1型多聚二磷酸腺苷核糖合成酶对去甲肾上腺素诱导心脏成纤维细胞表达基质金属蛋白酶的影响  

Involvement of poly(ADP-ribose) polymerase in norepinephrine-induced expression of MMPs/TIMPs in cultured cardiac fibroblasts

作　　者：黄丹[1] 黄恺[1] 王妍[1] 杨崇哲[1] 李小丽[1] 廖玉华[1] 

机构地区：[1]华中科技大学协和医院心内科华中科技大学同济医学院心血管病研究所,武汉430022

出　　处：《临床心血管病杂志》2008年第9期702-706,共5页Journal of Clinical Cardiology

基　　金：国家自然科学基金资助项目(No:30770881)

摘　　要：目的:探讨1型多聚二磷酸腺苷核糖合成酶(PARP-1)对去甲肾上腺素(NE)诱导培养的大鼠心脏成纤维细胞基质金属蛋白酶(MMP)-1,MMP-9及金属蛋白酶组织抑制剂(TIMP)-1表达的调节作用及其机制。方法:①培养乳鼠心脏成纤维细胞,10μmol/LNE刺激细胞24h,使用实时定量PCR法检测MMP-1、MMP-9及TIMP-1的基因表达水平;使用PARP抑制剂3-氨基苯甲酰胺(3-aminobenzamide,3AB)后,观察PARP-1对上述基因表达的影响。②检测心脏成纤维细胞内活性氧(ROS)水平、PARP酶活性的变化。③采用凝胶阻滞实验检测心脏成纤维细胞内转录因子AP-1的DNA结合能力,研究PARP-1对AP-1DNA结合能力的影响。结果:NE诱导心脏成纤维细胞内MMP-1,MMP-9及TIMP-1的基因表达水平明显增加。细胞内ROS产生增加,PARP酶被激活。核内转录因子AP-1的DNA结合能力明显增强。PARP抑制剂3AB可明显减少NE诱导的MMP-1、MMP-9及TIMP-1的基因表达水平,同时显著抑制AP-1的DNA结合能力。使用抗氧化剂vitC减少ROS产生,抑制了NE诱导的PARP-1活性增加及AP-1的DNA结合,进而显著降低了NE诱导的MMP-1、MMP-9及TIMP-1的基因表达水平。结论:NE刺激心脏成纤维细胞内ROS产生明显增多,大量的ROS激活了PARP使其酶活性显著增高,PARP通过调节转录因子AP-1的DNA结合调控了MMP-1,MMP-9及TIMP-1的基因表达。PARP可能是心脏纤维化过程中的重要调节机制之一。Objective： To investigate the role of poly （ADP-ribose） polymerase in NE-induced expression of MMPs/TIMPs in cultured rat cardiac fibroblasts. Method： Cultured rat cardiac fibroblasts were divided into 4 groups. Group 1 （control group） received no treatment. Group 2 received NE （10μmol/L） treatment alone for 24 hours. Group 3 was pretreated with 15 mmol/L PARP inhibitor 3-aminobenzamide （3AB） for 1 hours, followed by additional incubation with NE （10 μmol/L） for 24 hours. Group 4 was pretreated with vitamin C （1.4 mmol/ L） followed by NE treatment for 24 hours, lntracellular ROS level was measured with ROS-sensitive fluorophore DCF-DA by Fluoromax-3 spectrofluorimeter. PARP activity of cultured cells was measured with colorimetric PARP assay kit （Trevigen）. mRNA levels of MMP-1. MMP-9 and TIMP-1 were measured with real-time PCR. The DNA binding activity of AP-1 was measured with electrophoretic mobility shift assay （EMSA）. Result.. Both PARP inhibitor 3AB and vitC significantly prevented NE induced （1） increase in PARP activity; （2） increases in mRNA expression of MMP-1, MMP-9 and TIMP-1 in cardiac fibroblasts; （3） increase in ROS generation; （4） increase in the DNA binding activity of AP-1. Conclusion： Activation of PARP/PARP-1 contributes importantly to the fibrosis-related gene expression of MMP-1, MMP-9 and TIMP 1 in NE treated cardiac fibroblasts through promoting AP-1 DNA binding activity.

关 键 词：心肌纤维化 多聚ADP核糖聚合酶 去甲肾上腺素 基因表达 

分 类 号：R542.23[医药卫生—心血管疾病]