龙血通络胶囊对异丙肾上腺素诱导大鼠心肌缺血的保护作用

Protective effect of Longxuetongluo Capsule on isoproterenol-induced myocardial ischemia in rats

目的探究龙血通络胶囊对异丙肾上腺素(isoproterenol,ISO)诱导大鼠心肌缺血的保护作用。方法Wistar大鼠随机分为对照组、模型组、地奥心血康(62.50 mg/kg)组和龙血通络胶囊低、中、高剂量(103.12、206.25、412.50 mg/kg)组,每组10只。各组连续28 d ig给予相应药物,第27、28天给药1 h后,除对照组sc生理盐水外,其余各组sc ISO(150 mg/kg)。第1次sc ISO 48 h后摘取大鼠心脏,计算心脏指数;采用苏木素-伊红(HE)染色观察各组大鼠心肌组织病理变化;采用ELISA法检测各组大鼠血清中肌酸激酶(creatine kinase,CK)、肌酸激酶同工酶(creatine kinase-MB,CK-MB)和心肌钙蛋白-I(cardiac troponin-I,cTn-I)水平;采用试剂盒检测大鼠血清中乳酸脱氢酶(lactate dehydrogenase,LDH)、超氧化物歧化酶(superoxide dismutase,SOD)和天冬氨酸氨基转移酶(aspartate aminotransferase,AST)活性以及丙二醛(malondialdehyde,MDA)水平;采用TUNEL染色观察大鼠心肌细胞凋亡情况;采用免疫组化法检测心肌组织中核因子E2相关因子2(nuclear factor-erythroid 2-related factor 2,Nrf2)、血红素加氧酶-1(hemeoxygenase-1,HO-1)和剪切型半胱氨酸天冬氨酸蛋白酶-3(cleaved cystein-asparate protease-3,cleaved Caspase-3)蛋白表达。结果与对照组比较,模型组大鼠心脏指数显著升高(P<0.001),血清中cTn-I、CK-MB、CK、MDA水平以及LDH、AST活性显著升高(P<0.001),SOD活性降低(P<0.001),心肌组织出现明显的病理变化(P<0.001),心肌细胞凋亡率显著升高(P<0.001),心肌组织中cleaved Caspase-3、Nrf2和HO-1蛋白表达水平均显著升高(P<0.05、0.001)。与模型组比较,龙血通络胶囊组大鼠心脏指数明显降低(P<0.001),血清中cTn-I、CK-MB、CK、MDA水平以及LDH、AST活性显著降低(P<0.05、0.01、0.001),SOD活性显著升高(P<0.001);心肌组织的病理性改变得到改善(P<0.05);心肌组织中心肌细胞凋亡率显著降低(P<0.01);心肌组织中cleaved Caspase-3蛋白表达显著降低(P<0.05、0.01),Nrf2、HO-1蛋白表达显著升高(P<0.01、0.001)。结论龙血通络胶囊可改善ISO诱导大鼠心肌缺血损伤,其作用机制可能是通过调节Nrf2/HO-1通路,进而抑制氧化应激诱导的细胞凋亡。

Objective To investigate the protective effect of Longxuetongluo Capsule(龙血通络胶囊)on isoproterenol(ISO)-induced myocardial ischemia in rats.Methods Wistar rats were randomly divided into control group,model group,Di’ao Xinxuekang(62.50 mg/kg)group and Longxuetongluo Capsule low-,medium-and high-dose(103.12,206.25,412.50 mg/kg)groups,with 10 rats in each group.Each group was ig corresponding medication for 28 consecutive days.After 1 h of administration on 27th and 28th days,except for the control group with physiological saline,all other groups were sc ISO(150 mg/kg).The hearts of rats were collected and heart weight index was calculated 48 h after the first subcutaneous injection of ISO;Hematoxylin-eosin(HE)staining was used to observe the pathological changes of myocardial tissues in each group;The levels of creatine kinase(CK),creatine kinase-MB(CK-MB),cardiac troponin-I(cTn-I)were detected by ELISA;The activities of lactate dehydrogenase(LDH),superoxide dismutase(SOD)and aspartate aminotransferase(AST)and level of malondialdehyde(MDA)in serum were detected by kit;TUNEL staining was used to observe the apoptosis of cardiomyocytes in rats;The expressions of nuclear factor-erythroid 2-related factor 2(Nrf2),heme oxygenase-1(HO-1)and cleaved cystein-asparate protease-3(cleaved Caspase-3)in myocardial tissue were detected by immunohistochemistry.Results Compared with control group,heart weight index of rats in model group was significantly increased(P<0.001),levels of cTn-I,CKMB,CK,MDA and activities of LDH,AST in serum were significantly increased(P<0.001),SOD activity in serum was decreased(P<0.001);Obvious pathological changes were observed in myocardial tissue(P<0.001),cardiomyocyte apoptosis rate was increased(P<0.001);The expressions of cleaved Caspase-3,Nrf2 and HO-1 proteins in myocardial tissue were up-regulated(P<0.05,0.001).Compared with model group,heart weight index of rats in Longxuetongluo Capsule group was significantly decreased(P<0.001),levels of cTn-I,CK-MB,CK,MDA and activities of LDH,AST in serum were significantly decreased(P<0.05,0.01,0.001),SOD activity was significantly increased(P<0.001);The pathological changes of myocardial tissue were ameliorated(P<0.05);The cardiomyocyte apoptosis rate in myocardial tissue was significantly decreased(P<0.01);The expression of cleaved Caspase-3 in myocardial tissue was significantly decreased(P<0.05,0.01),the expressions of Nrf2 and HO-1 in myocardial tissue were significantly increased(P<0.01,0.001).Conclusion Longxuetongluo Capsule can ameliorate ISO-induced myocardial ischemia injury in rats,and its mechanism may be related to the regulation of Nrf2/HO-1 pathway,thereby inhibiting oxidative stress-induced apoptosis.