Immunohistochemical Analysis of Amelotin Expression in Ameloblastomas

Ameloblastomas are the most frequent odontogenic tumors from epithelial origin. Although considered benign, they are locally aggressive and present a high rate of recurrence. The greatest period of prevalence is observed between the 2^nd and the 5^th decades of life, and a slow-growth pattern is characteristic. The AMTN (Amelotin) gene was recently described as an ameloblast-specific gene, highly but transiently expressed in ameloblasts of the maturation stage enamel organ before tooth eruption. Although its expression pattern and preliminary functional studies in vivo suggest a fundamental role for this protein in the formation and mineralization of dental enamel, its specific function is unknown. Objectives: To determine the expression pattern of AMTN in clinical cases of human ameloblastoma to investigate a possible association of this protein with tumoral status of ameloblast cells. Methods: Immunohistochemistry analysis was performed in human ameloblastoma sections provided by the Department of Oral Pathology, University of Toronto (n=4). Sections of a case of ameloblastic fibro-odontome were used as a control. Rabbit polyclonal antibodies against two peptide sequences of human AMTN were utilized at a dilution of 1:1000. For the negative control, the primary antibody was replaced by pre-immune serum at the same concentration. Antigen retrieval methods were performed to enhance the signal. Results: Expression of AMTN was clearly marked in the nucleus of the cuboidal or columnar epithelial cells that constitute the tumor islands. The central mass of loosely connected cells or cystic areas showed a negative staining, and the fibrous stroma presented a faint signal. Conclusion: AMTN is clearly expressed by ameloblastoma epithelial cells. The nuclear localization in these cells was unexpected, since AMTN is a secreted protein that accumulates in the interface between mature ameloblasts and the enamel surface during normal development. Grants: Canadian Institutes of Health Research.