shRNA静默CD4+ T细胞AHNAK1基因对小鼠甲状腺相关性眼病进程的抑制作用
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国家自然科学基金青年项目(No.30801000)


Inhibition effect of the thyroid-associated ophthalmopathy in mice through the lentivirus expressing AHNAK1 shRNA on T cells
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National Natural Science Foundation of China(No.30801000)

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    摘要:

    目的:构建表达小鼠CD4+T细胞钙支架蛋白AHNAK1的短发夹RNA(short hairpin RNA ,shRNA)慢病毒载体,并研究其对小鼠甲状腺相关性眼病(thyroid-associated ophthalmopathy,TAO)的抑制效应。

    方法:设计并筛选对AHNAK1具有良好干扰效力的shRNA序列,慢病毒载体包装干扰序列,感染小鼠CD4+T细胞,检测AHNAK1静默对T细胞功能的抑制作用,采用实验动物模型观察AHNAK1体内抑制甲状腺相关性眼病的效果。

    结果:成功筛选出具有良好干扰效力的shRNA,并包装入慢病毒。病毒滴度为1.0×106TU/mL,转染慢病毒的CD4+T细胞展现出失能倾向,抑制炎症免疫反应; 在动物模型中抑制T细胞中AHNAK1表达可以有效控制甲状腺眼病的发生发展, 显著降低治疗组T细胞中IL-2、IL-1β和IFN-γ的表达。

    结论:成功构建了表达小鼠AHNAK1 shRNA的慢病毒,具有抑制T细胞分泌IL-2、IL-1β和IFN-γ的表达效应,能够有效抑制甲状腺眼病的发生发展。

    Abstract:

    AIM:To construct the mice model with lentivirus expressing AHNAK1 shRNA in CD4+ T cells, and to study its inhibitory effect on the thyroid-associated ophthalmopathy(TAO)in mice.

    METHODS: The shRNA sequence with good disturbing potency towards AHNAK1 was designed and selected; then the shRNA was packed into lentivirus; and the CD4+ T cells were infected. The infected CD4+ T cells of mice by the packed lentivirus were observed to detect the inhibition effect on T cells. And then the immunotherapeutic effects of AHNAK1-/- on TAO were observed by experimental animal model.

    RESULTS: The shRNA with good disturbing potency was successfully screened and correctly inserted into the lentivirus. The titer of the recombinant lentivirus was 1.0×106TU/mL. The CD4+ T cells infected by lentivirus showed the anergy trend and restrained the immune response of inflammation. Suppressing the expression of AHNAK1 in T cells of animal model can effectively control the occurring and proceeding of TAO, which can significantly reduce the expression of IL-2 /IL-1β/IFN-γ in the T cells of the control group.

    CONCLUSION: This paper successfully constructs mice model with the recombinant lentivirus expressing AHNAK1 shRNA, which has a favorable inhibitory effect on secretion of IL-2, IL-1β, IFN-γ by T cells. The recombinant lentivirus can effectively inhibit the occurring and proceeding of TAO in mice.

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陈华新,于少硕,周维明,等. shRNA静默CD4+ T细胞AHNAK1基因对小鼠甲状腺相关性眼病进程的抑制作用.国际眼科杂志, 2014,14(2):225-228.

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  • 收稿日期:2013-08-21
  • 最后修改日期:2013-12-31
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  • 在线发布日期: 2014-01-20
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