A novel in vivo model to study endochondral bone formation; HIF-1 alpha activation and BMP expression

Numerous growth and transcription factors have been implicated in endochondral bone fort-nation of the growth plate. Many of these factors are up-regulated during hypoxia and downstream of Hypoxia-Inducible Factor (HIF)-1 alpha 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-1 alpha and the possible roles of (HIF-1 alpha 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)-beta(1), Bone Morphogenetic Proteins (BMP)-2, -4, and -7, Indian Hedgehog (Ihh), Parathyroid Hormone-related Protein (PTHrP), Sox9, Runx2, HIF-1 alpha, 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-beta(1), BMP-2, -4, and -7, HIF-1 alpha, and POSTN were examined.

During the chondrogenic phase, the expression of Sox9, Ihh, and HIF-1 alpha was significantly up-regulated. TGF-beta(1) 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-beta(1), BMP-2, -4, and -7, HIF-1 alpha, and POSTN within the chondrocytes during chondrogenesis.

This discrepancy in gene expression between mRNA and protein level for TGF-beta(1) and the different BMPs is indicative of post-transcriptional regulation of protein synthesis. HTF-1 alpha 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. (c) 2006 Elsevier Inc. All rights reserved.