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Abstract：
This study aims to investigate the effects of 1,25-dihydroxyvitamin D3 (1,25(OH)2VitD3) on osteogenic differentiation of human periodontal ligament stem cells (hPDLSCs) and the activity of hPDLSC sheets and the differences in the tissue regeneration activity of hPDLSC sheets on tooth root fragment treated by different methods. Healthy caries-free premolars were collected. The hPDLSCs were obtained by enzymatic digestion. Surface markers of stem cells were analyzed by flow cytometry and the multidirectional differentiation ability of hPDLSCs was detected. During the osteogenic differentiation of hPDLSCs, 1,25(OH)2VitD3 was added and the effect of 1,25(OH)2VitD3 on osteogenic differentiation of hPDLSCs was assessed using Western blotting, quantitative reverse transcription-polymerase chain reaction (qRT-PCR), enzyme-linked immunosorbent assay, cell staining, and immunofluorescence. After hPDLSC sheets were prepared, histology and immunofluorescence analysis of the effect of 1,25(OH)2VitD3 on sheet activity were performed. In addition, root fragments were prepared and treated with scaling, 24% EDTA (ethylenediamide tetraacetic acid), and Er,Cr:YSGG lasers, respectively, and the tissue regeneration activity of hPDLSC sheets on different root fragments were observed. 1,25(OH)2VitD3 promoted the high gene and protein expressions of osteogenic markers ALP (alkaline phosphatase), Runx2, and OPN (osteopontin antibody) in hPDLSCs, along with enhanced ALP activity and staining, alizarin red staining, and immunofluorescence staining, indicating that the osteogenic differentiation ability of hPDLSCs was improved. Extracellular matrix secretion was increased in hPDLSC sheets, along with the positive expressions of the protein markers fibronectin and collagen I, suggesting that 1,25(OH)2VitD3 could enhance these effects. In addition, the root fragments treated by Er,Cr:YSGG laser were more suitable for the attachment and regeneration of hPDLSC sheets, demonstrating that 1,25(OH)2VitD3 could improve the tissue regeneration performance of these sheets. 1,25(OH)2VitD3 can promote osteogenic differentiation of hPDLSCs and thus plays an active role in hPDLSC sheet formation and tissue regeneration. In addition, the Er,Cr:YSGG laser can be used as the recommended treatment method for the root surface regenerated by hPDLSCs.
摘要：
本研究旨在探讨1,25-二羟维生素D3(1,25(OH)2VitD3)对人牙周膜干细胞(hPDLSCs)成骨分化和hPDLSC片活性的影响以及hPDLSC片对不同方法处理的牙根碎片组织再生活性的差异。收集健康的无龋齿前磨牙。通过酶消化获得hPDLSC。通过流式细胞术分析干细胞的表面标志物，并检测hPDLSCs的多向分化能力。在hPDLSCs成骨分化过程中,添加1,25(OH)2VitD3,并使用蛋白质印迹评估1,25(OH)2VitD3对hPDLSCs成骨分化的影响,定量逆转录聚合酶链反应（qRT-PCR），酶联免疫吸附测定，细胞染色，和免疫荧光。在制备hPDLSC片材之后,对1,25(OH)2VitD3对片活性的影响进行组织学和免疫荧光分析。此外,准备根碎片并用鳞片处理，24%EDTA（乙二胺四乙酸），呃，Cr:YSGG激光器，分别,并观察了hPDLSC片在不同根碎片上的组织再生活性。1,25(OH)2VitD3促进成骨标志物ALP(碱性磷酸酶)基因和蛋白的高表达,runx2和OPN（骨桥蛋白抗体）在hPDLSCs，随着增强的ALP活性和染色，茜素红染色，免疫荧光染色，表明hPDLSCs的成骨分化能力得到提高。hPDLSC片的细胞外基质分泌增加，随着蛋白标志物纤连蛋白和I型胶原的阳性表达，表明1,25(OH)2VitD3可以增强这些作用。此外,用Er处理的根碎片，Cr:YSGG激光器更适用于hPDLSC片材的附着和再生，证明1,25（OH）2VitD3可以改善这些薄片的组织再生性能。1,25(OH)2VitD3可以促进hPDLSCs的成骨分化,因此在hPDLSC片层形成和组织再生中起积极作用。此外,Er,Cr:YSGG激光可作为hPDLSCs再生根面的推荐处理方法。
