Vorschau

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Abstract

The major aim of this thesis is to create a novel in vitro model substrate for the controlled cell differentiation in pluripotent tissue sheets by fine-tuning the balance between cell-cell (in plane) and cell-matrix (out of plane) interactions. As the model substrate to culture tissue explants, planar lipid membranes deposited on solid substrates (supported membranes) were functionalized with the extracellular domain of Xenopus cadherin-11 (Xcad-11), expressed in embryos during the neural crest cell (NCC) differentiation. In Chapter 4, the quantitative functionalization of supported membranes with Xcad-11 was confirmed by the combination of specular X-ray reflectivity (XRR), grazing incidence X-ray fluorescence (GIXF), and quartz crystal microbalance with dissipation (QCM-D). In Chapter 5, "animal cap" tissue sheets isolated from blastula stage embryos of Xenopus laevis were placed on membranes functionalized with Xcad-11.Interactions of the isolated tissue with membranes sheets displaying Xcad-11 were investigated by the combination of reflection interference contrast microscopy (RICM) and fluorescence microscopy. The capability of such a model system to induce the NCC differentiation was demonstrated by the activation of NCC marker gene marker slug by enhanced green fluorescent signal of the fusion protein. In Chapter 6, the two opposite concentration gradients of two recombinant proteins was established by means of "membrane electrophoresis", utilizing the fluidic nature of supported membranes. This allows for the creation of one-dimensional gradients of morphogens (e.g. Wnt and BMP) on supported membranes, which can guide the development of three-dimensional tissues in vitro.