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。
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 andβ-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 andβ-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.
作者
申晓靖
李林林
路遥
王乐为
袁荣涛
郭庆圆
赵鹏
SHEN Xiao-jing;LI Lin-lin;LU Yao;WANG Le-wei;YUAN Rong-tao;GUO Qing-yuan;ZHAO Peng(Department of Stomatology,Qingdao Hospital,University of Health and Rehabilitation Sciences(Qingdao Municipal Hospital).Qingdao 266000,Shandong Province;Cadre Health Care Department,Qingdao Hospital of Traditional Chinese Medicine(Haici Medical Group).Qingdao 266000,Shandong Province;First People's Hospital of Wudu District,Gansu Longnan.Longnan 746000,Gansu Province,China)
出处
《中国口腔颌面外科杂志》
CAS
2023年第5期446-451,共6页
China Journal of Oral and Maxillofacial Surgery
基金
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)。
