崔冬冰,范安然,何志旭
CUI Dongbing,FAN Anran,HE Zhixu

• 1:贵州医科大学组织工程与干细胞实验中心

摘要(Abstract)：

目的:建立一种高效分离培养人羊膜间充质干细胞的方法。方法:分别用酶消化法、传统组织块贴壁法和改进的组织块贴壁法分离出h AMSCs,观察3种方法所获得h AMSCs细胞形态、获得时间,流式细胞学检测其免疫表型;并用不同的诱导体系将h AMSCs向成骨细胞、成神经细胞进行诱导分化,鉴定其分化潜能。结果:改进的组织块贴壁法获得h AMSCs细胞的纯度更高,传代次数更多,相同的培养时间里,获得的细胞数量是酶消化法的3~4倍,是传统组织块贴壁法的1~2倍;h AMSCs细胞高表达CD73、CD90、CD44和CD105,不表达CD11b、CD19、CD34、CD45、HLA-DR;茜素红染色和神经元特异性烯醇化酶检测证实h AMSCs细胞可诱导分化为成骨细胞和神经细胞。结论:改进的h AMSCs分离培养方法可获得较大数量细胞,且保留了向神经细胞或成骨细胞可分化潜能。
Objective: To investigate and improve the method of human amnion mesenchymal stem cellsseparation and culture( h AMSCs). Methods: The h AMSCs were established with the methods of trypsin,collagenase II,traditional and improved tissue attachment,respectively. The cell morphology and the growth time of h AMSCs were observed,and the immunophenotype were also detected with flow cytometry. In addition,h AMSCs were induced to differentiate into osteoblast and neuroblast with different system and the derived osteoblast and neuroblast were identified then. Results: The h AMSCs obtained with the improved separation method obtain higher purity and can be passaged more times,and the quantity of obtained h AMSCs can be reached up to 3 ~ 4 times of enzyme digestion,and 1 ~ 2times of traditional tissue attachment method within the same culture time; furthermore,the obtained h AMSCs highly express CD73,CD90,CD44 and CD105 while did not express CD11 b,CD19,CD34,CD45 and HLA-DR; the alizarin staining and neuron-specific enolase detection demonstrated that h AMSCs could be induced into osteoblast and neuroblast. Conclusions: The methods of h AMSCs separation and culture are improved and will provide the basis for the research and application of h AMSC.

关键词(KeyWords)： 人羊膜间充质干细胞;细胞培养;鉴定;诱导
human amnion mesenchymal stem cells(hAMSCs);cell culture;identification;induce

Abstract:

Keywords:

基金项目(Foundation): 贵州省科技厅贵州医科大学联合基金[黔科合LH字(2014)7078]

作者(Author): 崔冬冰,范安然,何志旭
CUI Dongbing,FAN Anran,HE Zhixu

DOI: 10.19367/j.cnki.1000-2707.2016.04.014

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