Osteogenesis in an in vitro coculture of human periodontal ligament fibroblasts and human microvascular endothelial cells.
BACKGROUND
Periodontal bone healing is a complex process involving many cells and processes that must function flawlessly for proper healing to occur. The exact progenitor cells that contribute to this process are not fully characterized. Periodontal fibroblasts and pericytes were postulated to be potential osteoprogenitor cells. This study describes a viable coculture model for the in vitro study of osteogenesis.


METHODS
Human microvascular endothelial cells (HMVEC) and human periodontal ligament (HPDL) fibroblasts were cocultured in a layered model and monitored for the development of runt-related transcription factor 2 (runx2) and desmin expression by real-time polymerase chain reaction. Conditions shown to be osteogenic (bone morphogenetic protein [BMP]-2 and enamel matrix derivative [EMD]) were compared to a control coculture that was unstimulated.


RESULTS
The HMVEC migrated into a layer of collagen containing only HPDL cells as monitored by fluorescent labeling. runx2 and desmin expressions were increased in stimulated cocultures in week 2 compared to controls. At week 3, the unstimulated control cocultures developed the expression of runx2 and desmin, and the cocultures that were stimulated with EMD and BMP-2 achieved significantly higher levels of these factors than any of the other conditions.


CONCLUSIONS
Signs of osteogenesis were present in the cocultures in unstimulated and stimulated conditions. However, in the stimulated condition, osteogenic markers were increased at earlier time points. As such, this model may provide a good method for the study of specific cellular processes that may lead to osteogenesis and eventually for understanding the regeneration of periodontal bone in vivo.
Neeley
Wendell W
WW
Department of Orthodontics, University of Texas Health Science Center, Dental School, San Antonio, TX 78229, USA.
Carnes
David L
DL
Cochran
David L
DL
D000704:Analysis of Variance; D055396:Bone Morphogenetic Protein 2 / Q000378:metabolism; D001861:Bone Regeneration / Q000502:physiology*; D002454:Cell Differentiation / Q000502:physiology*; D002478:Cells, Cultured; D018920:Coculture Techniques / Q000379:methods; D050659:Core Binding Factor Alpha 1 Subunit / Q000378:metabolism; D003746:Dental Enamel Proteins / Q000378:metabolism; D003893:Desmin / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology* / Q000502:physiology; D005347:Fibroblasts / Q000166:cytology* / Q000502:physiology; D006801:Humans; D055806:Microvessels / Q000166:cytology; D010012:Osteogenesis / Q000502:physiology*; D010513:Periodontal Ligament / Q000166:cytology*
C525718:BMP2 protein, human; D055396:Bone Morphogenetic Protein 2; D050659:Core Binding Factor Alpha 1 Subunit; D003746:Dental Enamel Proteins; D003893:Desmin; C493709:RUNX2 protein, human; C109069:enamel matrix proteins