Objective To investigate the reparative effect of 3D printed biomimetic bone scaffolds loaded with adipose stem cell-derived extracellular vesicles (ADSC-EVs) on long bone defects. Methods High-precision biomimetic bone
scaffolds were fabricated using digital light processing (DLP)3D printing with polycaprolactone triacrylate (PCLTA). ADSCEVs were drop-loaded onto the scaffold surface. Scanning electron microscopy was used to observe the morphology of the vesicles on the scaffold. Cell compatibility and osteogenic ability of the scaffold were evaluated using immunofluorescence staining with phalloidin and Alizarin Red. Critical bone defects (15 mm) were created in the bilateral radius of New Zealand white rabbits. Rabbits were randomly assigned to 3 groups: blank control group without scaffold implantation, control group with pure material scaffold implantation, and experimental group with scaffold loaded with ADSC-EVs. Samples were harvested at 12 weeks post-implantation for evaluation of osteogenesis and angiogenesis using X-ray, immunohistochemical staining, and histological analysis. Results Phalloidin staining showed uniform cell growth on the scaffold surface with normal cell morphology. Alizarin Red staining revealed that the vesicle-loaded scaffold had stronger osteoinductive ability (P< 0.05). Scanning electron microscopy demonstrated uniform distribution of vesicles on the scaffold surface and within the scaffold. X-ray images indicated that the experimental group had faster new bone formation and higher bone mass than the
control group (P<0.05) and blank control group (P<0.01), which was further confirmed by histological analysis with Hematoxylin-Eosin staining. Immunohistochemical staining of tissue sections showed that the expression of the endothelial cell-specific marker CD31 was higher in the experimental group than in the control group (P<0.05) and blank control group (P<0.01). Conclusion PCLTA biomimetic bone scaffolds loaded with ADSC-EVs exhibit excellent osteogenic and angiogenic abilities, effectively promoting the repair of long bone defects.