VEGFA from osteoblasts is not required for lamellar bone formation following tibial loading

The relationship between osteogenesis and angiogenesis is complex. Normal bone development requires angiogenesis, mediated by vascular endothelial growth factor A (VEGFA). Studies have demonstrated through systemic inhibition or genetic modification that VEGFA is indispensable for several types of bone repair, presumably via its role in supporting angiogenesis. But a direct role for VEGFA within osteoblasts, in the absence of angiogenesis, has also been suggested. To address the question of whether VEGFA from osteoblasts supports bone formation directly, we applied anabolic loading to induce lamellar bone formation in mice, a process shown to be independent of angiogenesis. We hypothesized that VEGFA from osteoblasts is required for lamellar bone formation. To test this hypothesis, we applied axial tibial compression to inducible Cre/LoxP mice from three lines. Vegfa^fl/fl mice were crossed with Ubiquitin C (UBC), Osterix (Osx) and Dentin-Matrix Protein 1 (DMP1) Cre-ERT2 mice to target all cells, (pre)osteoblast-lineage cells, and mature osteoblasts and osteocytes, respectively. Genotype effects were determined by comparing control (Vegfa^fl/fl) and Cre+ (VegfaΔ) mice for each line. At 5 months of age tamoxifen was injected for 5 days followed by a 3-week clearance prior to loading. Female and male mice (N = 100) were loaded for 5 days to peak forces to engender -3100 με peak compressive strain and processed for dynamic histomorphometry (day 12). Percent MS/BS increased 20-70 % as a result of loading, with no effect of genotype in Osx or Dmp1 lines. In contrast, the UBC groups had a significant decrease in relative periosteal BFR/BS in VegfaΔ vs. Vegfa^fl/fl mice. The UBC line did not have any cortical bone phenotype in non-loaded femurs. In summary, dynamic histomorphometry data confirmed that tibial loading induces lamellar bone formation. Contrary to our hypothesis, there was no decrease in loading-induced bone formation in the Osx or Dmp1 lines in the absence of VEGFA. There was a decrease in bone formation in the UBC line where all cells were targeted. This result indicates that VEGFA from a non-osteoblast cell source supports loading-induced lamellar bone formation, although osteoblast/osteocyte VEGFA is dispensable. These findings support a paracrine model whereby non-osteoblast VEGFA supports lamellar bone formation, independent of angiogenesis.