West China Journal of Stomatology ›› 2022, Vol. 40 ›› Issue (4): 377-385.doi: 10.7518/hxkq.2022.04.002

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Receptor activator of nuclear factor-κB ligand and tumor necrosis factor-α promotes osteoclast differentiation through the exosomes of inflammatory periodontal ligament stem cells

Dai Zhenning1(), Zheng Weihan2, Li Shiyu1,2()   

  1. 1.Dept. of Stomatology, Guangdong Second Traditional Chinese Medicine Hospital, Guangzhou 510095, China
    2.The Third Affiliated Hospital of Southern Medical University, Guangdong Medical Innovation Platform for Translation of 3D Printing Application, Guangzhou 510630, China
  • Received:2021-11-10 Revised:2022-05-08 Online:2022-07-25 Published:2022-07-27
  • Contact: Li Shiyu E-mail:409556338@qq.com;li421@126.com
  • Supported by:
    The National Natural Science Foundation of China(82073747);The Fellowship of China Postdoctoral Science Foundation(2021M701620);The President Foundation of The Third Affiliated Hospital of Southern Medical University(YQ2021001);Yingcai Project in Guangzhou Development District (2019);Guangdong Regional Joint Youth Fund Project(2021A1515111074)


Objective Pathological bone resorption is common in chronic periodontitis. However, the effect of exosomes (Exo) secreted by periodontal ligament stem cells (PDLSCs) on bone resorption is unclear. This study explored the Exo of inflammatory PDLSCs, their protein components, and their effects on osteoclast differentiation. Methods PDLSCs were isolated from the periodontal ligament tissues of orthodontic patients and those with chronic periodontitis. The surface markers of PDLSCs were detected by flow cytometry. Exo were characterized by Western blot, transmission electron microscope (TEM), bicinchoninic acid assay (BCA), nanosight tracking analysis (NTA). The protein components of Exo were detected by protein profiling. The expression levels of differentially expressed proteins tumor necrosis factor-α (TNF-α), receptor activator of nuclear factor-κB ligand (RANKL), interleukin (IL)-1α, transforming growth factor β (TGF-β), and bone morphogenetic protein 2 (BMP-2) were verified by enzyme linked immunosorbent assay (ELISA). Then, 10, 100, and 1 000 μg·mL-1 of Exo-CP or Exo-WT were added to RAW264.7 medium, and the expression levels of osteoclast-related indicators were detected by real time quantitative polymerase chain reaction (RT-qPCR), Western blot, and tartrate resistant acid phosphatase (TRAP) staining at 5 days. Experimental data were statistically analyzed using SPSS 24.0 software. Results The differentially expressed proteins enriched in Exo-CP were mainly related to the tumor necrosis factor (TNF) signaling, osteoclast differentiation, and nuclear transcription factor κB (NF-κB) signaling pathways. ELISA experiments confirmed Exo-CP had high expression of TNF-α, RANKL, and IL-1α and low expression of TGF-β1 and BMP-2 (P<0.05). Adding Exo-CP to RAW264.7 significantly increased the expression of mRNA and proteins related to osteoclast differentiation of cells. In a concentration-dependent manner, the effect of Exo-CP on osteoclast differentiation at concentrations of 100 and 1 000 μg·mL-1 was significantly higher than that on the 10 μg·mL-1 concentration group (P<0.05). Conclusion Pathological bone resorption of chronic periodontitis may be caused by the activation of Exo-CP to promote osteoclast differentiation. The main protein in Exo may be RANKL and TNF-α. This research provides a new perspective on pathological bone resorption in chronic periodontitis.

Key words: chronic periodontitis, periodontal ligament stem cells, exosomes, receptor activator of nuclear factor-κB ligand, tumor necrosis factor-α, osteoclast differentiation

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