Osteopontin expression in osteoblasts and osteocytes during bone formation under mechanical stress in the calvarial suture in vivo

Introduction: Bone formation is known to be stimulated by mechanical stress; however, molecules involved in stress-dependent regulation of bone formation have not yet been fully characterized. Extracellular matrix proteins such as osteopontin (OPN) could play a role in mediation of the mechanical stress signal to osteoblasts. However, the function of OPN in bone formation under mechanical force is not known. Therefore, we examined the expression and the role of OPN in bone formation in vivo under tensile mechanical stress. Materials and Methods: Sagittal sutures of mice were subjected to expansion mechanical stress by setting orthodontic spring wires, and OPN expression during bone formation within the suture gap was examined. Results: Expansion of the sutures resulted in bone formation at the edges of the parietal bones within the sagittal suture. Immunohistochemical analysis revealed abundant accumulation of OPN protein in the matrix of newly formed bone on the inner edge of the parietal bone within the mechanically expanded sutures. Osteoblasts forming bone within the suture subjected to tensile stress also exhibited high levels of OPN protein expression. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis indicated that OPN mRNA expression was enhanced in wild-type calvariae subjected to expansion force compared with the control calvariae where dead spring wires were set without expansion stress. In addition, type I collagen mRNA was also expressed in the calvariae under the mechanical stimuli. To understand the function of OPN, sagittal sutures in OPN-deficient mice were subjected the expansion stress, and bone formation within the suture to fill the expanded gap was compared with that observed in wild-type mice. OPN deficiency reduced bone formation at the edge of the parietal bone in contact with the expanded suture gap. Conclusions: These observations revealed that OPN plays a pivotal role in bone formation under tensile mechanical stress.