The exosomes from human exfoliated deciduous teeth (SHED-Exos) have exhibited potential therapeutic role in dental and oral disorders. The biological effects of exosomes largely depend on cellular origin and physiological status of donor cell. In the present study, we explored the influence of conditioned exosomes from SHED with osteogenic induction on periodontal ligament stem cells (PDLSCs) in vitro. Conditioned SHED-Exos from a 3-day osteogenic supernatant were applied during PDLSCs osteogenic differentiation. We found that conditioned SHED-Exos had no cytotoxicity on PDLSCs viability assessed by CCK-8 assay. These SHED-Exos promoted PDLSCs osteogenic differentiation with deep Alizarin red staining, high alkaline phosphatase (ALP) activity and upregulated osteogenic gene expression (RUNX2, OPN and OCN). We further found BMP/Smad signaling and Wnt/β-catenin were activated by enhanced Smad1/5/8 phosphorylation and increased nuclear β-catenin protein expression. Inhibiting these two signaling pathways with specific inhibitors (cardamonin and LDN193189) remarkably weakened the enhanced osteogenic differentiation. Furthermore, Wnt3a and BMP2 were upregulated in SHED and SHED-Exos. Silencing Wnt3a and BMP2 in SHED-Exos partially counteracts the enhanced osteogenic differentiation. Our findings indicate that conditioned SHED-Exos-enhanced PDLSCs osteogenic differentiation was partly due to its carrying Wnt3a and BMP2. These data provide new insights into the use of SHED-Exos in periodontitis-induced bone defects therapy.

人类脱落乳牙的外泌体（SHED-Exos）在牙齿和口腔疾病中表现出潜在的治疗作用。外泌体的生物学效应在很大程度上取决于细胞来源和供体细胞的生理状态。在本研究中，我们探讨了成骨诱导的SHED条件性外泌体对牙周膜干细胞（PDLSCs）的影响。在PDLSCs成骨分化期间应用来自3天成骨上清液的条件化SHED-Exos。我们发现条件性SHED-Exos对CDL-8检测的PDLSCs生存力没有细胞毒性。这些SHED-Exos通过深茜素红染色，高碱性磷酸酶（ALP）活性和上调的成骨基因表达（RUNX2，OPN和OCN）促进了PDLSCs的成骨分化。我们进一步发现BMP / Smad信号传导和Wnt /β-catenin通过增强的Smad1 / 5/8磷酸化和增加的核β-catenin蛋白表达而被激活。用特异性抑制剂（小豆蔻苷和LDN193189）抑制这两个信号通路显着减弱了增强的成骨分化。此外，SHED和SHED-Exos中的Wnt3a和BMP2上调。使SHED-Exos中的Wnt3a和BMP2沉默部分抵消了成骨分化的增强。我们的发现表明，条件性SHED-Exos增强的PDLSCs成骨分化部分是由于其携带Wnt3a和BMP2。这些数据为SHED-Exos在牙周炎诱导的骨缺损治疗中的使用提供了新的见解。