Engineering Novel Materials for Stem Cell Culture and Expansion

Stem cells are a unique class of cells residing in the body which can replicate indefinitely and develop into more specialized types of cells, a process known as differentiation. Specifically, pluipotent stem cells have the ability to turn into all types of cells found in the body. Embryonic stem cells (ESCs) are the only natural pluripotent stem cell, however much progress in the last decade has expanded the potential of generating induced pluripotent stem cells (iPSCs) from cells as basic as human skin cells. Stem cells, especially iPSCs, could be utilized for many potential therapeutic purposes including tissue engineering, cell replacement therapies, disease modeling, and drug screening. These technologies are not currently feasible for general use due to large amounts of cells required for each application, extremely large patient populations, and few cells surviving implantations. Current stem cell culturing methods do not generate cell numbers at a scalable rate or provide the appropriate biochemical and mechanical cues to aid in proliferation or differentiation to the desired cell types. Therefore, this dissertation focuses on designing and developing novel tunable systems to expand stem cell culture in a scalable manner./nThis work investigates two different material systems to culture stem cells in three-dimensional (3D) hydrogels. Stem cells exist in very complex microenvironments, receiving inputs from other cells, the extracellular matrix, as well as soluble factors. The systems described in this work aim to create in vitro environments which support cell viability, growth, replication, and potential differentiation by tuning the mechanical and biochemical properties of the hydrogels. Substrate modulus affects stem cell behavior in two-dimensional systems and could play a large role in stem cell behavior and differentiation in 3D systems as well. Therefore, both systems were optimized for the appropriate modulus to support iPSC growth, with the potential to generate gels of varying moduli for enhanced proliferation or differentiation of multiple types of stem cells. Both systems were also designed with specific chemical functionality in order to attach biochemical cues such as proteins or peptides, which may help to better mimic the in vivo environments and improve cell viability./nThese systems were both designed to easily encapsulate and recover cells by incorporating a thermoreversible component poly(N-isopropylacrylamide) (PNIPAAm). Thermoreversibility and encapsulation overcome the limitations of harsh recovery steps and shear stress associated with many current 3D cell culture methods. One hydrogel system used poly(ethylene glycol) (PEG) as the hydrophilic component due to its versatility, biocompatibility, and hydrophilicity. The other hydrogel system attached PNIPAAm to hydrophilic hyaluronic acid (HA), a natural polysaccharide found in the extracellular matrix which plays an important role in cell signaling and matrix organization. Both of these hydrogel systems successfully supported the growth and expansion of hPSCs, which maintained expression of pluripotency markers after growing within the gels./nIn summary, this dissertation presents work on designing and developing novel hydrogel materials to encapsulate and grow stem cells to aid in the production of stem cells for future therapeutics.

Advisor: Schaffer, David V
Segalman, Rachel A