Large bone defects caused by trauma or disease will not heal properly without specific treatment. Current therapeutic strategies such as autologous bone grafts, allogeneic bone grafts, and prosthetics have inherent limitations, which have led to alternative tissue-engineering approaches to improve the bone regeneration. In this regard, bone marrow-derived stem cells delivery has proven to be an effective therapy for bone regeneration by enhancing bone formation and improving union healing. Specifically, human mesenchymal stem cells (hMSCs) are commonly used since they are multipotent and can differentiate into osteoblasts under osteogenic signals. Typically, these cues include soluble factors (e.g., dexamethasone or bone morphogenic proteins) added to cell culture media, but there is a growing interest in the design of hMSC delivery vehicles whereby the osteogenic factors are presented locally to the cells. In this regard, the Anseth group has focused on poly(ethylene glycol) (PEG) based hydrogels for the encapsulation, 3D culture and delivery of hMSC. This PEG scaffold provides a basic environment that is permissive for 3D hMSC culture and the focus of my research aims to create a niche environment that promotes osteogenic differentiation. Since stem cell differentiation is comparatively slow, attention has focused on speeding up the differentiation process. The recent discovery of microRNAs (miRNAs) and their ability to control global gene expression has shed new light on methods to control stem cell differentiation. Studies about the effects of miRNA on stem cells indicate that miRNA transfection, together with chemical osteogenic cues can promote the osteogenesis of hMSCs. This discovery implies that by altering the cellular miRNA activity, one might induce stem cells differentiation, thus enhancing tissue formation. However, very little is known about the role of the cellular microenvironment in this process and mounting evidence suggests that the response of cells to differentiation cues depends on this external niche. In order to better understand how hMSCs respond to osteogenic cues in a 3D environment and to apply miRNAs in vivo to promote hMSCs' osteogenesis, we initially focused on synthetic strategies to functionalize PEG scaffolds with osteogenic chemical cues to study their potential synergistic effects with miRNAs. Our ultimate goal is to create a "cell free" in vivo therapeutic system that allows hMSCs from the host body to migrate into a biomaterial system, encounter appropriate osteogenic signals, and facilitate the regeneration of functional bone tissue. Thus, the overall objective of my research is to develop an osteoinductive, in vivo material system for hMSCs that will improve our understanding of how functionalization of the scaffold with chemical osteogenic cues and miRNAs can synergize together to promote the differentiation of hMSCs.