1163 Immunomodulation of Growth Factor Microenvironment Activates Bone Healing

Friday, March 23, 2012: 3:30 p.m. - 4:45 p.m.
Presentation Type: Poster Session
M.O. FREIRE, Craniofacial Biology, University of Southern California, Los Angeles, CA, A. NGUYEN, Department of Periodontics, University of Southern California, Los Angeles, CA, J. KOOK, Oral Biochemistry, Chosun University, Kwang-ju, Republic Of Korea, and H. ZADEH, Dept. of Periodontology, University of Southern California, Los Angeles, CA
Objective :The purpose of this study was to characterize the  biological and molecular mechanisms of tethered monoclonal antibodies (mAbs) in the extracellular microenvironment.

Methods : In vivo analyses was performed by immobilizing Anti-BMP-2 mAb , isotype control Ab (25 ug/ml), were adsorbed in each of the scaffolds by incubation at room temperature for 1 hour. We treated the critical size defects of both rat and rabbits with the matrices. . To investigate  protein expression of BMP-2,-4,-7 and presence of abs in early stages ( 1,3,7 days) after surgical implantation, confocal analysis was utilized. After 4 and 6 weeks, bone formation was evaluated morphologically by Micro-CT scan and histology, and molecularly by immunohistochemistry of BMP-2 and osteocalcin. To characterize Ab binding kinetics  in vitro, ElISA was developed. Epitope mapping analysis was studied by protein array.

Results:  Bone Regeneration  revealed by micro-CT and histological analysis  after4 and 6 weeks demonstrated that in both rat and rabbit wounds there was a significant increase  bone formation (p<0.001)  when compared to isotype controls. Confocal analyses of in situ  expression BMPs showed  increase in BMP-2,-4,-7 after 1 to 7 days. In addition, after 4 and6 weeks, BMP-2 and osteocalcin expression was increased in bone regeneration sites only. In vitro  cross reactivity of Abs by western blots was positive for BMP-2,-7. Kinetics Abs and BMPs interaction showed high dissociation constant  (kD=11nM). Protein array utilizing 12 overlapping peptides of each mature BMPs, showed that  the N-terminus region of BMP-2,-7 was a positive functional region of Ab binding.

Conclusion: This data provide the first molecular elucidation of the interaction of AMOR and BMP-2,-4,-7. These  antibodies modulate the extracellular matrix,  forming a complex  with BMPs in vivo, increasing its local concentration and mediate bone regeneration in vivo, suggesting a novel role of monoclonal antibodies in bioengineering.

Keywords: Antigens-antibodies, Bioengineering, Extracellular matrix molecules, Therapeutics and Wound healing