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外周血内皮祖细胞纯化与鉴定

发表时间:2017-09-15  浏览量:621  下载量:99
全部作者: 赵娴,魏韩笑,周箐竹,李杉,王松梅,刘流
作者单位: 昆明医科大学第一附属医院整形外科;云南大学生命科学院;昆明医科大学公共卫生学院
摘 要: 目的:从自体外周血提取纯化具有优异新生血管及管腔形成能力的内皮祖细胞(endothelial progenitor cells,EPCs),为组织工程血管化的研究提供优质、取材方便、血管化能力强的种子细胞。方法:抽取新西兰大耳白兔外周血,采用密度梯度离心法、贴壁法提取纯化外周血EPCs,在内皮细胞培养基(endothelial cell medium,ECM)内进行培养。取第三代细胞,利用免疫荧光染色的方法检测贴壁细胞CD34、CD133、vWF的表面标志,以及免疫组织化学双标染色鉴定EPCs. 结果:抽取的外周血用密度梯度离心法结合贴壁培养法提取纯化并进行扩增,获得高纯度的EPCs,第三代外周血EPCs抗原CD34、CD133、vWF免疫荧光鉴定均为阳性;EPCs的免疫组织化学双标染色(同时摄取乙酰化低密度脂蛋白及荆豆凝集素的免疫荧光复合染色)鉴定也证实,分离培养的细胞为血管EPCs. 结论:兔外周血经密度梯度离心分离及贴壁法纯化,可获得纯度较高的EPCs.
关 键 词: 生物医学工程学;外周血;内皮祖细胞;细胞纯化;免疫组织化学双标染色
Title: Purification and identification of EPCs derived from peripheral blood
Author: ZHAO Xian, WEI Hanxiao, ZHOU Qingzhu, LI Shan, WANG Songmei, LIU Liu
Organization: Department of Plastic Surgery, The First Affiliated Hospital of Kunming Medical University; School of Life Sciences, Yunnan University; College of Public Health, Kunming Medical University
Abstract: Objective: The endothelial progenitor cells (EPCs) derived from peripheral blood with excellent ability of vascularization were isolated and purified, to supply high quality and easily obtained seed cells for tissue engineering vascularization. Methods: EPCs were isolated from rabbits peripheral blood with density gradient centrifugation and cell attachment, and cultured in endothelial cell medium (ECM). The expressions of CD34, CD133 and vWF on the third generation adherent cells were detected with immunofluorescent staining, and EPCs were identified with immunohistochemical double staining. Results: Higher purity EPCs could be isolated with Ficoll density gradient centrifugation and cell attachment methods. The CD34, CD133 and vWF of the third generation peripheral blood derived EPCs were positive with immunofluorescent staining, and DiI labeled acetyl low-density lipoprotein cholesterol and fluorescein labeled lectin immunofluorescence double staining were positive, which were consistent with the EPCs characteristic. Conclusion: EPCs isolated from rabbits peripheral blood with Ficoll density gradient centrifugation and cell attachment were highly purified.
Key words: biomedical engineering; peripheral blood; endothelial progenitor cells; cell purification; immunohistochemical double staining
发表期数: 2017年9月第17期
引用格式: 赵娴,魏韩笑,周箐竹,等. 外周血内皮祖细胞纯化与鉴定[J]. 中国科技论文在线精品论文,2017,10(17):1934-1941.
 
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