上海交通大学学报(医学版) ›› 2019, Vol. 39 ›› Issue (9): 969-.doi: 10.3969/j.issn.1674-8115.2019.09.006

• 论著·基础研究 • 上一篇    下一篇

基于胆固醇-DNA的活细胞膜修饰研究

刘江波 1, 2,王丽华 1,左小磊 3   

  1. 1. 中国科学院上海应用物理研究所物理生物学研究室,上海光源生物成像中心,上海 201800;2. 中国科学院大学,北京 100049;3. 上海交通大学医学院分子医学研究院,上海交通大学医学院附属仁济医院,上海 200127
  • 出版日期:2019-09-28 发布日期:2019-11-02
  • 通讯作者: 左小磊,电子信箱:zuoxiaolei@sjtu.edu.cn。
  • 作者简介:刘江波(1992—),男,土家族,博士生;电子信箱: ljbsinap@163.com。
  • 基金资助:
    国家自然科学基金面上项目(31470960);上海市教育委员会高峰高原学科建设计划(20171913)

Research of live cell membranes modification based on cholesterol-DNA

LIU Jiang-bo1, 2, WANG Li-hua1, ZUO Xiao-lei3   

  1. 1. Bio-imaging Center, Shanghai Synchrotron Radiation Facility, Laboratory of Physical Biology, Shanghai Institute of Applied Physics, Chinese Academy of Sciences, Shanghai 201800, China; 2. University of Chinese Academy of Sciences, Beijing 100049, China; 3. Institute of Molecular Medicine, Renji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200127, China
  • Online:2019-09-28 Published:2019-11-02
  • Supported by:
    General Program of National Natural Science Foundation of China, 31470960; Shanghai Municipal Education Commission—Gaofeng Clinical Medicine Support, 20171913)。

摘要: 目的 ·探讨连接胆固醇的脱氧核糖核酸 (deoxyribonucleic acid,DNA)对活细胞膜的修饰作用。方法 ·以悬浮细胞 L1210和贴壁细胞 PC-12为研究对象,将每种细胞分为实验组(胆固醇 -DNA孵育)和对照组(磷酸盐缓冲液处理)。通过共聚焦显微镜获得 2组细胞的荧光强度,并对实验组细胞进行三维重构。采用扫描电子显微镜观察实验组细胞的形态。采用荧光漂白恢复实验检测胆固醇 -DNA修饰对细胞膜流动性的影响。结果 ·悬浮细胞和贴壁细胞获得了一致的结果。与对照组相比,实验组细胞表面的荧光强度有所加强(均 P0.000)。共聚焦显微镜的三维重构显示,实验组细胞的荧光分布在整个细胞表面。扫描电子显微镜结果显示,实验组细胞的形态经胆固醇 -DNA修饰后并未改变。经荧光漂白处理后,实验组 L1210细胞的相对荧光强度下降至 0.090,且在 110 s内恢复至 0.860。结论 ·胆固醇 -DNA能够对整个活细胞膜进行修饰,且修饰后的细胞膜仍具有流动性。该方法不仅能够用于悬浮细胞,还可应用于贴壁细胞。

关键词: 胆固醇, 脱氧核糖核酸, 细胞膜, 修饰

Abstract: Objective · To investigate the modification of live cell membranes with cholesterol linked deoxyribonucleic acid (cholesterol-DNA). Methods · The suspension L1210 cells and adherent PC-12 cells were included. L1210 cells and PC-12 cells were divided into the experimental group (incubated with cholesterol-DNA) and the control group (treated with phosphate buffer saline), respectively. The fluorescence intensity of the cells in the two groups was obtained and the experimental group cells were three-dimensional reconstructedconfocal microscopy. The morphology of the cells in the experimental group was observedscanning electron microscopy (SEM). The effect of cholesterol-DNA modification on cell membrane fluidity was detectedfluorescence recovery after photobleaching. Results · The results of suspension cells and adherent cells were consistent. Compared with the control group, the fluorescence intensity of cell surface in the experimental group was increased (both P0.000). Three-dimensional reconstruction of confocal microscopy showed that the fluorescence of the cells in the experimental group was distributed across the surface of the global cell. SEM showed that the morphology of the cells in the experimental group did not change with cholesterol-DNA modification. After fluorescence photobleaching, the relative fluorescence intensity of the L1210 cells in the experimental group was decreased to 0.090, and then recovered to 0.860 within 110 s. Conclusion · Cholesterol-DNA can modify the whole live cell membranes, and the modified cell membranes still have fluidity. This method can modify not only the suspension cells, but also the adherent cells.

Key words: cholesterol, deoxyribonucleic acid (DNA), cell membrane, modification

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