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Abstract
Mesenchymal Stem Cells (MSCs) have been investigated for several decades for use in tissue engineering and regenerative medicine. Initially, they were only thought of as a source of multipotent cells for tissue transplantation. However, difficulties in maintaining the potency of transplanted MSCs and safety and regulatory concerns have prompted a shift in focus to their secretome, which can have regenerative and immunosuppressive effects on endogenous cells. This dissertation explores various ways to manipulate MSCs for different therapeutic benefits. First, we evaluated the ability of lentiviral transduced MSCs to express bone morphogenetic protein-2 (BMP-2) and heal critical sized bone defects compared to delivery of rhBMP-2 in scaffolds. We found that BMP-2 MSCs released the protein over a longer period of time than BMP-2 loaded scaffolds, and they induced comparable healing of critical sized defects to the loaded scaffolds. We next investigated the use of a parainfluenza-5 derived amplifying virus like particle (AVLP) to induce transgene expression in MSCs. This involved examining the transduction efficiency and stability of expression over time of both an EGFP and BMP-2 vector. We found that while MSCs could be reliably transduced to express EGFP stably over time, transduction with the BMP-2 vector proved to induce much more variable expression. Finally, we compared the effect of acidic, hypoxic, and inflammatory culture on MSCs and their extracellular vesicles (EVs). We characterized the EVs produced in the various conditions and compared their ability to suppress T-cells to that of their parent cells. We found that EV secretion, size, and surface markers vary with the extracellular conditions of their secreting MSCs, and while MSCs are generally better at suppressing T-cell activation, EVs from acidic conditioned MSCs induce the formation of regulatory T-cells. We have identified various avenues for modifying the MSC secretome which could be of value for future therapeutics.