EPO/EPOR信号通路对成体脊髓神经干细胞分化作用的实验研究

Experimental study on the effect of EPO / EPOR signaling pathway on the differentiation of neural stem cells in adult spinal cord

目的研究促红细胞生成素(EPO)及其受体(EPOR)信号通路对大鼠成体脊髓神经干细胞(NSCs)分化的影响。方法取成年SD大鼠脊髓细胞,在培养基DMEM/F12中进行培养,取第3代NSCs作为研究对象:在实验组的培养液中再加入重组人促红细胞生成素(rhE PO),同时在培养液中使用等体积生理盐水作为对照组。采用免疫荧光染色方法和Western blot法,检测NSCs上EPOR蛋白表达情况;采用免疫荧光染色法检测NSCs的分化现象。结果免疫荧光染色结果显示实验组与对照组NSCs中均有EPOR蛋白表达,实验组较对照组增强;Western blot结果显示实验组EPOR蛋白表达水平较对照组增高,差异有统计学意义(P<0.01);免疫荧光染色法显示实验组NSCs分化为神经元的比例高于对照组,差异有统计学意义(P<0.01)。结论rhEPO能激活EPO/EPOR信号通路,从而提高成体脊髓NSCs向神经元分化的能力。

Objective To study the influence of the erythropoietin（ EPO） and erythropoietin receptor（ EPOR） signaling pathway on the differentiation of adult rat spinal cord neural stem cells（ NSCs）. Methods Adult SD rat spinal cord cells were cultured in DMEM / F12 medium and the 3rd generation of the NSCs was chosen as the experimental object： in the experimental group the recombinant human erythropoietin（ rh EPO） was further added into the culture medium,meanwhile in a control group only using saline with an equal volume. By using immunofluorescence staining and Western blot,we measured EPOR expression in the NSCs,while we observed the differentiation of NSCs by immunofluorescence staining. Results The results of immunofluorescence staining showed that EPOR expression could be measured in both NSCs of the experimental group and the control group,and the experimental group EPOR protein maintained at a higher level than that of the control group. Western blot results showed that the expression level of EPOR protein in the experimental group was higher than that in the control group,the difference was statistically significant（ P 〈0. 01）. Immunofluorescence staining results of differentiation showed the NSCs of experimental group could differentiate into neurons,whose different proportion was more than the control group,the difference was significant（ P 〈0. 01）. Conclusion rh EPO can activate EPO / EPOR signaling pathway,thereby enhancing the differentiate ability of adult neural stem cells of spinal cord into neurons.