Department of Orthopaedic Surgery and Musculoskeletal Research Center, Washington University in St. Louis, St. Louis, MO, United States of America.
Department of Orthopaedic Surgery and Musculoskeletal Research Center, Washington University in St. Louis, St. Louis, MO, United States of America; St. Louis University, St. Louis, MO, United States of America.
Bone. 2022 Oct;163:116502. doi: 10.1016/j.bone.2022.116502. Epub 2022 Jul 21.
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 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) 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 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.
成骨与血管生成之间的关系非常复杂。正常的骨骼发育需要血管内皮生长因子 A（VEGFA）介导的血管生成。研究通过系统抑制或基因修饰表明，VEGFA 对于几种类型的骨修复是必不可少的，可能是通过其支持血管生成的作用。但是，在没有血管生成的情况下，VEGFA 直接在成骨细胞中的作用也已被提出。为了解决成骨细胞中的 VEGFA 是否直接支持骨形成的问题，我们应用合成代谢负荷诱导小鼠板层骨形成，该过程已被证明与血管生成无关。我们假设成骨细胞中的 VEGFA 是板层骨形成所必需的。为了验证这一假设，我们应用轴向胫骨压缩来测试三条线的诱导型 Cre/LoxP 小鼠。Vegfa 小鼠与 Ubiquitin C（UBC）、Osterix（Osx）和 Dentin-Matrix Protein 1（DMP1）Cre-ERT2 小鼠交配，分别靶向所有细胞、（前）成骨细胞谱系细胞和成骨细胞和破骨细胞。通过比较每条线的对照（Vegfa）和 Cre+（VegfaΔ）小鼠，确定基因型效应。在 5 个月大时，用他莫昔芬注射 5 天，然后在负荷前清除 3 周。对 100 只雌性和雄性小鼠（N=100）进行 5 天的负荷，以产生-3100με 的峰值压缩应变，并进行动态组织形态计量学分析（第 12 天）。由于负荷，MS/BS 的百分比增加了 20-70%，但 Osx 或 Dmp1 线的基因型没有影响。相比之下，UBC 组的 VegfaΔ 与 Vegfa 小鼠的相对骨外膜 BFR/BS 显著降低。在未加载的股骨中，UBC 线没有任何皮质骨表型。总之，动态组织形态计量学数据证实胫骨负荷可诱导板层骨形成。与我们的假设相反，在 Osx 或 Dmp1 线中，VEGFA 缺失时，负荷诱导的骨形成并没有减少。在靶向所有细胞的 UBC 线中，骨形成减少。这一结果表明，非成骨细胞来源的 VEGFA 支持负荷诱导的板层骨形成，尽管成骨细胞/破骨细胞 VEGFA 是可有可无的。这些发现支持旁分泌模型，即非成骨细胞的 VEGFA 支持板层骨形成，与血管生成无关。
