Kim Hyun-Soo, Zheng Mingzhen, Kim Do-Kyung, Lee Won-Pyo, Yu Sang-Joun, Kim Byung-Ock
Department of Periodontology, Chosun University School of Dentistry, Gwangju, Korea.
Department of Stomatology, Affiliated Hospital of Yanbian University, Yanji, China.
J Periodontal Implant Sci. 2018 Feb 27;48(1):34-46. doi: 10.5051/jpis.2018.48.1.34. eCollection 2018 Feb.
The purpose of this study was to evaluate the effects of 1,25-dihydroxyvitamin D on the proliferation, differentiation, and matrix mineralization of MC3T3-E1 osteoblast-like cells
MC3T3-E1 osteoblastic cells and 1,25-dihydroxyvitamin D were prepared. Cytotoxic effects and osteogenic differentiation were evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, alkaline phosphatase (ALP) activity assay, ALP staining, alizarin red S staining, and reverse transcription-polymerase chain reaction (RT-PCR) for osteogenic differentiation markers such as ALP, collagen type I (Col-I), osteocalcin (OCN), vitamin D receptor (VDR), and glyceraldehyde 3-phosphate dehydrogenase.
The MTT assay showed that 1,25-dihydroxyvitamin D did not inhibit cell growth and that the rate of cell proliferation was higher than in the positive control group at all concentrations. ALP activity was also higher than in the positive control group at low concentrations of 1,25-dihydroxyvitamin D (10, 10, and 10 M). RT-PCR showed that the gene expression levels of ALP, Col-I, OCN, and vitamin D receptor (VDR) were higher at a low concentration of 1,25-dihydroxyvitamin D (10 M). Alizarin red S staining after treatment with 1,25-dihydroxyvitamin D (10 M) showed no significant differences in the overall degree of calcification. In contrast to the positive control group, formation of bone nodules was induced in the early stages of cell differentiation.
We suggest that 1,25-dihydroxyvitamin D positively affects cell differentiation and matrix mineralization. Therefore, it may function as a stimulating factor in osteoblastic bone formation and can be used as an additive in bone regeneration treatment.
本研究旨在评估1,25 - 二羟基维生素D对MC3T3 - E1成骨样细胞增殖、分化及基质矿化的影响。
制备MC3T3 - E1成骨细胞和1,25 - 二羟基维生素D。使用3-(4,5 - 二甲基噻唑 - 2 - 基)-2,5 - 二苯基四氮唑溴盐（MTT）法、碱性磷酸酶（ALP）活性测定、ALP染色、茜素红S染色以及针对成骨分化标志物如ALP、I型胶原（Col - I）、骨钙素（OCN）、维生素D受体（VDR）和甘油醛 - 3 - 磷酸脱氢酶的逆转录 - 聚合酶链反应（RT - PCR）来评估细胞毒性作用和成骨分化。
MTT法显示，1,25 - 二羟基维生素D不抑制细胞生长，且在所有浓度下细胞增殖率均高于阳性对照组。在低浓度的1,25 - 二羟基维生素D（10、10和10 μM）时，ALP活性也高于阳性对照组。RT - PCR表明，在低浓度的1,25 - 二羟基维生素D（10 μM）时，ALP、Col - I、OCN和维生素D受体（VDR）的基因表达水平更高。用1,25 - 二羟基维生素D（10 μM）处理后的茜素红S染色显示，钙化总体程度无显著差异。与阳性对照组相反，在细胞分化早期诱导形成了骨结节。
我们认为1,25 - 二羟基维生素D对细胞分化和基质矿化有积极影响。因此，它可能在成骨细胞骨形成中起刺激因子的作用，可作为骨再生治疗的添加剂。
