慢病毒载体介导红色荧光蛋白基因转染人脑胶质瘤干细胞的实验研究  

作　　者：陈金明[1] 周幽心[1] 陈桂林[1] 韦永新[1] 吴庭枫[1] 

机构地区：[1]苏州大学附属第一医院神经外科,215006

出　　处：《浙江临床医学》2012年第5期516-518,共3页Zhejiang Clinical Medical Journal

基　　金：江苏省卫生厅135重点学科建设基金资助（No.k201106）;苏州市科技计划（SYS201025）

摘　　要：目的探讨HIV-1来源的慢病毒载体介导红色荧光蛋白（RFP）基因转染人脑胶质瘤干细胞可行性和试验方法。方法用无血清干细胞培养基培养人脑胶质瘤干细胞，然后用细胞免疫荧光染色检测其干细胞特性表面标志物CDl33、Nestin表达情况。以HIV-1来源的慢病毒为载体，以RFP基因为目的的基因转染人脑胶质瘤干细胞，荧光显微镜下观察RFP阳性细胞表达情况及其转染率。MTT法检测转染后细胞与未转染RFP细胞组细胞增殖情况。再次用细胞免疫荧光染色检测转染后脑胶质瘤干细胞表面标志物CDl33、Nestin表达情况。结果红色荧光慢病毒转染胶质瘤干细胞体外连续培养后，RFP在干细胞中稳定表达，在荧光显微镜下即发出红色荧光，并且转染效率高。RFP．1entivirus转染后干细胞与未转染RFP组比较，生长曲线无明显差异，且转染后干细胞表面标志物仍表达阳性。结论采用HIV-1来源的慢病毒载体介导以RFP基因为生物标记物标记人脑胶质瘤干细胞是可行的，以慢病毒转染对干细胞的生长影响小，转染效率高。Objective To understand the feasibility and methods of red fluorescent protein （ RFP ） transfection in glioma stem cells by RFP-Lentivirus vector HIV-1. Methods The glioma stem cells were cultivated in serum free medium, then, immunostained with stem cell surface marker antibodies - CD133,Nestin. The glioma stem cells was transfected with the lentivirus vector HIV-1 containing gene. Cells expression of RFP were observed by fluorescent microscopy and RFP transfection ratio was detected; Cell proliferation of RFP untransfected and transfected group were assayed by MTT method. Finally, RFP-positive glioma stem cells were used for the detectinon of CD133, Nestin expression in immunofluorescence again. Results After glioma stem cells were transfected by RFP-Lentivirus vector HIV-1 and continuous passage cultured in vitro. RFP gene was stably highly expressed in glioma stem cells .Compared with RFP untransfected group, the cell growth curve of RFP transfected group had no significant difference （ P〉0.05 ） , RFP transfected group still expresses stem cell surface marker. Conclusion RFP can be highly effectively transfered into glioma stem cells under HIV-1 lentivirus vector mediation, and has no influence to cell growth.

关 键 词：胶质瘤干细胞 慢病毒 红色荧光蛋白 

分 类 号：R739.41[医药卫生—肿瘤]