Colocalization of c-Src (pp60src) and bone morphogenetic protein 2/4 expression during mandibular distraction osteogenesis: in vivo evidence of their role within an integrin-mediated mechanotransduction pathway.
Craniofacial distraction osteogenesis (DO) is an evolving reconstructive technique with expanding applications for the treatment of bony deficiencies of the facial skeleton. Mechanical force has been known to play a fundamental role in modulating sustained osteogenic response and therefore is believed to function as a critical regulator of DO. We hypothesize that key clustering components of an integrin-mediated signaling pathway, including c-Src (pp60), are necessary for mediating the response to mechanical force. The specific aim of this study is to demonstrate up-regulation of a key focal adhesion molecule, c-Src, selectively in new bone formation subject to the mechanical forces of distraction and to demonstrate a lack of that same up-regulation in new bone formation associated with simple fracture healing. An additional specific aim is to demonstrate colocalization of c-Src expression and bone morphogenetic protein (BMP 2/4) expression during mandibular DO. Using a rat model of mandibular DO, c-Src and BMP 2/4 expression were evaluated in critical size defects, subcritical size defects, and mandibles undergoing gradual distraction. Osseous regeneration was observed in the course of gradual distraction; this process was associated with increased expression of c-Src. Furthermore, the presence of BMP 2/4 closely approximated c-Src expression spatially and temporally, suggesting a link between cytoplasmic focal adhesion activation and the resultant nuclear regulation of osteogenic protein expression. In significant contradistinction, minimal c-Src expression was found in the subcritical-sized defects where the fractures healed secondarily but where no gradual distraction was performed. Instead, the new bone formation inherent in the secondarily healed subcritical-sized defects demonstrated expected BMP 2/4 expression but was devoid of an up-regulation of c-Src. Finally, as expected, minimal expression of both c-Src and BMP was found in fibrous nonunion specimens. C-src expression was observed during gradual distraction; furthermore, minimal c-Src expression was visualized during subacute and critical-size defect fracture healing. C-Src expression also closely approximated BMP expression during DO. These findings that c-Src expression is found primarily only during conditions of cyclic distraction forces strongly implicates that mechanical force during gradual distraction is associated with c-Src expression. These results provide in vivo support for previous in vitro evidence that mechanical force profoundly influences osseous regeneration during distraction osteogenesis by means of a c-Src dependent mechanotransduction pathway, resulting in increased expression of osteogenic proteins, including BMP 2/4.
Rhee
Samuel T
ST
Section of Plastic Surgery, University of Michigan Medical Center, Ann Arbor, MI, USA.
Buchman
Steven R
SR
D000818:Animals; D055396:Bone Morphogenetic Protein 2; D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins / Q000235:genetics*; D001861:Bone Regeneration / Q000502:physiology*; D051417:Focal Adhesion Kinase 1 / Q000235:genetics; D006984:Hypertrophy / Q000473:pathology; D007150:Immunohistochemistry; D016023:Integrins / Q000502:physiology*; D008334:Mandible / Q000473:pathology / Q000502:physiology*; D040542:Mechanotransduction, Cellular / Q000502:physiology*; D009434:Neural Pathways / Q000473:pathology*; D019857:Osteogenesis, Distraction; D011505:Protein-Tyrosine Kinases / Q000235:genetics; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000235:genetics*; D051381:Rats; D017207:Rats, Sprague-Dawley; D016212:Transforming Growth Factor beta / Q000235:genetics*
C525720:Bmp2 protein, rat; C525811:Bmp4 protein, rat; D055396:Bone Morphogenetic Protein 2; D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins; D016023:Integrins; D016212:Transforming Growth Factor beta; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D016392:Proto-Oncogene Proteins pp60(c-src); C496052:Ptk2 protein, rat