Hsa_circRNA_33287 promotes the osteogenic differentiation of maxillary sinus membrane stem cells via miR-214-3p/Runx3

Background: 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. Methods: 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. Results: 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. Conclusion: 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.