Ectopic Osteogenesis by Ex Vivo Gene Therapy Using Beta Tricalcium Phosphate as a Carrier

Abstract

Injuries and other damage to large bone can result in defects that do not heal spontaneously and lead to severe functional impairment. Better therapies are greatly needed to address this worldwide problem. The objective of the present study was to determine whether adenoviral delivery of modified human BMP2 gene (AdBMP2) using beta tricalcium phosphate (ß-TCP) as a carrier could promote osteoblastic differentiation of bone marrow mesenchymal stem cells (BMSCs) and ectopic bone formation. Rabbit BMSCs were separated from tibia aspirates and expanded in vitro. The BMSCs were then infected with AdBMP-2. Expression of BMP2, alkaline phosphatase, type I collagen, osteonectin, osteopontin, and mineralization of the cells confirmed secretion of active BMP2. Cells were observed to differentiate and maintain the osteoblast phenotype. For additional in vivo experiments, subcutaneous pockets were created on the backs of nude mice, which were then implanted with AdBMP2-BMSCs/ß-TCP, Adβgal-BMSCs/ß-TCP, BMSCs/ß-TCP, or ß-TCP alone. The nude mice were sacrificed after 4 weeks for histological evaluation. Adβgal-BMSCs/ß-TCP, BMSCs/ß-TCP, and ß-TCP did not show bone formation, although extensive fibrous tissue formed in the subcutaneous space in the rats implanted with ß-TCP. However, new bone tissue formation was observed on the inner walls of the pores of the ß-TCP-treated animals, and ectopic bone formation (mainly ‘‘cartilage-bone inducing’’) was observed in the AdBMP2-BMSCs/ß-TCP composite. These results confirmed the osteogenic potential of BMSCs after AdBMP2 transduction and revealed that AdBMP2-BMSC/ß-TCP composites could provide the capacity for bone formation and maturation during the more advanced stages of healing.