Osteoinduction with highly purified β-tricalcium phosphate in dog dorsal muscles and the proliferation of osteoclasts before heterotopic bone formation

The aim of the study was to examine the chronological histology of osteoinduction of highly purified β-tricalcium phosphate (β-TCP) implanted in dog dorsal muscles. Specimens were harvested on days 14, 28, 42, 56, 112 and 168 after implantation, and were analyzed by hematoxylin and eosin (HE) staining, tartrate-resistant acid phosphatase (TRAP) staining, immunohistochemistry, in situ hybridization, and silver impregnation. After day 28, abundant TRAP- and cathepsin K-positive multinucleated cells adhered to β-TCP, suggesting that these cells are osteoclasts that can resorb β-TCP. On day 56, new bone was formed and α1 chain of type I procollagen mRNA-positive osteoblasts lined the newly formed bone. Silver impregnation showed abundant collagen fibrils within the β-TCP micropores. These results suggest that micropores function as a storage space for extracellular matrix components, including collagen. Newly formed bone never degenerated in the late stage, suggesting that β-TCP has good biocompatibility and this material retains the conditions appropriate for osteointegration and bioresorption. In conclusion, β-TCP has osteoinductivity after implantation in dog dorsal muscles without use of bone marrow cells or osteoinductive cytokines. The appearance of a large number of active osteoclasts precedes new bone formation.