Objective: To investigate the effect and related mechanism of salvianolate on myocardial ischemia-reperfusion (I/R) injury in rats. Methods: Thirty-six adult Sprague-Dawley rats were divided into three groups (n=12 each) using random number table method: control group (coronary artery was not ligated) , I/R group (myocardial I/R model was established by ligation and opening of left anterior descending coronary artery) , and salvianolate+I/R group (5 mg/kg of salvianolate was injected through the tail vein at the time of reperfusion) .Triphenyltetrazolium chloride (TTC) stain was utilized to measure the myocardial infarct size. The ELISA method was used to detect myocardial necrotic markers, including cardiac troponin T (TnT) , creatine kinase-MB (CK-MB) and lactate dehydrogenase (LDH) . Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining was used to analyses the levels of apoptosis. The levels of cleaved Caspase-3 protein were analyzed with Western blot.Cold luminescence method was used to detect the ATP level of myocardial tissue. The levels of 8-hydroxy-2'-deoxyguanosine (8-OHdG) in myocardial tissue were detected by immunofluorescence. Results: (1) The infarct size in control group, I/R group and salvianolate+I/R group were 0, (23.90±5.66) %, and (12.06±5.97) %, respectively (P<0.01 or 0.05) . (2) The TnT level was (0.04±0.03) , (16.96±2.80) , and (6.95±2.31) ng/ml, the CK-MB level was (43.6±23.5) , (1 135.8±180.6) ,and (390.3±121.5) U/L, the LDH level was (119.0±58.6) , (1 838.6±543.8) , and (631.6±228.3) U/L in control group, I/R group and salvianolate+I/R group, all significantly lower in salvianolate+I/R group than in I/R group (all P<0.01) . (3) The rate of TUNEL positive myocardial cells were (1.07±1.16) %, (21.36±4.11) %,and (13.30±3.67) % in control group, I/R group,and salvianolate+I/R group (all P<0.01) . (4) The cleaved Caspase-3 expression was 0.11±0.05, 0.84±0.20,and 0.43±0.09 in control group, I/R group, and salvianolate+I/R group (all P<0.01) . (5) The ATP level of myocardial tissue was (100.0±0.0) %, (34.2±9.2) %,and (62.1±18.0) % respectively in control group, I/R group, and salvianolate+I/R group (all P<0.01) . (6) There was almost no 8-OHdG expression in the myocardial tissue of control group. The expression of 8-OHdG in the myocardial tissue of I/R group was greater than that of the control group. The expression of 8-OHdG in the myocardial tissue of salvianolate+I/R group was less than that of the I/R group. Conclusion: Salvianolate may alleviate myocardial I/R injury of rat through reducing the mitochondrial DNA oxidative damage, protecting mitochondrial function and inhibiting the apoptosis of myocardial cells.
目的： 探讨丹参多酚对大鼠心肌缺血再灌注损伤的作用及机制。 方法： 采用随机数字表法，将36只成年Sprague-Dawley大鼠分为对照组（不结扎冠状动脉）、缺血再灌注组（I/R组，通过结扎和打开冠状动脉左前降支建立大鼠心肌缺血再灌注损伤模型）和丹参多酚+I/R组（在缺血再灌注的同时，经尾静脉注射丹参多酚5 mg/kg），每组均为12只。通过2，3，5-氯代三苯基四氮唑（TTC）染色检测各组心肌梗死面积；采用ELISA法检测心肌坏死标志物，包括血清肌钙蛋白T（TnT）、肌酸激酶同工酶（CK-MB）和乳酸脱氢酶（LDH）水平；采用TUNEL法检测心肌组织的细胞凋亡情况；采用Western blot检测各组心肌组织中活化含半胱氨酸的天冬氨酸蛋白水解酶3（Caspase-3）蛋白表达水平；采用冷发光法检测心肌组织三磷酸腺苷（ATP）水平；采用免疫荧光法检测心肌组织8-羟基脱氧鸟苷（8-OHdG）水平。 结果： （1）对照组、I/R组和丹参多酚+I/R组的心肌梗死面积分别为0、（23.90±5.66）%和（12.06±5.97）%，I/R组大于对照组（P<0.01），而丹参多酚+I/R组小于I/R组（P<0.05）。（2）对照组、I/R组和丹参多酚+I/R组血清TnT水平分别为（0.04±0.03）、（16.96±2.80）和（6.95±2.31）ng/ml，CK-MB水平分别为（43.6±23.5）、（1 135.8±180.6）和（390.3±121.5）U/L，LDH水平分别为（119.0±58.6）、（1 838.6±543.8）和（631.6±228.3）U/L，I/R组均高于对照组，而丹参多酚+I/R组均低于I/R组（P均<0.01）。（3）对照组、I/R组和丹参多酚+I/R组TUNEL阳性心肌细胞比率分别为（1.07±1.16）%、（21.36±4.11）%和（13.30±3.67）%，I/R组高于对照组，丹参多酚+I/R组低于I/R组（P均<0.01）。（4）对照组、I/R组和丹参多酚+I/R组活化Caspase-3蛋白表达水平分别为0.11±0.05、0.84±0.20和0.43±0.09，I/R组高于对照组，丹参多酚+I/R组低于I/R组（P均<0.01）。（5）对照组、I/R组和丹参多酚+I/R组心肌组织ATP水平分别为（100.0±0.0）%、（34.2±9.2）%和（62.1±18.0）%，I/R组低于对照组，丹参多酚+I/R组高于I/R组（P均<0.01）。（6）对照组心肌组织中几乎无8-OHdG表达；I/R组心肌组织中8-OHdG表达多于对照组；丹参多酚+I/R组心肌组织中8-OHdG表达少于I/R组。 结论： 丹参多酚可能是通过减轻线粒体DNA氧化损伤，保护线粒体功能，抑制心肌细胞凋亡，从而减轻大鼠心肌的缺血再灌注损伤。.
Keywords: Mitochondria; Myocardial reperfusion injury; Salvianolate.