Circular RNA SIPA1L1 promotes osteogenesis via regulating the miR-617/Smad3 axis in dental pulp stem cells

BACKGROUND: Bone regeneration is preferred for bone loss caused by tumors, bone defects, fractures, etc. Recently, mesenchymal stem cells are considered as optimistic tools for bone defect therapy. Dental pulp stem cells (DPSCs) are a promising candidate for regenerative medicine and bone regeneration. Our previous study showed that upregulated circSIPA1L1 during osteogenesis of DPSCs is of significance. In this paper, the potential role of circSIPA1L1 in osteogenesis of DPSCs and its underlying mechanisms are explored. METHODS: The circular structure of circSIPA1L1 was identified by Sanger sequencing and PCR. Regulatory effects of circSIPA1L1 and miR-617 on mineral deposition in DPSCs were assessed by alkaline phosphatase (ALP) and alizarin red S (ARS) staining and in vivo bone formation assay were conducted to verify the biological influences of circSIPA1L1 on DPSCs. Western blot was performed to detect the protein expression of Smad3. Localization of circSIPA1L1 and miR-617 was confirmed by FISH. Dual-luciferase reporter assay and rescue experiments were conducted to investigate the role of the circSIPA1L1/miR-617/Smad3 regulatory axis in osteogenesis of DPSCs. RESULTS: Sanger sequencing and back-to-back primer experiments confirmed the closed-loop structure of circSIPA1L1. CircSIPA1L1 could promote the committed differentiation of DPSCs. MiR-617 was predicted to be the target binding circSIPA1L1 through MiRDB, miRTarBase, and TargetScan database analyses, which was further confirmed by dual-luciferase reporter assay. FISH results showed that circSIPA1L1 and miR-617 colocalize in the cytoplasm of DPSCs. MiR-617 exerted an inhibitory effect on the osteogenesis of DPSCs. Knockdown of circSIPA1L1 or upregulation of miR-617 downregulated phosphorylated Smad3. In addition, rescue experiments showed that knockdown of miR-617 reversed the inhibitory effect of circSIPA1L1 on osteogenesis of DPSCs. CONCLUSION: CircRNASIPA1L1 promotes osteogenesis of DPSCs by adsorbing miR-617 and further targeting Smad3.