Chemically Defined Platforms for Culturing Pluripotent Stem Cells

Pluripotent stem cells (PSCs) such as human embryonic stem cell (hESCs) and induced pluripotent stem cells (iPSCs) are of interest based on their unique ability to self renew indefinitely and to differentiate into any mature cell type in the human body. In addition to exposing stem cells to potential xenogenic contaminants, standard PSC culturing practices notoriously display heterogeneous PSC growth and variable differentiation. These limitations represent a significant barrier towards successfully bringing regenerative medicine approaches into mainstream practice. The research described in this dissertation addresses these concerns through the development of chemically defined stem cell culturing environments utilizing synthetic materials.;The first part of this dissertation examines how the physical and chemical properties of cell culture surfaces can influence the formation of common intermediates of PSC differentiation known as embryoid bodies (EBs). Methyl terminated materials with hydrophobic surfaces were identified to enrich for populations of EBs within a size range that displayed higher cellular viability, a lower degree of cell death, and enhanced differentiation potential. In addition, the interactions between hydrophobic substrates and components of cell culture media used for EB formation were examined to suggest a mechanism for the observed improvements in EB mediated differentiation. These studies traced the evolution of a model hydrophobic surface as it becomes both superhydrophilic and an effective material for use in the suspension culture of EBs.;The second theme to this research examines the design of a synthetic materials platform for improving the environment in which colonies of self-renewing PSCs are grown and maintained prior to the induction of differentiation. Spatial patterns of self-assembled monolayer (SAM) materials are fabricated with microcontact printing (μCP) techniques to sequester the growth of hESCs within spatial patterns with defined chemistries. This approach achieves unprecedented spatial control over stem cell growth and standardizes parameters such as colony size and shape. In addition, hESCs grown on these materials remain genetically stable and retain their pluripotency over multiple passages. Ultimately, this research introduces a robust and scalable stem cell culturing tool that provides a chemically defined alternative for the long-term maintenance of self-renewing PSC populations.