中国组织工程研究

中国组织工程研究 ›› 2025, Vol. 29 ›› Issue (5): 899-907.doi: 10.12307/2025.281

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miR-338-3p靶向核因子κB受体活化因子配体影响牙槽骨成骨细胞增殖及凋亡

朗么磋，张义林，汪  莉   

  1. 双流区第一人民医院(四川大学华西空港医院)口腔科，四川省成都市  621000
  • 收稿日期:2023-12-01 接受日期:2024-02-07 出版日期:2025-02-18 发布日期:2024-06-01
  • 通讯作者: 朗么磋，硕士，副主任医师，双流区第一人民医院(四川大学华西空港医院)口腔科，四川省成都市 621000
  • 作者简介:朗么磋，女，1983年生，四川省阿坝州人，藏族，硕士，副主任医师，主要从事错颌畸形矫正方面的研究。

MiR-338-3p affects proliferation and apoptosis of alveolar bone osteoblasts by targeting receptor activator of nuclear factor-kappaB ligand 

Lang Mecuo, Zhang Yilin, Wang Li   

  • Received:2023-12-01 Accepted:2024-02-07 Online:2025-02-18 Published:2024-06-01
  • Contact: Lang Mecuo, Department of Stomatology, The First People’s Hospital of Shuangliu District (West China Airport Hospital of Sichuan University), Chengdu 621000, Sichuan Province, China​
  • About author:Lang Mecuo, Master, Associate chief physician, Department of Stomatology, The First People’s Hospital of Shuangliu District (West China Airport Hospital of Sichuan University), Chengdu 621000, Sichuan Province, China

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摘要：




文题释义：
核因子κB受体活化因子配体(RANKL)：是成骨细胞产生的Ⅱ型跨膜蛋白，能够诱导破骨细胞前体细胞的分化和融合，促进破骨细胞的生成、成熟，并提高其活性。核因子κB受体活化因子配体含量增加，预示骨量减少，并伴有牙槽骨吸收的风险。
成骨细胞：是骨形成的主要功能细胞，在骨形成过程中需要经历细胞增殖、基质成熟、矿化和细胞凋亡4个主要阶段。促进牙槽骨成骨细胞的增殖并抑制其凋亡对牙槽骨骨形成具有重要意义。

背景：miR-338-3p能够抑制破骨细胞分化，下调核因子κB受体活化因子配体水平能够促进骨形成。然而miR-338-3p是否通过调控核因子κB受体活化因子配体表达影响牙槽骨成骨细胞增殖、凋亡尚不清楚。
目的：探究miR-338-3p靶向核因子κB受体活化因子配体对牙槽骨成骨细胞增殖、凋亡的影响及机制。
方法：分离人牙槽骨成骨细胞，对其进行细胞转染及Wnt-C59(Wnt/β-catenin通路抑制剂)处理，分为转染对照组、miR-338-3p组、miR-338-3p+对照组、miR-338-3p+核因子κB受体活化因子配体组和miR-338-3p+Wnt-C59组。双荧光素酶实验验证miR-338-3p对核因子κB受体活化因子配体的调控作用；CCK-8、EdU染色检测细胞增殖水平；流式细胞术检测细胞周期和凋亡水平；RT-qPCR检测细胞miR-338-3p、核因子κB受体活化因子配体、Wnt-3a、β-catenin、糖原合酶激酶3β mRNA表达水平；Western Blot检测细胞核因子κB受体活化因子配体、增殖细胞核抗原、Ki67、细胞周期蛋白D1、Bcl-2、Bax、天冬氨酸半胱氨酸蛋白酶3、Wnt-3a、β-catenin、糖原合酶激酶3β蛋白表达水平。
结果与结论：①miR-338-3p靶向调控核因子κB受体活化因子配体；②过表达miR-338-3p后，细胞存活率、EdU阳性细胞率、S期细胞比例升高，细胞凋亡率降低，miR-338-3p、增殖细胞核抗原、Ki67、细胞周期蛋白D1、Bcl-2、Wnt-3a、β-catenin mRNA和蛋白水平升高，Bax、天冬氨酸半胱氨酸蛋白酶3、核因子κB受体活化因子配体、糖原合酶激酶3β mRNA和蛋白水平降低(均P < 0.05)；③过表达核因子κB受体活化因子配体或Wnt-C59处理能够减弱过表达miR-338-3p对细胞增殖、凋亡的影响(均P < 0.05)；④结果表明，miR-338-3p靶向核因子κB受体活化因子配体表达可促进牙槽骨成骨细胞增殖，并抑制细胞凋亡，其可能通过激活Wnt/β-catenin信号通路发挥作用。
https://orcid.org/0009-0002-4080-9376(朗么磋)
中国组织工程研究杂志出版内容重点：组织构建；骨细胞；软骨细胞；细胞培养；成纤维细胞；血管内皮细胞；骨质疏松；组织工程

