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胡景伟,蔡 哲,周忠蜀,张 岚,舒 峻,潘 琳,黄小杰,徐 波,向 青,贺 春,徐 杨,高 艳,崔晓慧.体外培养人羊膜间充质细胞的神经生物学特点[J].中国康复医学杂志,2009,24(12):1070~1073
体外培养人羊膜间充质细胞的神经生物学特点    点此下载全文
胡景伟  蔡 哲  周忠蜀  张 岚  舒 峻  潘 琳  黄小杰  徐 波  向 青  贺 春  徐 杨  高 艳  崔晓慧
北京协和医学院研究生院,北京,100730
基金项目:国家自然科学基金(30471773);北京市科委重大研究课题(D07050701350704);国家863重大专项研究课题(2006AA02A114)
DOI:
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摘要:
      目的:探讨体外培养人羊膜间充质细胞(hAMCs)的神经生物学特征。方法:应用MTS分析法、BrdU掺入法和增殖细胞核抗原(PCNA)的免疫荧光染色,探讨体外培养hAMCs的增殖活性。利用免疫细胞化学法检测hAMCs 中间充质细胞标记(STRO-1、Vimentin)、神经干细胞标记蛋白(Nestin、PSA-NCAM)、神经细胞标记蛋白(β-tubulin-Ⅲ、TH)和神经分化相关蛋白(math-1、mash-1)的表达。结果: MTS分析法显示hAMCs在接种后第6—8天增殖速度最快,第8—24天活细胞数基本保持稳定;BrdU和PCNA的免疫荧光染色结果显示体外培养hAMCs中具有大量BrdU和PCNA阳性细胞存在;体外培养的hAMCs表达间充质干细胞特异性标记物STRO-1和Vimentin,也表达神经干细胞标记物Nestin和PSA-NCAM,同时还可见神经元特异性标记物β-tubulin-Ⅲ和TH,以及神经元分化相关蛋白math-1和mash-1的表达;体外培养hAMCs中存在有TH/BrdU和β-tubulin-Ⅲ/BrdU双阳性细胞。结论:体外培养的hAMCs表达间充质干细胞、神经干细胞、神经元的特异性标记蛋白,同时具有增殖活性;hAMCs表达神经元分化相关蛋白,提示hAMCs具有向神经元分化的潜能。
关键词:人羊膜间充质细胞  神经细胞  增殖
The neurobiology characteristics of human amnion mesenchymal cells culture in vitro    Download Fulltext
Graduate School of Beijing Union Medical College, Beijing, 100730
Fund Project:
Abstract:
      Objective: To evaluate the neurobiological features of human amnion mesenchymal cells (hAMCs) in vitro. Method: CellTiter 96?誖AQueous One Solution Cell Proliferation Assay (MTS) , BrdU incorporation and detection of proliferating cell nuclear antigen (PCNA) were applied to evaluate the self-proliferation ability of hAMCs. Immunocytochemistry was used to detect the expressions of markers characteristic of mesenchymal stem cell (STRO-1 and Vimentin), neural stem cells (Nestin and PSA-NCAM), neural cells (β-tubulin-Ⅲ and TH), and the proteins related to neural differentiation (math-1, mash-1). Result: ①MTS analysis showed that from the 6th d to the 8th d the amount of live hAMCs grew fast; from the 8th d to the 24th d, the amount of live hAMCs was relatively stable. Immunofluorescence demonstrated some of hAMCs were positively stained by anti-BrdU and anti-PCNA. ②Cultured hAMCs were able to express mesenchymal stem cell markers STRO-1 and Vimentin, neural stem cell markers Nestin and PSA-NCAM, neuron markers β-tubulin-Ⅲ and TH and neural differentiation related protein math-1 and mash-1. ③Some of hAMCs coexpressed TH/BrdU and β-tubulin-Ⅲ/BrdU. Conclusion: In vitro, hAMCs not only express phenotype of mesenchymal stem cells, neural stem cells and neurons, but also have the capacity of self-proliferation; hAMCs express the proteins related to neural differentiation and have the potential to differentiate into neural-like cells.
Keywords:human amnion mesenchymal cells  neural cells  proliferation
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