摘要
目的建立一种 PKH26体外荧光标记人骨髓间质干细胞(MSCs)的方法。方法培养人骨髓间质干细胞,按 PKH26标记程序进行标记后培养,采用激光共聚焦显微镜、流式细胞分析,观察标记后人 MSCs 生长状态、荧光强度变化和传代培养效果。应用逆转录-聚合酶链反应(RT-PCR)等方法观察标记细胞的生物学功能。结果标记后 MSCs 呈红色荧光,标记率达100%,体外连续传代培养7代后,荧光标记细胞荧光强度逐渐减弱,荧光标记细胞百分比逐渐减低。PKH26标记MSCs 的生长形态、生长活力、nucleostemin、三磷酸甘油醛脱氢酶(GAPDH)基因表达及体外诱导分化能力等生物学特性不发生改变。结论 PKH26荧光标记人 MSCs 是一种有效、实用的方法,该方法对MSCs 转归、可塑性及 MSCs 移植治疗研究具有重要的价值。
Objective To establish a method of labeling human mesenchymal stem cells （MSCs） with PKH26 in vitro. Methods MSCs were cultured and labeled with PKH26 according to the manufacturer＇s instruction. The growth, fluorescence intensity and serial subcultivation of labeled MSCs were analyzed with the confocal laser microscope and the flow cytometry. The biological characteristics of labeled MSCs were investigated by RT-PCR. Results The labeled MSCs appeared red fluorescence and the labeling rate was 100 percent. During serial subcultivation of labeled MSC from passage 1 to passage 7, the fluorescence intensity and the labeling rate of MSCs were gradually decreased. The biological features such as morphology, growth, expression level of nucleostemin and GAPDH gene and capability of differentiation into osteoblast in vitro were not affected by labeling. Conclusion Labeling the human MSCs with PKH26 is an effective and practical method, which can be used as an important tool in the study on the homing, plasticity and transplantation of MSCs.
出处
《中华检验医学杂志》
CAS
CSCD
北大核心
2006年第9期834-837,共4页
Chinese Journal of Laboratory Medicine
基金
国家自然科学基金(30471938)
卫生部科学研究基金(WKJ2005-2-024)
镇江市检验医学重点实验室项目(SH2006066)
江苏大学创新预研基金(04CX07)
江苏大学高级人才启动基金(04JDG028)
关键词
间质干细胞
荧光标记技术
骨髓
Mesenchymal stem cells
Fluorescence labeling
Bone marrow
