<p>Osteogenesis Imperfecta (OI) is a genetic disorder where mutations in Collagen I gene or other genes associated with defective collagen matrix lead to low bone density, bone brittleness with increased fracture risk. Current pharmacological treatments for OI do not address the underlying defect in collagen production. Cellular therapy represents a strategy to form healthy bone in OI, but systemic stem cell delivery approaches have not been successful. By contrast, locally delivered skeletal stem and progenitor cells (SSPCs) can engraft within OI bone, differentiate into matrix-producing osteoblasts, and generate normal collagen. However, without perturbing the marrow environment, endosteal engraftment is low. Irradiation used as preconditioning significantly enhances engraftment of donor progenitor cells. We aimed to determine what alterations in the bone marrow microenvironment influence donor SSPCs engraftment and their differentiation into mature lineages. Several preconditioning treatments and animal models were used to determine optimal conditions for local cell transplantation. Preconditioning with lethal irradiation (IRR) in osteogenesis imperfecta mice (OIM) exhibited high donor cell engraftment on the endosteal surface. Chemical myeloablation using busulfan and cyclophosphamide (BuCy) significantly increased donor cell engraftment, but the effect was less than that of IRR. Interestingly, BuCy caused similar initial changes in T and B cell composition as IRR, but proportions of NK, monocytes/macrophages and neutrophils were not as significant as with IRR. An additional preconditioning strategy involved ablation of recipient OI osteoblasts using OIM/Col2.3ΔTK/⁺ mice, but was not sufficient for mesenchymal donor cell engraftment. Finally, we investigated whether IRR affects cell therapy success via increasing bone turnover. We found that by reducing bone resorption with zoledronic acid, OIM mice had decreased donor cell engraftment. We separately observed increased engraftment rates in high bone turnover of OIM mice compared to WT. Lethal irradiation remained the most effective pretreatment for SSPCs engraftment and their differentiation into osteoblasts producing healthy matrix in OIM bone. Results indicate that increased bone turnover in OIM mice positively affect engraftment and differentiation into osteoblasts/osteocytes. Future cell transplantation research should include repurposing of existing treatments that can increase bone turnover.</p>