脂肪源性神经干细胞诱导分化为γ-氨基丁酸能神经元的体外研究  

Differentiation of adipose-derived neural stem cells into GABAergic neurons in vitro

作　　者：任红苗[1] 王宜南[1] 陈继川[1] 张世昌[1] 

机构地区：[1]第三军医大学大坪医院野战外科研究所耳鼻咽喉-头颈外科,重庆400042

出　　处：《第三军医大学学报》2012年第11期1067-1070,共4页Journal of Third Military Medical University

基　　金：重庆市自然科学基金(CSTC2009BA5015)~~

摘　　要：目的探讨诱导脂肪源性神经干细胞(neural stem cells,NSCs)分化为GABA能神经元的体外培养方法,为细胞移植修复神经系统损伤提供种子细胞。方法贴壁培养C57BL/6小鼠(附睾脂肪垫)脂肪来源的干细胞(adipose-derived stem cells,ADSCs),流式细胞仪进行表型鉴定(CD45、CD90、CD105)及标志物纤连蛋白(fibronectin)鉴定后,在碱性成纤维细胞生长因子(basic fibroblast growth factor,bFGF)、表皮生长因子(epidermal growth factor,EGF)、2%B27的Neuro-basal培养基中诱导成NSCs。用免疫荧光细胞染色法进行NSC特异性标记物Nestin、增殖能力BrdU检测,再次添加脑源性神经营养因子、骨形成蛋白2后,维甲酸(RA)诱导分化为γ-氨基丁酸能神经元,并进行其标志物GABA的检测。结果分离纯化的ADSCs CD90、CD105表达强阳性,CD45表达阴性;诱导后的干细胞球经Nestin及BrdU鉴定为NSCs,且具有较强的增殖能力;神经干细胞诱导分化后的神经元GABA抗体染色呈阳性。结论在特定神经营养因子作用下,ADSCs在体外诱导生成的NSCs可被诱导分化为GABA能神经元。Objective To provide the seed cells for the repair of nervous system after cell transplan-tation by studying how to induce the differentiation of adipose-derived neural stem cells（NSCs） into GABAergic neurons in vitro.Methods ADSCs,isolated from the epididymal fat pads in C57BL/6 mouse,were adherently cultured.After cell surface antigens（CD45,CD90 and CD105） and ADSCs marker（fibronectin） were identified by flow cytometry,NSCs were induced from neurobasal medium containing basic fibroblast growth factor（bFGF）,epidermal growth factor（EGF） and 2% B27.Nestin,a specific marker of NSC,and its self-proliferation ability（BrdU）were detected with immunofluorescence staining.Differentiation of ADNSC into GABAergic neurons was induced with retinoic acid（RA） by adding neurotrophic factor and bone-formation protein 2.Marker of GABAergic neurons was then detected.Results The CD90,CD105 and fibronectin were positively expressed while the CD45 was negatively expressed in isolated and purified ADSCs.The stem cells,identified as NSCs with nestin and BrdU,showed a potential of self-proliferation ability.The antibody to GABAergic neurons was positively stained.Conclusion ADSCs-derived NSCs in vitro can differentiate into GABAergic neurons under the action of specific nerve growth factors.

关 键 词：脂肪来源干细胞 神经干细胞 Γ-氨基丁酸能神经元 

分 类 号：R322.8[医药卫生—人体解剖和组织胚胎学] R329-33[医药卫生—基础医学]