当药苷调节兴奋-收缩耦联改善心肌缺血再灌注损伤的机制  

Sweroside Effectively Protects Against Ischemia/Reperfusion Injury by Regulating Excitation-contraction Coupling

作　　者：唐家杨 王青[1] 于雪[1] 魏小棋 于江 李帅[1] 李森 郭淑贞[1] TANG Jiayang;WANG Qing;YU Xue;WEI Xiaoqi;YU Jiang;LI Shuai;LI Sen;GUO Shuzhen(Beijing University of Chinese Medicine,Beijing 100029,China)

机构地区：[1]北京中医药大学,北京100029

出　　处：《中国实验方剂学杂志》2022年第15期85-93,共9页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家自然科学基金项目(81922075)。

摘　　要：目的:探索当药苷通过兴奋-收缩耦联信号通路对缺血再灌注损伤后心脏收缩/舒张功能的影响。方法:24只健康雄性SD大鼠随机分为正常组、模型组、10μmol·L^(-1)当药苷给药组与1μmol·L^(-1)地高辛给药组,并采用Langendorff系统结合左前降支冠状动脉结扎建立缺血再灌注损伤模型(I/R),2,3,5-氯化三苯基四氮唑(TTC)染色检测各组心脏梗死面积,Powerlab生理记录仪检测血流动力学参数,如左心室舒张压(LVDP)、左心室终末舒张压(LVEDP)、左心室终末收缩压(LVESP)、左心室内压最大上升速率(+dp/dt_(max))和左心室内压最大下降速率(-dp/dt_(max));提取分离乳鼠原代心肌细胞(NRCMs),建立缺氧/复氧(H/R)损伤模型,随机分为正常组、模型组、1μmol·L^(-1)当药苷给药组和10μmol·L^(-1)当药苷给药组,多功能成像细胞分析仪和激光共聚焦显微镜测定各组心肌细胞活力、心率、收缩幅度、收缩时程、达峰时程和舒张时程及钙瞬变峰值。根据前期转录组学测序结果及文献调研,利用实时荧光定量聚合酶链式反应(Real-time PCR)检测L型钙通道相关基因(Cacnb2),细胞色素C氧化酶相关基因(Cox6a2),肌钙蛋白(Tnnc1、Tnni3、Tnnt2)、肌动蛋白(Actc1)、肌球蛋白(Myh6、Myl2、Myl4)等兴奋-收缩耦联通路基因的mRNA表达并对差异基因进行聚类分析。结果:与正常组比较,模型组心肌梗死面积显著增加(P<0.01)、LVDP显著降低(P<0.01)、LVEDP显著上升(P<0.01)、LVESP明显下降(P<0.05)、+dp/dt_(max)有下降趋势和-dp/dt_(max)上升趋势及心肌细胞活力降低、心率降低、收缩幅度降低、收缩时程升高、达峰时程升高和舒张时程升高(P<0.01),而当药苷可以逆转上述指标(P<0.05)。此外,心肌细胞经H/R损伤后,Cacnb2、Cox6a2、Tnnc1、Tnni3、Tnnt2、Actc1、Myh6、Myl2、Myl4等兴奋-收缩耦联通路的基因表达下降(P<0.05、P<0.01)。而使用当药苷预处理后,可以增强上述基因的表达(Objective: To explore the effect of sweroside on the protection of cardiac systolic/diastolic function during ischemia/reperfusion (I/R) injury.Method: Twenty-four healthy male SD rats were randomly divided into control group, model group, 10 μmol·L^(-1) sweroside group and 1 μmol·L^(-1) digoxin group. The I/R injury was modeled by Langendorff and ligation of the left anterior descending coronary artery. The infarct size in each group was measured by 2,3,5-triphenyltetrazolium chloride (TTC) staining and hemodynamic parameters such as left ventricular diastolic pressure (LVDP), left ventricular end-diastolic pressure (LVEDP), left ventricular end-systolic pressure (LVESP), maximum rate of rising of left ventricular pressure (+dp/dt(max)) and maximum rate of decreasing of left ventricular pressure (-dp/dt_(max)) of rat isolated heart were detected by Powerlab. In addition, neonatal rat cardiomyocytes (NRCMs) were isolated and randomly divided into control group, model group, 1 μmol·L^(-1) sweroside group and 10 μmol·L^(-1) sweroside group. Hypoxia/reoxygenation (H/R) injury model was established. Cardiac systolic function and calcium transients were examined by multi-functional cell imaging analyzer and laser confocal microscope. Furthermore, real-time polymerase chain reaction(Real-time PCR) was used to verify the mRNA expression of excitation-contraction coupling genes such as L-type calcium channel (Cacnb2), cytochrome c oxidase subunit 6A2 (Cox6a2), troponin (Tnnc1, Tnni3, Tnnt2), actin (Actc1), and myosin (Myh6, Myl2, Myl4) according to the results of previous transcriptome sequencing and literature investigation. Differentially expressed genes were subjected to cluster analysis.Result: Compared with the conditions in the control group, increased cardiac infarction size (P<0.01) and LVEDP (P<0.01) and decreased LVDP (P<0.01) and LVESP (P<0.05) were observed in the model group, with +dp/dt_(max) of increasing trend while -dp/dt_(max) decreasing. Moreover, the cell viability, heart rate and contraction

关 键 词：当药苷 缺血再灌注损伤 收缩与舒张功能 CA^(2+) 兴奋收缩耦联通路 

分 类 号：R2-0[医药卫生—中医学] R33R285R541