title: Establishment and characterization of a primary calcifying epithelial odontogenic tumor cell population.
abstract: Calcifying epithelial odontogenic tumors (CEOTs) are rare neoplasms derived from dental tissue with the unique characteristic of calcifying amyloid-like material.
  
  
  OBJECTIVE
  To establish primary CEOT epithelial-derived cell populations, investigate the expression of enamel matrix proteins (EMPs), and identify potential ameloblastin (AMBN) and patched 1 (PTCH1) gene alterations.
  
  
  METHODS
  A 28-year-old patient with a lesion of the posterior maxilla, radiographically characterized by a radiolucency with well-defined borders containing mixed radiopacities, agreed to participate with informed consent. The patient's biopsy confirmed the diagnosis of CEOT, and a small representative tumor fragment was ascertained for cell culture. Explant cultures were established and used to establish primary cell populations. These were analyzed for morphology, cell proliferation, mineralization activity, expression of epithelial-associated markers (qRT-PCR and immunocytochemistry), and gene mutations of AMBN or PTCH1. DNA was extracted from tumor cells and gene coding and exon-intron boundaries overlapping fragments amplified. PCR products were bidirectional DNA sequenced and compared against reference sequence.
  
  
  RESULTS
  A CEOT cell population was established and proliferated in culture and could be maintained for several passages. Expression of EMPs, cytokeratin 14 and 17, and patched (PTCH1), as well as ALP activity, was detected. These cells also had the ability to mineralize, similar to the primary tumor. Two AMBN alterations were identified in the sample: c.1323G>A/A441A (rs7680880) and c.1344*+111delA. Two single-nucleotide polymorphisms were identified in the PTCH1 gene.
  
  
  CONCLUSIONS
  Our data support the establishment of a CEOT-derived cell population, which expresses known epithelial-associated proteins.
authors.affiliation: Institute of Oral Health Research, University of Alabama at Birmingham, Birmingham, AL, USA.
authors.forename: Hope M
authors.initials: HM
authors.lastname: Amm
authors.forename: Douglas L
authors.initials: DL
authors.lastname: Rollins
authors.forename: Changchun
authors.initials: C
authors.lastname: Ren
authors.forename: Juan
authors.initials: J
authors.lastname: Dong
authors.forename: Patricia
authors.initials: P
authors.lastname: DeVilliers
authors.forename: Helen
authors.initials: H
authors.lastname: Rivera
authors.forename: Mary
authors.initials: M
authors.lastname: MacDougall
mesh_terms: D000328:Adult; D000469:Alkaline Phosphatase / Q000032:analysis; D002114:Calcinosis / Q000473:pathology; D018929:Cell Culture Techniques; D049109:Cell Proliferation; D048430:Cell Shape; D002478:Cells, Cultured; D004273:DNA, Neoplasm / Q000235:genetics; D003746:Dental Enamel Proteins / Q000032:analysis / Q000235:genetics; D004847:Epithelial Cells / Q000473:pathology; D005091:Exons / Q000235:genetics; D006801:Humans; D007438:Introns / Q000235:genetics; D053547:Keratin-14 / Q000032:analysis; D053537:Keratin-17 / Q000032:analysis; D009154:Mutation / Q000235:genetics; D009808:Odontogenic Tumors / Q000737:chemistry / Q000235:genetics / Q000473:pathology*; D000072079:Patched Receptors; D000072081:Patched-1 Receptor; D011956:Receptors, Cell Surface / Q000032:analysis / Q000235:genetics; D017422:Sequence Analysis, DNA; D012878:Skin Neoplasms / Q000737:chemistry / Q000235:genetics / Q000473:pathology*
chemical_list: C490889:AMBN protein, human; D004273:DNA, Neoplasm; D003746:Dental Enamel Proteins; D053547:Keratin-14; D053537:Keratin-17; C000606236:PTCH1 protein, human; D000072079:Patched Receptors; D000072081:Patched-1 Receptor; D011956:Receptors, Cell Surface; C109069:enamel matrix proteins; D000469:Alkaline Phosphatase
keywords: ameloblastin; calcifying epithelial odontogenic tumor; enamel matrix proteins; odontogenic tumors; patched