人脐带间充质干细胞源外泌体对血管内皮细胞缺氧-复氧损伤的保护作用  

Protective effect of human umbilical cord mesenchymal stem cell-derived exosomes on vascular endothelial cells injured by hypoxia/reoxygenation

作　　者：朱华 赵翡 马玲 马海英 卢晓梅 张宇 于艳秋 ZHU Hua;ZHAO Fei;MA Ling;MA Hai-ying;LU Xiao-mei;ZHANG Yu;YU Yan-qiu(Department of Pathophysiology,Collage of Basic Medical Sciences,China Medical University,Shenyang 110122;Shenyang Cell Center Co.Ltd.,Shenyang 110196,China)

机构地区：[1]中国医科大学基础医学院病理生理教研室,辽宁沈阳110122 [2]沈阳细胞治疗工程技术研发中心有限公司,辽宁沈阳110196

出　　处：《解剖科学进展》2024年第3期225-228,232,共5页Progress of Anatomical Sciences

基　　金：国家自然科学基金(2600015005)。

摘　　要：目的研究人脐带间充质干细胞来源的外泌体(hUCMSCs-exo)对血管内皮细胞缺氧-复氧(H/R)损伤的保护作用。方法取足月新生儿脐带中的华通氏胶以组织块贴壁法提取人脐带间充质干细胞(hUCMSCs),流式细胞术检测hUCMSCs表面标志物CD34、CD45、CD73、CD90、CD105,成脂、成骨实验检测hUCMSCs的分化能力。超速离心法提取hUCMSCs培养上清液中的外泌体,电镜检测hUCMSCs-exo的形态及Western blot检测hUCMSCs-exo的标志蛋白CD9、CD63。人脐静脉内皮细胞(HUVECs)经缺氧(1%O_(2))培养12 h,然后常氧(95%空气)培养12 h,以建立H/R模型。实验分组:对照组、H/R组、hUCMSCs-exo组。台盼蓝染色检测细胞存活率;CCK-8检测HUVECs增殖能力;Transwell实验检测HUVECs的迁移能力、成血管实验检测HUVECs的成血管能力。结果自华通氏胶中提取的干细胞CD73、CD90、CD105标记物表达阳性,CD34、CD45表达阴性,具有成脂、成骨分化能力。外泌体标志物CD9、CD63呈阳性表达。与对照组相比,H/R组活率降低,增殖、迁移、成血管能力下降,而hUCMSCs-exo提高了H/R后HUVECs的存活率,改善增殖能力、迁移能力和管腔形成能力。结论hUCMSCs-exo对H/R导致的HUVECs的损伤具有保护作用。Objective To study the protective effect of human umbilical cord mesenchymal stem cell-derived exosomes(hUCMSCs-exo)on vascular endothelial cell injured by hypoxia/reoxygenation(H/R).Methods Human umbilical cord mesenchymal stem cells(hUCMSCs)were extracted from the Wharton′s jelly of full-term newborn umbilical cord by tissue adherence method.The surface markers CD34,CD45,CD73,CD90,CD105 of hUCMSCs were detected by flow cytometry.The differentiation ability of hUCMSCs was detected by adipogenic and osteogenic experiments.The exosomes in the culture supernatant of hUCMSCs were extracted by ultracentrifugation.The morphology of hUCMSCs-exo was detected by electron microscope and the marker proteins CD9 and CD63 of hUCMSCs-exo were detected by Western blot.HUVECs were cultured for 12 h with hypoxia(1%O_(2))and then incubated with normoxic(95%空气)for 12 h to establish an H/R model.Experimental groups were divided into control,H/R,hUCMSCs-exo groups.Typan blue staining was used to detect the cell viability of HUVECs,CCK-8 was used to detect the proliferation ability of HUVECs,Transwell assay was used to detect the migration ability of HUVECs,and tube formation assay was used to detect the angiogenic ability of HUVECs.Results The CD73,CD90 and CD105 markers of hUCMSCs extracted from Wharton′s jelly were positive,but negative for CD34 and CD45 markers,and hUCMSCs had the adipogenic and osteogenic ability.The vesicle morphological characteristics of exosomes were observed by electron microscope,and the positive expressions of CD9 and CD63 were detected by Western blot.The cell viability and ability of proliferation and migration and angiogenesis were lower in H/R group than in control group.hUCMSCs-exo improved the cell viability of HUVECs after H/R,ameliorated their proliferation ability,migration ability and tube formation ability.Conclusion hUCMSCs-exo has a protective effect on the damaged HUVECs by H/R injury.

关 键 词：人脐带间充质干细胞 外泌体 血管内皮细胞 缺氧-复氧 

分 类 号：R331.3[医药卫生—人体生理学]