Methods: Premolar tooth extraction and preparation of osseous defects were performed in the mandibles of beagle dogs. After 3 months, β-TCP was implanted into cylindrical artificial bone defects (4.5 x 8 mm). Dogs were sacrificed at 4, 7 and 14 days, after canned by microCT (μCT), total RNA was isolated from bone tissues using Fast Prep and RNeasy Mini Kit. Gene expression profiles were examined using Affymetrix GeneChip (Canine Genome 2.0 Array, ca. 38,000 genes) system and GeneSpring software was used for data analysis. PCR was used to confirm differences in transcription that were observed in the microarray analysis. We also examined IGF-I, IGF-II, IGF1R and IGFBPs protein expression levels in implanted tissue by immunohistochemistry.
Results: Genes were identified that were up regulated 2-fold or more by β-TCP at day 4 (2,130 genes), day 7 (2,020 genes), and day 14 (1,515 genes). Down-regulated genes were 1,279, 1,936 and 5,384 at day 4, 7, and 14, respectively. Among these, IGF-I, IGF-II, IGF1R, IGF2R and six IGF-binding proteins (IGFBP-1 ~ 6) were significantly up regulated by β-TCP. The results were also confirmed using PCR and immunohistochemistry.
Conclusion: These findings suggest that the regulation of proliferation and differentiation of osteoblast cells by β-TCP via IGF signaling involves the promotion of bone formation.