Department of Periodontology, School of Dentistry and Dental Research Institute, Seoul National University, Seoul, Korea.
Department of Molecular Genetics, School of Dentistry and Dental Research Institute, Seoul National University.
J Periodontol. 2017 Feb;88(2):e42-e48. doi: 10.1902/jop.2016.160294. Epub 2016 Sep 13.
In previous studies by the authors, it was demonstrated that a fibronectin (FN)-derived oligopeptide, termed F20, stimulates osteoblast differentiation in vitro and bone formation in vivo. However, the fundamental molecular mechanism by which F20 stimulates osteogenesis remains unknown. Therefore, in this study the molecular mechanism underlying the effect of F20 in osteoblast differentiation is investigated.
The role of F20 in osteoblast differentiation was examined using mouse bone-marrow-derived ST2 cell line. The effect of Smad1/5 was determined following small interfering RNA knockdown. Runt-related transcription factor (Runx) 2, alkaline phosphatase (Alp), and osteocalcin (Oc) mRNA levels were determined by quantitative real-time polymerase chain reaction, and their transcriptional activation was assessed using luciferase reporter assays. Extracellular signal-regulated kinase (ERK) phosphorylation was visualized via immunoblotting.
Synthetic oligopeptide F20 stimulated expression of bone marker genes Runx2, Alp, and Oc in ST2 cells via Smad and ERK or mitogen-activated protein kinase signaling pathways as did bone morphogenic protein 2 (BMP2). Furthermore, Runx2 acted as a transcription factor during F20-induced osteoblast differentiation.
Collectively, these results indicate that F20 induces osteoblast differentiation with a pattern similar to that mediated by BMP2 signaling pathway. The authors' previous data also showed that FN-derived oligopeptide improved wound healing, and it is suggested that F20 might serve as a therapeutic biomolecule to facilitate periodontal tissue regeneration.
作者之前的研究表明，一种纤维连接蛋白（FN）衍生的寡肽，称为 F20，可刺激体外成骨细胞分化和体内骨形成。然而，F20 刺激成骨的基本分子机制尚不清楚。因此，本研究旨在探讨 F20 刺激成骨作用的分子机制。
使用小鼠骨髓源性 ST2 细胞系研究 F20 在成骨细胞分化中的作用。采用小干扰 RNA 敲低法确定 Smad1/5 的作用。通过实时定量聚合酶链反应测定 runt 相关转录因子（Runx）2、碱性磷酸酶（Alp）和骨钙素（Oc）的 mRNA 水平，并通过荧光素酶报告基因测定评估其转录激活。通过免疫印迹法可视化细胞外信号调节激酶（ERK）磷酸化。
合成寡肽 F20 通过 Smad 和 ERK 或丝裂原活化蛋白激酶信号通路刺激 ST2 细胞中骨标志物基因 Runx2、Alp 和 Oc 的表达，骨形态发生蛋白 2（BMP2）也是如此。此外，Runx2 在 F20 诱导的成骨细胞分化过程中作为转录因子发挥作用。
综上所述，这些结果表明 F20 通过类似于 BMP2 信号通路介导的方式诱导成骨细胞分化。作者之前的数据还表明，FN 衍生的寡肽可改善伤口愈合，因此 F20 可能作为一种治疗性生物分子促进牙周组织再生。
