miR-93-5p在骨碎补总黄酮介导的兔骨髓间充质干细胞成骨分化中的作用及机制

doi:10.19438/j.cjoms.2023.05.004

中国口腔颌面外科杂志 ›› 2023, Vol. 21 ›› Issue (5): 446-451.doi: 10.19438/j.cjoms.2023.05.004

miR-93-5p在骨碎补总黄酮介导的兔骨髓间充质干细胞成骨分化中的作用及机制

申晓靖¹, 李林林², 路遥³, 王乐为³, 袁荣涛¹, 郭庆圆¹, 赵鹏¹

1.康复大学青岛医院（青岛市市立医院）口腔科,山东青岛 266000;
2.青岛市中医医院（海慈医疗集团）干部保健科,山东青岛 266000;
3.陇南市武都区第一人民医院,甘肃陇南 746000

收稿日期:2022-11-22 修回日期:2023-02-15 出版日期:2023-09-20 发布日期:2023-10-11
通讯作者: 赵鹏,E-mail：zhaopengyayi@163.com
作者简介:申晓靖（1986-）,女,硕士研究生,主治医师,E-mail：sxj18660288008@163.com
基金资助:
山东省中医药科技重点项目（Z-2022008）; 山东省医药卫生科技发展计划项目（202008030332）

The role and mechanism of miR-93-5p in osteogenic differentiation of rabbit bone marrow mesenchymal stem cells mediated by total flavonoids of rhizoma drynariae

SHEN Xiao-jing¹, LI Lin-lin², LU Yao³, WANG Le-wei³, YUAN Rong-tao¹, GUO Qing-yuan¹, ZHAO Peng¹

1. Department of Stomatology, Qingdao Hospital, University of Health and Rehabilitation Sciences(Qingdao Municipal Hospital). Qingdao 266000, Shandong Province;
2. Cadre Health Care Department, Qingdao Hospital of Traditional Chinese Medicine (Haici Medical Group). Qingdao 266000, Shandong Province;
3. First People's Hospital of Wudu District, Gansu Longnan. Longnan 746000, Gansu Province, China

Received:2022-11-22 Revised:2023-02-15 Online:2023-09-20 Published:2023-10-11

摘要/Abstract

摘要： 目的：探讨miR-93-5p在骨碎补总黄酮（TFRD）介导的兔骨髓间充质干细胞(rBMSCs)成骨分化中的作用。方法：采用成骨培养液对rBMSCs进行成骨诱导分化,通过茜素红S和油红O染色分别检测rBMSCs的成骨和成脂分化。运用荧光定量PCR（qPCR）检测miR-93-5p的表达,Western免疫印迹检测FZD6、ALP、Runx2和WNT通路相关蛋白（Dishevelled和β-catenin）的蛋白表达水平。双荧光素酶报告基因分析法检验miR-93-5p和FZD6的结合。采用SPSS 22.0软件包对数据进行统计学分析。结果：与对照组相比,TFRD促进rBMSCs的成骨分化,抑制miR-93-5p的表达,促进FZD6和成骨相关基因ALP和Runx2的表达。双荧光素酶报告基因分析表明,FZD6是miR-93-5p的下游靶基因。过表达miR-93-5p抑制rBMSCs的成骨分化和FZD6、ALP和Runx2的蛋白表达,而过表达FZD6逆转miR-93-5p的抑制作用。使用WNT通路抑制剂（KYA1797K）处理rBMSCs,可逆转TFRD对成骨分化的促进作用,导致成骨细胞分化减少,ALP、Runx2和WNT通路相关蛋白（Dishevelled和β-catenin）表达下调。结论：TFRD通过下调miR-93-5p,促进FZD6的表达,激活WNT信号通路,促进rBMSCs成骨分化,从而有助于口腔种植体骨结合。

关键词: 骨碎补总黄酮, miR-93-5p, FZD6, WNT通路

Abstract: PURPOSE: To investigate the role of miR-93-5p in osteogenic differentiation of rabbit bone marrow mesenchymal stem cells (rBMSCs) mediated by total flavonoids of rhizoma drynariae(TFRD). METHODS: The osteogenic differentiation of rBMSCs was induced by osteogenic medium. The osteogenic and adipogenic differentiation of rBMSCs were detected by Alizarin red S and oil red O staining, respectively. The expression of miR-93-5p was detected by fluorescence quantitative PCR (qPCR), and the expression level of FZD6, ALP, Runx2 and WNT pathway related proteins (Dishevelled and β-catenin) was detected by Western blotting. The binding of miR-93-5p and FZD6 was tested by double-luciferase reporter gene analysis. The data were analyzed with SPSS 22.0 software package. RESULTS: Compared with the control group, TFRD promoted the osteogenic differentiation of rBMSCs, inhibited the expression of miR-93-5p, and promoted the expression of FZD6 and osteogenic related genes ALP and Runx2. Double-luciferase reporter gene analysis showed that FZD6 was the downstream target gene of miR-93-5p. Overexpression of miR-93-5p inhibited the osteogenic differentiation of rBMSCs and the protein expression of FZD6, ALP and Runx2, while overexpression of FZD6 reversed the inhibitory effect of miR-93-5p. Treatment of rBMSCs with WNT pathway inhibition (KYA1797K) could reverse the promoting effect of TFRD on osteogenic differentiation, resulting in reduced osteoblast differentiation and downregulation of ALP, Runx2 and WNT pathway related proteins (Dishevelled and β-catenin). CONCLUSIONS: TFRD promote the osteogenic differentiation of rBMSCs by downregulating miR-93-5p and upregulating FZD6 to activate the WNT signaling pathway, which contributes to the osseointegration of dental implant.

