长链非编码 RNA 钾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 调节人牙周膜干细胞的增殖和成骨分化。
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.
机构信息
Dept. of Stomatology, The Fourth Affiliated Hospital of Guangxi Medical University, Liuzhou 545005, China.
Dept. of Stomatology, Beijing Rehabilitation Hospital of Capital Medical University, Beijing 100144, China.
出版信息
Hua Xi Kou Qiang Yi Xue Za Zhi. 2021 Oct 1;39(5):547-554. doi: 10.7518/hxkq.2021.05.008.
OBJECTIVES
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 (<0.05). The down-regulation of lncRNA KCNQ1OT1 inhibited cell proliferation and reduced the mRNA and protein expression levels of OCN, OPN, and ALP (<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 (<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 (<0.05).
CONCLUSIONS
Down-regulation of lncRNA KCNQ1OT1 inhibited the proliferation and osteogenic differentiation of hPDLSCs by targeting the up-regulated expression of miR-24-3p.
目的
本研究旨在探讨长链非编码 RNA（lncRNA）钾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其分子机制。
方法
分离培养正常牙周组织的 hPDLSCs，用矿化液诱导 hPDLSCs 成骨分化。下调 lncRNA KCNQ1OT1，转染抗 miR-24-3p 过表达，观察对 hPDLSCs 增殖及骨钙素（OCN）、骨桥蛋白（OPN）和碱性磷酸酶（ALP）水平的影响。实时定量聚合酶链反应（RT-qPCR）检测 lncRNA KCNQ1OT1、miR-24-3p、OCN、OPN 和 ALP 水平。噻唑蓝（MTT）法检测细胞活力和活性。MTT 法评价细胞增殖。Western blot 法检测蛋白表达。双荧光素酶实验检测 lncRNA KCNQ1OT1 与 miR-24-3p 的靶向关系。
结果
hPDLSCs 成骨过程中 lncRNA KCNQ1OT1 表达上调，miR-24-3p 表达下调（<0.05）。下调 lncRNA KCNQ1OT1 抑制细胞增殖，降低 OCN、OPN 和 ALP 的 mRNA 和蛋白表达水平（<0.05）。lncRNA KCNQ1OT1 靶向调控 miR-24-3p。过表达 miR-24-3p 抑制细胞增殖，降低 OCN、OPN 和 ALP 的 mRNA 和蛋白表达水平（<0.05）。抑制 miR-24-3p 逆转下调 lncRNA KCNQ1OT1 对细胞增殖及 OCN、OPN 和 ALP 的 mRNA 和蛋白表达水平的影响（<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。
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