促红细胞生成素对小鼠诱导多能干细胞源性神经干细胞向功能性神经元样细胞分化的影响

目的:探讨促红细胞生成素体外对小鼠诱导多能干细胞(iPSCs)来源的神经干细胞神经分化的影响。方法:神经干细胞培养基培养诱导iPSCs分化为神经干细胞,设4个浓度加入含促红细胞生成素的培养基,免疫细胞化学荧光鉴定神经元标志物微管相关蛋白2(MAP-2)、神经干细胞标志物巢蛋白的表达。FM 1-43染色技术观察FM 1-43染色颗粒消失的情况。结果:iPSCs在条件培养液的诱导下形成神经干细胞克隆,15 U/ml促红细胞生成素处理后神经干细胞克隆球向神经元方向分化,呈MAP-2阳性细胞。新分化的神经元有大量FM 1-43绿色阳性颗粒。结论:iPSCs体外诱导可获得神经干细胞,促红细胞生成素浓度为15 U/ml时可促进iPSCs源性神经干细胞向功能性的神经元方向分化。

Effect of erythropoietin on the differentiation of neural stem cells derived from mouse induced pluripotent stem cells into functional neuron-like cells

Objective:To study the effects of erythropoietin on differentiation of neural stem cells derived from mouse induced pluripotent stem cells into neuron-like cells.Methods:Primary neural stem cell culture medium was added to induce induced pluripotent stem cells(iPSCs)to neural stem cells,which were then inoculated in 6-well plate adherent culture,containing erythropoietin in the growing medium (4 subgroups).Morphological changes of the cells were observed under inverted microscope.Immunofluorescence staining technology was used to detect the expression of nestin and microtubule-associated protein 2 (MAP2),and FM 1-43 staining technique was used to observe the disappearance of the FM 1-43 staining particles, which showed the synaptic terminals with functional activity.Results:Neural stem cell clones were formed from iPSCs in the induction conditions.These clones could differentiated into neuron-like cells under treatment with 15 U/ml erythropoietin and these neuron-like cells could be marked by neuron-like marker MAP-2.Under depolarization,the activity of synapse was enhanced and a large number of FM 1-43 endocytosis particles were in axon terminal.Conclusion:iPSCs can be induced to differentiate into NSCs invitro,and erythropoietin (especially at 15 U/mL)can induce neural stem cells devived from mouse induced pluripotent stem cells to differentiate into functional neurons.