BACKGROUND AIMS

Previous studies identified a circulating human osteoblastic population that expressed osteocalcin (OCN), increased following fracture and pubertal growth, and formed mineralized colonies in vitro and bone in vivo. A subpopulation expressed CD34, a hematopoietic/endothelial marker. These findings led to our hypothesis that hematopoietic-derived CD34OCN cells exist in the circulation of mice and are modulated after fracture.

METHODS

Flow cytometry was used to identify CD34OCN cells in male B6.SJL-PtprcPepc/BoyJ and Vav-Cre/mTmG (VavR) mice. Non-stabilized tibial fractures were created by three-point bend. Fractures were longitudinally imaged by micro-computed tomography, and immunofluorescent staining was used to evaluate CD34OCN cells within fracture callus. AMD3100 (10 mg/kg) was injected subcutaneously for 3 days and the CD34OCN population was evaluated by flow cytometry.

RESULTS

Circulating CD34OCN cells were identified in mice and confirmed to be of hematopoietic origin (CD45; Vav1) using two mouse models. Both circulating and bone marrow-derived CD34OCN cells peaked three weeks post-non-stabilized tibial fracture, suggesting association with cartilage callus transition to bone and early mineralization. Co-expression of CD34 and OCN in the fracture callus at two weeks post-fracture was observed. By three weeks, there was 2.1-fold increase in number of CD34OCN cells, and these were observed throughout the fracture callus. AMD3100 altered CD34OCN cell levels in peripheral blood and bone marrow.

DISCUSSION

Together, these data demonstrate a murine CD34OCN circulating population that may be directly involved in fracture repair. Future studies will molecularly characterize CD34OCN cells, determine mechanisms regulating their contribution, and examine if their number correlates with improved fracture healing outcomes.