中国口腔颌面外科杂志

中国口腔颌面外科杂志 ›› 2025, Vol. 23 ›› Issue (5): 434-441.doi: 10.19438/j.cjoms.2025.05.002

• 论著 • 上一篇    下一篇

山柰酚-3-O-阿拉伯糖苷对骨髓间充质干细胞成骨分化的调控作用及机制探讨

张澔泽1, 任海宁1, 祝庆海1, 王晨星1, 叶金海1,2   

  1. 1.江苏省口腔疾病研究重点实验室,南京医科大学附属口腔医院 口腔颌面外科,江苏 南京 210029;
    2.复旦大学附属中山医院 口腔颌面外科,上海 200032
  • 收稿日期:2025-04-07 修回日期:2025-06-03 发布日期:2025-10-10
  • 通讯作者: 叶金海,E-mail：yjh98001@163.com
  • 作者简介:张澔泽（1999-）,男,硕士,E-mail：zhanghaoze19991126@163.com
  • 基金资助:
    “江苏省卫生健康委科研项目”重点项目（K2023061); 江苏省老年健康科研项目（面上项目KLM2023019)

Study on the regulation and mechanism of kaempferol-3-O-arabinoside on osteogenic differentiation of bone marrow mesenchymal stem cells

Zhang Haoze1, Ren Haining1, Zhu Qinghai1, Wang Chenxing1, Ye Jinhai1,2   

  1. 1. Jiangsu Key Laboratory of Oral Diseases; Department of Oral and Maxillofacial Surgery, The Affiliated Stomatological Hospital of Nanjing Medical University. Nanjing 210029, Jiangsu Province;
    2. Department of Oral and Maxillofacial Surgery, Zhongshan Hospital, Fudan University. Shanghai 200032, China
  • Received:2025-04-07 Revised:2025-06-03 Published:2025-10-10

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摘要： 目的: 分析山奈酚-3-O-阿拉伯糖苷(KA)通过刺激骨髓间充质干细胞（BMSCs)对成骨分化的调控及对主要靶点表皮生长因子受体（EGFR）、丝裂原活化蛋白激酶（MAPK）信号通路的作用。方法: 从健康年轻男性阻生牙拔牙创分离BMSCs,培育至第三代后,使用不同浓度KA处理。采用CCK-8测定细胞增殖活性,碱性磷酸酶（ALP）及茜素红（ARS）染色检测KA处理后BMSCs的成骨能力变化,利用ImageJ软件分析ALP活力。采用实时定量反转录PCR（RT-qPCR）及免疫蛋白印迹（Western blot）检测成骨分化基因Runt相关转录因子2（RUNX2）、I型胶原（COL1）和骨桥蛋白（OPN）基因及蛋白的表达变化,采用网络药理学分析KA的主要作用靶点和可能作用的相关通路。使用5 μmol/L EGFR抑制剂吉非替尼、10 μmol/L MEK1/2抑制剂U0126、10 μmol/L p38抑制剂SB203580处理BMSCs,抑制MAPK信号通路主要信号分子后,使用RT-qPCR、Western blot检测BMSCs成骨能力的变化。结果: CCK-8结果提示,0、10、20、40 μmol/L浓度的KA对BMSCs的活性无明显影响,80、160 μmol/L浓度的KA对BMSCs活性影响较大。ALP及ARS染色结果显示,10 μmol/L的KA成骨诱导效果最好。RT-qPCR及Western blot结果显示,10 μmol/L浓度的KA促进BMSC中RUNX2、OPN、COL1的基因及蛋白表达（P<0.05）。网络药理结果显示,KA可能通过EGFR-MAPK信号通路对BMSCs成骨分化发挥促进作用。Western blot结果显示,10 μmol/L浓度的KA能够促进BMSCs激活MAPK信号通路。使用吉非替尼、U0126、SB203580处理后,BMSCs的RUNX2、OPN、COL1基因和蛋白表达显著下降（P<0.05）。结论: 一定浓度的KA通过EGFR-MAPK-RUNX2信号通路,促进BMSCs的成骨分化能力。

关键词: 山奈酚-3-O-阿拉伯糖苷, 骨髓间充质干细胞, 成骨细胞, MAPK信号通路, 表皮生长因子受体, 成骨分化

Abstract: PURPOSE: To study the effect of kaempferol-3-O-arabinoside(KA) on osteogenic differentiation and epidermal growth factor receptor(EGFR), mitogen-activated protein kinase(MAPK) pathway by stimulating bone marrow mesenchymal stem cells (BMSCs). METHODS: BMSCs were isolated from the extraction sockets of impacted teeth in healthy young males, cultured until passage 3, and then treated with different concentrations of KA. CCK-8 assay was used to determine cell proliferation activity. Alkaline phosphatase (ALP) and alizarin red S (ARS) staining were employed to detect the changes in osteogenic capacity of BMSCs after KA treatment, and ImageJ software was used to analyze ALP activity. Real-time quantitative reverse transcription PCR (RT-qPCR) and Western blot were used to detect the changes in the gene and protein expressions of osteogenic differentiation-related genes including Runt-related transcription factor 2 (RUNX2), type I collagen (COL1), and osteopontin (OPN). Network pharmacology was adopted to analyze the main action targets and potential related signaling pathways of KA. BMSCs were treated with 5 μmol/L EGFR inhibitor gefitinib, 10 μmol/L MEK1/2 inhibitor U0126, and 10 μmol/L p38 inhibitor SB203580 to inhibit the main signaling molecules of the MAPK signaling pathway, and then RT-qPCR and Western blot were used to detect the changes in the osteogenic capacity of BMSCs. RESULTS: The results of CCK-8 showed that 80 and 160 μmol/L KA significantly affected the activity of BMSCs, and 10, 20 and 40 μmol/L KA were used for further experiments. The results of ALP and ARS staining indicated that 10 μmol/L KA had the best osteogenic induction effect. The results of RT-qPCR and Western blot showed that KA at 10 μmol/L had the best effect on the expression of mRNA and protein related to osteogenesis. The results of prediction analysis suggested that KA might promote osteogenic differentiation of BMSCs through EGFR- MAPK pathway. Western blot results showed that BMSCs activated MAPK signaling pathway after KA treatment, and the osteogenic promotion effect of BMSCs was decreased after treatment with MAPK signaling pathway-related inhibitors. CONCLUSIONS: Certain concentration of KA can stimulate and regulate BMSCs, activate EGFR-MAPK-RUNX2 signaling pathway and promote osteogenic differentiation.

Key words: Kaempferol-3-O-arabinoside, Bone marrow mesenchymal stem cells, Osteoblast, MAPK signaling pathway, Epidermal growth factor receptor, Osteogenic differentiation

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