685 Amelogenin Promotes Dentinogenesis And Cementogenesis Of Dental/periodontal Stem/progenitor Cells

Friday, March 23, 2012: 8 a.m. - 9:30 a.m.
Presentation Type: Oral Session
J. GU1, M. CHEN1, T. SHEU2, J. ZHOU1, S. WANG3, Y. SHEN1, C. LU1, R. YANG1, W. ZHAO1, and J. MAO1, 1Center for Craniofacial Regeneration, Columbia University, New York, NY, 2Department of Orthopaedics, University of Rochester School of Medicine and Dentistry, Rochester, NY, 3Stomatology hospital, Xi'an Jiaotong University, Xi'an, Shanxi, China
Objective: Amelogenin is the predominant matrix component of ameloblasts in the developing enamel. Interestingly, odontoblasts and periodontal ligament cells also synthesize amelogenin.  Recently, we showed that amelogenin acts as a signal molecule during mineralized tissue formation. Here, we further investigated the role of amelogenin in bioengineered odontogenesis and cementogenesis. 

Method: Mouse amelogenin gene was cloned into E. coli expression vector pDEST17 and mammalian expression vector pCMV6, respectively. Dental pulp (DP) and periodontal ligament (PDL) stem/progenitor cells were isolated per our prior methods. E.coli expressed recombinant mouse amelogenin (rM179) was purified using Cobalt-Sepharose6-resin affinity column. Wild-type DP and PDL cells were transfected with pCMV6-amelogenin vector and then subjected to G418 selection to obtain an exclusive population of amelogenin-expressing cells. The expression of amelogenin was confirmed with western-blot.  Differentiation was examined using ALP/von Kossa staining and real-time qPCR. In a separate experiment, DP or PDL cells were seeded in calcium phosphate scaffolds and implanted subcutaneously in the SCID mice. The tissue formation was analyzed by histology, histomorphometry and immunohistochemistry.

Result: 1) Recombinant amelogenin showed robust biological activity and induced the odontoblastic/osteogenic differentiation of PDL and DP cells as illustrated by calcified matrix deposition. Consistently, DSPP, DMP-1 were upregulated in DP cells, whereas CEMP-1 was up-regulated in PDL cells; 2) Compared to vector controls, DP and PDL cells constitutively expressing amelogenin (DP-AML, PDL-AML) showed enhanced calcified matrix deposition as well as increased marker gene expression; 3) The DP-AML cells became polarized and laid down DSPP positive matrix on the surface of calcium phosphate scaffold in vivo. The PDL-AML cells formed a cementum-like structure with cementum markers including CEMP-1 and CAP, in addition to putative Sharpey’s fibers. 

Conclusion: Amelogenin promotes dentinogenesis and cementogenesis both in tissue culture and in vivo. Additional large animal and human studies are warranted to investigate whether amelogenin orchestrates dentin and cementum regeneration.

This abstract is based on research that was funded entirely or partially by an outside source: NIH RC2DE020767 Grant to Jeremy J. Mao ADEA/William J. Gies Foundation Dental Research Scholarship to Jiafeng Gu

Keywords: Amelogenin, Dentin and Regeneration