中国神经再生研究(英文版)

中国神经再生研究(英文版) ›› 2014, Vol. 9 ›› Issue (22): 1968-1978.doi: 10.4103/1673-5374.145378

• 原著:周围神经损伤修复保护与再生 • 上一篇    下一篇

胎牛皮脱细胞真皮基质上间充质干细胞可形成神经组织工程支架

  

  • 收稿日期:2014-10-15 出版日期:2014-11-25 发布日期:2014-11-25

Differentiation of mesenchymal stem cells into neuronal cells on fetal bovine acellular dermal matrix as a tissue engineered nerve scaffold

Yuping Feng 1, 2, Jiao Wang 3, Shixin Ling 2, Zhuo Li 2, Mingsheng Li 2, Qiongyi Li 2, Zongren Ma 2, Sijiu Yu 1   

  1. 1 Animal Medicine College of Gansu Agriculture University, Lanzhou, Gansu Province, China
    2 Gansu Provincial Animal Cell Engineering Center; Key Laboratory of Bioengineering & Technology of State Ethnic Affairs Commission, Life
    Science and Engineering College of Northwest University for Nationalities, Lanzhou, Gansu Province, China
    3 Laboratory of Molecular Neurobiology, Institute of Systems Biology, Shanghai University, Shanghai, China
  • Received:2014-10-15 Online:2014-11-25 Published:2014-11-25
  • Contact: Sijiu Yu, Ph.D., Animal Medicine College of Gansu Agriculture University, Lanzhou 730070, Gansu Province, China, sjyu@163.com.
  • Supported by:

    This research was supported by a grant from Construction Project of Gansu Provincial Animal Cell Engineering Center, No. 0808NTGA013 and Program for Innovative Research Team in University of Ministry of Education of China, No. IRT13091.

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摘要：

实验旨在验证课题组提出的在胎牛皮脱细胞真皮基质支架上，人骨髓间充质干细胞向神经元样细胞分化及并能够构建组织工程神经。应用显微镜长期连续观察诱导分化细胞的形态、生长、增殖与分化现象，通过免疫组织化学和苏木精伊红染色和甲苯胺蓝染色技术，鉴定了分化细胞的神经元特异蛋白和尼氏小体，以确认骨髓间充质干细胞的神经元样分化。结果显示经神经分化培养基的诱导分化培养后，胎牛皮脱细胞真皮基质上骨髓间充质干细胞分化出具有单极、两级或多极神经突起的神经元特异形态，表达神经元特异的神经元胞体和神经突蛋白、beta-Ⅲ tubulin蛋白。长期诱导培养后分化细胞形成了纵横交错的多层神经网络样结构，高倍扫描电镜下可见长的神经纤维有多个微纤维构成，几个相邻的神经元形成了具有树突-树突、轴突-树突、树突-轴突突触联系的神经环路，并可见树突棘及带有丝状伪足的生长锥，可见尼氏小体。结果表明在胎牛皮脱细胞真皮基质上，骨髓间充质干细胞经神经分化诱导培养后可分化为具有神经元形态及功能的神经元，进而形成了组织工程化神经。

关键词: 神经再生, 周围神经损伤, 胎牛, 胎牛皮脱细胞真皮基质, 生物支架材料, 骨髓, 间充质干细胞, 神经分化, 神经元, 组织工程化神经

Abstract:

The purpose of this study was to assess fetal bovine acellular dermal matrix as a scaffold for supporting the differentiation of bone marrow mesenchymal stem cells into neural cells following induction with neural differentiation medium. We performed long-term, continuous observation of cell morphology, growth, differentiation, and neuronal development using several microscopy techniques in conjunction with immunohistochemistry. We examined specific neuronal proteins and Nissl bodies involved in the differentiation process in order to determine the neuronal differentiation of bone marrow mesenchymal stem cells. The results show that bone marrow mesenchymal stem cells that differentiate on fetal bovine acellular dermal matrix display neuronal morphology with unipolar and bi/multipolar neurite elongations that express neuronal-specific proteins, including βIII tubulin. The bone marrow mesenchymal stem cells grown on fetal bovine acellular dermal matrix and induced for long periods of time with neural differentiation medium differentiated into a multilayered neural network-like structure with long nerve fibers that was composed of several parallel microfibers and neuronal cells, forming a complete neural circuit with dendrite-dendrite to axon-dendrite to dendrite-axon synapses. In addition, growth cones with filopodia were observed using scanning electron microscopy. Paraffin sectioning showed differentiated bone marrow mesenchymal stem cells with the typical features of neuronal phenotype, such as a large, round nucleus and a cytoplasm full of Nissl bodies. The data suggest that the biological scaffold fetal bovine acellular dermal matrix is capable of supporting human bone marrow mesenchymal stem cell differentiation into functional neurons and the subsequent formation of tissue engineered nerve.

Key words: nerve regeneration, peripheral nerve defects, fetal bovine, acellular dermal matrix, biological scaffold, bone marrow mesenchymal stem cells, neuronal differentiation, neurons, tissue engineered nerve, neural regeneration