上海交通大学学报（医学版）

›› 2013, Vol. 33 ›› Issue (3): 263-.doi: 10.3969/j.issn.1674-8115.2013.03.002

• 论著（基础研究） • 上一篇    下一篇

重组杆状病毒载体介导钠碘转运体基因在胰岛细胞中表达的研究

刘 帅, 潘 昱, 张一帆   

  1. 上海交通大学 医学院附属瑞金医院核医学科, 上海 200025
  • 出版日期:2013-03-28 发布日期:2013-03-29
  • 通讯作者: 张一帆, 电子信箱: zhangyifan1992@yahoo.com.cn。
  • 作者简介:刘 帅（1987—）, 女, 硕士； 电子信箱: elaine_liu87@yahoo.com.cn。
  • 基金资助:

    国家自然科学基金(81171367, 30570525)；上海市教委科研创新项目(12YZ041)

Expression of recombinant baculovirus vector-mediated sodium iodine symporter gene in islet cells

LIU Shuai, PAN Yu, ZHANG Yi-fan   

  1. Department of Nuclear Medicine, Ruijin Hospital,Shanghai Jiaotong University School of Medicine, Shanghai 200025, China
  • Online:2013-03-28 Published:2013-03-29
  • Supported by:

    National Natural Science Foundation of China, 81171367, 30570525； Shanghai Education Committee Foundation, 12YZ041

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摘要：

目的 探讨以杆状病毒为载体，钠碘同向转运体（NIS） 基因作为报告基因监测胰岛细胞的可行性。方法 通过Bac-to-Bac杆状病毒表达系统制备绿色荧光蛋白（GFP）和NIS基因重组杆状病毒（Bac-GFP和Bac-NIS）。分别以不同感染复数（MOI）的Bac-GFP感染大鼠胰岛细胞，荧光显微镜观察胰岛细胞的感染百分比和荧光表达强度；以MOI为40的Bac-GFP感染胰岛细胞，荧光显微镜下观察荧光表达时间及细胞生存状态。以不同MOI的Bac-NIS感染胰岛细胞，测定感染细胞的碘摄取情况；以MOI为40的Bac-NIS感染胰岛细胞，动态观察感染细胞摄碘能力的变化以及高氯酸钠（NaClO4）对细胞摄碘的影响。结果 Bac-GFP可高效感染SD大鼠胰岛细胞，对细胞无毒性作用；感染的胰岛细胞百分比和荧光强度随MOI的提高而上升，细胞感染百分比可高达95%以上（MOI=40），且荧光表达时间可持续2周左右。Bac-NIS感染的胰岛细胞表现出摄碘功能增强和NaClO4抑制摄碘的特性,且感染细胞摄碘的放射性计数随MOI的提高而增加；MOI为40时，Bac-NIS感染细胞摄碘的放射性计数是空白对照的12倍。结论 杆状病毒介导的NIS 基因在胰岛细胞中高效表达，有效介导放射性碘摄取，可用对胰岛细胞移植情况的监测。

关键词: 杆状病毒, 钠碘同向转运体, 胰岛细胞, 报告基因

Abstract:

Objective To investigate the feasibility of monitoring islet cells with baculovirus as vector and sodium iodine symporter (NIS) gene as report gene. Methods Green fluorescent protein (GFP) and NIS gene baculovirus (Bac-GFP and Bac-NIS) were produced according to Bac-to-Bac baculovirus expression system. The rat islet cells were isolated and infected by Bac-GFP at different multiplicities of infection (MOI), and the infection percentage and fluorescence intensity were observed with fluorescence microscope. The islet cells were infected by Bac-GFP at MOI of 40, and the florescence protein expression time and cell survive state were observed with fluorescence microscope. The islet cells were infected by Bac-NIS at different MOI, and the iodide uptake of infected cells was determined. The islet cells were infected by Bac-NIS at MOI of 40, and the iodide uptake and effect of natrium perchloricum (NaClO4) on iodide uptake of infected cells were  dynamically observed. Results Bac-GFP could effectively infect SD rat islet cells, with no cytotoxicity. Both infection percentage and fluorescence intensity of islet cells increased with MOI, the infection percentage could be as high as 95% at MOI of 40, and fluorescence microscopy revealed that the florescence protein expression time could last for about two weeks. Bac-NIS infected islet cells exhibited enhanced iodide uptake function and inhibition property of NaClO4. The radioactive iodine uptake of infected cells increased with MOI. At MOI of 40, the radioactive count of iodide uptake of Bac-NIS infected cells was as 12 times as that of control group. Conclusion Recombinant NIS gene baculovirus can highly express in islet cells and effectively mediate iodine uptake of infected islet cells, which may provide a potent method to monitor the islet cell transplantation.

Key words: baculovirus, sodium iodine symporter, islet cell, report gene