Ding Rong, Chen Chen, Wang Ling, Wang Yijie, Chai Zhen, He Siyu, Zhang Qianqian, Cheng Shuli, Zou Rui
Key Laboratory of Shaanxi Province for Craniofacial Precision Medicine Research, College of Stomatology, Xi'an Jiaotong University, Xi'an, Shaanxi Province, China; Clinical Research Centre of Shaanxi Province for Dental and Maxillofacial Diseases, College of Stomatology, Xi'an Jiaotong University, Xi'an, Shaanxi Province, China; College of Stomatology, Xi'an Jiaotong University, Xi'an, Shaanxi Province, China.
Key Laboratory of Shaanxi Province for Craniofacial Precision Medicine Research, College of Stomatology, Xi'an Jiaotong University, Xi'an, Shaanxi Province, China; Clinical Research Centre of Shaanxi Province for Dental and Maxillofacial Diseases, College of Stomatology, Xi'an Jiaotong University, Xi'an, Shaanxi Province, China; College of Stomatology, Xi'an Jiaotong University, Xi'an, Shaanxi Province, China.
Int Dent J. 2025 May 1;75(4):100783. doi: 10.1016/j.identj.2025.02.022.
This study aimed to examine the influence of matrix stiffness on osteogenic differentiation via epigenetic mechanisms in human periodontal ligament stem cells (hPDLSCs), with implications for understanding orthodontic tooth movement.
hPDLSCs were cultured on substrates with varying stiffness (soft and stiff). Dot blot and immunofluorescence techniques were employed to measure global DNA methylation levels. RT-qPCR and alkaline phosphatase (ALP) activity assays were conducted to assess differences in DNA methylation and osteogenic potential. Additionally, ELISA was used to quantify DNA methyltransferase content and activity.
hPDLSCs on stiffer substrates exhibited increased 5-methylcytosine (5-mC) and higher global DNA methylation levels than those on soft substrates. With increased matrix stiffness, DNMT3A and DNMT3B mRNA expression levels rose. hPDLSCs on stiff matrices also showed elevated DNMT3B enzyme content and osteogenic activity. When global DNA methylation was reduced, mRNA levels of RUNX2, ALP, and Col-1 decreased, along with a notable reduction in ALP staining intensity in the inhibitor group.
Matrix stiffness is positively associated with global DNA methylation, with DNMT3B likely mediating this regulation in hPDLSCs. Furthermore, DNA methylation levels are positively linked to the osteogenic capability of hPDLSCs.
本研究旨在通过表观遗传机制研究基质硬度对人牙周膜干细胞（hPDLSCs）成骨分化的影响，以有助于理解正畸牙齿移动。
将hPDLSCs培养在具有不同硬度（软和硬）的基质上。采用斑点印迹和免疫荧光技术测量整体DNA甲基化水平。进行RT-qPCR和碱性磷酸酶（ALP）活性测定以评估DNA甲基化和成骨潜能的差异。此外，使用ELISA定量DNA甲基转移酶的含量和活性。
与软基质上的hPDLSCs相比，硬基质上的hPDLSCs表现出5-甲基胞嘧啶（5-mC）增加和更高的整体DNA甲基化水平。随着基质硬度增加，DNMT3A和DNMT3B mRNA表达水平升高。硬基质上的hPDLSCs还显示出DNMT3B酶含量和成骨活性升高。当整体DNA甲基化降低时，RUNX2、ALP和Col-1的mRNA水平下降，抑制剂组的ALP染色强度也显著降低。
基质硬度与整体DNA甲基化呈正相关，DNMT3B可能在hPDLSCs中介导这种调节。此外，DNA甲基化水平与hPDLSCs的成骨能力呈正相关。
