Son Jang-Ho, Park Bong-Soo, Kim In-Ryoung, Sung Iel-Yong, Cho Yeong-Cheol, Kim Jung-Soo, Kim Yong-Deok
Department of Oral and Maxillofacial Surgery, Ulsan University Hospital, University of Ulsan College of Medicine, Ulsan, South Korea.
Department of Oral Anatomy, School of Dentistry, Pusan National University, Yangsan, South Korea.
Lasers Med Sci. 2017 Apr;32(3):533-541. doi: 10.1007/s10103-017-2145-6. Epub 2017 Jan 13.
Melatonin has anabolic effects on the bone, even under hypoxia, and laser irradiation has been shown to improve osteoblastic differentiation. The aim of this study was to investigate whether laser irradiation and melatonin would have synergistic effects on osteoblastic differentiation and mineralization under hypoxic conditions. MC3T3-E1 cells were exposed to 1% oxygen tension for the hypoxia condition. The cells were divided into four groups: G1-osteoblast differentiation medium only (as the hypoxic condition), G2-treatment with 50 μM melatonin only, G3-laser irradiation (808 nm, 80 mW, GaAlAs diode) only, and G4-treatment with 50 μM melatonin and laser irradiation (808 nm, 80 mW, GaAlAs diode). Immunoblotting showed that osterix expression was markedly increased in the melatonin-treated and laser-irradiated cells at 48 and 72 h. In addition, alkaline phosphatase activity significantly increased and continued to rise throughout the experiment. Alizarin Red staining showed markedly increased mineralized nodules as compared with only melatonin-treated or laser-irradiated cells at day 7, which significantly increased by day 14. Moreover, when melatonin-treated cells were laser-irradiated, the differentiation and mineralization of cells were found to involve p38 MAPK and PRKD1 signaling mechanisms. However, the enhanced effects of laser irradiation with melatonin were markedly inhibited when the cells were treated with luzindole, a selective melatonin receptor antagonist. Therefore, we concluded that laser irradiation could promote the effect of melatonin on the differentiation and mineralization of MC3T3-E1 cells under hypoxic conditions, and that this process is mediated through melatonin 1/2 receptors and PKRD/p38 signaling pathways.
褪黑素对骨骼具有合成代谢作用，即使在缺氧条件下也是如此，并且已证明激光照射可改善成骨细胞分化。本研究的目的是探讨在缺氧条件下激光照射和褪黑素对成骨细胞分化和矿化是否具有协同作用。将MC3T3-E1细胞置于1%氧张力的缺氧条件下。细胞分为四组：G1组仅用成骨细胞分化培养基（作为缺氧条件），G2组仅用50 μM褪黑素处理，G3组仅用激光照射（808 nm，80 mW，砷化镓铝二极管），G4组用50 μM褪黑素和激光照射（808 nm，80 mW，砷化镓铝二极管）。免疫印迹显示，在48小时和72小时时，褪黑素处理组和激光照射组细胞中的osterix表达明显增加。此外，碱性磷酸酶活性显著增加，并在整个实验过程中持续上升。茜素红染色显示，与仅褪黑素处理组或激光照射组细胞相比，在第7天矿化结节明显增加，到第14天显著增加。此外，当对褪黑素处理的细胞进行激光照射时，发现细胞的分化和矿化涉及p38丝裂原活化蛋白激酶（p38 MAPK）和蛋白激酶D1（PRKD1）信号机制。然而，当用选择性褪黑素受体拮抗剂鲁辛朵治疗细胞时，激光照射与褪黑素的增强作用被显著抑制。因此，我们得出结论，激光照射可促进褪黑素对缺氧条件下MC3T3-E1细胞分化和矿化的作用，并且这一过程是通过褪黑素1/2受体和PKRD/p38信号通路介导的。
