活血解毒中药配伍抑制心肌细胞自噬改善大鼠冠脉无复流的作用机制研究

Study on the mechanism of Huoxue Jiedu decoction in inhibiting cardiomyocyte autophagy and improving no-reflow of coronary artery in rats

目的探讨活血解毒中药配伍改善大鼠冠脉无复流所致心肌损伤的作用机制。方法选择96只8~10周龄SPF级雄性Wistar大鼠,随机分为假手术组、冠脉无复流模型组、阳性药对照组、活血解毒中药低、中、高剂量组。假手术组和模型组给予等体积生理盐水灌胃,阳性药物组给予等体积阿司匹林灌胃,活血解毒中药低、中、高剂量组给予相应剂量的中药灌胃。干预14天后结扎大鼠冠状动脉左前降支4小时,再灌注8小时,建立冠脉无复流心肌损伤模型。腹主动脉取血,测定血清肌酸激酶同工酶-MB(creatine kinase-MB,CK-MB)和肌钙蛋白T(cardiac troponin T,cTnT)的含量;硝基四氮唑蓝染色测量心肌梗死面积、硫磺素S染色测量心肌无复流面积;Q-PCR和Western blot检测心肌组织中微管相关蛋白轻链3(microtubule associated protein light chain 3,LC3)、自噬效应蛋白Beclin-1、B淋巴细胞瘤2(B-cell lymphoma-2,Bcl 2)基因和蛋白的表达。结果与模型组相比,活血解毒中药低、中、高剂量组大鼠血清CK-MB、心肌梗死面积、心肌无复流面积均显著减少(P<0.01);活血解毒中药低、中、高剂量组大鼠心肌组织中自噬相关基因及蛋白LC3、Beclin-1表达显著降低(P<0.01)、Bcl2表达显著增加(P<0.01)。结论活血解毒中药配伍能够减轻大鼠冠脉无复流导致的心肌损伤,其机制与抑制心肌细胞的过度自噬有关。

Objective To explore the mechanism of promoting blood circulation and detoxification Chinese medicine to improve myocardial injury caused by no-reflow of coronary artery in rats.Methods A total of 96 SPF male Wistar rats aged 8~10 weeks were randomly divided into sham operation group,no coronary reflow model group,positive drug control group,and low,medium and high dose groups of Chinese medicine for promoting blood circulation and detoxification.The sham operation group and the model group were given an equal volume of normal saline by gavage,the positive drug group was given an equal volume of aspirin by gavage,and the low,medium and high dose groups of activating blood and detoxifying Chinese medicine were given corresponding doses of Chinese medicine.After 14 days,the left anterior descending coronary artery of the rat was ligated for 4 hours and reperfused for 8 hours to establish a coronary myocardial injury model without reflow.Blood was taken from the abdominal aorta to determine the serum creatine kinase isoenzyme(CK-MB)and troponin T(cTnT)content;Thioflavin S staining to measure myocardial no-reflow area;Q-PCR and Western blot to detect myocardial tissue LC3,Beclin-1,Bcl2 gene and protein expression.Results Compared with the model group,the blood serum CK-MB and myocardial no-reflow area of the rats in the low,medium,and high dose groups of the activating blood and detoxification drugs were significantly reduced(P<0.01);the rats in the low,medium and high dose groups of the activating blood and detoxification drugs The expression of autophagy-related genes and proteins LC3 and Beclin-1 in myocardial tissue were significantly reduced(P<0.01)and the expression of Bcl2 was significantly increased(P<0.01).Conclusion The combination of activating blood and detoxifying traditional Chinese medicine can reduce myocardial injury caused by no-reflow of coronary artery in rats,and its mechanism is related to the inhibition of excessive autophagy of myocardial cells.