人脐带间充质干细胞外泌体的分离鉴定及其溶血性能研究
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国家自然科学基金(31671202);南京医科大学科技发展基金(2016NJMUZD018)


Isolation and identification of exosomes derived from human umbilical cord mesenchymal stem cells and hemolysis effect of exosomes
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    摘要:

    目的:分离并鉴定人脐带间充质干细胞(human umbilical cord mesenchymal stem cells,HUCMSC)分泌的外泌体,研究其对人血的溶血性能。方法:通过超速离心法从脐带干细胞培养上清中分离外泌体,采用蛋白免疫印迹法、透射电镜和纳米颗粒跟踪分析法进行鉴定;制备人红细胞悬液,加入不同浓度外泌体,进行溶血试验,用试管法观察并用紫外分光光度法测定计算溶血度。结果:分离获得的外泌体特异富集蛋白CD9、CD81、Alix,不含有高尔基体蛋白Gm130;透射电镜下可见其呈杯盘状结构,粒径直径为40~120 nm;脐带干细胞外泌体对人血的溶血率小于5%。结论:HUCMSC分泌的外泌体具有外泌体的经典结构及典型特征,不引起人血溶血反应,为未来脐带干细胞外泌体作为载体进行药物递送提供了实验依据。

    Abstract:

    Objective:To isolate and identify exosomes derived from human umbilical cord mesenchymal stem cells (HUCMCS) and to testify the potential of hemolysis of exosomes. Methods:HUCMSC-derived exosomes were isolated and purified through ultracentrifugation. Western blot and transmission electron microscope were used to identify the characteristic of the exosome. Exosomes were incubated with human erythrocyte suspension to observe the hemolysis. Results:HUCMSCs derived exosomes expressed positive makers of CD9,CD81 and Alix,didn’t express the Golgi Body protein Gm130 as negative marker. The particles size displayed range from 40-120 nm. It can be seen a classical doughnut shape of exosomes through transmission electron microscope. And the exosome derived from HUCMSCs didn’t cause the hemolysis of human red cell. Conclusion:Exosomes secreted by human umbilical cord stem cells possessed classical characteristics of exosomes and had no effects on hemolysis,which provide the experimental basis of potential clinical use of HUCMSC exosomes for drug delivery.

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杨玮杰,李佳曌,陈良键,张 婧,刘凯鲁,张 驰,肖 月,李 晶.人脐带间充质干细胞外泌体的分离鉴定及其溶血性能研究[J].南京医科大学学报(自然科学版),2019,(2):165-170

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  • 收稿日期:2018-01-23
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  • 在线发布日期: 2019-03-06
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