脐血单个核细胞诱导的神经干细胞中Foxg1和Nestin基因的表达及其相互关系  

Expressions and Relationship between Foxg1 Gene and Nestin Gene in Neural Stem Cells Induced from Human Umbilical Cord Blood Mononuclear Cells

作　　者：王军[1] 范亚珍[1] 陈海[1] 朱登纳[1] 吴值荣[1] 侯艳艳[1] 

机构地区：[1]郑州大学第三附属医院脑瘫康复科,郑州450052

出　　处：《实用儿科临床杂志》2010年第11期848-850,868,共4页Journal of Applied Clinical Pediatrics

基　　金：河南省医学科技攻关计划项目(200703066)

摘　　要：目的探讨人脐血单个核细胞(UBC-MNCs)体外分离培养和定向诱导分化为神经干细胞(NSCs)的机制,观察培养细胞增殖分化情况,检测NSCs中Foxg1和Nestin基因mRNA的表达情况及相互关系。方法从脐血中分离出UBC-MNCs,用含人表皮细胞生长因子(hEGF)、碱性成纤维细胞生长因子(bFGF)和B27因子的Neurobasal培养基联合诱导其向NSCs方向分化,观察NSCs增殖分化及形态学特点;免疫组织化学法检测培养细胞中神经细胞标志抗原巢蛋白(Nestin)、神经元特异性烯醇化酶(NSE)、胶质纤维酸性蛋白(GFAP)的表达;5-溴-2-脱氧尿嘧啶核苷(Brdu)标记靶细胞,并测定增殖指数;RT-PCR法检测诱导前、培养3d、6d、9d、12d细胞中Foxg1和Nestin基因mRNA的表达变化。结果 UBC-MNCs可定向诱导分化为神经元样细胞,表达Nestin、NSE、GFAP标记抗原。Brdu标记细胞阳性率达90%。Foxg1mRNA在诱导前细胞中低表达,诱导后逐渐升高(P<0.05),6d达高峰,后逐渐下降(P<0.05);NestinmRNA表达逐渐升高(P<0.05)。结论体外诱导培养可获得UBC-MNCs源性NSCs,表达神经细胞特异性标志物Nestin、NSE和GFAP蛋白,Foxg1和Nestin基因表达明显增强,是NSCs增殖分化的关键调控因子。脐血能够成为NSCs的新来源。Objective To explore the mechanism of human umbilical cord blood mononuclear cells （ UBC - MNCs） that were isolated and cultured in vitro, induced and differentiated into neural stem cells （NSCs）. The proliferation and differentiation of cultured cells were ob- served. The expressions and relationship between Foxgl gene and Nestin gene mRNA,which appeared in NSCs,were detected. Methods The UBC- MNCs were isolated from human umbilical cord blood, cultured in Neurobasal medium added with human epidermal growth factor （hEGF） ,basic fibroblast growth factor （bFGF） and B27 factures in vitro,and the UBC - MNCs were induced and differentiated into NSCs. The characteristics of NSCs morphology ,proliferation and differentiation were observed. Cultured cells specific neuron antigen markers of Nes- tin, neuron specific enolase （ NSE）, glial ~oriliary acidic protein （GFAP） were detected by immunohistochemistry. Target cells were traced by 5 -bromo- 2 -deoxyuridine （Brdu） and cell proliferation indexes were determined. Reverse transcription polymerase chain reaction（ RT- PCR） for Foxgl gene and Nestin gene were performed to confirm their mRNA expressions in cells as before induction they were cultured for 3 d,6 d,9 d and 12 d,respectively. Results Most UBC - MNCs could be differentiated into neuron - like cells after oriented induction, ex- pressing labeled antigens of Nestin, NSE and GFAP. The positive rate of Brdu labeled cells went up to 90%. The expression of Foxgl gene mRNA was low at first, then gradually increased（ P 〈 0.05）, and came to its peak on the 6＇h day, then gradually declined（ P 〈 0.05 ）. Nestin gene mRNA gradually increased（ P 〈 O. 05 ）. Conclusions NSCs can be obtained from UBC - MNCs that is induced and cultured in vitro. Foxg1 gene and Nestin gene are the key proliferation and differentiation regulation factors of NSCs. Human cord blood can be the new source of NSCs.

关 键 词：Foxg1 NESTIN 脐血 神经干细胞 细胞培养 诱导分化 

分 类 号：R329[医药卫生—人体解剖和组织胚胎学]