancer and wound healing, a better understanding of leader cells in many cell types and environments will allow significant improvement to therapies and treatments for a wide variety of disease processes. In fact, more recent studies on collective migration and invasion have broadened the field to include other cell types, including mesenchymal cancer cells and fibroblasts. However, the proper technology for picking out dynamic, single cells within a moving and changing cell population over time has severely limited previous investigation into leader cell formation and influence over other cells. In line with these previous studies, we not only bring new technology capable of dynamically monitoring leader cell formation, but we propose that leader cell behavior is more than just an epithelial process, and that it is a critical physiological process in multiple cell types and diseases.