中国神经再生研究(英文版)

中国神经再生研究(英文版) ›› 2018, Vol. 13 ›› Issue (3): 497-501.doi: 10.4103/1673-5374.228734

• 原著:颅神经损伤修复保护与再生 • 上一篇    下一篇

体外培养前庭上皮细胞经Math1诱导分化为毛细胞过程中的细胞增殖

  

  • 收稿日期:2018-01-24 出版日期:2018-03-15 发布日期:2018-03-15
  • 基金资助:

     中国国家自然科学基金项目（81420108010, 81271084, 81370022, 81570920, 81000413, 81200740, 81200738, 81371093, 81400460, 81200739），973项目（2011CB504500，2011CB504506） ，上海市科委创新项目（11411952300），上海市卫生系统优秀人才培养项目（XYQ2013084）

Cell proliferation during hair cell regeneration induced by Math1 in vestibular epithelia in vitro

Yi-bo Huang1, 2, 3, Rui Ma1, 2, 3, Juan-mei Yang1, 2, 3, Zhao Han1, 2, 3, Ning Cong1, 2, 3, Zhen Gao1, 2, 3, Dongdong Ren1, 2 , 3, Jing Wang1, 2 , 3,Fang-lu Chi1, 2 , 3   

  1. 1 Department of Otology and Skull Base Surgery, EYE & ENT Hospital of Fudan University, Shanghai, China
    2 Shanghai Clinical Medical Center of Hearing Medicine, Shanghai, China
    3 Key Laboratory of Hearing Medicine, Ministry of Health, Shanghai, China
  • Received:2018-01-24 Online:2018-03-15 Published:2018-03-15
  • Contact: Fang-lu Chi, M.D., Ph.D.,chifanglu@126.com.
  • Supported by:

    The work was supported by the National Natural Science Foundation of China (NSFC), grant No. 81420108010, 81271084 to FLC,81370022, 81570920, 81000413 to DR, 81200740 to JMY, 81200738 to NC, 81371093 to ZH, 81400460 to ZG, 81200739 to JW; 973 Program,grant No. 2011CB504500 and 2011CB504506; The Innovation Project of Shanghai Municipal Science and Technology Commission, grant No. 11411952300 to FLC; and the Training Program of the Excellent Young Talents of the Shanghai Municipal Health System, grant No.XYQ2013084 to DR.

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摘要：

毛细胞再生是纠正因毛细胞损伤或缺失引起的听力损失和平衡障碍的根本方法。而如何促进毛细胞再生是当今研究的热点。哺乳动物耳蜗毛细胞不能再生，只有极少量前庭毛细胞可以自发再生。然而经Math1基因转染一些内耳细胞可以在体外或体内分化为毛细胞。而鸟类毛细胞可以通过支持细胞分化和转分化方式再生，且这一过程与细胞增殖有关。因此实验拟对哺乳动物中Math1诱导毛细胞再生和细胞分裂的关系进行分析。实验分离了小鼠大脑前庭，获得前庭上皮细胞，并经Ad-Math1-EGFP转染，为追踪毛细胞转化过程中的细胞分裂，在培养不同时间点加入BrdU来跟踪细胞增殖情况，通过免疫组化检测细胞分化与增殖结果。结果发现，当上皮细胞处于高增殖时期时，这些细胞可以通过Math1基因转染转化为毛细胞。而处于低增殖时期的细胞经Math1基因诱导后，未见BrdU阳性毛细胞。而在Ad-Math1-EGFP转染前和转染后对细胞进行BrdU标记，得出细胞分裂仅发生在Math1转染前，而Math1诱导的新毛细胞再生过程中及之后很少发生细胞分裂。结果说明高增殖能力的前庭上皮细胞可通过Math1基因转染分化为新的毛细胞，但这一过程与细胞增殖无关。

orcid:0000-0002-3243-0393(Fang-lu Chi)

关键词: 神经再生, 细胞增殖, 细胞分裂, Math1, 毛细胞, 细胞分化, 跨细胞分化, 前庭细胞

Abstract:

Hair cell regeneration is the fundamental method of correcting hearing loss and balance disorders caused by hair cell damage or loss. How to promote hair cell regeneration is a hot focus in current research. In mammals, cochlear hair cells cannot be regenerated and few vestibular hair cells can be renewed through spontaneous regeneration. However, Math1 gene transfer allows a few inner ear cells to be transformed into hair cells in vitro or in vivo. Hair cells can be renewed through two possible means in birds: supporting cell differentiation and transdifferentiation with or without cell division. Hair cell regeneration is strongly associated with cell proliferation. Therefore, this study explored the relationship between Math1-induced vestibular hair cell regeneration and cell division in mammals. The mouse vestibule was isolated to harvest vestibular epithelial cells. Ad-Math1-enhanced green fluorescent protein (EGFP) was used to track cell division during hair cell transformation.5-Bromo-2′-deoxyuridine (BrdU) was added to track cell proliferation at various time points. Immunocytochemistry was utilized to determine cell differentiation and proliferation. Results demonstrated that when epithelial cells were in a higher proliferative stage, more of these cells differentiated into hair cells by Math1 gene transfer. However, in the low proliferation stage, no BrdU-positive cells were seen after Math1 gene transfer. Cell division always occurred before Math1 transfection but not during or after Math1 transfection, when cells were labeled with BrdU before and after Ad-Math1-EGFP transfection. These results confirm that vestibular epithelial cells with high proliferative potential can differentiate into new hair cells by Math1 gene transfer, but this process is independent of cell proliferation.

Key words: nerve regeneration, cell proliferation, cell division, Math1, hair cells, hair cell renewal, supporting cell differentiation, vestibular cells, neural regeneration