IADR Abstract Archives
Osteocalcin-driven DMP1 Overexpression Affects Bone Development in a Mouse Model
Objectives: Investigate the influence of osteocalcin (OCN)-driven overexpression of dentin matrix protein 1 (DMP1) in a transgenic mouse model (OC-DMP1) on the biochemistry, architecture and function of bone during development.
Methods: Transgenic and WT femur and calvaria were dissected at 15, 30, 60, and 90-days of age. Samples (n=3) were demineralized, embedded, sectioned and immunofluorescence (IF) was performed (Ab: C-DMP1, Runx2, OCN, ALP, CD31). Bone total protein (n=4) was collected and analyzed by Western blot (Ab: C-DMP1, Runx2, OCN, FGF23, VEGF). Micro-CT analysis (n=8) was performed followed by embedding, sectioning and mineral analysis (n=5) via backscatter-SEM (BSE-SEM). Femurs mechanical and material properties were analyzed (n=12) by 3-point bend test. Statistical analysis by two-way ANOVA followed by Tukey’s post-hoc analysis (p<0.05).
Results: Protein expression varied significantly by age, genotype and bone. These data were supported by IF and quantification. Micro-CT revealed that at 30-days, femur cortical bone in OC-DMP1 mice was significantly thinner, while trabecular spacing was decreased at 30 and 60-days. Calvaria were significantly thinner at 30 and 60-days. BSE-SEM showed that OC-DMP1 femur mineral density was significantly less while mineral heterogeneity was greater at 60-days. OC-DMP1 calvaria mineral density and heterogeneity was significantly different at all time points, greater at 15 and 60-days while less at 30 and 90-days. Femur 3-point bend testing revealed that OC-DMP1 bones have significantly decreased stiffness and strength at 60-days, but undergo increased post-yield displacement at 90-days. At 30-days, femurs have significantly increased modulus while at 60 and 90-days the modulus and ultimate stress is less than WT.
Conclusions: OC-DMP1 femur and calvaria are uniquely influenced in a spatiotemporal manner biochemically during development by the transgene that affects the expression of osteogeneic and mineralizing proteins thereby perturbing mineralization and tissue architecture. Additionally, femur mechanical and material properties are influenced.
Methods: Transgenic and WT femur and calvaria were dissected at 15, 30, 60, and 90-days of age. Samples (n=3) were demineralized, embedded, sectioned and immunofluorescence (IF) was performed (Ab: C-DMP1, Runx2, OCN, ALP, CD31). Bone total protein (n=4) was collected and analyzed by Western blot (Ab: C-DMP1, Runx2, OCN, FGF23, VEGF). Micro-CT analysis (n=8) was performed followed by embedding, sectioning and mineral analysis (n=5) via backscatter-SEM (BSE-SEM). Femurs mechanical and material properties were analyzed (n=12) by 3-point bend test. Statistical analysis by two-way ANOVA followed by Tukey’s post-hoc analysis (p<0.05).
Results: Protein expression varied significantly by age, genotype and bone. These data were supported by IF and quantification. Micro-CT revealed that at 30-days, femur cortical bone in OC-DMP1 mice was significantly thinner, while trabecular spacing was decreased at 30 and 60-days. Calvaria were significantly thinner at 30 and 60-days. BSE-SEM showed that OC-DMP1 femur mineral density was significantly less while mineral heterogeneity was greater at 60-days. OC-DMP1 calvaria mineral density and heterogeneity was significantly different at all time points, greater at 15 and 60-days while less at 30 and 90-days. Femur 3-point bend testing revealed that OC-DMP1 bones have significantly decreased stiffness and strength at 60-days, but undergo increased post-yield displacement at 90-days. At 30-days, femurs have significantly increased modulus while at 60 and 90-days the modulus and ultimate stress is less than WT.
Conclusions: OC-DMP1 femur and calvaria are uniquely influenced in a spatiotemporal manner biochemically during development by the transgene that affects the expression of osteogeneic and mineralizing proteins thereby perturbing mineralization and tissue architecture. Additionally, femur mechanical and material properties are influenced.