Reduction of BMP6-induced bone formation by calcium phosphate in wild-type compared with nude mice

Nude mice have been extensively used to investigate the potency of tissue engineering strategies for bone repair. However, the contribution of pro-inflammatory and proregenerative stimuli of the host for the process of new bone formation and integration remains poorly understood. In this study, ectopic bone formation was investigated in nude (Nu) versus wild-type (WT) mice. Calcium phosphate (CaP) scaffolds (CopiOs [Zimmer] and Bio-Oss [Geistlich]) were loaded with different concentrations of rhBMP6 (40, 120, and 240 ng/mm³ rhBMP6) and implanted subcutaneously in Nu (BALB/c and NMR1) and WT (BALB/c and c57BL/6) mice. CaP scaffolds loaded with rhBMP6 did not form bone in WT mice. However, in Nu mice, 40 ng/mm³ rhBMP6 was sufficient to generate relevant volumes of new bone at 6 weeks after implantation. Looking into potential underlying mechanisms, TNF-α blocking antibodies were injected intraperitoneally but could not restore bone formation. Also, mouse periosteal cells (mPDCs) seeded in CopiOs loaded with rhBMP6 did not significantly improve the outcome. Abrogation of bone formation was associated with dense cellular infiltration, in particular with the presence of CD3+ T-lymphocytes. To probe a correlation between calcium ions and impaired bone formation in WT mice, type 1 collagen gels were loaded with rhBMP6 and calcium chloride and injected subcutaneously. These gels generated new bone in WT mice despite the increased percentage of CD3+ cells at Day 3 after implantation as compared with control gels. Overall, this study illustrated the negative effect of the inflammatory host response on the bone-forming capacity of rhBMP6 coated on bioceramic scaffolds.