Periodontal ligament stem cells (PDLSCs) have mesenchymal-stem-cells-like qualities, and are considered as one of the candidates of future clinical application in periodontal regeneration therapy. Enamel matrix derivative (EMD) is widely used in promoting periodontal regeneration. However, the effects of EMD on the proliferation and osteogenic differentiation of human PDLSCs grown on the Ti implant surface are still no clear. Therefore, this study examined the effects of EMD on human PDLSCs in vitro. Human PDLSCs were isolated from healthy participants, and seeded on the surface of Ti implant disks and stimulated with various concentrations of EMD. Cell proliferation was determined with Cell Counting Kit-8 (CCK-8). The osteogenic differentiation of PDLSCs was evaluated by the measurement of alkaline phosphatase (ALP) activity, Alizarin red staining, and real-time polymerase chain reaction (qRT-PCR) and Western blotting, respectively. The results indicated that EMD at concentrations (5–60 µg/ml) increased the viability and proliferation of PDLSCs. The treatment with 30 and 60 µg/ml of EMD significantly elevated ALP activity, augmented mineralized nodule formation and calcium deposition, and upregulated the mRNA and protein levels of Runx-2 and osteocalcin (OCN) in the PDLSCs grown on the Ti surface. Further investigation found that EMD treatment did not change the protein levels of phosphatidylinositol-3-kinase (PI3K), p-PI3K, Akt and mTOR, but significantly upregulated the phosphorylated levels of Akt and mTOR. Collectively, these results suggest that EMD stimulation can promote the proliferation and osteogenic differentiation of PDLSCs grown on Ti surface, which is possibly associated with the activation of Akt/mTOR signaling pathway.

牙周膜干细胞（PDLSC）具有间充质干细胞样特性，被认为是牙周再生治疗中未来临床应用的候选之一。牙釉质基质衍生物（EMD）被广泛用于促进牙周再生。但是，EMD对在Ti植入物表面生长的人PDLSC的增殖和成骨分化的影响尚不清楚。因此，本研究考察了EMD对体外人PDLSC的影响。从健康参与者中分离出人PDLSC，并将其接种在Ti植入盘的表面，并用各种浓度的EMD刺激。用细胞计数试剂盒8（CCK-8）测定细胞增殖。通过分别测量碱性磷酸酶（ALP）活性，茜素红染色和实时聚合酶链反应（qRT-PCR）和Western印迹来评估PDLSC的成骨分化。结果表明，EMD浓度（5–60 µg / ml）可提高PDLSC的活力和增殖。用30和60 µg / ml的EMD处理可显着提高ALP活性，增加矿化的结节形成和钙沉积，并上调Ti表面生长的PDLSC中Runx-2和骨钙素（OCN）的mRNA和蛋白水平。进一步的研究发现，EMD处理并没有改变磷脂酰肌醇3-激酶（PI3K），p-PI3K，Akt和mTOR的蛋白质水平，但显着上调了Akt和mTOR的磷酸化水平。总体而言，这些结果表明，EMD刺激可以促进在Ti表面生长的PDLSC的增殖和成骨分化，这可能与Akt / mTOR信号通路的激活有关。