华西口腔医学杂志 ›› 2021, Vol. 39 ›› Issue (5): 547-554.doi: 10.7518/hxkq.2021.05.008

• 基础研究 • 上一篇    

长链非编码RNA钾离子电压门控通道亚家族Q成员1重叠转录本1通过靶向miR-24-3p调控人牙周膜干细胞增殖和成骨分化

庞鸣1(), 韦红霞1, 陈茜2()   

  1. 1.广西医科大学第四附属医院口腔科,柳州 545005
    2.首都医科大学附属北京康复医院口腔科,北京 100144
  • 收稿日期:2020-09-19 修回日期:2021-07-05 出版日期:2021-10-01 发布日期:2021-10-11
  • 通讯作者: 陈茜 E-mail:pangmingqwe@163.com;9693079@qq.com
  • 作者简介:庞鸣,主治医师,硕士,E-mail:pangmingqwe@163.com

Long non-coding RNA potassium voltage-gated channel subfamily Q member 1 overlapping transcript 1 regulates the proliferation and osteogenic differentiation of human periodontal ligament stem cells by targeting miR-24-3p

Pang Ming1(), Wei Hongxia1, Chen Xi2()   

  1. 1.Dept. of Stomatology, The Fourth Affiliated Hospital of Guangxi Medical University, Liuzhou 545005, China
    2.Dept. of Stomatology, Beijing Rehabilitation Hospital of Capital Medical University, Beijing 100144, China
  • Received:2020-09-19 Revised:2021-07-05 Online:2021-10-01 Published:2021-10-11
  • Contact: Chen Xi E-mail:pangmingqwe@163.com;9693079@qq.com

摘要: 目的

探讨长链非编码RNA(lncRNA)钾离子电压门控通道亚家族Q成员1重叠转录本1(KCNQ1OT1)对人牙周膜干细胞(hPDLSCs)增殖和成骨分化的影响及分子机制。

方法

分离培养正常牙周组织的hPDLSCs,矿化液诱导hPDLSCs成骨分化,研究下调lncRNA KCNQ1OT1、过表达miR-24-3p对hPDLSCs增殖、骨钙素(OCN)、骨桥蛋白(OPN)、碱性磷酸酶(ALP)的影响。实时荧光定量聚合酶链反应(RT-qPCR)检测lncRNA KCNQ1OT1、miR-24-3p、OCN、OPN、ALP的表达水平;甲基噻唑基四唑(MTT)法检测细胞增殖能力;蛋白质印迹(Western blot)法检测蛋白表达;双荧光素酶报告实验检测lncRNA KCNQ1OT1和miR-24-3p的靶向关系。

结果

在hPDLSCs成骨诱导中,lncRNA KCNQ1OT1表达水平升高,miR-24-3p表达水平降低(P<0.05);下调lncRNA KCNQ1OT1表达,抑制细胞增殖,降低OCN、OPN和ALP的mRNA与蛋白表达水平(P<0.05)。lncRNA KCNQ1OT1靶向调控miR-24-3p,过表达miR-24-3p后,抑制细胞增殖,降低OCN、OPN和ALP的mRNA与蛋白表达水平(P<0.05)。抑制miR-24-3p可逆转下调lncRNA KCNQ1OT1对细胞增殖以及OCN、OPN和ALP的mRNA与蛋白表达水平的影响(P<0.05)。

结论

下调lncRNA KCNQ1OT1通过靶向上调miR-24-3p抑制hPDLSCs的增殖和成骨分化。

关键词: 钾离子电压门控通道亚家族Q成员1重叠转录本1, miR-24-3p, 人牙周膜干细胞, 增殖, 成骨分化

Abstract: Objective

This study aims to explore the effect and molecular mechanism of long non-coding RNA (lncRNA) potassium voltage-gated channel subfamily Q member 1 overlapping transcript 1 (KCNQ1OT1) on proliferation and osteogenic differentiation in human periodontal ligament stem cells (hPDLSCs).

Methods

The hPDLSCs of normal periodontal tissues were isolated and cultured. The mineralized solution induced the osteoblast differentiation of hPDLSCs. The down-regulation of lncRNA KCNQ1OT1, the overexpression of anti-miR-24-3p on the proliferation and the levels of osteocalcin (OCN), osteopontin (OPN) and alkaline phosphatase (ALP) of hPDLSCs were investigated. Real-time quantitative polymerase chain reaction (RT-qPCR) was used to detect the levels of lncRNA KCNQ1OT1, miR-24-3p, OCN, OPN, and ALP. Methyl thiazolyl tetrazolium (MTT) method was used to detect cell viability and activity. Cell proliferation was evaluated by MTT. Western blot was used to detect protein expression. The targeted relationship between lncRNA KCNQ1OT1 and miR-24-3p was detected by double-luciferase experiment.

Results

The expression level of lncRNA KCNQ1OT1 increased, and that of miR-24-3p decreased during the osteogenesis of hPDLSCs (P<0.05). The down-regulation of lncRNA KCNQ1OT1 inhibited cell proliferation and reduced the mRNA and protein expression levels of OCN, OPN, and ALP (P<0.05). LncRNA KCNQ1OT1 targeted and regulated miR-24-3p. The overexpression of miR-24-3p inhibited cell proliferation and reduced the mRNA and protein expression levels of OCN, OPN, and ALP (P<0.05). Inhibition of miR-24-3p reversed the effect of the down-regulation of lncRNA KCNQ1OT1 on cell proliferation and mRNA and protein expression levels of OCN, OPN, and ALP (P<0.05).

Conclusion

Down-regulation of lncRNA KCNQ1OT1 inhibited the proliferation and osteogenic differentiation of hPDLSCs by targeting the up-regulated expression of miR-24-3p.

Key words: potassium voltage-gated channel subfamily Q member 1 overlapping transcript 1, miR-24-3p, human periodontal ligament stem cells, proliferation, osteogenic differentiation

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