钙蛋白酶对血管紧张素Ⅱ诱导的心肌成纤维细胞增殖及纤维化模型的影响  

Effect of Calpain on Angiotensin II-induced Proliferation and Fibrosis in Cardiac Fibroblasts

作　　者：马越[1] 余洋[1] 贾岩岩[1] 陈少锐[1] 叶建涛[1] 马韵子 李卓明[1] 刘培庆[1] 

机构地区：[1]中山大学药学院,广东广州510006

出　　处：《中山大学学报（医学科学版）》2013年第4期491-497,共7页Journal of Sun Yat-Sen University:Medical Sciences

基　　金：国家自然科学基金(81000072;81072641)

摘　　要：【目的】研究钙蛋白酶在血管紧张素II(Ang II)诱导的心肌纤维化中的作用及作用机制。【方法】原代培养SD大鼠心肌成纤维细胞(CF),取传代2-4代细胞,,预先给予不同浓度calpain抑制剂PD150616,观察对Ang II诱导致纤维化模型的影响。实验分为空白对照组(control)、模型组(Ang II)、低浓度给药组(Ang II+PD150616_5μmol/L)、高浓度给药组(Ang II+PD150616_10μmol/L)。Edu检测CF增殖,Western blot检测纤维化因子纤连蛋白(FN)、结缔组织生长因子(CTGF)、α-平滑肌肌动蛋白(α-SMA)蛋白表达,同时检测calpain激活的下游产物Cleavedα-Fodrin以及胞浆中p-IκB与细胞核内P65、NFAT4的蛋白水平变化。【结果】PD150616显著抑制由Ang II诱导的CF增殖及FN、CTGF、α-SMA等纤维化因子表达上调。Ang II显著提高Cleavedα-Fodrin的表达,PD150616可阻断Ang II的作用,提示calpain受Ang II激活,是Ang II作用的下游因子。PD150616可抑制Ang II引起的胞内IκB磷酸化和胞核中p65、NFAT4的表达上调,提示calpain的作用与NF-κB、NFAT4转位入核的信号通路相关。【结论】calpain在Ang II刺激诱导心肌纤维化过程中发挥重要作用,其机制可能与NF-κB和NFAT信号通路的激活有关。[Objective] To investigate the effect of calpain on angiotensin Ⅱ （Ang Ⅱ）-induced myocardial fibrosis and the possible mechanisms. [Methods] Cardiac fibroblasts （CF） were primary cultured from hearts of Sprague-Dawley rats. Ang Ⅱ was treated to induce myocardial fibrosis and the effect of the pharmacological calpain inhibitor PD150616 was studied. Edu assay was used to test the proliferation of CF. Protein expression of fibronectin （FN）, connective tissue growth factor （CTGF）, α-smooth muscle actin （α-SMA）, Cleaved α-Fodrin which indicates the activation of calpain, the cytoplasmic p-IgB, as well as the nuclear p65 and NFAT4, were measured by Western blot analysis. [ Results ] PD 150616 significantly inhibited Ang II-induced CF proliferation and up- regulation of FN, CTGF and α-SMA, suggesting that calpain plays a role in myocardial fibrosis. The expression of cleaved α-Fodrin was remarkably increased by Ang II, which was reversed by PD150616, suggesting that calpain was downstreamly activated by Ang II. PD150616 inhibited Ang II-induced cytoplasmic IKB phosphrylation and up-regulation of nuclear p65 and NFAT4, indicating that the effect of calpain on myocardial fibrosis is probably through the nuclear translocation of NF-KB and NFAT4. [ Conclusion] Calpain, when activated by Ang II, can induce myocardial fibrosis, probably through the activation of NF-KB and NFAT signaling pathways.

关 键 词：钙蛋白酶 心肌纤维化 血管紧张素Ⅱ 

分 类 号：R965.2[医药卫生—药理学]