title: Transient changes in oxygen tension inhibit osteogenic differentiation and Runx2 expression in osteoblasts.
abstract: Vascular disruption following bony injury results in a hypoxic gradient within the wound microenvironment. Nevertheless, the effects of low oxygen tension on osteogenic precursors remain to be fully elucidated. In the present study, we investigated in vitro osteoblast and mesenchymal stem cell differentiation following exposure to 21% O(2) (ambient oxygen), 2% O(2) (hypoxia), and <0.02% O(2) (anoxia). Hypoxia had little effect on osteogenic differentiation. In contrast, short-term anoxic treatment of primary osteoblasts and mesenchymal precursors inhibited in vitro bone nodule formation and extracellular calcium deposition. Cell viability assays revealed that this effect was not caused by immediate or delayed cell death. Microarray profiling implicated down-regulation of the key osteogenic transcription factor Runx2 as a potential mechanism for the anoxic inhibition of differentiation. Subsequent analysis revealed not only a short-term differential regulation of Runx2 and its targets by anoxia and hypoxia, but a long-term inhibition of Runx2 transcriptional and protein levels after only 12-24 h of anoxic insult. Furthermore, we present evidence that Runx2 inhibition may, at least in part, be because of anoxic repression of BMP2, and that restoring Runx2 levels during anoxia by pretreatment with recombinant BMP2 rescued the anoxic inhibition of differentiation. Taken together, our findings indicate that brief exposure to anoxia (but not 2% hypoxia) down-regulated BMP2 and Runx2 expression, thus inhibiting critical steps in the osteogenic differentiation of pluripotent mesenchymal precursors and committed osteoblasts.
authors.affiliation: Department of Surgery, Stanford University School of Medicine, Stanford, California 94305-5148, USA.
authors.forename: Ali
authors.initials: A
authors.lastname: Salim
authors.forename: Randall P
authors.initials: RP
authors.lastname: Nacamuli
authors.forename: Elise F
authors.initials: EF
authors.lastname: Morgan
authors.forename: Amato J
authors.initials: AJ
authors.lastname: Giaccia
authors.forename: Michael T
authors.initials: MT
authors.lastname: Longaker
mesh_terms: D016475:3T3 Cells; D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins / Q000494:pharmacology; D002454:Cell Differentiation / Q000502:physiology; D002470:Cell Survival / Q000502:physiology; D024042:Collagen Type I / Q000235:genetics; D050659:Core Binding Factor Alpha 1 Subunit; D015536:Down-Regulation; D015870:Gene Expression / Q000187:drug effects / Q000502:physiology; D000860:Hypoxia / Q000378:metabolism / Q000503:physiopathology*; D008648:Mesoderm / Q000166:cytology; D051379:Mice; D009363:Neoplasm Proteins / Q000235:genetics* / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D010006:Osteoblasts / Q000166:cytology* / Q000378:metabolism*; D015675:Osteocalcin / Q000235:genetics; D010100:Oxygen / Q000378:metabolism*; D012886:Skull / Q000166:cytology; D013234:Stem Cells / Q000166:cytology / Q000502:physiology; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects / Q000502:physiology; D016212:Transforming Growth Factor beta
chemical_list: C525719:Bmp2 protein, mouse; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins; D024042:Collagen Type I; D050659:Core Binding Factor Alpha 1 Subunit; D009363:Neoplasm Proteins; D014157:Transcription Factors; D016212:Transforming Growth Factor beta; D015675:Osteocalcin; D010100:Oxygen