Department of Oral and Maxillofacial Surgery, The First Affiliated Hospital, Sun Yat-sen University & Guangdong Key Laboratory of Stomatology, Guangdong, China.
Dental Implant Department, Affiliated Zhongshan Hospital, Sun Yat-sen University, Zhongshan, Guangdong, China.
Biomed Pharmacother. 2019 Jan;109:1709-1717. doi: 10.1016/j.biopha.2018.10.159. Epub 2018 Nov 20.
Circular RNAs (circRNAs) comprise a novel class of noncoding RNAs that play important roles in a variety of diseases. However, the mechanism by which circRNAs regulate the osteogenic differentiation of maxillary sinus membrane stem cells (MSMSCs) remains largely unclear.
Microarray analysis was used to explore the expression profiles of circRNAs during the osteogenic differentiation of normal and BMP2 induced-MSMSCs. CircRNA_33287 was identified by agarose electrophoresis, quantitative real-time PCR (qRT-PCR), and western blotting. The function of circRNA_33287 was assessed by loss- and gain-of-function techniques and Alizarin red staining. Potential miRNA binding sites for circRNA_33287, and the target genes of miR-214-3p, were predicted by using online bioinformatics analysis tools. The relationships among the regulatory roles played by circRNA_33287, miR-214-3p, and Runt-related transcription factor 3 (Runx3), during the osteogenic differentiation of MSMSCs were verified by use of the dual luciferase reporter assay, qRT-PCR, and western blotting techniques, respectively. In addition, the molecular sponge potential of circRNA_33287 for miRNA was assessed via in vivo ectopic bone formation and a histological analysis performed after hematoxylin and eosin staining.
Expression of circRNA_33287 was confirmed to be up-regulated during the osteogenic differentiation of MSMSCS. Overexpression and silencing of circRNA_33287 increased and decreased the expression levels of key markers of osteogenesis, respectively, including Runx2, OSX, and ALP. Furthermore, circRNA_33287 acted as a molecular sponge for miR-214-3p, which regulated Runx3 expression by targeting its 3'UTR. Moreover, circRNA_33287 protected Runx3 from miR-214-3p-mediated suppression. In addition, circRNA_33287 was shown to increase ectopic bone formation in vivo and displayed the strongest ability to stimulate bone formation when co-transfected with a miR-214-3p inhibitor.
The novel pathway circRNA_33287/miR-214-3p/Runx3 was found to play a role in regulating the osteoblastic differentiation of MSMSCs in the posterior maxilla.
环状 RNA（circRNAs）是一类新型的非编码 RNA，在多种疾病中发挥重要作用。然而，circRNAs 调节上颌窦膜干细胞（MSMSCs）成骨分化的机制在很大程度上仍不清楚。
利用微阵列分析技术研究正常和 BMP2 诱导的 MSMSCs 成骨分化过程中 circRNAs 的表达谱。通过琼脂糖电泳、实时定量 PCR（qRT-PCR）和 Western blot 鉴定 circRNA_33287。通过缺失和获得功能技术以及茜素红染色评估 circRNA_33287 的功能。利用在线生物信息学分析工具预测 circRNA_33287 的潜在 miRNA 结合位点和 miR-214-3p 的靶基因。通过双荧光素酶报告基因检测、qRT-PCR 和 Western blot 技术分别验证 circRNA_33287、miR-214-3p 和 Runt 相关转录因子 3（Runx3）在 MSMSCs 成骨分化过程中的调控作用关系。此外，通过体内异位骨形成和苏木精-伊红染色后的组织学分析评估 circRNA_33287 作为 miRNA 的分子海绵的潜力。
证实 circRNA_33287 在 MSMSCS 成骨分化过程中表达上调。circRNA_33287 的过表达和沉默分别增加和降低了成骨关键标志物的表达水平，包括 Runx2、OSX 和 ALP。此外，circRNA_33287 作为 miR-214-3p 的分子海绵，通过靶向其 3'UTR 调节 Runx3 的表达。此外，circRNA_33287 保护 Runx3 免受 miR-214-3p 介导的抑制。此外，circRNA_33287 显示出在体内增加异位骨形成的能力，并且当与 miR-214-3p 抑制剂共转染时表现出最强的刺激骨形成的能力。
发现新型通路 circRNA_33287/miR-214-3p/Runx3 在调节上颌窦后骨 MSMSCs 的成骨分化中发挥作用。
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