Ball, Alyssa N., authorGoodrich, Laurie R., advisorDonahue, Seth W., committee memberMcIlwraith, C. Wayne, committee memberEhrhart, Nicole P., committee memberKawcak, Christopher E., committee memberBark, David L., Jr., committee member2019-06-142019-06-142019https://hdl.handle.net/10217/195326Optimizing the environment of complex bone healing and improving treatment of catastrophic bone fractures and segmental bone defects remains an unmet clinical need both human and equine veterinary medical orthopedics. Animal models of fracture repair often involve small rodents, as historically significant large animal models, like the dog, continue to gain favor as companion animals. This trend continues despite the well documented limitations in comparing fracture repair in humans, as few similarities exist. Study design, number of studies, and availability of funding also continues to limit large animal studies. Osteoinduction is often therapeutically targeted to incite new bone growth. Osteoinduction with recombinant BMP-2 (rhBMP-2) results in robust bone formation; although, long-term quality is scrutinized due to poor bone mineral quality. Gene therapy continues to gain popularity among researchers to augment bone healing, and with the approval of the first cell-based gene therapy treatment in South Korea, the clinical reputation of gene therapy is under scrutiny. Progenitor cell therapies and the content variation of patient-side treatments (e.g. PRP and BMAC) are being studied in humans, while the immunologic properties of autologous and allogeneic treatments are being studied in the horse. The objective of this Masters Thesis was to determine whether scAAV-equine-BMP-2 transduced cells would induce osteogenesis in equine bone marrow derived mesenchymal stem cells (BMDMSCs) in vitro, and if these cells could be cryopreserved in an effort to osteogenically prime them as an "off-the-shelf" gene therapeutic approach for fracture repair. Our study found that transgene expression is altered by cell expansion, as would be expected by a transduction resulting in episomal transgene expression, and that osteoinductive levels could still be achieved five days after recovery. Further, protein expression continues up to fourteen days after initial transduction. This is the first evidence that cryopreservation of genetically modified BMDMSCs would not alter the osteoinductive potential or clinical use of allogeneic donor cells in cases of equine fracture repair. Future directions should include in vivo pilot data aimed at elucidating whether or not these cells expedite clinical healing. Other genes and gene combinations (e.g. PTH 1-34 and BMP-2) could be introduced to the expression vector backbone and compared to the vector utilized in this study.born digitalmasters thesesengCopyright and other restrictions may apply. User is responsible for compliance with all applicable laws. For information about copyright law, please see https://libguides.colostate.edu/copyright.Text