Key words: Total flavonoids of rhizoma drynariae, miR-93-5p, FZD6, WNT pathway

中图分类号:

R783

申晓靖, 李林林, 路遥, 王乐为, 袁荣涛, 郭庆圆, 赵鹏. miR-93-5p在骨碎补总黄酮介导的兔骨髓间充质干细胞成骨分化中的作用及机制[J]. 中国口腔颌面外科杂志, 2023, 21(5): 446-451.

SHEN Xiao-jing, LI Lin-lin, LU Yao, WANG Le-wei, YUAN Rong-tao, GUO Qing-yuan, ZHAO Peng. The role and mechanism of miR-93-5p in osteogenic differentiation of rabbit bone marrow mesenchymal stem cells mediated by total flavonoids of rhizoma drynariae[J]. China J Oral Maxillofac Surg, 2023, 21(5): 446-451.

参考文献

[1] Papaspyridakos P, Chen CJ, Singh M, et al.Success criteria in implant dentistry: a systematic review[J]. J Dent Res, 2012, 91(3): 242-248.
[2] Brånemark PI, Adell R, Breine U, et al.Intra-osseous anchorage of dental prostheses. I. experimental studies[J]. Scand J Plast Reconstr Surg, 1969, 3(2): 81-100.
[3] Bosshardt DD, Chappuis V,Buser D.Osseointegration of titanium, titanium alloy and zirconia dental implants: current knowledge and open questions[J]. Periodontol 2000, 2017, 73(1): 22-40.
[4] Perez JR, Kouroupis D, Li DJ, et al. Tissue engineering and cell-based therapies for fractures and bone defects[J]. Front Bioeng Biotechnol, 2018, 6: 105.e1-105.e23.
[5] Hu Y, Mu P, Ma X, et al.Rhizoma drynariae total flavonoids combined with calcium carbonate ameliorates bone loss in experimentally induced Osteoporosis in rats via the regulation of Wnt3a/β-catenin pathway[J]. J Orthop Surg Res, 2021, 16(1): 702-710.
[6] Zhao K, Chen M, Liu T, et al.Rhizoma drynariae total flavonoids inhibit the inflammatory response and matrix degeneration via MAPK pathway in a rat degenerative cervical intervertebral disc model[J]. Biomed Pharmacother, 2021, 138: 111466.
[7] Chen GY, Chen JQ, Liu XY, et al.Total flavonoids of rhizoma drynariae restore the MMP/TIMP balance in models of osteoarthritis by inhibiting the activation of the NF-κB and PI3K/AKT pathways[J]. Evid Based Complement Alternat Med, 2021: 6634837.
[8] Mu P, Hu Y, Ma X, et al.Total flavonoids of Rhizoma Drynariae combined with calcium attenuate osteoporosis by reducing reactive oxygen species generation[J]. Exp Ther Med, 2021, 21(6): 618-628.
[9] He N, Zhang Y, Zhang Y, et al.Increasing fracture risk associates with plasma circulating microRNAs in aging people's sarcopenia[J]. Front Physiol, 2021, 12: 678610.
[10] Hui S, Yang Y, Li J, et al.Differential miRNAs profile and bioinformatics analyses in bone marrow mesenchymal stem cells from adolescent idiopathic scoliosis patients[J]. Spine J, 2019, 19(9): 1584-1596.
[11] Xue H, Tu Y, Ma T, et al.miR-93-5p attenuates IL-1β-induced chondrocyte apoptosis and cartilage degradation in osteoarthritis partially by targeting TCF4[J]. Bone, 2019, 123: 129-136.
[12] Zhang Y, Wei QS, Ding WB, et al.Increased microRNA-93-5p inhibits osteogenic differentiation by targeting bone morphogenetic protein-2[J]. PLoS One, 2017, 12(8): e0182678.
[13] Zhang Y, Zhuang Z, Wei Q, et al.Inhibition of miR-93-5p promotes osteogenic differentiation in a rabbit model of trauma-induced osteonecrosis of the femoral head[J]. FEBS Open Bio, 2021, 11(8): 2152-2165.
[14] Abuna RPF, Oliveira FS, Adolpho LF, et al.Frizzled 6 disruption suppresses osteoblast differentiation induced by nanotopography through the canonical Wnt signaling pathway[J]. J Cell Physiol, 2020, 235(11): 8293-8303.
[15] Hang K, Ye C, Xu J, et al.Apelin enhances the osteogenic differentiation of human bone marrow mesenchymal stem cells partly through Wnt/β-catenin signaling pathway[J]. Stem Cell Res Ther, 2019, 10(1): 189-198.
[16] Lu Y, Liu YK, Wan FY, et al.CircSmg5 stimulates the osteogenic differentiation of bone marrow mesenchymal stem cells by targeting the miR-194-5p/Fzd6 axis and beta-catenin signaling[J]. Environ Toxicol, 2022, 37(3): 593-602.
[17] Li S, Zhou H, Hu C, et al.Total flavonoids of rhizoma drynariae promotes differentiation of osteoblasts and growth of bone graft in induced membrane partly by activating Wnt/β-catenin signaling pathway[J]. Front Pharmacol, 2021, 12: 675470.

miR-93-5p在骨碎补总黄酮介导的兔骨髓间充质干细胞成骨分化中的作用及机制

The role and mechanism of miR-93-5p in osteogenic differentiation of rabbit bone marrow mesenchymal stem cells mediated by total flavonoids of rhizoma drynariae

PDF (PC)

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 1

编辑推荐

Metrics

本文评价