关键词: miR-338-3p, 核因子κB受体活化因子配体, 牙槽骨成骨细胞, Wnt/β-catenin信号通路

Abstract:

BACKGROUND: MiR-338-3p could inhibit osteoclast differentiation, and downregulation of receptor activator of nuclear factor-κB ligand level could promote bone formation. However, it is unclear whether miR-338-3p can affect the proliferation and apoptosis of alveolar bone osteoblasts by regulating the receptor activator of nuclear factor-κB ligand level.
OBJECTIVE: To explore the effect and mechanism of miR-338-3p on proliferation and apoptosis of alveolar bone osteoblasts by targeting receptor activator of nuclear factor-κB ligand. 
METHODS: Human alveolar bone osteoblasts were isolated, transfected and treated with Wnt-C59 (Wnt/β-catenin pathway inhibitor), and divided into transfection control group, miR-338-3p group, miR-338-3p+control group, miR-338-3p+receptor activator of nuclear factor-κB ligand group and miR-338-3p+Wnt-C59 group. The dual luciferase report experiment was used to verify the regulatory effect of miR-338-3p on receptor activator of nuclear factor-κB ligand. Cell counting kit-8 and 5-Ethynyl-2’-deoxyuridine staining were used to detect cell proliferation levels. Flow cytometry was used to detect cell cycle and apoptosis levels. RT-qPCR was used to detect miR-338-3p, receptor activator of nuclear factor-κB ligand, Wnt-3a, β-Catenin, glycogen synthase kinase-3β mRNA levels. Western blot was used to detect RANKL, proliferating cell nuclear antigen, Ki67, CyclinD1, B-cell lymphoma/leukemia-2, B-cell lymphoma-2 related X protein, Caspase3, Wnt-3a, β-catenin, glycogen synthase kinase-3β protein levels. 
RESULTS AND CONCLUSION: miR-338-3p could target the regulation of receptor activator of nuclear factor-κB ligand. After overexpression of miR-338-3p, cell survival rate, 5-Ethynyl-2’-deoxyuridine positive cell rate, proportion of S-phase cells were increased, and apoptosis rate was decreased. The mRNA and protein levels of miR-338-3p, proliferating cell nuclear antigen, Ki67, CyclinD1, B-cell lymphoma/leukemia-2, Wnt-3a, and β-catenin were increased, while the mRNA and protein levels of B-cell lymphoma-2 related X protein, Caspase3 protein, receptor activator of nuclear factor-κB ligand, and glycogen synthase kinase-3β were decreased (all P < 0.05). Overexpression of receptor activator of nuclear factor-κB ligand or Wnt-C59 could weaken the effects of overexpression of miR-338-3p on cell proliferation and apoptosis (all P < 0.05). Overall, miR-338-3p promotes alveolar bone osteoblast proliferation and inhibits apoptosis by targeting receptor activator of nuclear factor-κB ligand, which may act through activation of the Wnt/β-catenin signaling pathway.

Key words: miR-338-3p, receptor activator of nuclear factor-κB ligand, alveolar bone osteoblast, Wnt/β-catenin signaling pathway

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