Bone formation in a rat critical sized bone defect upon implantation of an intrinsically osteoinductive calcium phosphate ceramic, with or without rhBMP-2

Objectives: The aim of this study was to investigate the ability of an osteinductive calcium phosphate ceramic alone or in combination with rhBMP-2 to bridge a critical sized calvarial defect in rats. The hypothesis was that the use of intrinsically osteoinductive ceramic could minimize the dosage of rhBMP-2. Methods: 120 male Wistar Han rats were used upon approval by institutional animal ethics committee (IMI,Zg,HR). 14mm critical sized circular defects were created on calvaria of animals by using a dental drill. Animals received a PTFE ring Ø14mm h 4mm, that was filled with either only a biphasic calcium phosphate ceramic BCP, particles 150-500 µm, consisting of ß tricalcium phosphate and hidroxyapatite in 20:80 ratio, or BCP loaded with 5µg of rhBMP-2, or 2.5µg of rhBMP-2 or left empty (sham). Animals were sacrificed at 3, 6 and 12 weeks post-implantation. Bone formation was quantitatively evaluated using µCT (SkyScan 1076, BE).3D reconstruction was performed by CTAn software, different tresholding values were used to distinguish ceramic particles from the newly formed bone and to determine BV% in total volume of interest BV/TV. ANOVA with Duncan's post hoc test was used to compare different conditions, in addition qualitative histological analysis on Goldner trichrome stained tissue sect. Results: Macroscopically, implants appeared firmly attached to the calvaria. In sham groups only limited new bone formation was observed 9.5 ±1.1%, 7.2±1.7%, 9.6±2.8%, 3,6,12 weeks respectively. The newly formed bone extended from host bone bed to the center of the defect, between particles of implanted ceramics (Xpand ) in all test groups with or without growth factor. Newly formed bone tissue was observed in all cases as early as 3 weeks with %BV/TV of 24.33±0.9% in BCP, 27.9 ±1.2% in BCP/rhBMP-2 2.5µg and 24.5±1.7% in BCP/rhBMP-2 5.0µg. At 6 weeks, a slight increase in the amount of bone was observed in all conditions with a % BV/TV of 36.4 ±3.8%, 31.0±3.8% and 35.0±5.0% in BCP, BCP/rhBMP-2 2.5 µg and BCP/rhBMP-2 5.0µg respectively, while a decrease between 6 and 12 weeks, with %BV/TV values of 24.7±3.9%, 32.54±1.2% and 31.3±2.8% for BCP, BCP/rhBMP-2 2.5-µg and BCP/rhBMP-2 5.0µg, respectively. No statistical differences among the groups were observed at any time point. Conclusions: Study results showed no statistically significant differences in bone formation between the BCP alone and loaded with 2.5 or 5 µg rhBMP-2 in a critical sized calvarial deffect in rats.While it is generally thought that CaP biomaterials are not osteoinductive, BCP used here was previously shown to possess intrinsic osteoinductivity which may explain comparable performance in this study in terms of dynamic bone formation and the total ammount of bone formed in presence and in absence of rhBMP-2. Keywords: bone regeneration, BCP, growth factor