上海口腔医学

上海口腔医学 ›› 2026, Vol. 35 ›› Issue (1): 35-41.doi: 10.19439/j.sjos.2026.01.006

• 论著 • 上一篇    下一篇

LncRNA PVT1靶向miR-18b-5p参与牙髓间充质干细胞成牙本质分化的作用机制

郑郁1,2, 翟桐1,3, 崔敏1, 刘飞1   

  1. 1.西安交通大学口腔医院 陕西省颅颌面精准医学研究重点实验室,陕西 西安 710006;
    2.西安建筑科技大学医院 口腔科,陕西 西安 710055;
    3.西安交通大学口腔医院 第三门诊部, 陕西 西安 710000
  • 收稿日期:2025-02-13 修回日期:2025-03-17 出版日期:2026-03-12 发布日期:2026-03-12
  • 通讯作者: 翟桐,E-mail：zhaitongsien@163.com
  • 作者简介:郑郁（1983-）,主治医师,硕士,E-mail：keuk42@163.com
  • 基金资助:
    西安市创新能力强基计划-医学研究项目（21YXYJ0122）

Exploration of the role of LncRNA PVT1 targeting miR-18b-5p in the differentiation of dental pulp mesenchymal stem cells into dentin

Zheng Yu1,2, Zhai Tong1,3, Cui Min1, Liu Fei1   

  1. 1. Shaanxi Provincial Key Laboratory of Craniofacial Precision Medicine Research, Hospital of Stomatology, Xi'an Jiaotong University. Xi'an 710006;
    2. Department of Stomatology, Xi'an University of Architecture and Technology Hospital. Xi'an 710055;
    3. Department of the Third Clinic, Hospital of Stomatology, Xi'an Jiaotong University. Xi'an 710000, Shaanxi Province, China
  • Received:2025-02-13 Revised:2025-03-17 Online:2026-03-12 Published:2026-03-12

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摘要： 目的 分析长链非编码核糖核酸（LncRNA）浆细胞瘤多样异位基因1（PVT1）靶向微小核糖核酸-18b-5p（miR-18b-5p）参与牙髓间充质干细胞（dental pulp mesenchymal stem cells,DPMSCs）成牙本质分化的作用机制。方法 分离培养人DPMSCs,油红O、茜素红、阿利新蓝染色鉴定,流式细胞术检测细胞表面标记分子。以2×104个细胞/cm2的密度将细胞接种于24孔板,分为PVT1上调+miR-18b-5p下调组（转染pcDNA-PVT1和antagomiR-18b-5p）、PVT1下调+miR-18b-5p上调组（转染si-PVT1和agomiR-18b-5p）、空载1组（转染NC pcDNA）、空载2组（转染ago-NC）、PVT1上调+空载2组（转染pcDNA-PVT1和空载体2）、空载1+miR-18b-5p下调组（转染空载体1和antagomiR-18b-5p）和空白组。7天后茜素红染色观察细胞钙矿化情况,试剂盒检测碱性磷酸酯酶（ALP）活性。实时荧光定量PCR（RT-qPCR）检测各组LncRNA PVT1、miR-18b-5p与牙本质涎磷蛋白（DSPP）、骨钙素（OCN）、Runt相关转录因子2（RUNX2）基因表达。免疫印迹法（Western blot）检测各组DSPP、OCN和RUNX2蛋白表达。双荧光素酶报告基因检测实验探讨LncRNA PVT1与miR-18b-5p的靶向关系。结果 成功分离人DPMSCs并经油红O、茜素红、阿利新蓝染色和流式细胞术鉴定。PVT1上调+miR-18b-5p下调组成牙本质分化活性最强（P<0.05）,PVT1下调+miR-18b-5p上调组则最弱（P<0.05）。PVT1上调+miR-18b-5p下调组LncRNA PVT1、DSPP、OCN和RUNX2表达最高（P<0.05）,miR-18b-5p表达最低（P<0.05）,PVT1下调+miR-18b-5p上调组相反（P<0.05）。LncRNA PVT1与miR-18b-5p存在结合位点,且LncRNA PVT1可负反馈靶向miR-18b-5p。结论 上调LncRNA PVT1可靶向抑制miR-18b-5p,促进人DPMSCs成牙本质分化,推测与上调DSPP、OCN和RUNX2表达有关。

关键词: 长链非编码核糖核酸, 浆细胞瘤多样异位基因1, 微小核糖核酸-18b-5p, 牙髓间充质干细胞, 成牙本质分化

Abstract: PURPOSE: To analyze on the role of long-chain non coding ribonucleic acid (LncRNA) plasmacytoma variant translocation 1 (PVT1) in the differentiation of dental pulp mesenchymal stem cells (DPMSCs) into dentin by targeting microribonucleic acid 18b-5p(miR-18b-5p). METHODS: Human DPMSCs were isolated and cultured, then identified by staining with oil red O, Alizarin red, and Alisin blue, and detected cell surface marker molecules by flow cytometry. Inoculate cells onto a 24 well plate, 2×104/cm2 density, grouping, including PVT1 upregulation+miR-18b-5p downregulation group (transfected with pcDNA-PVT1 and antomiR-18b-5p), PVT1 downregulation+miR-18b-5p upregulation group (transfected with si-PVT1 and agomiR-18b-5p), empty 1 group (transfected with NC pcDNA), empty 2 group (transfected with ago-NC), PVT1 upregulation + empty 2 group (transfected with pcDNA-PVT1 and empty 2), empty1+miR-18b-5p downregulation group (transfected with empty1 and antomiR-18b-5p) and a blank group. After 7 days, Alizarin red staining was used to observe the calcium mineralization of cells, and alkaline phosphatase (ALP) activity was detected using a reagent kit. Real time fluorescence quantitative polymerase chain reaction(RT-qPCR) was used to detect the expressions of LncRNA PVT1, miR-18b-5p, dentin salivary phosphoprotein(DSPP), osteocalcin(OCN), and Runt related transcription factor 2 (RUNX2) genes in each group. Western blot(WB) was used to detect the expressions of DSPP, OCN, and RUNX2 proteins in each group. Dual luciferase reporter gene detection experiment was used to explore the targeting relationship between LncRNA PVT1 and miR-18b-5p. RESULTS: This study successfully isolated human DPMSCs and identified them by staining with oil red O, Alizarin red, Alishin blue and flow cytometry. The PVT1 upregulation+miR-18b-5p downregulation group had the strongest dentin differentiation activities (P<0.05), while that of PVT1 downregulation+miR-18b-5p upregulation group was the weakest(P<0.05). The LncRNA PVT1, DSPP, OCN and RUNX2 expressions were highest in the PVT1 upregulation+miR-18b-5p downregulation group(P<0.05), while miR-18b-5p expression was lowest (P<0.05). The PVT1 downregulation+miR-18b-5p upregulation group showed the opposite trend(P<0.05). There were binding sites between LncRNA PVT1 and miR-18b-5p, and LncRNA PVT1 could negatively feedback and target miR-18b-5p. CONCLUSIONS: Upregulation of LncRNA PVT1 can target the inhibition of miR-18b-5p and promote the differentiation of human DPMSCs into dentin, which may be related to the upregulation of DSPP, OCN and RUNX2 expressions.

Key words: Long-chain non coding ribonucleic acid, Plasmacytoma variant translocation 1, Microribonucleic acid 18b-5p, Dental pulp mesenchymal stem cells, Odontogenic differentiation

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