Mobilization of endothelial progenitor cells in fracture healing and distraction osteogenesis.
BACKGROUND
Fracture healing and distraction osteogenesis (DO) are unique postnatal bone formation processes, and neovascularization is critically required for successful bone regeneration. We investigated endothelial progenitor cell (EPC) mobilization during bone regeneration, and the possible contribution of EPCs to increased vascularization and new bone formation, especially in DO.


METHODS
Mouse tibia fracture and rat tibia DO models were used in this study. The proportion of EPCs among the peripheral and splenic mononuclear cells (MNCs) was determined by examining the endothelial lineage staining characteristics and EPC cell surface markers. Messenger RNA expression of molecules related to EPC mobilization and homing at the fracture site were analyzed by ribonuclease protection assay and reverse-transcription polymerase chain reaction. In the rat tibia DO model, we measured blood flow during DO, and determined the distribution of ex vivo-expanded and intravenously-infused EPCs.


RESULTS
The proportion of EPCs among the peripheral and splenic MNCs increased after fracture, peaked on post-fracture day 3, and returned to basal levels during the healing period. Messenger RNA expression of EPC mobilizing cytokines such as vascular endothelial growth factor (VEGF), stem cell factor, monocyte chemoattractant protein-1, and stromal cell-derived factor-1, were upregulated at the fracture callus. The plasma VEGF levels peaked prior to the increase in the EPC proportion. Adhesion molecules involved in EPC homing were expressed at the fracture callus. In the DO model, the temporal pattern of the increase in the EPC proportion was similar to that in the fracture healing model, but the EPC proportion increased again during the distraction and consolidation phases. The distraction gap was relatively ischemic during the distraction phase and blood flow increased profusely later in the consolidation phase. The number of EPCs homing to the bone regeneration site in the DO model correlated with the number of transplanted EPCs in a dose-dependent manner.


CONCLUSIONS
These findings suggest that signals from the bone regeneration site mobilize EPCs from the bone marrow into the peripheral circulation. Increased EPC mobilization and homing may contribute to neovascularization and thus to new bone formation in fracture healing and DO.
Lee
Dong Yeon
DY
Department of Orthopaedic Surgery, Seoul National University College of Medicine, South Korea. orthop@dreamwiz.com
Cho
Tae-Joon
TJ
Kim
Jin A
JA
Lee
Hye Ran
HR
Yoo
Won Joon
WJ
Chung
Chin Youb
CY
Choi
In Ho
IH
D000071916:AC133 Antigen; D000818:Animals; D015703:Antigens, CD / Q000032:analysis; D001861:Bone Regeneration / Q000502:physiology; D015815:Cell Adhesion Molecules / Q000378:metabolism; D002465:Cell Movement / Q000502:physiology*; D016207:Cytokines / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology* / Q000378:metabolism / Q000637:transplantation; D005260:Female; D017102:Fracture Healing / Q000502:physiology*; D006023:Glycoproteins / Q000032:analysis; D007963:Leukocytes, Mononuclear / Q000166:cytology / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D018919:Neovascularization, Physiologic; D019857:Osteogenesis, Distraction; D010455:Peptides / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D051116:Receptors, Scavenger / Q000032:analysis; D012039:Regional Blood Flow / Q000502:physiology; D033581:Stem Cell Transplantation; D013234:Stem Cells / Q000166:cytology* / Q000378:metabolism; D013977:Tibia / Q000098:blood supply / Q000254:growth & development; D013978:Tibial Fractures / Q000378:metabolism / Q000503:physiopathology; D042461:Vascular Endothelial Growth Factor A / Q000097:blood; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000032:analysis
D000071916:AC133 Antigen; D015703:Antigens, CD; D015815:Cell Adhesion Molecules; D016207:Cytokines; D006023:Glycoproteins; D010455:Peptides; D051116:Receptors, Scavenger; D042461:Vascular Endothelial Growth Factor A; C467480:vascular endothelial growth factor A, mouse; D040301:Vascular Endothelial Growth Factor Receptor-2