Zhang Jian, Wu Jiankun, Lin Xiuya, Liu Xueli
Dental Clinic, Cangzhou Central Hospital, No.16 Xinhua West Road, Cangzhou 061000, Hebei, China.
Curr Stem Cell Res Ther. 2023;18(4):560-567. doi: 10.2174/1574888X17666220704092411.
Pulp regeneration is a promising strategy that promotes the continued development of young permanent teeth with immature apical foramen. Platelet-rich fibrin (PRF) was found to stimulate the proliferation and differentiation of osteoblasts, but its effects on osteoblast/odontoblast differentiation of human dental pulp stem cells (hDPSCs) are unknown.
The hDPSCs were isolated and identified using known surface markers by flow cytometry. The CCK-8 assay and the expression of Ki67 and PCNA were used to examine hDPSC proliferation. After 7 days of culture in an osteo-/odontoblastic induction medium with various concentrations of liquid PRF (0, 10% and 20%), the early stage of osteogenesis-intracellular alkaline phosphatase (ALP) was checked. After 21 days of culture, matrix mineralization was checked using Alizarin Red S and quantified. The mRNA and protein levels of osteo-/odontoblastic genes, including RUNX2, DSPP, DMP1 and BSP, were measured by qRT-PCR. The notch signal was checked by Western blot to analyze three key proteins (Notch 1, Jagged 1 and Hes 1).
PRF-treated groups showed higher expression of Ki-67 and PCNA, higher ALP activity, and the higher dose showed a stronger induction. PRF promoted osteo-/odontoblastic differentiation of hDPSCs indicated by elevated protein levels and mRNA levels of the expression of osteo-/odontoblastic markers. The three key proteins in Notch signaling showed an increase compared with the control group and increased as the PRF concentration increased.
PRF can promote the proliferation and osteo-/odontoblastic differentiation of hDPSC, which may be through the Notch signaling pathway.
牙髓再生是一种很有前景的策略，可促进根尖孔未成熟的年轻恒牙继续发育。富血小板纤维蛋白（PRF）被发现可刺激成骨细胞的增殖和分化，但其对人牙髓干细胞（hDPSCs）成骨细胞/成牙本质细胞分化的影响尚不清楚。
采用已知表面标志物通过流式细胞术分离和鉴定hDPSCs。使用CCK-8法以及Ki67和PCNA的表达来检测hDPSC的增殖。在含有不同浓度液体PRF（0、10%和20%）的成骨/成牙本质诱导培养基中培养7天后，检测成骨早期细胞内碱性磷酸酶（ALP）。培养21天后，使用茜素红S检测基质矿化并进行定量。通过qRT-PCR测量包括RUNX2、DSPP、DMP1和BSP在内的成骨/成牙本质基因的mRNA和蛋白质水平。通过蛋白质印迹法检测Notch信号，分析三种关键蛋白（Notch 1、Jagged 1和Hes 1）。
PRF处理组显示Ki-67和PCNA表达更高，ALP活性更高，且较高剂量显示出更强的诱导作用。PRF促进了hDPSCs的成骨/成牙本质分化，表现为成骨/成牙本质标志物表达的蛋白质水平和mRNA水平升高。与对照组相比，Notch信号通路中的三种关键蛋白有所增加，且随着PRF浓度的增加而增加。
PRF可促进hDPSC的增殖和成骨/成牙本质分化，这可能是通过Notch信号通路实现的。
