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目的 探究加入成骨诱导前后脂肪间充质干细胞来源（dipose mesenchymal stem cells,ADSCs）与脐带间充质干细胞来源（umbilical cord mesenchymal stem cells,UCMSCs）的细胞外囊泡（extracellular vesicles,EVs）对骨髓间充质干细胞（bone marrow mesenchyml stem cells,BMSCs）的成骨能力是否存在差异。方法 对大鼠ADSCs和UCMSCs成骨诱导培养和普通培养，采用超速离心法提取不同培养方式大鼠ADSCs和UCMSCs来源EVs并进行鉴定，实验分为5组：BMSCs未添加EVs（对照组）、BMSCs+普通ADSC-EVs培养组（A组）、BMSCs+成骨ADSCsEVs培养组（O-A组）、BMSCs+普通UCMSC-EVs培养组（U组）、BMSCs+成骨UCMSCs-EVs培养组（O-U组）。CCK-8检测EVs对BMSCs增殖能力的影响，通过碱性磷酸酶染色、茜素红染色、qRT-PCR及Western blot检测添加不同培养条件EVs对BMSCs成骨能力的影响。结果 A组、O-A组、U组、O-U组较对照组增殖能力明显增强，A组和U组增殖能力明显高于其他组（P<0.05）。碱性磷酸酶染色及定量分析显示，O-A组和O-U组较对照组染色面积大且颜色深，碱性磷酸酶活性显著升高（P<0.01）；茜素红染色显示，O-A组和O-U组较对照组矿化结节明显，OD值明显大于对照组（P<0.01）。qRT-PCR和Western blot结果分别显示，O-A组、O-U组成骨相关因子在mRNA水平和蛋白水平均显著增强（P<0.05）。结论 成骨培养后的UCMSC-EVs、ADSC-EVs能促进BMSCs向成骨细胞方向分化。
Abstract：Objective To explore whether there are differences in the osteogenic ability of bone marrow mesenchymal stem cells after the addition of extracellular vesicles derived from adipose mesenchymal stem cells and umbilical cord mesenchymal stem cells before and after osteogenic induction. Methods The EVs derived from rat ADSCs and UCMSCs were extracted by ultracentrifugation and identified. The experiments were divided into five groups: BMSCs group(control),BMSCs+ normal ADSC-EVs culture group(A), BMSCs+ osteogenic ADSCs-EVs culture group(O-A), BMSCs+ normal UCMSC-EVs culture group(U), BMSCs+ osteogenic UCMSCs-EVs culture group(O-U). The CCK-8 assay was used to evaluate the impact of five distinct groups on the proliferative capacity of BMSCs. Alkaline phosphatase staining(ALP), alizarin red S staining(ARS), qRT-PCR and western blot detection were performed to detect BMSCs osteogenic differentiation. Results The proliferation capacity of Group A, Group O-A, Group U, and Group O-U was significantly enhanced compared with the control group, and the proliferation capacity of Group A and Group U was significantly higher than that of the other groups(P<0.05).ALP staining and activity analysis showed that the groups O-A and O-U had larger staining area and darker color than the control group, and ALP activity was significantly increased(P<0.01).ARS staining showed that the mineralized nodules in groups O-A and O-U were more obvious than those in control group,and their OD values were significantly higher than those in control group(P <0.01). Meanwhile, the results of qRTPCR and western blot analysis demonstrated a significant enhancement in the expression of osteogenic factors at both the m RNA and protein levels in groups O-A and O-U(P<0.05). Conclusion After osteogenic induction, ADSC-EVs and UCMSC-EVs could significantly enhance the osteogenic differentiation ability of BMSCs.
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基本信息:
DOI：10.19749/j.cn.cjgd.1672-2973.2025.03.001
中图分类号:R329.2
引用信息:
[1]李柔,徐宝华,郭燕宁,等.成骨培养条件下干细胞来源细胞外囊泡对骨髓间充质干细胞成骨性能分析[J].中华老年口腔医学杂志,2025,23(03):161-168.DOI:10.19749/j.cn.cjgd.1672-2973.2025.03.001.
基金信息:
中央高水平医院临床科研业务费资助项目（2022-NHLHCRF-ZSYX-02,2023-NHLHCRF-YXHZ-MS-09）
2025-05-20
2025-05-20