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首页|期刊导航|临床口腔医学杂志|miR-548d-5p通过TLR2/Myd88/NF-κB信号通路促进牙周膜干细胞细胞增殖和成骨分化的研究

miR-548d-5p通过TLR2/Myd88/NF-κB信号通路促进牙周膜干细胞细胞增殖和成骨分化的研究OACSTPCD

miR-548d-5p contributes to cell proliferation and osteogenic differentiation via TLR2/Myd88/NF-κB axis in periodontal ligament stem cells

中文摘要英文摘要

目的:探讨TLR2 负性上游调节因子miR-548d-5p对人牙周膜干细胞(human periodontal ligament stem cells,hPDLSCs)增殖和成骨分化的影响及作用机制.方法:构建TLR2 过表达载体TLR2 OE.TLR2 OE和miR-548d-5p模拟物miR-548d-5p mimic 分别转染 hPDLSCs,RT-qPCR 评估转染效率.CCK-8 法和细胞克隆形成实验检测hPDLSCs增殖能力.茜素红染色和蛋白免疫印迹检测OCN蛋白表达水平,评价其成骨分化能力.蛋白免疫印迹测定TLR2/MyD88/NF-κB信号通路分子Myd88 和NF-κB表达水平.结果:转染TLR2 OE可显著降低hPDLSCs细胞存活率和克隆形成数目(P<0.05),显著降低hPDLSCs矿化结节形成数量和OCN蛋白表达水平(P<0.05),同时显著增加hPDLSCs Myd88 和NF-κB分子表达水平(P<0.05).转染miR-548d-5p mimic,可显著增加hPDLSCs细胞存活率和克隆形成数目(P<0.05),显著增加hPDLSCs矿化结节形成数量和OCN蛋白表达水平(P<0.05),同时显著降低hPDLSCs Myd88 和 NF-κB 分子表达水平(P<0.05).与空白组相比,TLR2 OE 和 miR-548d-5p mimic 共转染后,hPDLSCs存活率和克隆形成数目无显著性差异,hPDLSCs矿化结节形成数量和OCN蛋白表达水平,及Myd88 和NF-κB分子表达水平均无显著性差异.结论:TLR2 抑制hPDLSCs的增殖和成骨分化,miR-548d-5p则增强hPDLSCs的增殖和成骨分化.miR-548d-5p可拮抗TLR2 过表达对hPDLSCs增殖和成骨分化的影响.miR-548d-5p通过TLR2/Myd88/NF-κB信号通路调节牙周膜干细胞的细胞增殖和成骨分化.

Objective:To explore the effect and mechanism of TLR2 negative upstream regulatory factor miR-548d-5p on the proliferation and osteogenic differentiation of human periodontal ligament stem cells(hPDLSCs).Methods:Construct TLR2 overexpression vector TLR2 OE.TLR2 OE and miR-548d-5p mimic were transfected into hPDLSCs respectively.The transfection efficiency was evaluated by RT-qPCR.Proliferation ability of hPDLSCs was detected by CCK-8 assay and clone formation experiment.Alizarin red staining and Western blotting were used to determine the expression level of OCN protein and evaluate the osteogenic differentiation ability of hPDLSCs.TLR2/MyD88/NF-κB signaling pathway molecules Myd88 and NF-κB expression levels were determined by Western blot.Results:The survival rate,clone formation numbers and num-bers of mineralized nodules formed by hPDLSCs were significantly reduced after transfection with TLR2 OE,while the ex-pression levels of Myd88 and NF-κB molecules were significantly incresed(P<0.05).The survival rate,clone formation num-bers and numbers of mineralized nodules formed by hPDLSCs were significantly incresed after transfection with miR-548d-5p mimic,while the expression levels of Myd88 and NF-κB molecules were significantly reduced(P<0.05).Compared with the control group,there was no significant difference in survival rate,clone formation numbers,numbers of mineralized nodules formed by hPDLSCs and Myd88 and NF-κB expression levels in TLR2 OE and miR-548d-5p mimic co-transfection group.Conclusion:Overexpression of TLR2 inhibited the proliferation and osteogenic differentiation of hPDLSCs.miR-548d-5p en-hanced the proliferation and osteogenic differentiation ability of hPDLSCs.MiR-548d-5p could antagonize the effect of TLR2 on the proliferation and osteogenic differentiation of hPDLSCs through regulating TLR2/Myd88/NF-κB signaling pathway.

卢妍君;李文霞;于飞

重庆医科大学第一临床学院 重庆 400016华中科技大学同济医学院附属同济医院口腔医学中心 湖北 武汉 430030

生物工程

Toll样受体2牙周膜干细胞miRNA

Toll like receptor 2Periodontal ligament stem cellsmiRNA

《临床口腔医学杂志》 2024 (001)

8-12 / 5

10.3969/j.issn.1003-1634.2024.01.003

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