人脐血间充质干细胞来源的外泌体:分离鉴定及生物学特性

Exosomes derived from human umbilical cord blood mesenchymal stem cells: isolation, identification and biological characteristics

背景:外泌体是间充质干细胞分泌的膜性小囊泡,越来越多的研究表明,间充质干细胞可能通过旁分泌外泌体而发挥对组织损伤的修复作用,目前人脐血间充质干细胞来源的外泌体分离鉴定尚未见报道。目的:分离纯化人脐血间充质干细胞来源的外泌体,并鉴定其生物学特性。方法:收集人脐血间充质干细胞培养上清,采用超滤及梯度离心法分离并纯化外泌体,通过透射电镜观察其形态,流式细胞术检测其表面标志CD63、CD81、CD90、CD73、CD105、CD29、CD166。结果与结论:人脐血来源的间充质干细胞能够分泌外泌体,电镜下呈椭圆或碟状的囊泡结构,直径40-100 nm。外泌体表达其共性表达标志CD63、CD81及间充质干细胞表面黏附分子CD90、CD73、CD105、CD29、CD166。结果表明采用超滤及梯度离心法可以成功从间充质干细胞培养上清中分离纯化外泌体,间充质干细胞分泌的外泌体具有外泌体的胞膜蛋白组分。

BACKGROUND：Exosomes are membrane vesicles secreted by mesenchymal stem cells. Increasing studies have shown that mesenchymal stem cells can secrete exosomes via paracrine function to play a role in tissue injury. However, reports on how to isolate and identify exosomes derived from human umbilical cord blood mesenchymal stem cells are few. OBJECTIVE：To extract, purify and identify exosomes derived from human umbilical cord blood mesenchymal stem cells. METHODS：The cellculture supernatant of human umbilical cord blood mesenchymal stem cells was col ected. Exosome was extracted and purified with ultrafiltration and gradient centrifugation methods. The morphology of exosome was observed by transmission electronic microscope, and the expressions of CD63, CD81, CD90, CD73, CD105, CD29, and CD166 in exosome of mesenchymal stem cells were analyzed by fluorescent activated cellsorting. RESULTS AND CONCLUSION：Mesenchymal stem cells from human umbilical cord blood secreted exosome＆amp;nbsp;which exhibited el iptic or saucer-like shape and its diameter ranged from 40 to 100 nm with membrane structure. Exosome could express the common surface adhesion molecules CD63, CD81 and the surface adhesion molecules CD90, CD73, CD105, CD29, CD166 of mesenchymal stem cells. These findings indicate that exosome may be secreted by mesenchymal stem cells of human umbilical cord blood, which contains plasma membrane proteins of umbilical cord blood mesenchymal stem cells.