程氏舒心汤调控NRG1/ErbB信号通路减轻大鼠心肌缺血再灌注损伤的机制研究

Chengshi Shuxin Decoction regulates the NRG1/ErbB signaling to reduce myocardial ischemia reperfusion injury in rats

目的探讨程氏舒心汤对大鼠心肌缺血再灌注损伤(MIRI)的疗效及分子机制。方法选取40只Sprague Dawley大鼠按随机数字表法分为假手术组、模型组、程氏舒心汤低剂量组及程氏舒心汤高剂量组,每组10只。通过结扎冠状动脉前降支构建MIRI大鼠模型,予不同剂量程氏舒心汤灌胃治疗。通过检测血清肌酸激酶同工酶(CK-MB)、肌钙蛋白I(cTnI)、乳酸脱氢酶(LDH)等心肌损伤标志物及心脏超声评估心功能,HE染色观察心肌病理损伤,采用透射电镜观察心肌超微结构改变,应用Western blot实验检测相关蛋白表达水平。结果假手术组大鼠建模后7 d全部存活,模型组大鼠仅存活6只,程氏舒心汤干预组大鼠各存活8只。与假手术组相比,模型组血清CK-MB[(1925.19±141.98)U/L比(919.83±104.74)U/L,P<0.05]、cTnI[(15.20±2.97)ng/mL比(0.29±0.08)ng/mL,P<0.05]、LDH[(1730.00±91.74)U/L比(780.70±78.31)U/L,P<0.05]等心肌损伤标志物显著升高,左心室射血分数(LVEF)[(45.38±3.86)%比(81.42±2.39)%,P<0.05]、左心室短轴缩短率(LVFS)[(23.60±2.11)%比(52.68±2.92)%,P<0.05]等心功能指标显著降低;HE染色发现,模型组心肌细胞变性坏死,心肌纤维断裂,出现明显炎症细胞浸润;透射电镜显示,心肌线粒体肿胀,线粒体嵴破坏,肌丝排列紊乱,横纹消失;Western blot检测发现,神经调节蛋白1(NRG1)、酪氨酸激酶受体2(ErbB2)及酪氨酸激酶受体4(ErbB4)蛋白表达降低;与模型组相比,程氏舒心汤低、高剂量组大鼠血清CK-MB[(1354.13±88.45)U/L、(1283.65±115.51)U/L比(1925.19±141.98)U/L,P<0.05]、cTnI[(10.92±2.63)ng/mL、(7.29±1.91)ng/mL比(15.20±2.97)ng/mL,P<0.05]、LDH[(1409.13±75.75)U/L、(1345.97±68.46)U/L比(1730.00±91.74)U/L,P<0.05]等心肌损伤标志物显著降低,LVEF[(53.70±2.50)%、(62.45±1.69)%比(45.38±3.86)%,P<0.05]、LVFS[(32.84±3.19)%、(39.48±4.15)%比(23.60±2.11)%,P<0.05]等心功能指标显著升高,且上述心肌病理损伤情况得到明显改善,NRG1、ErbB2及ErbB4蛋白表达升高。结论程氏舒心汤可减轻大鼠MIRI,其机制可能与调控NRG1/ErbB通路,促进NRG1、ErbB蛋白表达有关。

Objective To investigate the efficacy and molecular mechanism of Chengshi Shuxin Decoction on myocardial ischemia and reperfusion injury(MIRI)in a rat model.Methods Forty Sprague-Dawley rats were purchased and randomly divided into sham,model,Chengshi Shuxin Decoction-low dose,and Chengshi Shuxin Decoction-high dose groups(n=10 per group).The MIRI model of rats was constructed by ligation of the left anterior descending coronary artery and then treated with a high or low dose of Chengshi Shuxin Decoction via oral gavage.Cardiac function was then assessed by detecting serum level of creatine kinase isoenzyme(CK-MB),troponin I(cTnI),lactate dehydrogenase(LDH),and cardiac ultrasound,while the myocardial damage was observed by using H&E staining of tissue sections,myocardial ultrastructural change was observed by using the transmission electron microscopy,and level of different proteins was detected by using Western blot.Results All 10 rats in the sham operation group survived 7 days after modeling,but there were only six rats in the model group survived,and eight rats in high or low dose Chengshi ShuXin Decoction intervention groups survived.Compared with rats in the sham operation group,serum markers of myocardial injury,e.g.,CK-MB(1925.19±141.98 vs.919.83±104.74 U/L;P<0.05),cTnI(15.20±2.97 vs.0.29±0.08 ng/mL;P<0.05),and LDH(1730.00±91.74 vs.780.70±78.31 U/L;P<0.05)in the model group were all significantly increased.The left ventricular ejection fraction(LVEF,45.38±3.86 vs.81.42±2.39%;P<0.05)and the left ventricular fraction shortening(LVFS,23.60±2.11 vs.52.68±2.92%;P<0.05)were all decreased.H&E staining data showed that myocardial cells in the model group of rats were degenerated and necrotic and myocardial fibers were broken with obvious inflammatory cell infiltration.The transmission electron microscope showed myocardial mitochondrial swelling,mitochondrial ridge destruction,myofilament arrangement disorder,and transverse lines disappeared.Western blot data revealed that expression of NRG1,ErbB2 and ErbB4 proteins was also decreased.Compared with the model rats,serum level of myocardial injury,like CK-MB(1354.13±88.45 and 1283.65±115.51 vs.1925.19±141.98 U/L;all P<0.05),cTnI(10.92±2.63 and 7.29±1.91 vs.15.20±2.97 ng/mL;all P<0.05),LDH(1409.13±75.75 and 1345.97±68.46 vs.1730.00±91.74 U/L;all P<0.05)were significantly reduced after both high and low doses of Chengshi Shuxin Decoction treatment.The cardiac function indexes such as LVEF(53.70±2.50 and 62.45±1.69 vs.45.38±3.86%;all P<0.05),LVFS(32.84±3.19 and 39.48±4.15 vs.23.60±2.11%;all P<0.05)were all significantly induced after the treatment and the above-named myocardial injuries were all significantly improved,while expressions of NRG1,ErbB2 and ErbB4 proteins was upregulated.Conclusion Chengshi Shuxin Decoction can protect rats from MIRI,which is probably through regulation of NRG1/ErbB pathway by promoting expression of NRG1 and ErbB protein.