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丹酚酸对心肌缺血再灌注损伤大鼠的影响及相关机制

[The effects and related mechanism of salvianolate on rats with myocardial ischemia-reperfusion injury].

作者信息

Yue R C, Yang X L, Zhang R Y, Liu S, Liu J, Zeng J, Liang H, Wang W, Hu H X, Zeng C Y

机构信息

Department of Cardiology, North Sichuan Medical College First Affiliated Hospital, Nanchong 637000, China.

出版信息

Zhonghua Xin Xue Guan Bing Za Zhi. 2017 Dec 24;45(12):1072-1077. doi: 10.3760/cma.j.issn.0253-3758.2017.12.012.

Abstract

To investigate the effect and related mechanism of salvianolate on myocardial ischemia-reperfusion (I/R) injury in rats. Thirty-six adult Sprague-Dawley rats were divided into three groups (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. (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 (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 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 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 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 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. 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.

摘要

探讨丹酚酸对大鼠心肌缺血再灌注(I/R)损伤的影响及相关机制。将36只成年Sprague-Dawley大鼠采用随机数字表法分为三组(每组12只):对照组(冠状动脉未结扎)、I/R组(通过结扎并开放左冠状动脉前降支建立心肌I/R模型)、丹酚酸+I/R组(再灌注时经尾静脉注射5mg/kg丹酚酸)。采用氯化三苯基四氮唑(TTC)染色法测量心肌梗死面积。采用酶联免疫吸附测定(ELISA)法检测心肌坏死标志物,包括心肌肌钙蛋白T(TnT)、肌酸激酶同工酶MB(CK-MB)和乳酸脱氢酶(LDH)。采用末端脱氧核苷酸转移酶介导的dUTP缺口末端标记(TUNEL)染色法分析凋亡水平。采用蛋白质免疫印迹法分析裂解的半胱天冬酶-3蛋白水平。采用冷发光法检测心肌组织的三磷酸腺苷(ATP)水平。采用免疫荧光法检测心肌组织中8-羟基-2'-脱氧鸟苷(8-OHdG)水平。(1)对照组、I/R组和丹酚酸+I/R组的梗死面积分别为0、(23.90±5.66)和(12.06±5.97)(P<0.01或P<0.05)。(2)对照组、I/R组和丹酚酸+I/R组的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)、(1135.8±180.6)和(390.3±121.5)U/L,LDH水平分别为(119.0±58.6)、(1838.6±543.8)和(631.6±2,28.3)U/L,丹酚酸+I/R组均显著低于I/R组(均P<0.01)。(3)对照组、I/R组和丹酚酸+I/R组TUNEL阳性心肌细胞率分别为(1.07±1.16)、(21.36±4.11)和(13.30±3.67)(均P<0.01)。(4)对照组、I/R组和丹酚酸+I/R组裂解的半胱天冬酶-3表达分别为0.11±0.05、0.84±0.20和0.43±0.09(均P<0.01)。(5)对照组、I/R组和丹酚酸+I/R组心肌组织的ATP水平分别为(100.0±0.0)、(34.2±9.2)和(62.1±18.0)(均P<0.01)。(6)对照组心肌组织几乎无8-OHdG表达。I/R组心肌组织8-OHdG表达高于对照组。丹酚酸+I/R组心肌组织8-OHdG表达低于I/R组。丹酚酸可能通过减轻线粒体DNA氧化损伤、保护线粒体功能及抑制心肌细胞凋亡来减轻大鼠心肌I/R损伤。

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