慢病毒载体转染人骨髓间充质干细胞后其分化潜能的研究  

Differentiation Potential of EGFP Gene-Modified Mesenchymal Stem Cells Transfected by Lentiviral Vector

作　　者：陈慧玲[1,2] 白海[2] 

机构地区：[1]第四军医大学西京医院血液科,陕西西安710032 [2]兰州军区总医院血液病研究所,甘肃兰州730050

出　　处：《中国实验血液学杂志》2009年第2期400-403,共4页Journal of Experimental Hematology

摘　　要：本研究探讨携带增强型绿色荧光蛋白(EGFP)的慢病毒载体(lentiviral vector,LV)转染人骨髓间充质干细胞(mesenchymal stem cells,MSC)后的分化能力。从骨髓中分离、培养MSC,通过携带EGFP标记的慢病毒载体体外转染人MSC,观测绿色荧光蛋白的表达情况。最后,应用成脂诱导培养基(胰岛素、吲哚美辛、甘油-3-磷酸酰基转移酶IBMX、地塞米松)定向诱导分化已转染的MSC为脂肪细胞,苏丹黑染色观察诱导情况。结果表明,转染72小时后的MSC在荧光显微镜下显现弱荧光;经脂肪诱导培养基培养21天后在显微镜下可观察到细胞胞浆内形成高度折光性的脂滴,苏丹黑染色呈橘红色,且转染组与未转染组差异不大(p>0.05)。结论:成功应用携带EGFP基因的慢病毒载体对MSC进行基因修饰,使转染后的MSC仍具有分化能力,能被诱导为脂肪细胞。MSC可作为基因治疗的一种靶向性细胞。This study was aimed to investigate the differentiation potential after EGFP gene-modified mesenchymal stem cells （MSCs） transfected by lentiviral vector（LV）. After isolated, cultured and identified, MSCs were transfected with the lentiviral vector carrying EGFP gene in vitro. The transfection efficiency was measured by observing the expres- sion of green fluorescence protein. At last, the transfected MSCs were induced into adipocytes in adipogenesis supple- ment medium, the induction level was detected by Sudan fat stain. The results indicated that after transfection for 72 hours, weak fluorescence in MSCs was observed under fluorescence microscope. After 21 days, many lipid droplets with high refractivity occurred in cytoplasm of transfected MSCs, and showed orange in Sudan black stain. There were no significantly differences between transfected and non-transfected cells （p 〉 0.05 ）. It is concluded that MSCs were successfully transfected by LV carrying EGFP gene. The transfected MSCs maintain multiple differentiation and proliferation potential. In the adipogenesis supplement medium, transfected MSCs also can be induced to differentiate into adipocytes. MSCs can act as target cells for gene therapy.

关 键 词：间充质干细胞 慢病毒载体 增强型绿色荧光蛋白 基因转染 细胞分化 

分 类 号：R329.28[医药卫生—人体解剖和组织胚胎学] R730.54[医药卫生—基础医学]