Daisuke Noro1, Yoshihito Kurashige2, Kai Shudo2, Ayumi Takahashi2, Yoshihiro Abiko3, Masato Saitoh4. 1. Division of Pediatric Dentistry, School of Dentistry, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Tobetsu, Hokkaido 061-0293, Japan. Electronic address: noro@hoku-iryo-u.ac.jp. 2. Division of Pediatric Dentistry, School of Dentistry, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Tobetsu, Hokkaido 061-0293, Japan. 3. Division of Oral Medicine and Pathology, School of Dentistry, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Tobetsu, Hokkaido 061-0293, Japan. 4. Division of Pediatric Dentistry, School of Dentistry, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Tobetsu, Hokkaido 061-0293, Japan. Electronic address: msaitoh@hoku-iryo-u.ac.jp.
Abstract
BACKGROUND AND OBJECTIVE: The epithelial rests of Malassez (ERM) are developmental residues of Hertwig's epithelial root sheath, and they are present throughout life in the periodontal ligament. Previous studies have shown that ERM induce cementogenesis or inhibit cement-osteogenesis. This study investigated how ERM cells are involved in osteoblast mineralization in an in vitro coculture system. MATERIALS AND METHODS: ERM cells were isolated from porcine periodontal ligament by an outgrowth method. Osteoblast-like MC3T3-E1 cells were cocultured with ERM or gingival epithelium (GE) cells in six-well Transwell units. Mineralization of MC3T3-E1 cells was confirmed by Alizarin Red staining after 30 days of culture. Alkaline phosphatase (ALP) activity was measured in the culture media. To examine whether enamel matrix proteins are involved in the mineralization of MC3T3-E1 cells, an anti-amelogenin antibody was added to the culture media. RESULTS: The staining by Alizarin Red of MC3T3-E1 cells cocultured with ERM cells was clearly weaker than that in the GE cell coculture. The ALP activity in the ERM cell coculture was significantly lower than that in the GE cell coculture (p < 0.05). Alizarin Red staining of MC3T3-E1 cells in the ERM cell coculture with anti-amelogenin antibody was clearly stronger than in those cocultures without the antibody at 30 days. The ALP activity in the ERM cell coculture with anti-amelogenin antibody was significantly higher than in those cultures without the antibody (p < 0.05). CONCLUSION: This study demonstrated that ERM cells inhibit the calcification of osteoblast-like cells in a Transwell coculture system and that this inhibition is reversed with an anti-amelogenin antibody.
BACKGROUND AND OBJECTIVE: The epithelial rests of Malassez (ERM) are developmental residues of Hertwig's epithelial root sheath, and they are present throughout life in the periodontal ligament. Previous studies have shown that ERM induce cementogenesis or inhibit cement-osteogenesis. This study investigated how ERM cells are involved in osteoblast mineralization in an in vitro coculture system. MATERIALS AND METHODS: ERM cells were isolated from porcine periodontal ligament by an outgrowth method. Osteoblast-like MC3T3-E1 cells were cocultured with ERM or gingival epithelium (GE) cells in six-well Transwell units. Mineralization of MC3T3-E1 cells was confirmed by Alizarin Red staining after 30 days of culture. Alkaline phosphatase (ALP) activity was measured in the culture media. To examine whether enamel matrix proteins are involved in the mineralization of MC3T3-E1 cells, an anti-amelogenin antibody was added to the culture media. RESULTS: The staining by Alizarin Red of MC3T3-E1 cells cocultured with ERM cells was clearly weaker than that in the GE cell coculture. The ALP activity in the ERM cell coculture was significantly lower than that in the GE cell coculture (p < 0.05). Alizarin Red staining of MC3T3-E1 cells in the ERM cell coculture with anti-amelogenin antibody was clearly stronger than in those cocultures without the antibody at 30 days. The ALP activity in the ERM cell coculture with anti-amelogenin antibody was significantly higher than in those cultures without the antibody (p < 0.05). CONCLUSION: This study demonstrated that ERM cells inhibit the calcification of osteoblast-like cells in a Transwell coculture system and that this inhibition is reversed with an anti-amelogenin antibody.