绿色荧光蛋白转基因小鼠骨髓间充质干细胞的多向分化潜能研究
Study on Multi-Lineage Potential of Bone Marrow Mesenchymal Stem Cells Derived from Green Fluorescent Protein
Transgenic Mice
作者：李志勇,田卫东,刘磊,陈希哲,林云锋,闫征斌,陈玲,李声伟
Author：LI Zhi-yong, TIAN Wei-dong, LIULei, CHENXi-zhe, LIN Yun-feng, YAN Zheng-bin, CHENLing, LI Sheng-wei
收稿日期：2005-04-25 年卷（期）页码：2005,23(02):152-
期刊名称：华西口腔医学杂志
Journal Name：West China Journal of Stomatology
关键字：绿色荧光蛋白,转基因小鼠,骨髓间充质干细胞,多向分化潜能,
Key words：green fluorescent protein,transgenic mice,mensenchymal stem cells,multi-lineage potential,
基金项目：
国家重大基础研究前期研究专项资助项目(2002CC C00700);教育部高校优秀青年教师教学科研奖励计划资助项目 (2003682)
中文摘要
目的 研究绿色荧光蛋白(GFP)转基因小鼠骨髓间充质干细胞(MSCs)体外定向诱导下的多向分化潜能。
方法 取6周龄GFP转基因小鼠1只,用密度梯度离心法结合贴壁法体外培养获得GFP转基因小鼠骨髓MSCs有
限细胞系(GFP-MSCs)。取第3代GFP-MSCs进行体外多向分化诱导:成骨诱导后进行碱性磷酸酶增殖活性检测及
茜素红钙盐染色;成脂肪诱导20 d后进行油红O染色鉴定;成神经诱导6 h后用免疫组织化学方法检测神经元烯
醇化酶(NSE)的表达。结果 GFP-MSCs经成骨诱导后ALP表达较对照组明显升高,20 d后茜素红染色可见有橘红
色钙盐沉积;经成脂诱导后油红O染色阳性;经成神经诱导后,细胞形态由梭形转变为星状细胞,NSE染色呈强阳
性表达。结论 GFP转基因小鼠骨髓MSCs在体外诱导下具有多向分化能力,对GFP稳定表达无影响,可以作为研
究MSCs多向分化潜能机制的一个有效示踪工具。
英文摘要
Objective To study the multi-lineage potential of bone marrow mesenchymal stem cells (MSCs) derived from transgenic mice with green fluorescent protein (GFP) genein vitro.Methods A6-week-old GFP transgenic mouse was executed by dislocation of cervical vertebra, and the marrow in tibia and thighbone was washed outwith asepsis. The limited cell strains of MSCs derived from GFP transgenic mice(GFP-MSCs)were obtained with density gradient centrifugation. The passage 3 GFP-MSCs were induced to differentiate into osteoblast, adippcyte, neuron with solution of calcium induction medium, adipogenic medium and neural induction medium respectively. After being calcium-induced, the activity of alkaline phosphatase on GFP-MSCs was determined by micro-plate reader, and alizarin red staining was performed to test the formation of calcium concentration. The adipo-induced MSCs were detected with oil red O staining. Immunocytochemical stainingwas performed to detect the expression of NSE on neuron-induced MSCs.Results The ALP activity of GFP-MSCs heightened gradually alongwith being calcium-induced, and alizarin red staining showed positive. Oil red O staining of adipo-induced cells and NSE immunocytochemical staining of neu- ron-induced cells demonstrated positive.Conclusion The limited cell strain of GFP-MSCs possessesmulti-lineage potential,which can be used as an efficient tracking facility for studying the mechanism of multi-lineage potential on the MSCs.
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