Growth pattern of ameloblasts in bell stage of human deciduous tooth germ in-vitro culture system

AIM:To set up a culture system to observe the growth pattern of human ameloblast-like cells,in order to understand the initiation and differentiation of ameloblast.METHODS:A mandible of 19 weeks human fetus was obtained and studied with approval of Ethic Committee of Peking University.X-ray digital image was taken.Then the consecutive sections were prepared with one half of the mandible,while with the other half,enamel organ epithelials were separated and digested with collagenase.The ameloblast-like cells were selectively cultured by differential adhering method and in K-SFM media.The expression of enamel matrix proteins was analyzed by RT-PCR and immunocytochemisty.RESULTS:Bell stage tooth germs were recognized.In this stage,the differentiated ameloblast aligned orderly and secreted large amounts of extracellular matrix.Mineralization initiated from enamel-dentinal junction.Cells isolated from the tooth organ,mostly are ameloblast-like cells,proliferated and differentiated in K-SFM.The ameloblast-like cells were immunopositive for cytokeratin 14.Enamelin,amelogenin and ameloblastin were detected by RT-PCR.CONCLUSION: K-SFM media could be used selectively to culture ameloblast-like cells.This study demonstrated that in bell stage the ameloblast secreted enamel matrix,and the earliest mineralization occurred in the enamel-dentinal junction.