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骨髓间充质干细胞培养及向神经细胞诱导分化的实验研究
引用本文:朱光荣,周小玉,周建伟,吴雨洁,汪承亚.骨髓间充质干细胞培养及向神经细胞诱导分化的实验研究[J].南京医科大学学报,2003,23(2):100-102,F002.
作者姓名:朱光荣  周小玉  周建伟  吴雨洁  汪承亚
作者单位:[1]南京医科大学第一附属医院血液科,江苏南京210029 [2]南京医科大学公共卫生学院分子毒理室,江苏南京210029
基金项目:江苏省卫生厅重点课题基金资助项目 (H20013),南京医科大学创新基金资助项目(CX2001004)
摘    要:目的:体外培养骨髓间充质干细胞(BM—MSC),诱导BM—MSC向神经细胞分化。方法:采用DMEM培养液加胎牛血清(DMEM/FBS)或无血清培养基体外培养人BM—MSC,测定细胞倍增时间;免疫组织化学法分析BM—MSC的表面标记;纤维母细胞集落形成(CFU—F)试验检测BM—MSC增殖能力;应用二甲基亚砜/羟丁苯甲酯(DMS0/BHA)化学因子诱导BM—MSC向神经细胞分化。结果:BM—MSC为单层贴壁生长,呈现规则的集束辐射状排列。在对数生长期,细胞倍增时间约为46h。CFU—F试验表明,体外培养的MSC的集落形成能力为正常骨髓单个核细胞(MNC)的15000倍以上。免疫组化分析表明,BM—MSC为CDl3和CDl05阳性,CD34阴性。DMS0/BHA化学因子能诱导BM—MSC分化为神经细胞,但是分化后即失去增殖能力,并于48h后逐渐死亡。结论:通过体外培养可以获得大量高度均一的BM—MSC。BM—MSC可向神经细胞诱导分化,但尚缺乏实际应用的可能性。

关 键 词:骨髓间充质干细胞  细胞培养  诱导分化  神经细胞  干细胞移植
文章编号:1007-4368(2003)02-0100-03

BM-MSC Culture and Induction into Neurons
ZHU Guang-rong,ZHOU Xiao-yu,ZHOU Jian-wei,WU Yu-jie,WANG Cheng-ya.BM-MSC Culture and Induction into Neurons[J].Acta Universitatis Medicinalis Nanjing,2003,23(2):100-102,F002.
Authors:ZHU Guang-rong  ZHOU Xiao-yu  ZHOU Jian-wei  WU Yu-jie  WANG Cheng-ya
Abstract:Objective: To culture bone marrow mesenchymal stem cells (BM-MSC), in vitro, and to induce them into neurons. Methods: Human BM-MSC from healthy donors and primary diagnosed patients with hematological diseases were cultured in DMEM/FBS or MesenCult serum free media. BM-MSC doubling time was determined at exponential stage. CD13, CD105 and CD34 of cell surface markers were tested with histoimmunochemistry assay. Colony forming potent of the expanded BM-MSC was analyzed by CFU-F test. Conditioned media containing BHA/DMSO was applied for induction of BM-MSC into neurons. Results: BM-MSCs grew into monolayer cells attached on the bottom of culture flask. The cultured cells arranged regularly with a uniform radiation. At the exponential stage, cell doubling time was around 46 h in serum free culture system. BM-MSC was highly potential to proliferate. The colony-forming ability of the cultured MSC was 15 000 folds higher than bone marrow mononuclear cell(MNC) in CFU-F assay. Histoimmunochemistry analysis indicated that all the cultured BM-MSCs were CD13( + ), CD105( + ) and CD34( - ) . After incubation of BM-MSC in BHA/DMSO conditioned media, neurons were developed, which was identified by cell morphology and detecting of neurofilament with histoimmuno-chemical assay. However, the induced neurons lost their proliferation ability and were dying after 48h of induction. Conclusion: BM-MSC is a sort of stem cell of bone marrow different from HSC. A great number of MSC with high purity could be obtained by culture in vitro. Although induction of neurons from BM-MSC by DMSO/BHA was achieved in vitro, there is very little practical significance at present yet.
Keywords:bone marrow mesenchymal stem cell  cell culture  cell differentiation
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