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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
1

Influence of substrate topography and materials on behaviour of biological cells

Murray, Lynn Michelle January 2012 (has links)
A cell’s interaction with its extracellular environment is critical to tissue structure and function. This work investigates the effect of substrate topography on selective cell adhesion and morphology. Alterations in cell response to micro- and nanoscale signals and cues can cause changes in downstream functions of proteins and complexes such as invasive and metastatic motility of malignant tumour cells and the differentiation direction of stem cells. Biomaterial surfaces can be modified to provide different chemical and topographical cues and encourage controlled cell-substrate interaction. At the protein level, template substrates have shown and increased affinity for selective binding of the imprinted antigen or antibody. Topography of a cell’s microenvironment may be replicated as a permanent polymer mould by bioimprinting technology, which was developed at University of Canterbury. The resulting high resolution methacrylate polymer samples have been used for imaging and analysis, but have not previously been investigated as cell culture substrates. This work investigates the effect of bioimprint and photolithographic substrate patterning on cell behaviour in culture. Optimisation of a methacrylate co-polymer resulted in a 6:3:1 ethylene glycol dimethacrylate: methacrylic acid: photoinitiator polymer mixture cured by 240 seconds of UV exposure. The polymer was used to replicate cell membrane features into a permanent polymer mould [a bioimprint]. The resulting high resolution methacrylate bioimprints were cleaned and sterilised for use as a secondary cell culture substrate. Ishikawa endometrial cancer cells were cultured on bioimprinted methacrylate polymer substrates. Preliminary results showed preferential cell adhesion to bioimprinted areas over flat areas and also showed three dimensional spheroid growth instead of lateral two dimensional monolayer spreading. At higher seeding densities, preferential adhesion was similarly noted as well as peeling artefacts of shear stresses and cell size variation on flat methacrylate substrate regions. Fluorescent imaging and cell culture stencilling highlighted the association of secondary cells with bioimprint substrate features. To determine whether preferential cell adhesion effects were due to bioimprint features or general topography modification, secondary cancer cells were cultured on comparable photolithographically-defined, geometrically-patterned substrates. Methods for transferring regular pattern arrays into methacrylate polymer substrates were developed. No organisation or preferential adhesion effects were observed in association with pillar and hole patterns between 5-30 µm. However, artefact incidence in methacrylate polymer replication techniques led to development and adaptation of polystyrene patterning techniques. Experimental analysis of substrate-dependent effects on cell culture adhesion and organisation was extended to a non-cancerous cell line model. C2C12 mouse skeletal muscle cells were chosen for these investigations because of their ability to differentiate further, into myocytes or myofibrils. C2C12 myoblasts seeded on common cell culture substrates showed a notable morphology variation and extent of differentiation between cells grown on tissue culture polystyrene [TCPS] and polydimethylsiloxane [PDMS]. Myoblasts were plated on geometrically-patterned polystyrene and PDMS substrates. Significant alignment to grated pattern features was observed on both substrate types, before and after driven differentiation. Peeling artefacts of confluent tissue-like culture from PDMS surfaces which were observed were unreported previously in literature. The results reported in this thesis provide a foundation for potential research and commercial application for surface modification methods. The biomimetic topography provided by bioimprinted substrates can be used to identify and investigate cell activities, including for example the mechanisms of cell adhesion and separation in metastatic and invasive cancer research. Altering the material of the bioimprinted substrates may attune substrate topographies as scaffolds to direct specific stem cell differentiation for regenerative tissue engineering applications.
2

