上海口腔医学

上海口腔医学 ›› 2022, Vol. 31 ›› Issue (3): 237-242.doi: 10.19439/j.sjos.2022.03.003

• 论著 • 上一篇    下一篇

miR-31-5p对牙髓干细胞HIF-1α/BNIP3信号通路及成骨相关因子表达的影响

付洪海1, 孙乐刚1, 丁昌成2, 马向瑞1, 黄玉梅3   

  1. 1.滨州医学院附属医院 口腔颌面外科,2.药学部, 3.口腔修复科, 山东 滨州 256603
  • 收稿日期:2020-07-30 修回日期:2020-11-07 出版日期:2022-06-25 发布日期:2022-07-07
  • 通讯作者: 黄玉梅, E-mail：hymbyfy@163.com
  • 作者简介:付洪海（1983-）,男,硕士,主治医师,E-mail: byfyfhh@163.com
  • 基金资助:
    山东省自然科学基金（ZR2018PH023）; 滨州医学院徐荣祥再生医学发展计划（BY2020XRX011）

Effects of miR-31-5p on HIF-1α/BNIP3 signaling pathway and the expression of osteoblast-related factors of dental pulp stem cells

FU Hong-hai1, SUN Le-gang1, DING Chang-cheng2, MA Xiang-rui1, HUANG Yu-mei3   

  1. 1. Department of Oral and Maxillofacial Surgery, 2. Department of Pharmacy, 3. Department of Prosthodontics, Binzhou Medical College. Binzhou 256603, Shandong Province, China
  • Received:2020-07-30 Revised:2020-11-07 Online:2022-06-25 Published:2022-07-07

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摘要： 目的: 探讨微小RNA-31-5p（miR-31-5p）对牙髓干细胞（DPSCs）低氧诱导因子1α（HIF-1α）/Bcl-2/腺病毒E1B 19-kDa相互作用蛋白3（BNIP3）信号通路及成骨相关因子表达的影响。方法: 体外培养人DPSCs,分为对照组（不转染）、mimic NC组（转染negative control-miR-31-5p）、miR-31-5p mimic组（转染hsa-miR-31-5p mimic）、siRNA NC组（转染nonsense siRNA）和miR-31-5p siRNA组（转染miR-31-5p siRNA）。实时荧光定量PCR（qRT-PCR）检测各组DPSCs细胞中miR-31-5p、HIF-1α、BNIP3、碱性磷酸酶（ALP）、Runt相关转录子2（Runx2）mRNA的表达情况,MTT法检测各组DPSCs细胞增殖情况,ALP活性测定试剂盒检测各组DPSCs细胞ALP活性,蛋白印迹（WB）法检测各组DPSCs细胞中HIF-1α、BNIP3、Runx2蛋白的表达情况。采用SPSS 24.0软件包对数据进行统计学分析。结果: 与对照组、mimic NC组相比,miR-31-5p mimic组DPSCs细胞A值、ALP mRNA表达水平及活性、Runx2 mRNA及蛋白表达水平显著降低（P<0.05）,ALP染色明显减弱,miR-31-5p mRNA、HIF-1α、BNIP3 mRNA及HIF-1α、BNIP3、Beclin1蛋白表达水平显著升高（P<0.05）。与对照组、siRNA NC组相比,miR-31-5p siRNA组DPSCs细胞A值、ALP mRNA表达水平及活性、Runx2 mRNA及蛋白水平显著升高（P<0.05）,ALP染色明显增强,miR-31-5p mRNA、HIF-1α、BNIP3 mRNA及HIF-1α、BNIP3、Beclin1蛋白表达水平显著降低（P<0.05）。结论: miR-31-5p可以激活HIF-1α/BNIP3信号通路,抑制DPSCs细胞的成骨相关因子表达。

关键词: 微小RNA-31-5p, 牙髓干细胞, HIF-1α, BNIP3, 成骨相关因子表达

Abstract: PURPOSE: To investigate the effects of microRNA-31-5p (miR-31-5p) on the signal pathway of hypoxia inducible factor-1α (HIF-1α)/Bcl-2/adenovirus E1B 19-kDa-interacting protein 3(BNIP3) and the expression of osteoblast-related factors of dental pulp stem cells(DPSCs). METHODS: Human dental pulp stem cells (DPSCs) were cultured in vitro and divided into the control group (no transfection), mimic NC group (transfected with negative control-miR-31-5p), miR-31-5p mimic group (transfected with hsa-miR-31-5p mimic), siRNA NC group (transfected with nonsense siRNA) and miR-31-5p siRNA group (transfected with miR-31-5p siRNA).The expressions of miR-31-5p, HIF-1α, BNIP3, alkaline phosphatase(ALP) and Runt-related transcription factor-2(Runx2) mRNA in DPSCs were detected by real-time fluorescence quantitative PCR; the proliferation of DPSCs was detected by MTT; ALP activity of DPSCs was detected by ALP activity test kit; and the protein expressions of HIF-1α, BNIP3 and Runx2 in DPSCs were detected by Western blot. Statistical analysis was carried out with SPSS 24.0 software package. RESULTS: Compared with the control group and mimic NC group, the A value, ALP mRNA expression level and activity, Runx2 mRNA and protein expression levels of DPSCs in miR-31-5p mimic group were significantly lower (P<0.05), ALP staining decreased significantly, and the expression levels of miR-31-5p mRNA, HIF-1α, BNIP3 mRNA and HIF-1α, BNIP3, Beclin1 protein were significantly higher (P<0.05). Compared with the control group and siRNA NC group, the A value, ALP mRNA expression level and activity, Runx2 mRNA and protein expression levels of DPSCs in miR-31-5p siRNA group were significantly higher (P<0.05), ALP staining enhanced significantly, and the expression levels of miR-31-5p mRNA, HIF-1α, BNIP3 mRNA and HIF-1α, BNIP3, Beclin1 protein were significantly lower(P<0.05). CONCLUSIONS: MiR-31-5p may inhibit the expression of osteoblast-related factors of DPSCs, and activating HIF-1α/BNIP3 signaling pathway.

Key words: microRNA-31-5p, Dental pulp stem cells, HIF-1α, BNIP3, Expression of osteoblast-related factors

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