miR-34a对人颌骨骨髓间充质干细胞成骨分化的影响  

Effects of miR-34a on osteogenic differentiation of humanorofacial bone marrow mesenchymal stem cells

作　　者：张勃昕 吉爱红 曹正垚 孙秋望月 邓威 郭松松[1] 王晨星[1] 李怀奇[1] 叶金海[1] ZHANG Boxin;JI Aihong;CAO Zhengyao;SUN Qiuwangyue;DENG Wei;GUO Songsong;WANG Chenxing;LI Huaiqi;YE Jinhai(Jiangsu Key Laboratory of Oral Diseases,Department of Oral and Maxillofacial Surgery,The Affiliated Stomatological Hospital of Nanjing Medical University,Nanjing 210029,China;Zhangqiu Stomatological Hospital,Jinan 250001,China)

机构地区：[1]南京医科大学口腔疾病研究江苏省重点实验室南京医科大学附属口腔医院口腔颌面外科,江苏南京210029 [2]章丘区口腔医院,山东济南250001

出　　处：《口腔生物医学》2019年第1期1-5,共5页Oral Biomedicine

基　　金：国家自然科学基金(81371123);"科教强卫工程"医学重点人才项目;江苏高校优势学科建设工程资助项目(2018-87)

摘　　要：目的:研究miR-34a对人颌骨骨髓间充质干细胞(hOBMSCs)成骨分化的影响。方法:实时定量RT-PCR检测miR-34家族在hOBMSCs成骨诱导过程中的表达情况;采用碱性磷酸酶及茜素红染色检测miR-34a对于hOBMSCs成骨分化能力的影响;Western blot检测hOBMSCs成骨分化过程中miR-34a对于成骨相关蛋白的影响,同时检测hOBMSCs在正常培养条件下miR-34a对于DKK1蛋白表达的影响;双荧光素酶报告基因检测miR-34a与DKK1的靶向关系;通过裸鼠皮下成骨实验检测miR-34a对于hOBMSCs体内成骨的影响。结果:在hOBMSCs成骨诱导过程中,miR-34家族的miRNA表达量均显著上升(P<0.05),其中miR-34a最为明显;过表达miR-34a可显著提高hBMSCs的体外成骨能力;miR-34a可以负向调节DKK1,同时双荧光素酶报告基因实验证实DKK1为miR-34a的靶基因;体内实验同样证实miR-34a可以促进hOBMSCs的成骨分化。结论:miR-34a可能通过抑制DKK1的表达,促进hOBMSCs的成骨分化。Objective:To investigate the effect of miR-34a on osteogenic differentiation of hOBMSCs.Methods:Real-time quantitative RT-PCR was used to detect the expression of miR-34a/miR-34b/miR-34c in hOBMSCs during osteogenic induction.The effect of miR-34a on osteogenic differentiation of hOBMSCs were detected by alkaline phosphatase staining and alizarin red staining.The regulation role of miR-34a on the expression of osteogenic markers(RUNX2,OSX,OPN,OCN,COL1)during osteogenic induction and the effect of miR-34a on DKK1 protein expression under normal cell culture conditions were detected by westernblot.The target relationship of miR-34a and DKK1 was measured by dual luciferase reporter assay.The effect of miR-34a on osteogenesis of hOBMSCs was examined by subcutaneous osteogenesis in nude mice.Results:MiR-34a was demonstrated to be upregulated during the osteogenic differentiation of hOBMSCs.Overexpression of miR-34a significantly increased alkaline phosphatase activity,mineralization capacity,and the expression of osteogenesis-associated genes in hOBMSCs in vitro.Further investigations revealed that miR-34a inhibited the expression of DKK1,and dual-luciferase reporter gene assay confirmed that DKK1 was the target gene of miR-34a.In vivo experiments also confirmed that miR-34a could promote osteogenic differentiation of hOBMSCs.Conclusions:MiR-34a promoted hOBNSCs osteogenic differentiation via down-regulating the expression of DKK1.

关 键 词：MIR-34A DKK1 人颌骨骨髓间充质干细胞 成骨分化 

分 类 号：R780.2[医药卫生—口腔医学]