The Impact of sub-µm Surface Topography on Pluripotent Stem Cells

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Induced pluripotent stem cells (iPSCs) can differentiate into every cell type of the human body, but little is known about how specific surface patterns impact on pluripotent state or guide lineage specific cellular differentiation. We have recently used multi-beam laser technology to generate groove-ridge structures in polyimide with a periodicity in the submicrometer range that induce elongation of iPSC colonies, guide the orientation of apical actin fibers, and direct the polarity of cell division. In continuation we want to explore the possibility of using this technology to modulate the function of iPSCs with the following specific aims: (1) Tailored sub-micrometer structured biomaterials. We will investigate how multi-beam interference can be applied to generate more complex and homogeneous surface patterns. To this end ultrashort pulsed lasers and modified optical setups will be investigated for the generation of suitable surface textures on different materials with minimized material degradation, including polystyrene (PS) – that could also be applied to structure conventional PS tissue culture plates. (2) Impact of surface patterns on pluripotency. We will further determine how morphology, motility, cell divisions and spatial heterogeneity of iPSC colonies are affected by sub-micrometer patterns. Furthermore, effects on reprogramming efficiency, cytoskeletal organization, and gene expression profiles will be analyzed. (3) Impact of topography on differentiation of iPSCs. We will follow the hypothesis that sub-µm structures can influence lineage-specific differentiation. To this end, we will treat iPSCs with morphogens to study early differentiation events and spatial reorganization within iPSC colonies, and we will induce unbiased multi-lineage differentiation or lineage-specific differentiation on substrates. (4) The role of YAP/TAZ in recognizing topographic cues. The YAP/TAZ pathway plays a central role in mechanotransduction. We will modulate expression of YAP and TAZ with CRISPR-Cas9n technology to determine the sequel on cellular response to surface topography. Furthermore, the impact on relevant signal cascades will be addressed by single-cell RNA-sequencing. This proposal combines expertise of laser technology and stem cell research. It provides new perspectives to unravel effects of surface topography on pluripotent stem cells, which may ultimately support directed differentiation for regenerative medicine and drug screening.