促红细胞生成素对脊髓源性神经干细胞分化能力的影响  

Effect of erythropoietin on the differentiation of adult spinal cord-derived neural stem cells in vitro

作　　者：李小波[1] 张辉[1] 王少滨[2] 刘杰[1] 余涛[1] 杜怡斌[1] 尹宗生[1] 

机构地区：[1]安徽医科大学第一附属医院骨科,合肥230022 [2]安徽医科大学第四附属医院骨科,合肥230022

出　　处：《安徽医科大学学报》2016年第1期14-17,共4页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金(编号:81171173)

摘　　要：目的探讨成体大鼠脊髓源性神经干细胞(NSCs)在促红细胞生成素(EPO)的作用下体外分化的形态学表现,为成体脊髓源性NSCs移植治疗脊髓损伤提供理论基础。方法利用无血清悬浮培养,体外分离培养SD大鼠脊髓源性NSCs,随后以含血清培养基贴壁诱导分化培养7 d,行免疫细胞化学染色,以抗巢蛋白(Nestin)鉴定NSCs,以抗微管相关蛋白2(MAP2)和胶质纤维酸性蛋白(GFAP)免疫荧光染色检测NSCs的分化情况。选取第3代NSCs,向培养基中添加EPO,使EPO的终浓度为10 IU/ml,并设不添加EPO的对照组,免疫荧光共聚焦显微镜下分别观察并计数各组神经元/细胞总数得出百分率。结果 SD大鼠脊髓组织体外悬浮培养可获得大量神经球,所获得的神经球均表达Nestin蛋白,以含血清培养基贴壁诱导分化培养,可检测出MAP2和GFAP阳性细胞;EPO组诱导分化后,表达MAP2阳性细胞较对照组显著提高(P<0.05)。结论 SD大鼠脊髓体外培养可获得NSCs,EPO可促进成体脊髓源性NSCs向神经元方向分化。Objective To investigate the promotion of the differentiation of erythropoietin（EPO） on the differentia- tion of the neural stem cells（NSCs） in vitro from spinal cord of SD rats,and to provide experimental basis for the treatment of spinal cord injury by transplantating neural stem cells. Methods The spinal cord was isolated and cul- tured in serum-free suspension, then a differentiation suspension cultured in serum culture medium for 7 days was applied to the NSCs. Nestin was used to detect the NSCs, the differentiation ratio of NSCs into neurons and glia cells was detected by immunofluorescent histochemistry for mierotubule-associated protein 2 （MAP2） and glia fibrillary acid protein （GFAP）. After obtained the third passage （ P3 ） of NSCs, EPO was added to the medium on a final concentration of 10 IU/ml ,control group without EPO, the differentiation proportion of MAP2-positive cells to total cells was identified by immunofluoreseence staining. Results The results showed that the separated cells from spi- nal cord of SD rats formed neurospheres, in which nestin-positive cells were detected. GFAP and MAP2-positive ceils were detected after differentiation culture. The proportion of MAP2-positive cells in the EPO group showed an significant increase compared with the control group （P 〈 0.05 ）. Conclusion The results indicate that NSCs can be obtained from the separated spinal cord of SD rats; EPO can increase the differentiation rate of NSCs into neu- rons.

关 键 词：神经干细胞 促红细胞生成素 神经元 分化 

分 类 号：R651.21[医药卫生—外科学]