Abstract: BACKGROUND: A large number of studies have confirmed that exosomes can promote osteogenesis and vascularization. However, simple exosome therapy has problems such as poor targeting, and the content of loaded molecules cannot reach the therapeutic concentration.
OBJECTIVE: To load exosomes into injectable gluconolactone-sodium alginate β-tricalcium phosphate-polyethylene glycol hydrogel, and observe the effect of the hydrogel on peri-implant bone defect in vivo and in vitro. 
METHODS: Exosomes were extracted from bone marrow mesenchymal stem cells and wrapped in injectable gluconolactone-sodium alginate β-tricalcium phosphate-polyethylene glycol hydrogel. (1) In vitro experiment: The hydrogel loaded with exosomes and the hydrogel without exosomes were cocultured with endothelial progenitor cells, and exosomes uptake experiment, tubule formation experiment, cell proliferation, migration ability, and angiogenic gene detection were carried out. (2) In vivo experiment: Twelve male New Zealand white rabbits were used to prepare two standard implant cavities and corresponding bone defects in the long axis of one femur. A hydrogel loaded with exosomes was implanted in the bone defect after an implant was implanted in a cavity at the proximal end of the implant (experimental group), and an unloaded exosome hydrogel was implanted in the bone defect after an implant was implanted in a cavity at the distal end of the implant (control group). At 3, 6 and 9 weeks after operation, bone defects with implants were removed and stained with hematoxylin-eosin staining and Masson staining. Simultaneously, osteogenic and angiogenic genes were detected at 9 weeks after operation.  
RESULTS AND CONCLUSION: (1) In vitro experiment: Exosomes could enter endothelial progenitor cells. The proliferation, migration, angiogenesis and gene (CD31, vascular endothelial growth factor and basic fibroblast growth factor) expression of endothelial progenitor cells in the hydrogel-loaded group were higher than those in the hydrogel-unloaded group (P < 0.05). (2) In vivo experiment: Hematoxylin-eosin staining and Masson staining showed that at 3 weeks after operation, only a small amount of new bone was found in the two groups, and the material was partially degraded. At 6 weeks after operation, the amount of new bone in the two groups increased, and a large amount of new bone was found in the experimental group, with obvious calcium deposition. At 9 weeks after operation, compared with the control group, a large number of bone trabeculae thicker than mature were found in the experimental group, calcium salt deposition was more obvious, and a large number of osteoblasts were found around the bone trabeculae. The protein expressions of CD31, vascular endothelial growth factor, basic fibroblast growth factor, bone morphogenetic protein 2, type I collagen and osteocalcin in the experimental group were higher than those in the control group at 9 weeks after operation (P < 0.05). (3) The exosome-loaded gluconolactone-sodium alginate β-tricalcium phosphate-polyethylene glycol hydrogel could promote the proliferation, migration and angiogenic differentiation of endothelial progenitor cells and promote the repair and regeneration of bone defects around implants.

Key words: peri-implant bone defect, hydrogel, exosome, sodium alginate, β-tricalcium phosphate, polyethylene glycol