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[2]	顾 霞, 赵 敏, 王平义, 李一梅, 李文华. 低氧诱导因子1α与低氧相关疾病信号通路的关系[J]. 中国组织工程研究, 2021, 25(8): 1284-1289.
[3]	侯婧瑛, 于萌蕾, 郭天柱, 龙会宝, 吴 浩. 缺氧预处理激活HIF-1α/MALAT1/VEGFA通路促进骨髓间充质干细胞生存和血管再生[J]. 中国组织工程研究, 2021, 25(7): 985-990.
[4]	史洋洋, 秦英飞, 吴福玲, 何 潇, 张雪静. 胎盘间充质干细胞预处理预防小鼠毛细支气管炎[J]. 中国组织工程研究, 2021, 25(7): 991-995.
[5]	梁学奇, 郭黎姣, 陈贺捷, 武 杰, 孙雅琪, 邢稚坤, 邹海亮, 陈雪玲, 吴向未. 泡状棘球绦虫原头蚴抑制骨髓间充质干细胞向成纤维细胞的分化[J]. 中国组织工程研究, 2021, 25(7): 996-1001.
[6]	樊全宝, 罗惠娜, 王丙云, 陈胜锋, 崔连旭, 江文康, 赵明明, 王静静, 罗冬章, 陈志胜, 白银山, 刘璨颖, 张 晖. 低氧培养犬脂肪间充质干细胞的生物学特性[J]. 中国组织工程研究, 2021, 25(7): 1002-1007.
[7]	耿 瑶, 尹志良, 李兴平, 肖东琴, 侯伟光. hsa-miRNA-223-3p调控人骨髓间充质干细胞成骨分化的作用[J]. 中国组织工程研究, 2021, 25(7): 1008-1013.
[8]	伦志刚, 金 晶, 王添艳, 李爱民. 过氧化物还原酶6干预骨髓间充质干细胞增殖及体外向神经谱系诱导分化[J]. 中国组织工程研究, 2021, 25(7): 1014-1018.
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Mesenchymal stem cells (MSCs) have been considered as a good source of stem cells^[1-5] due to tremendous self-renewal and multi-lineage differentiation potential. Some curative effects have been made in animal experiments using MSCs to treat ischemic brain injury^[6]. Buzanska et al^[7-8] reported that umbilical cord blood-derived MSCs (UCBMSCs) can be induced to express the marker of neurons and glial cells. But the methods used to regulate the in vitro differentiation of MSCs into high proportion of nerve-like cells have been rarely reported. This study observed the induced differentiation of human UCBMSCs into nerve-like cells using different concentrations of brain derived neurotrophic factor (BDNF) combined with ciliary neurotrophic factor (CNTF) and further investigated the methods and mechanisms underlying the induced differentiation of UCBMSCs, hopefully providing experimental evidence and theoretical acknowledge for repair of central nervous system injury.

This study observed the effects and efficiency of BDNF, CNTF or their combination in induced differentiation of UCBMSCs into nerve-like cells. Immunohistochemistry of NSE and GFAP molecular markers^[10] revealed that under the condition of in vitro culture, UCBMSCs could differentiate into nerve-like cells^[11] and confirmed the feasibility of acquiring high proportion of neuronal-like cells in vitro. Results from this study demonstrated that BDNF or CNTF could induce UCBMSCs to differentiate into nerve-like cells^[12] and produced similar effects in promoting cell differentiation; BDNF combined with CNTF could acquire high proportion of nerve-like cells, and more than 80% of cells expressed NSE and GFAP markers. After 3 days of induction by BDNF, CNTF or their combination, nerve-like cells formed, as confirmed by NSE (a specific marker for neuron) and GFAP (a specific marker for glial cell) expression. Such cells appeared in the sporadic manner initially and in a gradual increasing tendency thereafter. These findings indicate that this protocol is feasible and efficient, and that to enhance the differentiation proportion of nerve-like cells, some improvements should be made in induction regulation in vitro. Nerve growth factors in the culture of cells and tissues have been confirmed to be able to promote neuronal survival and differentiation and regulate axonal and dendrite growth^[13]. Some scholars used epidermal growth factors, retinoic acid, and BDNF to induce bone marrow-derived MSCs and found that MSCs could differentiate into neuronal-like cells and differentiated cells expressed nestin, as confirmed by immunohistochemistry. Based on this, this study used BDNF combined with CNTF to induce UCBMSCs and acquired high proportion of neuronal-like cells and immunohistochemistry confirmed that the differentiated cells showed nerve-like morphology and NSE and GFAP expression. Quirici et al^[15] reported that there was low-affinity nerve growth factor receptor on the surface of MSCs. All growth factor receptors have tyrosinase activity. Once growth factors bind to correspondent receptors, a series of protein kinases would cause the phosphorylation of many protein substrates, and lead to immediate early gene transcription, which causes gene duplication, transcription and finally cell proliferation and differentiation^[16].  BDNF is a basic protein harvested from brain tissue extract, with a relative molecular mass of 12 000. It is primarily distributed in the central nervous system, such as cerebral cortex, medulla, cerebellum, and hippocampus, and in the peripheral heart, lung, skeletal muscle, and sciatic nerve^[17-18]. It provides nutrition to neurons in vivo and in vitro, maintains neuronal survival, promotes axonal elongation, and plays an important role in regulating synapse functions in the hippocampus and visual cortex. BDNF has been considered feasible in treating neurodegenerative diseases due to its role in nervous system development, adult nerve plasticity, and nerve structure integrity^[19-20].  BDNF binding to Trk extracellular region activates tyrosine kinase, produces two important second messengers IP3 and DG, which induce Ca²⁺ release and activate protein kinase C, respectively^[21-23]. CNTF is initially extracted from chicken ciliary body and named by its role in maintaining the survival of chicken parasympathetic ganglions^[24-25]. CNTF possesses multiple functions and can promote the survival of various nerve cells. It is the first neurotrophic factor found to maintain the survival of spinal cord motor neurons and the growth of processes in vivo and in vitro and plays an extremely important role in nervous system development, differentiation and nerve injury repair^[26-27]. CNTF also can induce neuronal differentiation, inhibit sympathetoblast proliferation and promote its differentiation^[28]. Following central nerve system injury, rapidly synthesized CNTF is released into extracellular space and provides nutrition to neurons and glial cells in the injured region, so CNTF is considered a repair factor for central nervous system injury^[29-30]. Results from this study also suggest that BDNF combined with CNTF can produce a synergic effect on differentiation of UCBMSCs and yields a higher proportion of neuronal-like cells than BDNF or CNTF induction.    To conclude, under the condition of in vitro culture, UCBMSCs can differentiate into nerve-like cells and high proportion of neuronal-like cells can be acquired.

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