Engineering Matrix Properties for Stem Cell Culture and Gene Delivery

Tissue engineering aims to regenerate lost or damaged tissues and frequently combines therapeutic cells, biomaterial scaffolds, and therapeutic molecules in efforts to do so. One of the primary considerations in such therapies is to orchestrate the interactions between these components to regulate the desired cellular responses. This thesis focuses on tuning the proliferation of and exogenous protein expression from precursor cells, multipotent stem cells, and pluripotent stem cells with the mechanical and chemical properties of hydrogel-based synthetic extracellular matrices (ECMs). First, a synthetic stem cell niche with controlled stiffness and density of cell adhesion peptides was developed for spermatogonial stem cell (SSC) culture (Chapter 2). The synthetic stem cell niche allowed for the growth of SSCs in both two-dimensional (2D) and three-dimensional (3D) cultures, and the proliferation of SSCs was influenced by the density of cell adhesion ligands but not by substrate stiffness. Separately, the effects of substrate stiffness on non-viral gene delivery were evaluated for stem and precursor cells commonly used in tissue engineering or frequently present at gene delivery sites (Chapter 3). The cellular uptake and expression of non-viral genes for fibroblasts, bone marrow stromal cells (BMSCs), and myoblasts were regulated by the stiffness of the synthetic ECM. Interestingly, the effects of matrix stiffness on non-viral gene delivery were dependent on cell type, likely due to differences in cellular sensitivity to matrix mechanics. Finally, the results from the first two sections were used to build an implantable microvascular stamp consisting of cells transfected with plasmid DNA encoding vascular endothelial growth factor (VEGF) and a hydrogel matrix with tunable stiffness and permeability (Chapter 4). Overall, the results from this thesis may be useful in understanding the role of the extracellular microenvironment in tuning a variety of cellular activities. Furthermore, the information gained from these studies may be instrumental in the design and development of a novel biomaterial to significantly enhance the quality of tissue regeneration therapies