2025年4月24日 周四
首页 > 按月查看>2022年第10月 >890-895. DOI:10.20039/j.cnki.1007-3949.2022.10.009
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基于胞质分裂的增殖心肌细胞探针检测方法的建立
作者:
作者单位:

(1.上海大学附属南通医院(南通市第六人民医院) 上海大学老年医学研究院,江苏省南通市 226011;2.上海大学上海器官修复工程技术研究中心 上海大学心血管研究所,上海市 200444)

作者简介:

王静,博士,副教授,硕士研究生导师,研究方向为运动保护心脏的分子机制,E-mail:han_x_i_a_o@163.com。通信作者肖俊杰,博士,教授,博士研究生导师,研究方向为运动保护心脏的分子机制,E-mail:junjiexiao@shu.edu.cn。

基金项目:

国家重点研发计划政府间国际科技创新合作重点专项(2018YFE0113500)；上海市科学技术委员会上海器官修复工程技术研究中心(20DZ2255400)；国家自然科学基金重点国际(地区)合作研究项目(82020108002)；国家自然科学基金青年基金项目(81900250)


The establishment of a probe method for the detection of proliferating cardiomyocytes based on mitosis with complete cytokinesis
Author:
  • WANG Jing 1,2

    WANG Jing

    Institute of Geriatrics of Shanghai University ＆ Affiliated Nantong Hospital of Shanghai University the Sixth Peoples Hospital of Nantong, Nantong, Jiangsu 226011, China;Institute of Cardiovascular Sciences ＆ Shanghai Engineering Research Center of Organ Repair, Shanghai University, Shanghai 200444, China
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  • MENG Xinxiu 1,2

    MENG Xinxiu

    Institute of Geriatrics of Shanghai University ＆ Affiliated Nantong Hospital of Shanghai University the Sixth Peoples Hospital of Nantong, Nantong, Jiangsu 226011, China;Institute of Cardiovascular Sciences ＆ Shanghai Engineering Research Center of Organ Repair, Shanghai University, Shanghai 200444, China
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  • LIU Shuqin 1,2

    LIU Shuqin

    Institute of Geriatrics of Shanghai University ＆ Affiliated Nantong Hospital of Shanghai University the Sixth Peoples Hospital of Nantong, Nantong, Jiangsu 226011, China;Institute of Cardiovascular Sciences ＆ Shanghai Engineering Research Center of Organ Repair, Shanghai University, Shanghai 200444, China
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  • ZHAO Xuan 1,2

    ZHAO Xuan

    Institute of Geriatrics of Shanghai University ＆ Affiliated Nantong Hospital of Shanghai University the Sixth Peoples Hospital of Nantong, Nantong, Jiangsu 226011, China;Institute of Cardiovascular Sciences ＆ Shanghai Engineering Research Center of Organ Repair, Shanghai University, Shanghai 200444, China
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  • XIAO Junjie 1,2

    XIAO Junjie

    Institute of Geriatrics of Shanghai University ＆ Affiliated Nantong Hospital of Shanghai University the Sixth Peoples Hospital of Nantong, Nantong, Jiangsu 226011, China;Institute of Cardiovascular Sciences ＆ Shanghai Engineering Research Center of Organ Repair, Shanghai University, Shanghai 200444, China
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Affiliation:

1.Institute of Geriatrics of Shanghai University ＆ Affiliated Nantong Hospital of Shanghai University (the Sixth Peoples Hospital of Nantong), Nantong, Jiangsu 226011, China;2.Institute of Cardiovascular Sciences ＆ Shanghai Engineering Research Center of Organ Repair, Shanghai University, Shanghai 200444, China)

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    摘要:

    目的]基于双荧光探针方法,探究全新的在活细胞水平检测心肌细胞增殖的实验方法,该方法能排除多核心肌细胞核复制带来的假阳性细胞,为精准探测心肌细胞增殖提供实验参考。 [方法]以细胞内细胞分裂周期蛋白20(CDC20)和痉挛性截瘫蛋白20(SPG20)作为探针靶点,构建分子探针,采用脂质体转染试剂2000(Lipo-2000)作为探针载体,将探针转染到细胞中,在活细胞水平检测CDC20和SPG20双阳性的细胞,即为发生胞质分裂的增殖细胞。 [结果]细胞不发生增殖时,荧光探针无法检测到目的序列,细胞不发荧光；细胞核复制分裂但不发生胞质分裂时,两个荧光探针不能同时检测到序列,细胞不同时发两种荧光；细胞发生胞质分裂的增殖时,细胞同时发两种荧光。 [结论]使用CDC20/SPG20双荧光探针方法,能够准确检测发生胞质分裂的增殖的心肌细胞,从而排除因核增殖产生的细胞增殖假阳性问题,使细胞增殖的检测精准度更高。

    Abstract:

    Aim To explore a novel experimental method for detecting cardiomyocyte proliferation at the live cell level, and to exclude false positive cells caused by multinucleate cardiomyocytes, so as to provide experimental reference for accurate detection of cardiomyocytes proliferation. Methods Use the cell division cyclin 20 (CDC20) and spastic paraplegia 20 (SPG20) as the molecular beacons targets. These molecular beacons allow for CDC20 and SPG20 detection in live cells, through Lipo-2000 transfection. The double positive cells are the proliferating ones that undergo cytokinesis. Results When the cells did not proliferate, the probes could not detect the target sequences and the cells cannot be detected by the fluorescence. When the nuclei replicated and divided without cytokinesis, the sequences could not be detected by both two probes at the same time, and the cells also could not be detected. When cells underwent mitosis with complete cytokinesis proliferation, both two fluorescence probes could be detected simultaneously. Conclusion The molecular beacons-based approach targeting CDC20 and SPG20 identified a proliferative subset of cardiomyocytes, which could exclude false positive problems of cell proliferation caused by nuclear proliferation and improve the accuracy of cell proliferation detection.

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引用本文

王静,孟新秀,刘书琴,赵璇,肖俊杰.基于胞质分裂的增殖心肌细胞探针检测方法的建立[J].中国动脉硬化杂志,2022,30(10):890~895.

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  • 收稿日期:2021-11-24
  • 最后修改日期:2022-05-02
  • 在线发布日期: 2022-10-09

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