Ayumi Takahashi1, Takao Morita2, Kaori Murata3, Erika Minowa1, Azmeree Jahan3, Masato Saito1, Akihiko Tanimura4. 1. Department of Pediatric Dentistry, School of Dentistry, Health Sciences University of Hokkaido, Ishikari-Tobetsu, Hokkaido 061-0293, Japan. 2. Department of Biochemistry, Nippon Dental University, School of Life Dentistry at Niigata, Niigata, 1-8, Hamauracho, Chuo-ku, Niigata-Shi, Niigata, 951-8580, Japan. 3. Department of Pharmacology, School of Dentistry, Health Sciences University of Hokkaido, Ishikari-Tobetsu, Hokkaido 061-0293, Japan. 4. Department of Pharmacology, School of Dentistry, Health Sciences University of Hokkaido, Ishikari-Tobetsu, Hokkaido 061-0293, Japan. Electronic address: tanimura@hoku-iryo-u.ac.jp.
Abstract
BACKGROUND AND OBJECTIVE: Amelogenins are major components of extracellular matrix proteins in developing teeth, and regulate the growth of enamel crystals. They also function as signaling molecules in cell differentiation. This study aimed to determine the biological effects of amelogenins on the differentiation of HAT-7 dental epithelial cells and MC3T3-E1 pre-osteoblastic cells using full-length recombinant human amelogenin (rh-AMEL). DESIGN: rh-AMEL was expressed in a mammalian cell line (Expi293F™) and was purified by DDK agarose beads. Effects of rh-AMEL on differentiation were evaluated by Mineralization and Alkaline phosphatase (ALP) activity using Alizarin Red S staining and colorimetric substrate p-nitrophenol, respectively. RESULTS: Western blotting and silver staining confirmed the successful purification of rh-AMEL. Mineralization and ALP activity in HAT-7 cells were significantly higher after treatment with 4 μg/mL rh-AMEL, but not after treatment with Emdogain® (EMD). In MC3T3-E1 cells, on the other hand, rh-AMEL showed biphasic effects on differentiation. Treatment with low concentrations of rh-AMEL (0.001-0.1 μg/mL) and EMD (0.01-1 μg/mL) increased mineralization and ALP activity in MC3T3-E1 cells, whereas treatment with high concentrations of rh-AMEL (4 μg/mL) and EMD (100 μg/mL) had the opposite effect. CONCLUSION: High concentrations of rh-AMEL and EMD decreased the differentiation of MC3T3-E1 cells. By contrast, a high concentration of rh-AMEL, but not that of EMD, promoted the differentiation of HAT-7 cells. This study demonstrates that the effects of rh-AMEL on cell differentiation differ between HAT-7 and MC3T3-E1 cells, and suggests that different regions on AMEL may induce the differentiation of these cell types.
BACKGROUND AND OBJECTIVE: Amelogenins are major components of extracellular matrix proteins in developing teeth, and regulate the growth of enamel crystals. They also function as signaling molecules in cell differentiation. This study aimed to determine the biological effects of amelogenins on the differentiation of HAT-7 dental epithelial cells and MC3T3-E1 pre-osteoblastic cells using full-length recombinant human amelogenin (rh-AMEL). DESIGN:rh-AMEL was expressed in a mammalian cell line (Expi293F™) and was purified by DDK agarose beads. Effects of rh-AMEL on differentiation were evaluated by Mineralization and Alkaline phosphatase (ALP) activity using Alizarin Red S staining and colorimetric substrate p-nitrophenol, respectively. RESULTS: Western blotting and silver staining confirmed the successful purification of rh-AMEL. Mineralization and ALP activity in HAT-7 cells were significantly higher after treatment with 4 μg/mL rh-AMEL, but not after treatment with Emdogain® (EMD). In MC3T3-E1 cells, on the other hand, rh-AMEL showed biphasic effects on differentiation. Treatment with low concentrations of rh-AMEL (0.001-0.1 μg/mL) and EMD (0.01-1 μg/mL) increased mineralization and ALP activity in MC3T3-E1 cells, whereas treatment with high concentrations of rh-AMEL (4 μg/mL) and EMD (100 μg/mL) had the opposite effect. CONCLUSION: High concentrations of rh-AMEL and EMD decreased the differentiation of MC3T3-E1 cells. By contrast, a high concentration of rh-AMEL, but not that of EMD, promoted the differentiation of HAT-7 cells. This study demonstrates that the effects of rh-AMEL on cell differentiation differ between HAT-7 and MC3T3-E1 cells, and suggests that different regions on AMEL may induce the differentiation of these cell types.
Authors: Kristóf Kádár; Viktória Juhász; Anna Földes; Róbert Rácz; Yan Zhang; Heike Löchli; Erzsébet Kató; László Köles; Martin C Steward; Pamela DenBesten; Gábor Varga; Ákos Zsembery Journal: Int J Mol Sci Date: 2021-04-13 Impact factor: 5.923
Authors: Anna Földes; Thanyaporn Sang-Ngoen; Kristóf Kádár; Róbert Rácz; Ákos Zsembery; Pamela DenBesten; Martin C Steward; Gábor Varga Journal: Front Pharmacol Date: 2021-06-02 Impact factor: 5.810