新型神经营养因子TAT-BDNF对胚胎干细胞分化的影响  

In vitro differentiation of embryonic stem cells with a neotype neurotrophin TAT-BDNF

作　　者：蒋晖[1] 金大地[1] 瞿东滨[1] 鞠躬[2] 

机构地区：[1]南方医科大学南方医院脊柱外科中心,广东广州510515 [2]第四军医大学神经科学研究所,陕西西安710032

出　　处：《中国骨与关节外科》2008年第2期139-144,共6页Chinese Journal of Bone and Joint Surgery

基　　金：国家自然科学基金(30271322)

摘　　要：目的探讨合成的新型神经营养因子TAT-BDNF对胚胎干细胞(ESCs)分化的影响,为进一步TAT-BDNF联合干细胞移植治疗脊髓损伤提供实验依据。方法取ESCs E14进行体外培养,在全反视黄酸诱导分化的基础上加入20μg/LTAT-BDNF,通过免疫细胞化学染色观察此融合蛋白对ESC分化的影响。结果TAT-BDNF能促使E14胚胎干细胞分化为多巴胺能神经元和γ氨基丁酸能神经元。随着作用时间的延长,表达成熟神经元NF-200的细胞比例逐步提高(7 d时约37.4%±2.7%、21 d时82.3%±7.6%);TH阳性的细胞比例也逐渐增多(7 d时约7.9%±1.6%,21 d时44%±3.5%),而且神经元的轴突也更加细长。结论新型神经营养因子TAT-BDNF能促进ESCs细胞向神经元分化并促进神经元成熟。Objectives To investigate the contribution of a neotype neurotrophin TAT-BDNF on embryonic stem cell differentiation.Methods Embryonic stem cells were cultured in the absence of leukemia inhibitory factor and MEF for 4d and formed embryonic bodies(EB).EB was exposed to RA(10^(-6) mol/L) for 4 days.Cells allowed to differentiate in DF12/N2 medium for 7,14,or 21 day.In treatment group,BDNF and TAT-BDNF were added to induce differentiation. The production of specific cells from differentiating ES cells was evaluated by measuring the NF200,which is a marker of mature neuron,and the expressions of tyrosine hydroxylase(TH) and glutamic acid decarboxylase(GAD), which are markers of dopaminergic and GABAergic neurons.Immunofluorescence assays was performed to evaluate the effect of TAT-BDNF on ES cells differentiation.Results After induced by RA and added with TAT-BDNF during the differentiation stages,embryonic stem cells differentiated into opaminergic neurons and GABA neurons in vitro.Along with the prolong of effecting time,the proportion of mature neuron increased(37.4%±2.7%on day 7 and 82.3%±7.6% on day 21).Conclusions TAT-BDNF can enhance ESCs differentiation of neuron and promote neuron mature.

关 键 词：胚胎干细胞 脑源性神经营养因子 分化 脊髓损伤 

分 类 号：R329.1[医药卫生—人体解剖和组织胚胎学]