Human pluripotent stem cells (hPSCs) have great potential in regenerative medicine, disease modeling, and drug screening applications due to their capacity for indefinite self-renewal and their ability to differentiate to any somatic cell type. They are especially promising for applications in the study and treatment of central nervous system (CNS) disorders and injuries, for which there is a lack of expandable populations of human cells for transplantation and in vitro models. However, a key

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... mitation is that most studies of hPSCs have been carried out in two-dimensional (2-D) culture systems, which fail to recapitulate the three-dimensional (3-D) microenvironment that these cells inhabit in vivo during embryonic development. The design of 3-D systems for hPSCs could improve our understanding of the role of the microenvironment in directing hPSC self-renewal and neuronal commitment. In this thesis, we engineered 3-D microenvironments for hPSCs by electrospinning synthetic polymers into scaffolds with micro-scale geometries, and examined their ability to direct hPSC self-renewal and conversion to neuronal cells. This thesis has two key aims. The first is to develop methods to adapt undifferentiated hPSCs to synthetic microenvironments and to examine the conditions required to maintain hPSC self-renewal under defined and synthetic substrate conditions. We have identified that 3-D scaffold geometry is a critical parameter that influences hPSC survival and self-renewal in fully synthetic substrates. The second aim was to reprogram hPSCs to "induced" neuronal cells by overexpressing neuronal transcription factors and to examine the neuronal conversion and maturation of these cells within 3-D scaffolds. We identified that enriched populations of reprogrammed human neuronal cells can be generated in 3-D, and that the fiber architecture can be tuned to accelerate neuronal maturation and functionality. Notably, we found that fibrous scaffolds with geometries that permit cellular infiltration and high levels [...]