Effect of recombinant human bone morphogenetic protein delivered by chitosan microspheres on ectopic osteogenesis in rats

In the present study, the effect of recombinant human bone morphogenetic protein-2 (rhBMP-2) delivered by chitosan (CS) microspheres on ectopic osteogenesis was investigated in a rat model. rhBMP-2-loaded CS microspheres and blank CS microspheres were prepared. A total of 24 male Sprague Dawley rats were divided into 4 groups with 6 rats in each group: The CS/rhBMP-2 group, the rhBMP-2 group, in which rhBMP-2 was directly implanted (rhBMP-2 dose in either group, 1 mg), the CS blank group and the control group. X-ray was performed at 4 weeks after ectopic osteogenesis surgery and micro-computed tomography (CT) examination was scheduled at 1, 2, 3 and 4 weeks after the surgery to determine ectopic osteogenesis in the different groups. Histological analysis, and determination of alkaline phosphatase (ALP) activity and calcium content were also performed. The mean diameter of the osteoid tissues was 1.1±0.3 cm (range, 0.8–1.4 cm) in the CS/rhBMP-2 group, which was significantly bigger than that in the rhBMP-2 group (0.3±0.1 cm; range, 0.1–0.4 cm) at 4 weeks after the surgery. X-ray analysis and micro-CT scan indicated that the area of high-density tissues and the radionuclide intensity, as well as bone volume in the 3-dimensional reconstruction were greatest in the CS/rhBMP-2 group, followed by those in the rhBMP-2 group. All parameters, including bone mineral density, tissue mineral density, tissue mineral content and bone volume fraction, were significantly higher in the CS/rhBMP-2 group at 3 and 4 weeks after the surgery, compared with those in the rhBMP-2 group. The histological analysis, ALP activity analysis and determination of calcium content revealed that the CS/rhBMP-2 system had the greatest ability to induce osteoblast differentiation. In conclusion, the CS/rhBMP-2 microsphere delivery system significantly enhanced the induction and promotion effects of rhBMP-2 regarding ectopic osteogenesis. The present study enhances the basic data available for future application of the CS/rhBMP-2 microspheres delivery system and provides a deeper understanding of the role of BMP-2 in bone regeneration.