Abstract
BackgroundHuman dental pulp stromal cells (hDPSCs) are promising sources of mesenchymal stem cells (MSCs) for bone tissue regeneration. Circular RNAs (circRNAs) have been demonstrated to play critical roles in stem cell osteogenic differentiation. Herein, we aimed to investigate the role of circAKT3 during osteogenesis of hDPSCs and the underlying mechanisms of its function.MethodsWe performed circRNA sequencing to investigate the expression profiles of circular RNAs during osteogenesis of hDPSCs. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) was performed to detect the expression pattern of circAKT3 and miR-206 in hDPSCs during osteogenesis. We knocked down circAKT3 and interfered the expression of miR-206 to verify their regulatory role in hDPSC osteogenesis. We detected hDPSCs mineralization by alkaline phosphatase (ALP) and Alizarin Red S (ARS) staining and used dual-luciferase reporter assay to validate the direct binding between circAKT3 and miR-206. To investigate in vivo mineralization, we performed subcutaneous transplantation in nude mice and used hematoxylin and eosin, Masson’s trichrome, and immunohistochemistry staining.ResultsTotally, 86 circRNAs were differentially expressed during hDPSC osteogenesis, in which 29 were downregulated while 57 were upregulated. circAKT3 was upregulated while miR-206 was downregulated during hDPSC osteogenesis. Knockdown of circAKT3 inhibited ALP/ARS staining and expression levels of osteogenic genes. circAKT3 directly interacted with miR-206, and the latter one suppressed osteogenesis of hDPSCs. Silencing miR-206 partially reversed the inhibitory effect of circAKT3 knockdown on osteogenesis. Connexin 43 (CX43), which positively regulates osteogenesis of stem cells, was predicted as a target of miR-206, and overexpression or knockdown of miR-206 could correspondingly decrease and increase the expression of CX43. In vivo study showed knockdown of circAKT3 suppressed the formation of mineralized nodules and expression of osteogenic proteins.ConclusionDuring osteogenesis of hDPSCs, circAKT3 could function as a positive regulator by directly sponging miR-206 and arresting the inhibitive effect of miR-206 on CX43 expression.
Highlights
Human dental pulp stromal cells are promising sources of mesenchymal stem cells (MSCs) for bone tissue regeneration
The intensity of alkaline phosphatase (ALP) staining was significantly increased after 7 days (OM 7D), and calcified nodules of Alizarin Red S (ARS) staining were apparently spotted after osteogenic induction for 14 days (OM 14D) (Fig. 1a, b)
The mRNA expression levels of ALP, Runt-related transcription factor 2 (RUNX2), and OCN were significantly upregulated in the group of osteogenic-induction medium (OM) 7D and OM 14D compared to OM 0D (Fig. 1c)
Summary
Human dental pulp stromal cells (hDPSCs) are promising sources of mesenchymal stem cells (MSCs) for bone tissue regeneration. Discovered by Gronthos et al [4], human dental pulp stromal cells (hDPSCs) are characterized by high levels of self-renewal and proliferation and multilineage differentiation capability [5]. They manifest similar clonogenic and proliferation properties to human bone marrow mesenchymal stromal cells (BMSCs) [4]. They can be more and non-invasively gained from removed teeth in comparison with BMSCs. Accumulating evidence has proved the potential of DPSCs in generating bone-like tissues and repairing bone loss [6,7,8]. HDPSCs represent rising candidates for therapies of bone tissue repair
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
