Zhang Wei, Yuan Xiao
Department of Orthodontics II, The Affiliated Hospital of Qingdao University, Qingdao 266000, China; School of Stomatology of Qingdao University, Qingdao 266003, China.
Department of Orthodontics II, The Affiliated Hospital of Qingdao University, Qingdao 266000, China.
Arch Oral Biol. 2022 Jun;138:105414. doi: 10.1016/j.archoralbio.2022.105414. Epub 2022 Mar 24.
In this study, we aimed to explore the mechanism underlying microRNA-20a (miR-20a) in viability, migration and osteogenic/odontoblastic differentiation of dental pulp stem cells (DPSCs).
The results of Targetscan unveiled that interleukin (IL)- 8 might bind to miR-20a, which was confirmed by dual-luciferase reporter assay. MiR-20a expression was up-regulated or down-regulated in DPSCs, followed by measurement of cell viability and migration via MTT and wound healing assays. Then, IL-8 was over-expressed in miR-20a mimic-transfected DPSCs with or without nuclear factor (NF)-κB inhibitor. The effect of miR-20a on osteogenic/odontoblastic differentiation potential of DPSCs was determined by alkaline phosphatase and alizarin red S staining. The expression levels of IL-8, osteogenic differentiation indicators and NF-κB/p65 signaling pathway in cells were detected through quantitative real-time polymerase chain reaction (qRT-PCR) or western blot RESULTS: Contrary to the down-regulated miR-20a, up-regulated miR-20a not only reinforced the viability and migration of DPSCs, but also promoted cell osteogenic/odontoblastic differentiation potential and related gene expressions (alkaline phosphatase, osteocalcin, and dentin sialophosphoprotein (DSPP)). Furthermore, IL-8 was verified to be the target of miR-20a. IL-8 over-expression effectively suppressed the osteogenic/odontoblastic differentiation potential as well as inhibited the activation of NF-κB pathway in DPSCs induced by miR-20a mimic, but these effects could be counteracted by NF-κB inhibitor.
MiR-20a up-regulation accelerated the viability, migration and osteogenic/odontoblastic differentiation potential of DPSCs by regulating NF-κB/p65 signaling pathway via targeting IL-8, which might be a potential treatment target for dental diseases.
在本研究中,我们旨在探索微小RNA-20a(miR-20a)在牙髓干细胞(DPSCs)的活力、迁移和成骨/成牙本质细胞分化中的潜在机制。
Targetscan的结果显示白细胞介素(IL)-8可能与miR-20a结合,这通过双荧光素酶报告基因检测得到证实。在DPSCs中上调或下调miR-20a的表达,然后通过MTT和伤口愈合试验测量细胞活力和迁移。接着,在转染了miR-20a模拟物的DPSCs中过表达IL-8,同时添加或不添加核因子(NF)-κB抑制剂。通过碱性磷酸酶和茜素红S染色确定miR-20a对DPSCs成骨/成牙本质细胞分化潜能的影响。通过定量实时聚合酶链反应(qRT-PCR)或蛋白质免疫印迹法检测细胞中IL-8、成骨分化指标和NF-κB/p65信号通路的表达水平。
与下调的miR-20a相反,上调的miR-20a不仅增强了DPSCs的活力和迁移能力,还促进了细胞的成骨/成牙本质细胞分化潜能及相关基因表达(碱性磷酸酶、骨钙素和牙本质涎磷蛋白(DSPP))。此外,证实IL-8是miR-20a的靶标。IL-8的过表达有效抑制了miR-20a模拟物诱导的DPSCs的成骨/成牙本质细胞分化潜能,并抑制了NF-κB通路的激活,但这些作用可被NF-κB抑制剂抵消。
上调miR-20a通过靶向IL-8调节NF-κB/p65信号通路,加速了DPSCs的活力、迁移和成骨/成牙本质细胞分化潜能,这可能是牙科疾病的一个潜在治疗靶点。