缺氧条件下Hsp90抑制剂17-AAG对RPE细胞整联蛋白连接激酶表达的影响
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:黑龙江省教育厅科学技术研究项目(No.12531389)


Effect of 17-AAG, a Hsp90 inhibitor, on the expression of ILK in RPE cells under hypoxic condition
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Heilongjiang Provincial Department of Education Science and Technology Research(No.12531389)

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    摘要:

    目的:研究热休克蛋白(heat shock protein 90,Hsp90)抑制剂17-AAG对缺氧诱导的视网膜色素上皮(retinal pigment epithelial,RPE)细胞整联蛋白连接激酶(integrin - linked kinase,ILK)表达的影响。

    方法:利用200μmol/L氯化钴(cobalt chloride, CoCl2)建立体外RPE细胞的化学缺氧模型。不同浓度的Hsp90抑制剂17-AAG预处理RPE细胞1h后给予12h缺氧处理。实验分为缺氧对照组、二甲基亚砜(DMSO)对照组和17-AAG预处理组。其中17-AAG预处理组根据浓度不同又分为6组:0.01,0.10,0.50,1.00,5.00,10.00μmol/L。应用RT-PCR和Western blot 方法检测ILK表达变化情况。

    结果:缺氧对照组、DMSO对照组和17-AAG预处理组ILK mRNA与内参β-actin mRNA光密度比值分别为1.32±0.04,1.29±0.03,0.93±0.06,0.70±0.05,0.53±0.03,0.44±0.04,0.32±0.04,0.30±0.03; ILK 蛋白与内参β-actin蛋白光密度比值分别为2.16±0.04,2.13±0.04,1.65±0.04,1.13±0.05,0.74±0.03,0.41±0.06,0.35±0.04,0.35±0.03。与缺氧对照组比较,17-AAG预处理组ILK mRNA和蛋白的表达明显降低(P<0.05),呈浓度依赖性。

    结论:Hsp90抑制剂17-AAG能够降低缺氧条件下RPE细胞ILK的表达。

    Abstract:

    AIM: To investigate the effect of 17-allylamio-17-demethoxygeldanamycin(17-AAG), an heat shock protein 90(Hsp90)inhibitor, on the expression of integrin- linked kinase(ILK)induced by hypoxia in retinal pigment epithelium(RPE)cells.

    METHODS: RPE cells were cultured with 200μmol/L cobalt chloride(CoCl2)for 12 hours to imitate chemical hypoxia. Pretreatment of 17-AAG was 1 hour prior to hypoxic insult. Experiment was divided into hypoxic control group, Dimethyl sulfoxide(DMSO)control group and 17-AAG pretreatment group(0.01, 0.10, 0.50, 1.00, 5.00 and 10.00μmol/L). RT-PCR and Western blot analysis were used to examine the expression of ILK in cultured RPE cells.

    RESULTS: The density ratios of ILK mRNA and β-actin mRNA of hypoxic control group, DMSO control group and 17-AAG pretreatment groups were 1.32±0.04,1.29±0.03,0.93±0.06,0.70±0.05,0.53±0.03,0.44±0.04,0.32±0.04,0.30±0.03; and the density ratios of ILK protein and β-actin protein were 2.16±0.04, 2.13±0.04, 1.65±0.04, 1.13±0.05, 0.74±0.03, 0.41±0.06, 0.35±0.04, 0.35±0.03. The ILK expression in 17-AAG pretreated groups were inhibited compared to hypoxic control group(P < 0.05), and the decrease was in concentration-dependent manner.

    CONCLUSION: Hsp90 inhibitor 17-AAG was effective to inhibit the expression of ILK induced by hypoxia in RPE cells.

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王业青,王健,吕勇,等.缺氧条件下Hsp90抑制剂17-AAG对RPE细胞整联蛋白连接激酶表达的影响.国际眼科杂志, 2014,14(5):811-814.

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  • 收稿日期:2013-12-28
  • 最后修改日期:2014-04-14
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  • 在线发布日期: 2014-04-21
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