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SIRT3 siRNA对ox-LDL刺激的人主动脉内皮细胞ICAM-1、E-selectin表达的影响

丁玲1,2， 巩会平1，杜贻萌1，王欣1，张岩3，王旭平2，鹿庆华1   

  1. 1. 山东大学第二医院心血管内科，山东 济南 250033；
    2. 山东大学齐鲁医院教育部与卫生部心血管重构和功能研究重点实验室，山东 济南 250012；
    3. 中国医学科学院北京协和医院心血管内科，北京 100730
  • 收稿日期:2013-09-04 出版日期:2014-04-10 发布日期:2014-04-10
  • 通讯作者: 鹿庆华 E-mail:lqhzhy@163.com
  • 基金资助:
    山东省科学发展计划项目（2009GG20002034)；山东省自然科学基金（ZR2010HL029，ZR2012HL51)；山东省科技攻关项目（2009GG20002077)

Effect of SIRT3 siRNA on ICAM-1 and E-selectin expressions in HAEC stimulated by ox-LDL

DING Ling1,2,  GONG Huiping1, DU Yimeng1, WANG Xin1, ZHANG Yan3, WANG Xuping2, LU Qinghua1   

  1. 1. Department of Cardiovascular, the Second Hospital of Shandong University, Jinan 250033, Shandong, China;
    2. Key Laboratory of Cardiovascular Remodeling and Function Research, Chinese Ministry of Education and
    Ministry of Health, Qilu Hospital of Shandong University, Jinan 250012, Shandong, China;
    3. Department of Cardiovascular, Chinese Academy of Medical Sciences ＆
    Peking Union College Hospital, Beijing 100730, China
  • Received:2013-09-04 Online:2014-04-10 Published:2014-04-10

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摘要： 目的  探讨SIRT3基因对氧化型低密度脂蛋白（ox-LDL)刺激的人主动脉内皮细胞（HAEC)炎性因子细胞间黏附分子-1（ICAM-1）、E-选择素（E-selectin）表达的影响。方法  将生长状态良好的细胞随机分为空白对照组、转染试剂对照组、阴性对照组及SIRT3 siRNA（1、2、3、4）组。筛选出最佳序列后，细胞分为空白对照组、siRNA组、（20、30、40μg/mL）ox-LDL组、siRNA+（20、30、40μg/mL）ox-LDL组。转染24h、ox-LDL刺激6h。qRT-PCR检测SIRT3 mRNA水平，Western blotting检测SIRT3蛋白表达，流式细胞术检测E-selectin、ICAM-1抗体的平均荧光强度值(MFI)。结果  SIRT3 siRNA2序列在mRNA水平的干扰效率为88.62%，蛋白水平为80.37%，干扰效果明显。与空白对照组相比，ox-LDL各组的SIRT3 mRNA相对表达量差异有统计学意义（P均＜0.05）；随ox-LDL浓度增加，SIRT3 mRNA相对表达量逐渐降低（P均＜0.05）。与siRNA组相比，siRNA+ox-LDL各组ICAM-1、E-selectin的MFI差异有统计学意义（P均＜0.05）；随ox-LDL浓度增加，MFI逐渐升高（P均＜0.05）。与空白对照组相比，siRNA组ICAM-1、E-selectin 的MFI无统计学差异（P均＞0.05）。结论  SIRT3基因与ox-LDL刺激的人主动脉内皮细胞ICAM-1、E-selectin表达升高相关，可能参与并调控动脉内皮细胞损伤的炎症反应。

关键词: 氧化型低密度脂蛋白, SIRT3, siRNA, E-选择素, 人主动脉内皮细胞, 细胞间黏附分子-1

Abstract: Objective  To investigate the effect of SIRT3 gene on the expressions of inflammatory factors intercellular cell adhesion molecule-1 (ICAM-1) and E-selectin in human aortic endothelial cell (HAEC) stimulated by oxidized-low density lipoprotein(ox-LDL). Methods  The cells were randomly divided into normal control group, transfection reagent control group, negative control group, and SIRT3 siRNA (1, 2, 3 and 4) groups. When the most effective interference sequence was selected out, the cells were divided into normal control group, siRNA group, (20, 30 and 40μg/mL) ox-LDL groups and siRNA+(20, 30 and 40μg/mL) ox-LDL groups. The cells were transfected for 24h and stimulated by ox-LDL for 6h. SIRT3 mRNA level was detected by qRT-PCR and its protein level was analyzed by Western blotting.  Mean fluorescence intensities (MFIs) of ICAM-1 and E-selectin antibody were detected by flow  cytometry. Results  SIRT3siRNA2 was the most effective interference sequence. The interference efficiency of SIRT3siRNA2 was 88.62% in mRNA level and 80.37% in protein level. Expressions of SIRT3 mRNA had significant differences in ox-LDL groups (all P＜0.05) compared with that in normal control group; with the increase of ox-LDL concentration, the expression of SIRT3 mRNA gradually declined (all P＜0.05). MFI of ICAM-1 and E-selectin had statistical differences in siRNA+ox-LDL groups compared with that in siRNA group (all P＜0.05); with the increase of ox-LDL concentration, the MFI was enhanced obviously (all P＜0.05). MFI of ICAM-1 and E-selectin in siRNA group had no significant difference compared with that in normal control group (P＞0.05). Conclusion  The SIRT3 gene has association with the expressive increments of  E-selectin and ICAM-1 in HAEC stimulated by ox-LDL, and may regulate the inflammatory reaction in artery endothelial cell injury process.

Key words: E-selectin, siRNA, SIRT3, Human aortic endothelial cell, Oxidized-low density lipoproteinSIRT3, Intercellular cell adhesion molecule-1

中图分类号: 

  • R541.4
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