摘要
背景:羊水干细胞是一种新的干细胞来源,具有不同于成体间充质干细胞的生物特性。目前有关羊水干细胞的研究多数以孕中期羊水为基础,对孕足月羊水所知甚少。目的:探讨孕足月羊水干细胞向神经元分化的可能性。设计、时间及地点:细胞学体外观察,于2007-10/2008-08在解放军总医院妇产科及解放军军事医学科学院干细胞及再生医学研究室完成。材料:孕足月羊水来自在解放军总医院行剖宫产、排除胎儿畸形的孕38～40周健康孕妇。方法:贴壁法从羊水中体外分离培养羊水干细胞,胰酶-EDTA消化传代。取第四五代细胞,用含1mmol/Lβ-巯基乙醇、体积分数为20%胎牛血清、10μg/L碱性成纤维生长因子的低糖培养基预诱导24h,去除培养液,洗涤后加入含5mmol/Lβ-巯基乙醇的无血清低糖培养基向神经元方向诱导4.0～5.0h;对照组不加任何诱导剂。主要观察指标:孕足月羊水干细胞的形态及免疫表型,诱导后向神经元样细胞的分化。结果:培养的羊水干细胞呈梭形,漩涡状排列,间质干细胞来源标志CD105,CD29及I类组织相容性抗原HLA-ABC均呈阳性表达,胚胎干细胞标志性基因SSEA-4亦呈阳性表达,不表达造血干细胞标志。诱导后,细胞形态发生明显改变,胞浆回缩,出现类似轴突的细长的两极或多极突起,免疫荧光染色神经元特异性烯醇化酶呈阳性表达。结论:在血清剥夺与β-巯基乙醇联合诱导条件下,孕足月羊水干细胞具有向神经元样细胞分化的潜力。
BACKGROUND： Amniotic fluid stem cells are a new source of stem cells, and have the biological property of adult mesenchymal stem cells （MSCs）. Presently, most studies of amniotic fluid stem cells are on the amniotic fluid during middle-term pregnant. The studies concerning full-term amniotic fluid are few. OBJECTIVE： To investigate the possibility of inducing full-term amniotic fluid stem cells to differentiate into neurons in vitro. DESIGN, TIME AND SETTING： The cytology in vitro study was performed at the Department of Gynaecology and Obstetrics, General Hospital of Chinese PLA and Research Room of Stem Cells and Regeneration Medicine, Academy of Military Medical Sciences of Chinese PLA from October 2007 to August 2008. MATERIALS： Full-term amniotic fluid was collected from healthy pregnant women （38-40 weeks） undergoing Caesarean section at the General Hospital of Chinese PLA, with the exception of fetal anomaly. METHODS： Stem cells were obtained from amniotic fluids by adherence method in vitro, and digested by trypsin-EDTA. Cells at passages 4 to 5 were induced in low-glucose medium containing 1 mmol/L β -mercaptoethanol, fetal bovine serum of 20% volume fraction and 10 μg/L basic fibroblast growth factors for 24 hours. After removal of medium, samples were washed and incubated in serum-free low-glucose medium containing 5 mmol/L β-mercaptoethanol for 4-5 hours. Cells in the control group were left intact. MAIN OUTCOME MEASURES： Morphology and immunophenotype of full-term amniotic fluid stem cells, and the differentiation of stem cells into neurons after induction were measured. RESULTS： Amniotic fluid stem cells were fibroblast-like, with the presence of whirlpool-shape, and positive for CD105, CD29 and HLA-ABC, as well as SSEA-4, but negative for the markers of hemopoietic stem cells. After induction, there were significant changes in cell morphology. Cytoplasm retracted and thin processes with two poles or multiple poles appeared. Cells were positive for neuron specific enolase by immunofluorescent staining. CONCLUSION： Full-term amniotic fluid-derived stem cells can differentiate into neuron-like cells in vitro under serum deprivation and β- mercaptoethanol induction.
出处
《中国组织工程研究与临床康复》
CAS
CSCD
北大核心
2009年第19期3680-3684,共5页
Journal of Clinical Rehabilitative Tissue Engineering Research
基金
全军医药卫生科研基金资助项目(06G101)~~
