SP1基因在sublytic C5b⁃9促大鼠肾小球系膜细胞增殖中的作用
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国家自然科学基金(31470853,81471626);江苏省高等学校大学生创新训练计划项目(201710312020Z)


The role of SP1 gene in sublytic C5b⁃9⁃induced GMC proliferation
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    摘要:

    目的:检查特异性蛋白1(specificity protein 1,SP1)在Thy-1肾炎(Thy-1 nephritis,Thy-1N)大鼠肾组织中和亚溶解型C5b-9(sublytic C5b-9)刺激的大鼠肾小球系膜细胞(glomerular mesangial cells,GMCs)中的表达情况,并探讨SP1表达上调对sublytic C5b-9促大鼠GMCs增殖的作用。方法:复制大鼠Thy-1N模型,同时用sublytic C5b-9刺激体外培养的大鼠GMCs,分别用real-time PCR和Western blot检查Thy-1N大鼠肾组织(体内)和sublytic C5b-9刺激的大鼠GMCs(体外)中SP1基因的mRNA和蛋白表达水平及其表达时相。与此同时,构建SP1基因的表达质粒(pEGFP-N1/SP1-His)和发夹状小干扰RNA质粒(SP1 shRNA,shSP1)。将上述质粒分别转染大鼠GMCs(同时设对照质粒转染组),再给予或不给予sublytic C5b-9刺激。用real-time PCR和Western blot检查SP1表达量,并行CCK-8实验检查GMCs的增殖情况。结果:在Thy-1N大鼠肾组织和sublytic C5b-9刺激的大鼠GMCs中,SP1 mRNA和蛋白表达水平均显著上调,且其体内外表达时相较为一致。体外过表达SP1可显著促进GMCs增殖,而沉默SP1可明显抑制sublytic C5b-9诱导的GMCs增殖反应。结论:Sublytic C5b-9刺激大鼠GMCs后能通过上调SP1表达促进GMCs增殖。

    Abstract:

    Objective:To detect the expression of specificity protein 1(SP1)in the renal tissues of rats with Thy-1 nephritis(Thy-1N)and in the glomerular mesangial cells(GMCs)stimulated with sublytic C5b-9,and investigate the role of SP1 expression in sublytic C5b-9-induced GMC proliferation. Methods:Rat Thy-1N was induced,and the cultured rat GMCs were stimulated with sublytic C5b-9. The expression of SP1 in the renal tissues of rats with Thy-1N(in vivo)and in the GMCs stimulated with sublytic C5b-9(in vivo)was detected by real-time PCR and Western blot,respectively. Meantime,SP1 expression plasmid(pEGFP-N1/SP1-His)and SP1 shRNA(shSP1)were constructed. Above-mentioned plasmids and control plasmids were transfected into rat GMCs followed by sublytic C5b-9 stimulation or not. The expression levels of SP1 were evaluated by real-time PCR and Western blot,and the cellular proliferation was determined by CCK-8. Results:The expression levels of SP1 in the renal tissues of rats with Thy-1N and in the GMCs stimulated with sublytic C5b-9 were markedly increased with similar time points. SP1 overexpression could markedly promote GMCs proliferation,while SP1 knockdown could obviously reduce sublytic C5b-9-induced GMCs proliferation. Conclusion:Sublytic C5b-9 promotes rat GMCs proliferation through up-regulation of SP1.

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朱宇峰,蔡梦媛,刘昌伟,王庆林,张恩瑞,钱宝梅,张志伟,赵 聃,邱 文,王迎伟. SP1基因在sublytic C5b⁃9促大鼠肾小球系膜细胞增殖中的作用[J].南京医科大学学报(自然科学版),2018,(11):1499-1505

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  • 收稿日期:2018-05-14
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  • 在线发布日期: 2018-12-03
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