Repetitive recombinant human bone morphogenetic protein 2 injections improve the callus microarchitecture and mechanical stiffness in a sheep model of distraction osteogenesis

Evidence suggests that recombinant human bone morphogenetic protein 2 (rhBMP-2) increases the mechanical integrity of callus tissue during bone healing. This effect may be either explained by an increase of callus formation or a modification of the trabecular microarchitecture. Therefore the purpose of the study was to evaluate the potential benefit of rhBMP-2 on the trabecular microarchitecture and on multidirectional callus stiffness. Further we asked, whether microarchitecture changes correlate with optimized callus stiffness. In this study a tibial distraction osteogenesis (DO) model in 12 sheep was used to determine, whether percutaneous injection of rhBMP-2 into the distraction zone influences the microarchitecture of the bone regenerate. After a latency period of 4 days, the tibiae were distracted at a rate of 1.25 mm/day over a period of 20 days, resulting in total lengthening of 25 mm. The operated limbs were randomly assigned to one treatment groups and one control group: (A) triple injection of rhBMP-2 (4 mg rhBMP-2/injection) and (B) no injection. The tibiae were harvested after 74 days and scanned by mu CT (90 mu m/voxel). In addition, we conducted a multidirectional mechanical testing of the tibiae by using a material testing system to assess the multidirectional strength. The distraction zones were tested for torsional stiffness and bending stiffness antero-posterior (AP) and medio-lateral (ML) direction, compression strength and maximum axial torsion. Statistical analysis was performed using multivariate analysis of variance (ANOVA) followed by student's t-test and Regression analysis using power functions with a significance level of P<0.05. Triple injections of rhBMP-2 induced significant changes in the trabecular architecture of the regenerate compared with the control: increased trabecular number (Tb. N.) (treatment group 1.73 mm/1 vs. control group 1.2 mm/1), increased cortical bone volume fraction (BV/TV) (treatment group 0.68 vs. control group 0.47), and decreased trabecular separation (Tb. Sp.) (treatment group 0.18 mm vs. control group 0.43 mm). The analyses of the mechanical strength of regenerated bone showed significant differences between treatment group (A) and the control group (B). The bending stiffness anterior- posterior (treatment group 17.48 Nm vs. control group 8.3 Nm), medial-lateral (treatment group 18,9 Nm vs. control group 7.92 Nm) and the torsional stiffness (treatment group 41.17N/degrees vs. control group 16.41N/degrees) are significantly higher in the treatment group than in the control group. The regression analyses revealed significant non-linear relationships between BV/TV, TB. N., Tb. Sp. and all mechanical properties. Maximal correlation coefficients were found for the Tb. Sp. vs. the bending stiffness AP and ML with R-2=0.69 and R-2=0.70 (P<0.0001). There was no significant relation between Connectivity and the compression strength and the maximum axial torque. This study suggests that rhBMP-2 optimizes the trabecular microarchitecture of the regenerate, which might explain the advanced mechanical integrity of newly formed bone under rhBMP-2 treatment.