摘要
目的通过构建H9c2心肌细胞缺氧模型,观察丹蒌片对心肌缺血损伤及心肌细胞钙超载的影响,并探讨其作用机制。方法取对数生长期的大鼠H9c2心肌细胞,采用加入连二亚硫酸钠(Na2S2O4)的方法建立心肌细胞缺氧损伤模型,通过外源性给予丹蒌片含药血清,采用细胞形态学观察法,MTT法检测细胞活性,采用激光共聚焦法检测各组心肌细胞内游离Ca2+浓度,采用蛋白免疫印迹技术检测钙超载磷酸化钙调蛋白依赖性激酶(p-CaMKⅡ)、钙调蛋白激酶(CaMKⅡ)的表达。结果通过细胞形态观察及MTT量效实验,确定采用5mmol/LNa2S2O4缺氧处理2h以建立稳定的心肌细胞缺氧损伤模型。与对照组比较,模型组心肌细胞内游离Ca^2+水平显著升高,P-CaMKⅡ表达增加;与模型比较,抑制剂组及丹蒌片组心肌细胞内游离Ca^2+水平均显著降低,P-CaMKⅡ表达降低(均P<0.05或P<0.01)。结论丹蒌片对心肌缺血损伤的保护作用可能与其减少外钙内流、降低内钙释放,调节P-CaMKⅡ表达有关。
Objective:To observe the effects of Danlou Tablets on myocardial ischemia injury and calcium overload in myocardial cells by constructing the hypoxic model of H9c2 myocardial cells,and to explore its mechanism. Methods:Rat H9c2 cardiomyocytes in logarithmic growth phase were taken and hypoxic injury model of cardiomyocytes was established by adding Na2S2O4. By exogenous administration of drug-containing serum,cell morphological observation,MTT assay and laser confocal method were used to detect intracellular free Ca2+ concentration in cardiac myocytes of each group. The expression of p-CaMKⅡ and CaMKⅡ in calcium overload was detected by Western blotting. Results:Through cell morphology observation and MTT dose-effect experiment,5 mmol/L Na2S2O4 hypoxic treatments was used for 2 hours to establish a stable hypoxic injury model of cardiac myocytes. Compared with the control group,the level of free Ca^2+ and the expression of P-CaMKⅡ in myocardial cells of the model group increased significantly. Compared with the model,the level of free Ca^2+ and the expression of P-CaMKⅡ in myocardial cells of the inhibitor group and Danlou Tablets group decreased significantly. Conclusion:The protective effect of Danlou Tablets on myocardial ischemia injury may be related to the decrease of calcium influx,release of calcium and regulation of P-CaMKⅡ expression.
作者
谭亚芳
梁玮婷
潘文君
祁建勇
张敏州
Tan Yafang;Liang Weiting;Pan Wenjun(The Second Affiliated Hospital of Guangzhou University of Chinese Medicine,Guangdong,Guangzhou 510120,China.)
出处
《中国中医急症》
2019年第6期945-949,共5页
Journal of Emergency in Traditional Chinese Medicine
基金
国家自然科学基金面上项目(81473471)
广东省中医院中医药科学技术研究专项(YN2016QJ19)
关键词
丹蒌片
心肌缺血
钙超载
Danlou Tablets
Myocardial ischemia
Calcium overload
