Department of Oral and Maxillofacial Pathology, School of Dentistry, Kyung Hee University, Seoul 02447, Korea.
National Institute for Korean Medicine Development, Gyeongsan 38540, Korea.
Int J Mol Sci. 2021 Mar 2;22(5):2483. doi: 10.3390/ijms22052483.
Root bark of Turcz. has been widely used as a traditional medicine and is a well-known anti-inflammatory agent. We isolated limonoid triterpene, obacunone (Obac) from the dried root bark of . Obac has been reported to exhibit varieties of biological activities including anti-inflammatory, anti-cancer, and anti-oxidant effects. This study aimed to investigate the beneficial effects and biological mechanisms of Obac in osteoblast differentiation and bone matrix mineralization. In the present study, Obac at concentrations ranging from 1 to 100 μM showed no proliferation effects in MC3T3-E1. The treatment of Obac (1 and 10 μM) increased wound healing and migration rates in a dose-dependent manner. Alkaline phosphatase (ALP) staining and activity showed that Obac (1 and 10 μM) enhanced early osteoblast differentiation in a dose-dependent manner. Obac also increased late osteoblast differentiation in a dose-dependent manner, as indicated by the mineralized nodule formation of ARS staining. The effects of Obac on osteoblast differentiation was validated by the levels of mRNAs encoding the bone differentiation markers, including , bone sialoprotein (), osteopontin (), and osteocalcin (). Obac increased the expression of bone morphogenetic protein (BMP), and the phosphorylation of smad1/5/8, and the expression of runt-related transcription factor 2 (RUNX2); Obac also inhibited GSK3β and upregulated the protein level of β-catenin in a dose-dependent manner during osteoblast differentiation. Obac-mediated osteoblast differentiation was attenuated by a BMP2 inhibitor, Noggin and a Wnt/β-catenin inhibitor, Dickkopf-1 (Dkk1) with the abolishment of RUNX2 expression and nuclear accumulation by Obac. Taken together, the findings of this study demonstrate that Obac has pharmacological and biological activates to promote osteoblast differentiation and bone mineralization through BMP2, β-catenin, and RUNX2 pathways, and suggest that Obac might be a therapeutic effect for the treatment and prevention of bone diseases such as osteoporosis and periodontitis.
Turcz. 的根皮被广泛用作传统药物，是一种著名的抗炎剂。我们从干燥的 Turcz. 根皮中分离出了五环三萜类化合物，obacunone（Obac）。Obac 已被报道具有多种生物活性，包括抗炎、抗癌和抗氧化作用。本研究旨在探讨 Obac 在成骨细胞分化和骨基质矿化中的有益作用和生物学机制。在本研究中，浓度为 1 至 100 μM 的 Obac 对 MC3T3-E1 没有增殖作用。Obac（1 和 10 μM）的处理以剂量依赖性方式增加了伤口愈合和迁移率。碱性磷酸酶（ALP）染色和活性表明，Obac（1 和 10 μM）以剂量依赖性方式增强了早期成骨细胞分化。Obac 还以剂量依赖性方式增加了晚期成骨细胞分化，通过 ARS 染色的矿化结节形成来指示。Obac 对成骨细胞分化的作用通过编码骨分化标志物的 mRNA 水平得到验证，包括 、骨涎蛋白（）、骨桥蛋白（）和骨钙素（）。Obac 增加了骨形态发生蛋白（BMP）的表达，以及 smad1/5/8 的磷酸化和 runt 相关转录因子 2（RUNX2）的表达；Obac 还以剂量依赖性方式抑制 GSK3β，上调 β-连环蛋白的蛋白水平，在成骨细胞分化过程中。Obac 介导的成骨细胞分化被 BMP2 抑制剂 Noggin 和 Wnt/β-连环蛋白抑制剂 Dickkopf-1（Dkk1）减弱，Obac 消除了 RUNX2 的表达和核积累。总之，本研究的结果表明，Obac 具有药理学和生物学活性，通过 BMP2、β-连环蛋白和 RUNX2 途径促进成骨细胞分化和骨矿化，并表明 Obac 可能是治疗骨质疏松症和牙周炎等骨骼疾病的一种治疗效果。
