Emans Pieter J, Spaapen Frank, Surtel Don A M, Reilly Keryn M, Cremers Andy, van Rhijn Lodewijk W, Bulstra Sjoerd K, Voncken Jan Willem, Kuijer Roel
Department of Orthopedic Surgery, University Hospital Maastricht, Maastricht, The Netherlands.
Bone. 2007 Feb;40(2):409-18. doi: 10.1016/j.bone.2006.08.005. Epub 2006 Sep 18.
Numerous growth and transcription factors have been implicated in endochondral bone formation of the growth plate. Many of these factors are up-regulated during hypoxia and downstream of Hypoxia-Inducible Factor (HIF)-1alpha activation. However, the specific function of these factors, in the context of oxygenation and metabolic adaptation during adult periosteal endochondral bone formation, is largely unknown. Here, we studied HIF-1alpha and the possible roles of (HIF-1alpha related) growth and transcription factors in a recently developed in vivo model for adult periosteal endochondral bone formation. At different phases of periosteal endochondral bone formation, mRNA levels of Transforming Growth Factor (TGF)-beta1, Bone Morphogenetic Proteins (BMP)-2, -4, and -7, Indian Hedgehog (Ihh), Parathyroid Hormone-related Protein (PTHrP), Sox9, Runx2, HIF-1alpha, Vascular Endothelial Growth Factor (VEGF), periostin (POSTN), and Glyceraldehyde-3-Phophate Dehydrogenase (GAPDH) were evaluated with RT-real time-PCR. Also protein levels of TGF-beta1, BMP-2, -4, and -7, HIF-1alpha, and POSTN were examined. During the chondrogenic phase, the expression of Sox9, Ihh, and HIF-1alpha was significantly up-regulated. TGF-beta1 mRNA levels were rather constant, and the mRNA levels of BMPs were significantly lower. Immunohistochemical detection of corresponding gene products, however, revealed the presence of the proteins of TGF-beta1, BMP-2, -4, and -7, HIF-1alpha, and POSTN within the chondrocytes during chondrogenesis. This discrepancy in gene expression between mRNA and protein level for TGF-beta1 and the different BMPs is indicative of post-transcriptional regulation of protein synthesis. HIF-1alpha activation and up-regulation of GAPDH support a hypoxia-induced metabolic shift during periosteal chondrogenesis. Our model recapitulates essential steps in osteochondrogenesis and provides a new experimental system to study and ultimately control tissue regeneration in the adult organism.
许多生长因子和转录因子与生长板的软骨内骨形成有关。这些因子中的许多在缺氧期间以及缺氧诱导因子（HIF）-1α激活的下游被上调。然而，在成人骨膜软骨内骨形成过程中的氧合和代谢适应情况下，这些因子的具体功能在很大程度上尚不清楚。在这里，我们在最近开发的成人骨膜软骨内骨形成体内模型中研究了HIF-1α以及（与HIF-1α相关的）生长因子和转录因子的可能作用。在骨膜软骨内骨形成的不同阶段，通过RT-实时PCR评估转化生长因子（TGF）-β1、骨形态发生蛋白（BMP）-2、-4和-7、印度刺猬因子（Ihh）、甲状旁腺激素相关蛋白（PTHrP）、Sox9、Runx2、HIF-1α、血管内皮生长因子（VEGF）、骨膜蛋白（POSTN）和甘油醛-3-磷酸脱氢酶（GAPDH）的mRNA水平。还检测了TGF-β1、BMP-2、-4和-7、HIF-1α和POSTN的蛋白水平。在软骨形成阶段，Sox9、Ihh和HIF-1α的表达显著上调。TGF-β1的mRNA水平相当恒定，而BMPs的mRNA水平显著较低。然而，相应基因产物的免疫组织化学检测显示，在软骨形成过程中软骨细胞内存在TGF-β1、BMP-2、-4和-7、HIF-1α和POSTN的蛋白质。TGF-β1和不同BMPs在mRNA和蛋白质水平上的基因表达差异表明蛋白质合成存在转录后调控。HIF-1α的激活和GAPDH的上调支持骨膜软骨形成过程中缺氧诱导的代谢转变。我们的模型概括了骨软骨形成的基本步骤，并提供了一个新的实验系统来研究并最终控制成体生物中的组织再生。
