Characterizing the BMP pathway in a wild type mouse model of distraction osteogenesis

Abstract

Distraction osteogenesis (DO) is a well established surgical technique for limb lengthening and replacement of bone loss due to trauma, infection or malignancies. Although the technique is widely used, one of its limitations is the long period of time required for the newly formed bone to consolidate. We have previously shown that exogenous application of bone morphogenetic proteins (BMPs) can increase bone formation during DO, however, exogenous BMPs have many drawbacks. An alternative method for accelerating the rate of bone formation may be to modulate the intrinsic BMP signaling pathway. The aim of the current study was to analyze the expression of various genes involved in the BMP pathway at various time periods during DO in order to identify potential targets for therapeutic manipulation. DO was applied to the right tibia of 80 adult wild type mice. Distraction began after a latency period of 5 days at a rate of 0.2 mm/12 h for 2 weeks. Mice were sacrificed in groups of 12 at the following times post surgery: day 5 (latency), days 11 and 17 (distraction) and days 34 and 51 (consolidation). Specimens were examined using radiology, microCT, histology, RT 2PCR, immunohistochemistry and Western analysis. Genes involved in the BMP pathway including the BMP ligands, receptors, antagonists and downstream effectors were examined. A significant upregulation of BMPs 2, 4 and 6 was observed using both PCR and immunohistochemistry during the distraction phase. The expression of BMP7 remained constant throughout the distraction and consolidation process. Surprisingly, the only receptors which were upregulated significantly were the Activin Receptor Type 1 (ActR1) during distraction and Activin Receptor Type 2b (ActR2b) during consolidation. Most interestingly, simultaneously with the ligands, an increase in the expression of the antagonists, Noggin, Chordin, Inhibin and BMP3 was observed. This study provides a clearer understanding of expression patterns during DO, which is a valuable resource for finding therapeutic options to stimulate bone formation. The results suggest that blocking BMP inhibitors may be a possible method for increasing the function of intrinsic growth factors involved in bone regeneration.