苓桂养心汤对扩张型心肌病大鼠心肌保护作用及肌浆网钙调控蛋白的影响

Protective Effect of Linggui Yangxin Decoction on Rats with Dilated Cardiomyopathy and Effect of Calcium Regulation-associated Protein of Myocardial Sarcoplasmic Reticulum

目的:明确苓桂养心汤对扩张型心肌病(DCM)大鼠心功能、心肌的保护作用;观察苓桂养心汤对DCM大鼠损伤心肌肌浆网(SR)钙调控相关蛋白的影响,探讨其可能的作用机制。方法:以60只SD大鼠腹腔注射阿霉素1.0 mg/kg/次,每周2次,连续6周,总量12 mg·kg-1建立DCM模型,另设正常组(10只),观察2周,8周后将存活DCM模型大鼠随机分为培哚普利组、苓桂养心汤组、模型组,连续灌胃给药8周。治疗后对上述各组大鼠行心脏超声检查;采血测定大鼠血清B型尿钠肽(BNP)水平及肌钙蛋白-Ⅰ(cTnI);取大鼠左心室心肌进行苏木素-伊红(HE)染色和透射电子显微镜检测大鼠心肌病理学形态。采用逆转录聚合酶链式反应(RT-PCR)和蛋白质免疫印迹法(Western blot)分别测定大鼠心肌肌浆网钙泵(SERCA2a),钙释放通道(RyR2),受磷蛋白(PLB),肌集钙蛋白(Calsequestrin),L-型钙离子通道(CAV1.3)的mRNA及蛋白表达水平。结果:与模型组比较,苓桂养心汤组可以改善DCM模型大鼠的心腔大小、心功能、心肌损伤及心肌病理学形态。与模型组比较,苓桂养心汤组SERCA2a,RyR2,CAV1.3,Calsequestrin mRNA表达升高;PLB mRNA表达下降。与模型组比较,苓桂养心汤组SERCA2a及Calsequestrin蛋白表达升高;PLB蛋白表达下降。结论:苓桂养心汤能缩小DCM大鼠左心室内径,提高左心室射血分数和缩短率,降低血清BNP和cTnI水平,保护DCM大鼠损伤心肌,改善心功能。苓桂养心汤可部分调控DCM大鼠心肌肌浆网钙泵,兰尼碱受体,受磷蛋白,肌集钙蛋白,L-型钙离子通道的mRNA基因和蛋白表达。苓桂养心汤治疗DCM大鼠的作用机制,可能与通过调节DCM大鼠心肌肌浆网钙调控相关蛋白的基因及蛋白表达,改善心肌细胞钙转运及心脏舒缩功能有关。

Objective： To clarify the protective effect of Linggui Yangxin decoction（ LGYXD） on myocardial injury and heart function,and observe the effect of LGYXD on calcium regulation associated protein of myocardial sarcoplasmic reticulum（ SR） in rats with Adriamycin-induced dilated cardiomyopathy（ DCM）,in order to explore its possible mechanism. Method： A total of 60 SD rats were randomly divided into normal group and DCM group. The latter was further divided into five groups： DCM control,perindopril and LGYXD groups. DCM was induced through intraperitoneal injection with Adriamycin（ 1. 0 mg·kg-1,twice a week） for 6 weeks. The treatment lasted for 8 weeks. At the 9thweek,echocardiography and myocardial metabolism imaging were performed on DCM rats to evaluate the efficiency. Serum levels of brain natriuretic peptide（ BNP） and cardiac troponin-I（ c Tn I） were measured by enzyme-linked immunosorbent assay. Cardiac histological changes were examined by hematoxylin and eosin（ HE） staining and electron microscopy. The mRNA and protein expressions of sarcoplasmic reticulum Ca2 ＋-ATPase（ SERCA2a）,ryanodine receptor（ RyR2）,phospholamban（ PLB）,Calsequestrin,L-type Ca2 ＋channel（ CAV1. 3） in rat hearts from each group were determined by reverse transcription-polymerase chain reaction（ RT-PCR） and western blotting. Result： Compared with the DCM control group,LGYXD can improve cardiac cavity size,and alleviate cardiac function,myocardial injury and cardiac histological changes in DCM rats.Compared with the DCM control group,expressions of SERCA2 amRNA,RyR2 mRNA,CAV1. 3mRNA,and Calsequestrin mRNA in myocardial tissues of LGYXD group were increased,and expression of PLB mRNA was decreased. Compared with the DCM control group,the protein expressions of SERCA2 aand Calsequestrin of LGYXD group were increased,and the protein expression of PLB was decreased. Conclusion： LGYXD can effectively reduce the dilated left ventricular diameter of DCM rat,improve EF and FS,decrease the serum levels of BNP and c Tn I,protect myocardium and improve heart function. LGYXD can partially regulate mRNA and protein expressions of PLB,SERCA2 a,RyR2,CAV1. 3,Calsequestrin in rats with dilated cardiomyopathy. The mechanism of LGYXD in treating DCM rats may be correlated with the regulation of the mRNA and protein expressions of calcium regulation-associated protein of myocardial sarcoplasmic reticulum in rats with dilated cardiomyopathy,and the improvement of Ca2 ＋transport function of myocardial cells and systolic-diastolic function of heart.