miR-150靶向β-catenin调控人牙周膜干细胞成骨分化能力的机制研究  

Study on effect of miR-150 for targeting β-catenin on osteogenic differentiation of human periodontal ligament stem cells

作　　者：冯保静[1] 赵西博 张伟祥 FENG Bao-jing;ZHAO Xi-bo;ZHANG Wei-xiang(Department of Prosthodontic,the Six-People’s Hospital of Anyang City,Anyang 455000,China)

机构地区：[1]安阳市第六人民医院,455000

出　　处：《中华老年口腔医学杂志》2023年第3期142-147,162,共7页Chinese Journal of Geriatric Dentistry

基　　金：河南省科技攻关项目(222102310681)。

摘　　要：目的 体外研究miR-150对人牙周膜干细胞(human periodontal ligament stem cells,hPDLSCs成骨分化能力的调控作用及其分子机制。方法 分离培养hPDLSCs并进行成骨向分化诱导,q-PCR检测miR-150及成骨相关基因(RUNX2、OCN、OPN、ALP)的表达;hPDLSCs中转染miR-150沉默(inhibitor)及过表达模拟物(mimics),q-PCR检测miR-150和β-catenin的表达,茜素红染色实验检测hPDLSCs成骨分化能力的变化;qPCR及Western blot检测沉默及过表达miR-150后β-catenin表达水平的变化;双荧光素酶报告基因实验验证miR-150的潜在靶基因;在hPDLSCs中过表达miR-150并加入SKL2001(Wnt/β-catenin通路激动剂),检测β-catenin表达及hPDLSCs成骨分化能力的变化。结果 RUNX2、OCN、OPN、ALP基因在hPDLSCs成骨向分化诱导后表达明显上调,而miR-150的表达则显著下调;q-PCR及茜素红染色结果显示抑制miR-150可明显上调成骨相关基因的表达并促进hPDLSCs成骨向分化,而过表达miR-150则结果相反;生物信息学分析显示β-catenin为miR-150的潜在靶基因,双荧光素酶报告进一步验证β-catenin为miR-150的靶基因;而SKL2001激活β-catenin后可逆转miR-150 mimics对hPDLSCs成骨分化的抑制作用。结论 miR-150可靶向β-catenin调控hPDLSCs的成骨向分化能力,提示miR-150在牙周组织工程损伤修复中发挥重要作用。Objective To investigate the regulation and molecular mechanism of miR-150 on the osteogenic differentiation ability of human periodontal ligament stem cells(hPDLSCs).Methods h PDLSCs were isolated and cultured for osteogenic differentiation induction.The expressions of miR-150 and osteogenic related genes(RUNX2、OCN、OPN、ALP)was detected by q-PCR.hPDLSCs were transfected with miR-150 inhibitors and mimics.Then the expression of miR-150 and β-catenin was examined by q-PCR.The changes of osteogenic differentiation ability of hPDLSCs were detected by alizarin red staining.β-catenin as the potential target genes of miR-150 was confirmed by dual luciferase report gene assay.The expression of β-catenin was evaluated by q-RCR and Western blot after silencing and overexpression of miR-150.The expression of β-catenin and the osteogenic differentiation of hPDLSCs were detected by q-PCR and alizarin red staining after co-stimulation by miR-150 mimics and SKL2001(Wnt/β-catenin pathway agonist).Results The expressions of RUNX2,OCN,OPN and ALP genes were significantly up-regulated after induction of osteogenic differentiation of hPDLSCs,while the expression of miR-150 was significantly down-regulated.miR-150 mimics significantly inhibited the osteogenic differentiation and osteogenic related genes expression in hPDLSCs,and vice versa.β-catenin was the potential target gene of miR-150,which confirmed by bioinformatics analysis and dual luciferase report gene assay.The miR-150 mimics and SKL2001 co-stimulation experimental results showed that activation of β-catenin reverse the inhibitory effect of miR-150 mimics on the osteogenic differentiation of hPDLSCs.Conclusion miR-150 inhibits hPDLSCs osteogenic differentiation ability via suppressing the expression of β-catenin,suggesting that miR-150 plays a vital role in the repair of periodontal tissue engineering damage.

关 键 词：miR-150 人牙周膜干细胞(hPDLSCs) Β-CATENIN 成骨分化 

分 类 号：R780.2[医药卫生—口腔医学]