Methods: Mandibles from quail embryos at Hamburger and Hamilton (HH) stage 23 were surgically extracted, placed on transwell membranes, and treated with agarose beads equilibrated in BMP4 or BMP7, using bovine serum albumin as contralateral controls. Mandibles were cultured in differentiation medium for three to six days and histologically assayed for bone. Representative sections were digitized and bone volume was estimated from the number of pixels comprising domains of stained matrix and condensations using the equation for a conical frustum: BV=1/3h[(Ai+Ai+1)+SQRT(AiAi+1)].
Results: Regardless of treatment, HH23 mandibles cultured for three and four days showed little to no histological evidence of bone. HH23 mandibles cultured for six days and treated with BMP4 or BMP7 exhibited an average 3-fold (p<.002) and 1.4-fold (p<.037) increase in bone condensation volume, respectively. Interestingly, four of those BMP4 mandibles developed no bone on the untreated side.
Conclusions: While time-dependent epithelial-mesenchymal interactions are known to regulate intramembranous ossification, our results reveal TGFß family members as candidate mediators of this process. The ability of exogenous BMPs to induce bone formation indicates that molecular-based therapies have strong potential to promote regeneration of skeletal tissues following trauma or degenerative skeletal disease.
Support: NIH/NIDCR/COHORT Training Grant T32/DE07306-09 to S.W.; NIDCR R03 DE014795-01 and R01 DE016402-01, and March of Dimes 5-FY04-26 to R.A.S.