Method: Transgenic mice overexpressing DMP1 via the OC promoter (OC-DMP1 mice) were generated, sacrificed at 15, 30, 60, and 90-days post-natal, and the femurs were dissected. Gene expression of 12 osteogenic genes was analyzed by RT-PCR. DMP1, OC, Runx2, and FGF23 protein was quantified and localized by Western blotting and immunohistochemistry, respectively. Cortical and trabecular bone architecture was quantitatively analyzed using Micro-CT. Mid-shaft cortical bone mineral density (BMD) was verified by backscatter SEM. Mechanical properties of the femurs were investigated by 3-point bend test. All data are expressed as mean ± SD and were analyzed by two-way ANOVA followed by Student’s t-test post-hoc analysis (α=0.05).
Result: RT-PCR revealed that osteogenic gene expression is altered in OC-DMP1 femurs in a time-dependent fashion (n=4). Protein analysis revealed that DMP1 is significantly increased at 15 and 30-days of age in OC-DMP1 femurs, specifically in the trabeculae (n=3). OC, Runx2, and FGF23 protein revealed a spatio-temporal dependence within the femurs growth plate and endocortical regions (n=3). Micro-CT revealed a difference in trabecular bone architecture at 30 and 60-days of age in OC-DMP1 femurs (n=8). Backscatter SEM verified there was no difference in cortical BMD between WT and OC-DMP1 femurs (n=5). No significant difference in mechanical properties was seen between WT and transgenic femurs by 3-point bend test (n=12).
Conclusion: These data suggest that osteocalcin-driven DMP1 overexpression influences bone biology and trabecular structure. There were several alterations in osteogenic gene expression and protein production within the distal growth plate, which lead to changes in the composition of the trabecular bone and its mineral architecture. These changes did not, however have an impact on the physiological mechanical properties of the transgenic mouse femurs.