口腔医学研究

口腔医学研究 ›› 2025, Vol. 41 ›› Issue (5): 426-431.DOI: 10.13701/j.cnki.kqyxyj.2025.05.012

• 口腔生物学研究 • 上一篇    下一篇

富血小板血浆小细胞外囊泡促进骨修复的动物实验研究

高阳1,2, 高林波3, 史春3, 吴大雷1*   

  1. 1.大连医科大学附属第二医院口腔科 辽宁 大连 116000;
    2.大连市友谊医院口腔科 辽宁 大连 116000;
    3.大连医科大学口腔医学院 辽宁 大连 116000
  • 收稿日期:2024-07-15 出版日期:2025-05-28 发布日期:2025-05-26
  • 通讯作者: *吴大雷,E-mail:weekendkiller@126.com
  • 作者简介:高阳(1982~ ),女,辽宁人,硕士,副主任医师,主要从事口腔医学方面的研究。
  • 基金资助:
    辽宁省自然科学基金(编号:20180550563)

Small Extracellular Vesicles of Platelet-rich Plasma Promoted Bone Tissue Repair

GAO Yang1,2, GAO Linbo3, SHI Chun3, WU Dalei1*   

  1. 1. Department of Stomatology, the Second Affiliated Hospital of Dalian Medical University, Dalian 116000, China;
    2. Department of Stomatology, Dalian Friendship Hospital, Dalian 116000, China;
    3. School of Stomatology, Dalian Medical University, Dalian 116000, China
  • Received:2024-07-15 Online:2025-05-28 Published:2025-05-26

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摘要： 目的: 本文通过提取小鼠富血小板血浆的小细胞外囊泡(platelet-rich plasma derived small extracellular vesicles,PRP-sEV),检测PRP-sEV对成骨细胞骨标志因子的表达影响,以及体内观察PRP-sEV对小鼠颅骨缺损模型的骨修复作用,为探讨PRP-sEV对骨缺损修复研究提供依据。方法: 提取小鼠PRP-sEV,通过纳米颗粒追踪分析、透射电子显微镜以及Western blot对PRP-sEV大小、结构以及标志蛋白进行鉴定。采用PKH67检测MC3T3-E1细胞对PRP-sEV的摄取情况,Western blot检测成骨标志蛋白的表达情况。建立小鼠颅骨缺损模型,通过苏木精-伊红(hematoxylin-eosin, HE)染色及Masson染色观察PRP-sEV对骨缺损修复作用,通过免疫组织化学实验检测骨形态发生蛋白2(bone morphogenetic protein 2, BMP-2)及骨桥蛋白(osteopontin, OPN)在PRP-sEV作用的颅骨缺损模型中的表达情况。结果: 纳米颗粒追踪分析结果显示,PRP-sEV大小约为120 nm, 透射电子显微镜结果显示,PRP-sEV具有典型的脂质双分子层结构,Western blot结果显示,小细胞外囊泡阳性标志蛋白CD63、CD81、凋亡连接基因-2-相互作用蛋白X(ALG-2-interacting protein X,ALIX)均呈阳性,微囊表面标记物CD40呈阴性。PKH67检测MC3T3-E1细胞对PRP-sEV正常摄取,Western blot结果显示PRP-sEV促进成骨标志蛋白BMP-2及OPN的表达,差异有统计学差异(P<0.05)。HE染色及Masson染色结果显示,PRP-sEV可促进骨缺损的修复,免疫组织化学结果显示BMP-2及OPN在PRP-sEV作用的颅骨缺损模型的表达高于对照组,差异有统计学差异(P<0.05)。结论: PRP-SEV可以促进骨组织的修复。

关键词: 颅骨缺损模型, 富血小板血浆, 小细胞外囊泡, 骨修复

Abstract: Objective: To investigate the impact of platelet-rich plasma derived small extracellular vesicles (PRP-sEV) on the expression of bone markers in osteoblasts, as well as to observe the bone repair effect of PRP-sEV in a mouse calvarial defect model. Methods: The small extracellular vesicles were extracted from mouse platelet-rich plasma. These PRP-sEV were then analyzed for size, structure, and marker proteins using nanoparticle tracking analysis, transmission electron microscopy, and western blot. PKH67 was used to assess the uptake of these vesicles by MC3T3-E1 cells and used western blot to examine the expression of osteogenic marker proteins. A mouse cranial defect model was established and evaluated the bone repair effects of PRP-sEV through hematoxylin-eosin (HE) staining and Masson staining. Immunohistochemistry was performed to assess the expression of bone morphogenetic protein 2 (BMP-2) and osteopontin (OPN) in PRP-sEV in the cranial defect model. Results: The size of PRP-sEV was around 144 nm. Transmission electron microscopy revealed that these vesicles had a typical lipid bilayer structure. Western blot results indicated that the positive marker proteins CD63, CD81, and ALG-2-interacting protein X (ALIX) were present in the vesicles, while the surface marker protein CD40 was absent. PKH67 testing demonstrated that MC3T3-E1 cells effectively incepted the PRP-sEV. Western blot results showed that these vesicles enhanced the expression of bone formation markers BMP-2 and OPN (P<0.05). HE staining and Masson staining results indicated that the PRP-sEV promoted bone defect repair. Immunohistochemistry results demonstrated that the expression of BMP-2 and OPN in the cranial bone defect model treated with the vesicles was higher than that of control group (P<0.05). Conclusion: The extracellular vesicles in platelet-rich plasma could promote bone tissue repair.

Key words: cranial bone defect model, platelet-rich plasma, extracellular vesicles, bone repair