摘要
目的:比较全骨髓贴壁法与密度梯度离心法分离培养小鼠内皮祖细胞(EPCs)的差异。方法:取8周龄雄性近交系昆明小鼠,提取原代EPCs,分别采用全骨髓贴壁法和密度梯度离心法分离培养细胞,观察细胞形态、细胞摄取DiI-acLDL、结合FITC-UEA-I双阳性率及细胞生长增殖情况。结果:全骨髓贴壁法培养第4天,可见较多贴壁细胞,呈圆形、短梭形,部分为长梭形;第6~8天贴壁细胞多为梭形及类圆形;随着培养时间的延长,梭形细胞数量渐渐增多,可出现细胞集落,10d后EPCs大量增殖,14d细胞长满培养瓶底部。密度梯度离心法接种24h后可见圆形贴壁细胞,并逐渐增多变形;培养至第4~7天,可以观察到贴壁细胞明显增多且体积变大,细胞为类圆形、梭形或多边形,第7~9天,细胞呈集落生长,多数细胞呈多边形或长梭型;培养至第2周可见贴壁细胞生长呈"铺路石"样外观。全骨髓贴壁法细胞摄取DiI-acLDL和结合FITC-UEA-I双阳性率高于密度梯度离心法(P<0.05),但两种方法细胞增殖率比较无明显差异(P>0.05)。结论:全骨髓贴壁法与密度梯度离心法均可培养出EPCs,且两种方法培养的细胞增殖活性相当,但前者更简单易行,EPCs培养成功率更高。
Objective:To compare the effects of whole bone marrow culture method and density gradient cen-trifugation method on isolating mouse endothelial progenitor cells(EPCs).Methods:Primary EPCs were ob-tained from male KM mice(8 weeks old)and isolated by whole bone marrow culture method and density gradient centrifugation method.Cell morphology,the abilities of uptaking Dil-acLDL and binding FITC-UEA-I,and cell proliferation were detected.Results:Primary cells isolated with whole bone marrow culture method showed round,short spindle shape or long spindle shape on day 4,and adherent cells mainly dis-played similar circular shape and spindle shape on day 6 to 8.Primary cell cultured with density gradient centrifugation method showed round,polygonal and long spindle shapes,and"cobblestone"morphology.The immunofluorescence showed that double staining positive rate of Dil-acLDL and FITC-UEA-I in cells cultured with whole bone marrow culture method was higher compared with density gradient centrifugation method(P〈0.05),but no significant difference was found in cell proliferation rate between the two meth-ods(P>0.05).Conclusion:The proliferation ability of EPCs cultured with whole bone marrow culture method and density gradient centrifugation method was similar,but the former was more easy to conduct and higher success rate.
作者
赵文飘
戴霞
刘玉花
翟露
林媛
罗组纯
Zhao Wenpiao;Dai Xia;Liu Yuhua;Zhai Lu;Lin Yuan;Luo Zuchun(Guanxi Medical University,Nanning 530021,China;The Fist Affiliated Hospital of Guanxi Medical University,Nanning 530021,China)
出处
《广西医科大学学报》
CAS
2018年第3期286-289,共4页
Journal of Guangxi Medical University
基金
国家自然科学基金资助项目(No.81660147)
关键词
内皮祖细胞
细胞培养
全骨髓贴壁法
密度梯度离心法
mouse
endothelial progenitor cells
cell culture
whole bone marrow adherence method
densi ty gradient centrifugation
