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Biomechanical Regulation of Cell Rearrangement and Fate Patterning Under Geometrical Confinement

Geometrical confinement or micropatterning techniques have been widely used to investigate cell migration, chirality, polarity, epithelial-mesenchymal transition, and stem cell differentiation with a potential for high-throughput screening. In this dissertation, geometrical confinement techniques are employed to study the biomechanical mechanisms in cell rearrangement and spatial patterning of embryonic cell fates. In chapter 2, I find that both cell contractility and actin gradient contributed to the radial alignment of rat embryonic fibroblasts. Combined with a Voronoi-cell model developed by our collaborator, our results demonstrate that the combined global tissue prestretch and differential cell stiffness between the inner and boundary cells can sufficiently lead to radial alignment. In chapter 3, I demonstrate that human pluripotent stem cells can self-organize to concentric rings of all major cell types in human ectoderm when cultured on micropatterned surfaces in a chemically defined condition. I reveal that modulating the dynamics of NODAL, BMP, and WNT signals is sufficient to control the spatial order of different cell types. The mathematical model developed by our collaborator suggests that changes in wavelength and phase of signaling patterns formed via reaction-diffusion may be the mechanism by which temporal information is translated into spatial information. In chapter 4, I generate an early human midbrain and hindbrain tissue with anteroposterior (AP) patterning using human pluripotent stem cells that are induced with BMP, WNT, RA, and SHH signals under fully defined culture conditions. I find that the cells self-organize into spatially patterned midbrain (OTX2+) and hindbrain (HOXB4+) progenitors after 6 days of induction. To investigate the mechanism of AP cell fates patterning, I find that SHH is not required in midbrain and hindbrain patterning while the reaction-diffusion of BMP/Noggin plays a critical role in AP regionalization. Drug treatment experiments show that valproic acid inhibits both midbrain and hindbrain development while isotretinoin disrupts AP patterning of the midbrain and hindbrain. In summary, I have employed geometrical confinement techniques to study various cell behaviors and demonstrated that geometrically confined cell-based models have great potential in fundamental research of various cell behaviors and applications in disease modeling and drug discovery.
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