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Abstract

Stem cells are undifferentiated building blocks of life that can self-renew and differentiate, which is distinct from specialized cells with limited division potentials. It is well-established that stem cell functions in vivo are tightly regulated by biochemical and physical cues from their surrounding microenvironments called stem cell niche. Recently, increasing numbers of studies have shown that the stem cell niche is never uniform or static, suggesting a strong demand of in vitro platforms possessing mechanical anisotropy and dynamically tunable elasticity. The primary aim of this thesis is the design and fabrication of microstructured polymer materials with non-conventional mechanical properties for regulation of human mesenchymal stem cells (hMSCs) derived from bone marrow. In Section 5, a new class of mechanical metamaterials for controlling cell behavior, called bio-metamaterials, was designed and fabricated by two-photon 3D laser printing. This approach allowed the fabrication of materials with anisotropic mechanical properties, such as Young’s modulus and Poisson’s ratio, by the rational design and arrangement of unit cells. Live-cell imaging and immunohistochemical staining demonstrated that the shape of hMSCs and their order of focal adhesions and cytoskeletons can be mechanically controlled by bio-metamaterials. Thereby it is important that the unit cells are smaller than the cell size and that they are deformable by cellular forces. It is notable that hMSCs did not show any differential response when the same metamaterial structures were made out of stiff base materials, which clearly validated the materials design strategy. In Section 6, dynamic 3D stem cell microenvironments for hMSCs were designed and fabricated by the combination of stimulus responsive, supramolecular hydrogels and 3D printed molds. To functionalize not only the top surface but also the side walls and bottoms of 3D microstructures, N-hydroxysuccinimde groups were integrated into the hydrogels. This enabled a one-step uniform functionalization of all surfaces with various extracellular matrix proteins. Confocal microscopy images of hMSCs in microcavities demonstrated that the dynamic mechanical environments could be switched through modulation of the density of non covalent supramolecular crosslinks by adding/removing small additives. In Section 7, a new photosensitive monomer was synthesized and integrated into supramolecular hydrogels. The introduction of photoswitches in polymer-based stem cell niches is of high interest, because the local stiffening and softening of materials can be realized on demand by illuminating any position of interest. To overcome the phototoxicity of near UV light (λ ≈ 350 - 365 nm) widely used for photoswitching, bulky and electron-rich methoxy groups were coupled to all four ortho positions of azobenzene, which minimized the damage to cells by isomerization at 470 nm and 620 nm. The reversible switching of material stiffness (Young’s modulus) was confirmed by nanoindentation by atomic force microscopy, and the cellular response was monitored by phase contrast imaging. The obtained results demonstrate that the combination of advanced 3D printing technology, new surface chemistry and photoswitchable monomers is a promising strategy towards the mechanical control of stem cells, both, in space and time.