title: Inhibitory effects of ameloblastin on epithelial cell proliferation.
abstract: OBJECTIVE
  Ameloblastin is an enamel matrix protein expressed in several tissues. Many potential mechanisms have been identified by which ameloblastin functions as an extracellular matrix protein. However, the biological effects of ameloblastin on gingival epithelial cells remain unclear. In the present study, we established a novel system to purify recombinant human ameloblastin and clarified its biological functions in epithelial cells in vitro.
  
  
  METHODS
  Recombinant human ameloblastin was isolated from COS-7 cells overexpressing HaloTag-fused human ameloblastin by the HaloTag system and then purified further by reverse-phase high-performance liquid chromatography. SCC-25 cells, derived from human oral squamous cell carcinoma, were treated with recombinant ameloblastin and then cell survival was assessed by a WST-1 assay. Cell cycle analysis was performed by flow cytometry.
  
  
  RESULTS
  The novel purification system allowed effective recovery of the recombinant ameloblastin proteins at a high purity. Recombinant ameloblastin protein was found to suppress the proliferation of SCC-25 cells. Flow cytometric analysis showed that ameloblastin treatment induced cell cycle arrest G1 phase.
  
  
  CONCLUSIONS
  We developed a procedure for production of highly purified recombinant human ameloblastin. Biological analyses suggest that ameloblastin induces cell cycle arrest in epithelial cells and regulates the progression of periodontitis.
authors.affiliation: Division of Infections and Molecular Biology, Department of Health Promotion, Kyushu Dental University, 2-6-1 Manazuru, Kokurakita-ku, Kitakyushu, Fukuoka 803-8580, Japan; Division of Pulp Biology, Operative Dentistry and Endodontics, Department of Cariology and Periodontology, Kyushu Dental University, 2-6-1 Manazuru, Kokurakita-ku, Kitakyushu, Fukuoka 803-8580, Japan.
authors.forename: Noriko
authors.initials: N
authors.lastname: Saito
authors.affiliation: Division of Infections and Molecular Biology, Department of Health Promotion, Kyushu Dental University, 2-6-1 Manazuru, Kokurakita-ku, Kitakyushu, Fukuoka 803-8580, Japan. Electronic address: arikichi@kyu-dent.ac.jp.
authors.forename: Wataru
authors.initials: W
authors.lastname: Ariyoshi
authors.affiliation: Division of Infections and Molecular Biology, Department of Health Promotion, Kyushu Dental University, 2-6-1 Manazuru, Kokurakita-ku, Kitakyushu, Fukuoka 803-8580, Japan.
authors.forename: Toshinori
authors.initials: T
authors.lastname: Okinaga
authors.affiliation: Genenet Co., Ltd, 5-22-8, Tanotsu, Higashi-Ku, Fukuoka 813-0034, Japan.
authors.forename: Mariko
authors.initials: M
authors.lastname: Kamegawa
authors.affiliation: Genenet Co., Ltd, 5-22-8, Tanotsu, Higashi-Ku, Fukuoka 813-0034, Japan.
authors.forename: Miho
authors.initials: M
authors.lastname: Matsukizono
authors.affiliation: Genenet Co., Ltd, 5-22-8, Tanotsu, Higashi-Ku, Fukuoka 813-0034, Japan.
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Akebiyama
authors.affiliation: Division of Pulp Biology, Operative Dentistry and Endodontics, Department of Cariology and Periodontology, Kyushu Dental University, 2-6-1 Manazuru, Kokurakita-ku, Kitakyushu, Fukuoka 803-8580, Japan.
authors.forename: Chiaki
authors.initials: C
authors.lastname: Kitamura
authors.affiliation: Division of Infections and Molecular Biology, Department of Health Promotion, Kyushu Dental University, 2-6-1 Manazuru, Kokurakita-ku, Kitakyushu, Fukuoka 803-8580, Japan.
authors.forename: Tatsuji
authors.initials: T
authors.lastname: Nishihara
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002294:Carcinoma, Squamous Cell / Q000188:drug therapy*; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D003746:Dental Enamel Proteins / Q000494:pharmacology*; D004847:Epithelial Cells / Q000378:metabolism*; D005434:Flow Cytometry; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D008807:Mice, Inbred BALB C; D009062:Mouth Neoplasms / Q000188:drug therapy*; D011994:Recombinant Proteins / Q000494:pharmacology*
chemical_list: D003746:Dental Enamel Proteins; D011994:Recombinant Proteins; C109069:enamel matrix proteins
keywords: Ameloblastin; Apoptosis; Cell cycle; Epithelial cell proliferation; Periodontitis; Protein purification