CircRNA Lrp6 promotes cementoblast differentiation via miR-145a-5p/Zeb2 axis
CircRNA Lrp6 promotes cementoblast differentiation via miR-145a-5p/Zeb2 axis
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DOI：
10.1111/jre.12933
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发表时间：
2021-09-07
影响因子：
3.5
通讯作者：
He, Hong
中科院分区：
文献类型：
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作者：
Li, Mengying;Du, Mingyuan;He, Hong
背景与目的牙骨质是牙周组织的一部分，是牙周韧带与牙槽骨的固定剂。成牙骨质细胞负责通过基质沉积和随后的矿化形成牙骨质。因此，探索成牙骨质细胞功能的新机制有助于牙骨质损伤的治疗。最近，circRNA Lrp 6（circLRP 6）由于其在细胞分化中的积极作用而受到关注，但其在成牙骨质细胞分化中的潜在作用仍不清楚。在此，我们试图阐明circLRP 6在成牙骨质细胞分化中的作用，并阐明任何相关的机制。材料与方法采用qRT-PCR检测circLRP 6、miR-145 a-5 p、锌指E-box结合同源框2（Zeb 2）、Runx 2、骨桥蛋白（Opn）和骨唾液蛋白（Bsp）的mRNA表达。Western blot检测Zeb 2蛋白表达水平。使用生物信息学分析和双荧光素酶报告基因测定来测试miR-145 a-5 p的潜在结合靶标。采用碱性磷酸酶（ALP）染色、ALP活性测定、茜素红S（ARS）染色及定量分析等方法评价成牙骨质细胞的分化潜能。结果在本研究中，circLRP 6在成牙骨质细胞分化中显著上调。此外，circLRP 6敲低抑制ALP水平，减少钙结节形成和Runx 2，Opn和Bsp的表达。在机械上，生物信息学分析和双荧光素酶报告基因测定证实miR-145 a-5 p是circLRP 6的潜在结合靶标。miR-145 a-5 p可负调控成牙骨质细胞分化。随后，生物信息学分析和双荧光素酶报告基因测定证实Zeb 2是潜在的miR-145 a-5 p靶标。miR-145 a-5 p过表达导致Zeb 2下调。此外，Zeb 2抑制部分逆转了成牙骨质细胞分化过程中circLRP 6的作用。结论circLRP 6可能通过拮抗miR-145 a-5 p的功能调节成牙骨质细胞分化，从而增加Zeb 2的表达。这项研究为促进牙骨质形成的方法的潜在发展奠定了基础。
Background and Objective Cementum is a part of the periodontium and anchors periodontal ligaments to the alveolar bone. Cementoblasts are responsible for the cementum formation via matrix deposition and subsequently mineralization. Thus, exploring novel mechanisms underlying the function of cementoblast contributes to the treatment of cementum damage. Recently, circRNA Lrp6 (circLRP6) has been of interest due to its active role in cell differentiation, but its potential role in cementoblast differentiation remains unclear. Herein, we attempted to elucidate the role of circLRP6 in cementoblast differentiation and clarify any associated mechanisms. Material and Methods The mRNA expressions of circLRP6, miR-145a-5p, zinc finger E-box binding homeobox 2 (Zeb2), runt-related transcription factor 2 (Runx2), osteopontin (Opn), and bone sialoprotein (Bsp) were evaluated by qRT-PCR. The protein levels of Zeb2 were measured by Western blot. Bioinformatic analysis and dual-luciferase reporter assays were used to test the potential binding targets of miR-145a-5p. The differentiation potentials of the cementoblasts were assessed by Alkaline phosphatase (ALP) staining, ALP activity assay, Alizarin red S (ARS) staining, and quantification. Results In this study, circLRP6 was significantly upregulated in cementoblast differentiation. Furthermore, circLRP6 knockdown inhibited ALP levels, reduced calcium nodule formation and the expression of Runx2, Opn, and Bsp. Mechanically, bioinformatic analysis and dual-luciferase reporter assays confirmed miR-145a-5p was a potential binding target of circLRP6. miR-145a-5p can negatively regulate cementoblast differentiation. Subsequently, bioinformatic analysis and dual-luciferase reporter assays confirmed Zeb2 was a potential miR-145a-5p target. miR-145a-5p overexpression resulted in a downregulation of Zeb2. Furthermore, Zeb2 inhibition partially reversed the effect of circLRP6 during cementoblast differentiation. Conclusion Taken together, circLRP6 appears to modulate cementoblast differentiation by antagonizing the function of miR-145a-5p, thereby increasing Zeb2. This study serves as a stepping stone for the potential development of an approach to promote cementum formation.