氧化型低密度脂蛋白对人脐静脉内皮细胞Fractalkine表达的影响及机制分析
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山西省科技攻关项目(20090311059-3)


The Effect of ox-LDL on the Expression of Fractalkine in Cultured Human Umbilical Vein Endothelial Cells and Its Mechanism
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    目的观察氧化型低密度脂蛋白对人脐静脉内皮细胞Fractalkine表达的影响并探讨其可能作用机制。方法采用胰蛋白酶消化法原代培养人脐静脉内皮细胞,取2~5代用于实验,随机分为:①对照组用RPMI1640培养基;②氧化型低密度脂蛋白不同浓度(5、25、50和75mg/L)组;③氧化型低密度脂蛋白作用不同时间(6、12、24、48和72h)组;④氧化型低密度脂蛋白+p38MAPK特异性阻断剂SB203580组;⑤p38MAPK特异性阻断剂SB203580组。用RT-PCR及ELISA法检测Fractalkine mRNA及蛋白表达水平,用Westernblot法检测p38MAPK磷酸化表达水平。结果①氧化型低密度脂蛋白在一定浓度范围及作用时间呈时间-剂量依赖方式诱导Fractalkine mRNA及蛋白表达增加,最佳作用时间为48h,最佳作用浓度为50mg/L;②与对照组比较,氧化型低密度脂蛋白诱导组人脐静脉内皮细胞p38MAPK磷酸化表达水平显著增加(P<0.05);使用SB203580干预后p38MAPK磷酸化水平明显下降,与氧化型低密度脂蛋白诱导组相比差异有统计学意义(P<0.05)。③预先用p38MAPK特异性阻断剂SB203580(20μmol/L)与内皮细胞共同孵育60min后,再加入氧化型低密度脂蛋白作用48h后,Fractalkine表达水平明显降低,与氧化型低密度脂蛋白组比较有统计学差异(P<0.05)。结论氧化型低密度脂蛋白可诱导内皮细胞Fractalkine表达增加,其作用机制可能是通过p38MAPK信号转导通路。

    Abstract:

    Aim To investigate the effect of oxidized low density lipoprotein(ox-LDL) on Fractalkine expression in cultured human umbilical vein endothelial cells (HUVEC) and explore its mechanism.Methods HUVECs were isolated and cultured,the passage 2~5 cells were used in experiment.HUVEC were coincubated with different concentrations of ox-LDL (5,25,50 and 75 mg/L) for 48 h or with 50 mg/L ox-LDL for different times(6,12,24,48 and 72 h).SB203580(20 μmol/L),an inhibitor of p38MAPK activation,was pretreated for 60min before HUVEC were coincubated with 50 mg/L ox-LDL for 48 h.Fractalkine mRNA and protein expression were detected by RT-PCR or ELISA.The phosphorylation of p38MAPK was determined by Western blot.Results The expressions of fractalkine in HUVEC were upregulated by ox-LDL(5,25 and 50 mg/L) in a concentration-dependent manner and were increased by ox-LDL(50 mg/L) for 6,12,24,48 and 72 h in a time-dependent manner;Compared with control group,the phosphorylation of p38MAPK was increased significantly in ox-LDL group (P<0.05),SB203580 (20 μmol/L) decreased ox-LDL induced fractalkine expression.Conclusion ox-LDL stimulates the expression of fractalkine in a concentration and time dependent manner,p38MAPK activation may mediate ox-LDL induced fractalkine expression.

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边云飞,杨慧宇,杨志明,张娜娜,高奋,肖传实.氧化型低密度脂蛋白对人脐静脉内皮细胞Fractalkine表达的影响及机制分析[J].中国动脉硬化杂志,2009,17(10):802~806.

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  • 收稿日期:2009-06-19
  • 最后修改日期:2009-09-12
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