The interaction of healthy and cancerous cells with nano- and microtopography

Davidson, Patricia 28 June 2011 (has links) (PDF)
This thesis deals with the differential response of healthy and cancerous cells to surface topography at the nanoscale and the microscale. Using a statistical method we developed we studied the interactions of cells with grooves of nanoscale depth. We demonstrate that healthy cells have a greater ability to align with deeper grooves, whereas cancerous cells are more sensitive to shallow grooves. Analysis reveals that the nucleus follows the alignment of the cell body more closely in cancerous cells, and that the nucleus of cancerous cells is more sensitive to shallow grooves.On microscale pillars we demonstrate for the first time that osteosarcoma cells deform to adopt the surface topography and that the deformation extends to the interior of the cell and in particular to the nucleus. We show that healthy cells only deform during the initial stages of adhesion and that immortalized cells show intermediate deformation between the healthy and cancerous cells. When the spacing between the pillars is reduced, differences in the deformation of different cancerous cell lines are detected. Deformation was also found to be related to the malignancy in keratinocytes, and related to the expression of Cdx2 in adenocarcinoma. The mechanism of deformation is tentatively attributed to the cytoskeleton and attempts to identify the main actors of deformation were performed using confocal microscopy and cytoskeleton inhibitors. Live cell imaging experiments reveal that the deformed cells are very mobile on the surfaces, loss of deformation is necessary for mitosis to occur and deformation after mitosis is more rapid than initial deformation upon adhesion to surfaces.
3

The interaction of healthy and cancerous cells with nano- and microtopography / L'interaction de cellules saines et cancéreuses avec la micro et la nanotopographie de surface

Davidson, Patricia 28 June 2011 (has links)
L'objet de cette thèse est l'étude comparative de la réponse de cellules saines et malignes à la micro- et la nano-topographie de surface. L'interaction avec des stries de profondeur nanométrique est étudiée grâce à une méthode statistique. Nous démontrons que les cellules saines s'alignent plutôt sur des stries profondes, et que les cellules cancéreuses sont plus sensibles aux stries peu profondes. L'analyse des noyaux révèle qu’ils suivent l'alignement des corps cellulaires plus fidèlement dans le cas des cellules cancéreuses et que les noyaux de ces dernières sont plus sensibles aux stries de faible profondeur. Sur des micro-piliers nous démontrons que les cellules d’ostéosarcomes sont capables de se déformer et de faire adopter à leurs noyaux la forme de l'espace entre les piliers. Ceci ne se produit que durant la phase initiale d'adhésion pour les cellules saines. Les cellules immortalisées présentent un niveau intermédiaire de déformation. Quand l'espacement entre piliers est réduit, des différences de déformation sont révélées entre les lignées cancéreuses testées. La déformation est aussi liée au caractère cancéreux de kératinocytes et à l'expression de Cdx2 dans des lignées d'adénocarcinomes. Nous avons tenté d'expliquer ce mécanisme de déformation en l'attribuant au cytosquelette grâce à des analyses en microscopie confocale et avec des inhibiteurs du cytosquelette. L'imagerie de cellules vivantes a permis d'observer que les cellules sont très mobiles même quand elles sont déformées, que la mitose nécessite la perte de la déformation et que la déformation après mitose est plus rapide que la déformation pendant l'adhésion initiale des cellules. / This thesis deals with the differential response of healthy and cancerous cells to surface topography at the nanoscale and the microscale. Using a statistical method we developed we studied the interactions of cells with grooves of nanoscale depth. We demonstrate that healthy cells have a greater ability to align with deeper grooves, whereas cancerous cells are more sensitive to shallow grooves. Analysis reveals that the nucleus follows the alignment of the cell body more closely in cancerous cells, and that the nucleus of cancerous cells is more sensitive to shallow grooves.On microscale pillars we demonstrate for the first time that osteosarcoma cells deform to adopt the surface topography and that the deformation extends to the interior of the cell and in particular to the nucleus. We show that healthy cells only deform during the initial stages of adhesion and that immortalized cells show intermediate deformation between the healthy and cancerous cells. When the spacing between the pillars is reduced, differences in the deformation of different cancerous cell lines are detected. Deformation was also found to be related to the malignancy in keratinocytes, and related to the expression of Cdx2 in adenocarcinoma. The mechanism of deformation is tentatively attributed to the cytoskeleton and attempts to identify the main actors of deformation were performed using confocal microscopy and cytoskeleton inhibitors. Live cell imaging experiments reveal that the deformed cells are very mobile on the surfaces, loss of deformation is necessary for mitosis to occur and deformation after mitosis is more rapid than initial deformation upon adhesion to surfaces.

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