Einfluss von Mikro- und Nanostrukturen auf Zellwachstum und -migration

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LEHNERT, Dirk, 2003. Einfluss von Mikro- und Nanostrukturen auf Zellwachstum und -migration [Dissertation]. Konstanz: University of Konstanz

@phdthesis{Lehnert2003Einfl-8044, title={Einfluss von Mikro- und Nanostrukturen auf Zellwachstum und -migration}, year={2003}, author={Lehnert, Dirk}, address={Konstanz}, school={Universität Konstanz} }

deu 2011-03-24T17:39:29Z terms-of-use Lehnert, Dirk Influence of micro- and nanostructures on cell adhesion and migration application/pdf 2011-03-24T17:39:29Z 2003 Einfluss von Mikro- und Nanostrukturen auf Zellwachstum und -migration Cell adhesion and migration play a fundamental role during development and maintenance of multicellular organisms. Cells and their behavior have to be regulated amongst other things by the extracellular matrix (ECM) to perform their tasks reliably. Any disruption of this regulation leads to malfunctions and abnormal development. In the presented work, the influence of ECM-geometry on adhesive cells was investigated. Structured substrates used in this study consist of a multitude of small, squared structures covered with adhesive ECM-protein which are embedded in an anti-adhesive surface of several mm² in size. The size of these regularly arranged protein-covered islands ( dots ) was varied between 12 µm² and 0.1 µm², the distance between dots from 1 µm to 30 µm. Using different combinations of these geometries, various in vivo ECM-structures like e.g. basal lamina or mesenchyme tissue were imitated and projected into two dimensions.<br />To produce the microstructured substrates, the technique of microcontact printing (µCP) was used and advanced. The smallest reproducable protein-covered structures are 0.1 µm² in size. The side lengths of these dots are 300 nm and therefore near the physical limits of optical resolution. In the present study, the full potential of microcontact printing could be exploited for the first time.<br />Three different fibroblast-like cell lines were plated on microstructured subtrates and their behavior during the first hour after plating was analysed. Cells spread over several ECM-dots and form functional cell-matrix-contacts restricted to the positions of dots. In addition, they show a rectangular morphology as well as straight edges, strictly orientating along the underlying dot-pattern. Variation of the patterns allowed presentation of different surface-coverages with ECM-protein and quantitative analysis of geometrical limits of cell adhesion, cell spreading and migration. It turned out, that the number of adherent cells ist positively correlated with the presented amount of protein and that optimal cell adhesion is achieved at 20% surface coverage with ECM-protein. Another positive correlation was found between cell size and ECM-protein surface coverage with a coverage of 15% being necessary for optimal cell spreading. Astonishingly, on the tested subtrate patterns, these effects are correlated to the surface coverage and not to the geometric pattern of the substrate. Since surface coverage is a function of dot sizes and distances, the influence of anti-adhesive distances on cell spreading was investigated. It turned out that all three tested cell lines can bridge distances of 25 µm. Variation of dot sizes showed, that dots of 0.1 µm² are able to induce intracellular signaling. Furthermore, they support cell spreading when distances between dots are smaller than 4 µm. At larger distances, these small dots are removed from the substrate and internalized into the cells. Cells use the complete surface of dots smaller than 3 µm² for contact formation. Dots of 12 µm² are only used partially and especially at the edges of the dots.<br />These results let to new models of cell-matrix-interactions. Furthermore, using the advanced technique of substrate patterning by microcontact printing, existing hypotheses can be experimentally tested more precisely now. This will result in a better understanding of cellular behavior, wich can contribute e.g. to surface optimisation of medical implants. Lehnert, Dirk

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