Department of Orthopedics, Washington University, St. Louis, MO 63110, USA.
Bone. 2011 Feb;48(2):250-8. doi: 10.1016/j.bone.2010.09.005. Epub 2010 Sep 22.
Osteogenesis occurs by formation of woven or lamellar bone. Little is known about the molecular regulation of these two distinct processes. We stimulated periosteal bone formation at the ulnar mid-diaphysis of adult rats using a single bout of forelimb compression. We hypothesized that loading that stimulates woven bone formation induces higher over-expression of genes associated with cell proliferation, angiogenesis and osteogenesis compared to loading that stimulates lamellar bone formation. We first confirmed that a single bout of 100 cycles of loading using either a rest-inserted (0.1 Hz) or haversine (2 Hz) waveform (15 N peak force) was non-damaging and increased lamellar bone formation (LBF loading). Woven bone formation (WBF loading) was stimulated using a previously described, damaging fatigue loading protocol (2 Hz, 1.3 mm disp., 18 N peak force). There were dramatic differences in gene expression levels (based on qRT-PCR) between loading protocols that produced woven and lamellar bone. In contrast, gene expression levels were not different between LBF loading protocols using a rest-inserted or haversine waveform. Cell proliferation markers Hist4 and Ccnd1 were strongly upregulated (5- to 17-fold) 1 and 3 days after WBF loading, prior to woven bone formation, but not after LBF loading. The angiogenic genes Vegf and Hif1a were upregulated within 1 h after WBF loading and were strongly up on days 1-3 (3- to 15-fold). In sharp contrast, we observed only a modest increase (<2-fold) in Vegfa and Hif1a expression on day 3 following LBF loading. Consistent with these relative differences in gene expression, vascular perfusion 3 days after loading revealed significant increases in vessel number and volume following WBF loading, but not after LBF loading. Lastly, bone formation markers (Runx2, Osx, Bsp) were more strongly upregulated for woven (4- to 89-fold) than for lamellar bone (2-fold), consistent with the differences in new bone volume observed 10 days after loading. In summary, robust early increases both molecularly and histologically for cell proliferation and angiogenesis precede woven bone formation, whereas lamellar bone formation is associated with only a modest upregulation of molecular signals at later timepoints.
成骨作用是通过编织骨或板层骨的形成发生的。关于这两个不同过程的分子调节，人们知之甚少。我们通过单次前肢压缩刺激成年大鼠尺骨干骺端的骨膜骨形成。我们假设，与刺激板层骨形成的负荷相比，刺激编织骨形成的负荷会引起与细胞增殖、血管生成和成骨相关的基因更高的过度表达。我们首先通过使用休息插入（0.1 Hz）或 haversine（2 Hz）波形（15 N 峰值力）进行的单次 100 次循环加载证实，这种加载是非破坏性的，并增加了板层骨形成（LBF 加载）。使用先前描述的破坏性疲劳加载方案（2 Hz、1.3 mm 位移、18 N 峰值力）刺激编织骨形成（WBF 加载）。产生编织骨和板层骨的加载方案之间的基因表达水平（基于 qRT-PCR）存在显著差异。相比之下，使用休息插入或 haversine 波形进行 LBF 加载时，基因表达水平没有差异。细胞增殖标记物 Hist4 和 Ccnd1 在 WBF 加载后 1 天和 3 天（在编织骨形成之前）强烈上调（5 至 17 倍），但在 LBF 加载后没有上调。血管生成基因 Vegf 和 Hif1a 在 WBF 加载后 1 小时内上调，并在第 1 天至第 3 天（3 至 15 倍）强烈上调。与此形成鲜明对比的是，我们仅观察到 LBF 加载后第 3 天 Vegfa 和 Hif1a 表达适度增加（<2 倍）。与基因表达的这些相对差异一致，负荷后 3 天的血管灌注显示 WBF 负荷后血管数量和体积显著增加，但 LBF 负荷后没有增加。最后，与板层骨（2 倍）相比，编织骨形成的骨形成标志物（Runx2、Osx、Bsp）上调更为强烈（4 至 89 倍），这与负荷后 10 天观察到的新骨体积差异一致。总之，细胞增殖和血管生成的分子和组织上的早期快速增加先于编织骨形成，而板层骨形成仅与稍后时间点的分子信号的适度上调相关。
