中国组织工程研究

中国组织工程研究 ›› 2018, Vol. 22 ›› Issue (17): 2692-2698.doi: 10.3969/j.issn.2095-4344.0482

• 脐带脐血干细胞 umbilical cord blood stem cells • 上一篇    下一篇

鼠神经生长因子联合脑源性神经营养因子诱导人脐血间充质干细胞向神经元样细胞分化

陈 俊，杨自金，李红梅   

  1. 徐州医科大学附属连云港医院(连云港市第一人民医院)儿内科，江苏省连云港市 222002
  • 修回日期:2017-11-25 出版日期:2018-06-18 发布日期:2018-06-18
  • 通讯作者: 杨自金，主任医师，硕士生导师，徐州医科大学附属连云港医院(连云港市第一人民医院)儿内科，江苏省连云港市 222002
  • 作者简介:陈俊，男，1991年生，湖北省随州市人，汉族，徐州医科大学在读硕士，主要从事脐血间充质干细胞相关研究。
  • 基金资助:

    江苏省连云港市科技局重点科室建设(SH1117)

Human umbilical cord blood-derived mesenchymal stem cells differentiate into neuron-like cells induced by combination of mouse nerve growth factor and brain-derived neurotrophic factor

Chen Jun, Yang Zi-jin, Li Hong-mei   

  1. Department of Pediatrics, Lianyungang Hospital Affiliated to Xuzhou Medical University (First People’s Hospital of Lianyungang), Lianyungang 222002, Jiangsu Province, China
  • Revised:2017-11-25 Online:2018-06-18 Published:2018-06-18
  • Contact: Yang Zi-jin, Chief physician, Master’s supervisor, Department of Pediatrics, Lianyungang Hospital Affiliated to Xuzhou Medical University (First People’s Hospital of Lianyungang), Lianyungang 222002, Jiangsu Province, China
  • About author:Chen Jun, Master candidate, Department of Pediatrics, Lianyungang Hospital Affiliated to Xuzhou Medical University (First People’s Hospital of Lianyungang), Lianyungang 222002, Jiangsu Province, China
  • Supported by:

    the Key Division Construction of Lianyungang Municipal Science and Technology Department, No. SH1117

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摘要：

文章快速阅读：

文题释义：
鼠神经生长因子：是从小鼠颌下腺分离纯化出的一种细胞因子，能够促进神经系统损伤后的修复，对中枢及周围神经元的生长、分化、再生和功能特性的表达均具有重要的调控作用。
脑源性神经营养因子：是1982年Barde等首先在猪脑中发现的一种具有神经营养作用的蛋白质。脑源性神经营养因子及其受体在神经系统广泛表达，其中海马和皮质的含量最高。

 

摘要
背景：作者在前期研究中发现鼠神经生长因子具备诱导脐血间充质干细胞向神经元样细胞分化的能力，为进一步探讨提高诱导效率的方法，拟尝试使用多种细胞生长因子进行联合诱导。
目的：探讨鼠神经生长因子联合脑源性神经营养因子在体外条件下对人脐血间充质干细胞向神经元样细胞诱导分化的作用。
方法：复苏获赠的原代人脐血间充质干细胞并进行体外扩增培养，取第5代脐血间充质干细胞分为5组，其中前4组为预诱导组，在加入成纤维生长因子和表皮生长因子预诱导24 h后，再用鼠神经生长因子、脑源性神经营养因子及二者联合对脐血间充质干细胞进行诱导，对照组仅使用等量细胞培养基培养；最后一组为非预诱导组，先用细胞培养基培养24 h后，再加入鼠神经生长因子和脑源性神经营养因子进行联合诱导。倒置显微镜下观察诱导后细胞形态变化，并用免疫细胞化学法检测神经元特异性烯醇化酶、胶质纤维酸性蛋白的表达，实时荧光定量PCR技术检测神经元特异性烯醇化酶、胶质纤维酸性蛋白在mRNA水平的相对表达量。
结果与结论：①脐血间充质干细胞形态为大小不一的长梭形、纺锤形，经诱导后细胞胞体逐渐回缩变圆，并向一侧或多侧发出突起，呈现出神经元样细胞改变；②免疫细胞化学及实时荧光定量PCR结果显示各实验组细胞均阳性表达神经元特异性烯醇化酶、胶质纤维酸性蛋白，联合诱导组神经元特异性烯醇化酶、胶质纤维酸性蛋白阳性率及二者mRNA表达量均高于单独诱导组、对照组以及非预诱导组；③鼠神经生长因子和脑源性神经营养因子均能诱导脐血间充质干细胞向神经元样细胞分化，二者在诱导过程中具有协同作用，联合使用能够有效提高诱导效率。

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程
ORCID: 0000-0002-3762-1586(陈俊)

关键词: 干细胞, 脐血, 间充质干细胞, 鼠神经生长因子, 脑源性神经营养因子, 神经元样细胞, 诱导分化

Abstract:

BACKGROUND: We have found that mouse nerve growth factor has the ability to induce differentiation of umbilical cord blood mesenchymal stem cells (UCB-MSCs) into neurons in vitro. In order to further explore the method of improving the induction efficiency of nerve cells, we attempt to combine a variety of cell growth factors for cell induction.
OBJECTIVE: To explore the effect of mouse nerve growth factor combined with brain-derived neurotrophic factor on the differentiation of UCB-MSCs into neuron-like cells in vitro.
METHODS: After the donated primary UCB-MSCs were resuscitated and cultured, the passage 5 UCB-MSCs were divided into five groups. The first four groups served as pre-induced groups, and fibroblast growth factor and epidermal growth factor were added to pre-induce cells for 24 hours, and mouse nerve growth factor and brain-derived neurotrophic factor, alone or in combination, were used thereafter to induce UCB-MSCs, while in control group, only the same amount of cell medium was added. The last group was non-pre-induced group, in which the cells were cultured in the cell culture medium for 24 hours, and then mouse nerve growth factor and brain-derived neurotrophic factor were both added to induce UCB-MSCs. The morphological changes of cells were observed under inverted microscope. The expression of neuron-specific enolase (NSE) and glial fibrillary acidic protein (GFAP) was detected by immunocytochemistry technique. Real-time qPCR was used to detect the relative expression of NSE and GFAP at mRNA level.
RESULTS AND CONCLUSION: (1) The cell morphology of UCB-MSCs was in long shuttle shape and spindle shape with unequal size. After induction, the cell bodies gradually retracted and became rounded, and the projections extended to one-side or multi-sides, presenting with the neuron-like changes. (2) Immunocytochemistry and real-time qPCR results showed that NSE and GFAP were positive in each experimental group, and the positive rate and mRNA expression of NSE and GFAP in the combined induction group were higher than those in the other groups. (3) Either mouse nerve growth factor or brain-derived neurotrophic factor could induce UCB-MSCs to differentiate into neuron-like cells. Moreover, there was a cumulative effect between the two cytokines, and their combined use could effectively improve the efficiency of induction.

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

Key words: Fetal Blood, Mesenchymal Stem Cells, Nerve Growth Factor, Brain-Derived Neurotrophic Factor, Cell Differentiation, Neurons, Tissue Engineering

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