丹参饮颗粒抑制NLRP3炎症小体抗大鼠心肌缺血再灌注损伤

Danshenyin Granules Inhibit NLRP3 Inflammasome Activation to Attenuate Myocardial Ischemia Reperfusion Injury in Rats

目的:观察丹参饮颗粒对心肌缺血再灌注损伤的作用及其通过氧化应激和NLRP3炎症小体活化途径的作用机制。方法:采用冠状动脉左前降支结扎制备大鼠心肌缺血再灌注模型,随机分为模型对照组、丹参饮颗粒1 g/kg组、3 g/kg组和9 g/kg组,缺血1 h后进行再灌注,另设假手术对照组。于再灌注前0.5 h和再灌注20 h时分别灌胃给予丹参饮颗粒溶液或纯水。再灌注24 h后检测大鼠心功能,测定心肌梗死范围、血清心肌酶谱、心肌炎症因子、超氧化物歧化酶(SOD)活性、丙二醛(MDA)含量和总抗氧化能力(T-AOC),心肌冰冻切片染色观察活性氧(ROS)含量,免疫印迹分析NLRP3炎症小体活化水平。结果:与假手术对照组比较,模型对照组LVESD明显升高,而EF和FS亦急剧降低,梗死区面积明显增加,血清中AST、LDH、CK-MB、MDA含量明显升高(P<0.05或P<0.01),SOD活性和T-AOC活力显著降低,心肌组织TNF-α、IL-1β和IL-6含量显著升高,ROS水平明显升高,NLRP3、凋亡相关斑点样蛋白(ASC)、成熟型半胱天冬酶-1(cleaved Caspase-1)和白介素-1β(cleaved IL-1β)蛋白表达显著上调(P<0.05或P<0.01)。与模型对照组比较,丹参饮颗粒3 g/kg、9 g/kg组大鼠心功能明显提高,心肌梗死范围减小,血清心肌酶水平、心肌组织炎症因子和ROS水平显著降低,MDA含量降低,而SOD和T-AOC活力升高(P<0.05或P<0.01),还明显下调NLRP3、ASC、cleaved Caspase-1和cleaved IL-1β蛋白水平。结论:丹参饮颗粒具有显著的抗心肌缺血再灌注损伤作用,可能与其抑制氧化应激、抑制NLRP3炎症小体活化和缓解炎症有关。

Objective: To investigate the anti-myocardial ischemia-reperfusion injury(MIRI) effect of Danshenyin granules, and study its oxidative stress mechanism and the activation of NLRP3 inflammasome. Methods: Rat myocardial ischemia reperfusion model was established by the method of ligation of the left anterior descending coronary artery. Then model rats were randomly divided into I/R group, 1 g/kg, 3 g/kg and 9 g/kg Danshenyin granules groups. Reperfusion was performed 1 h after ischemia, Danshenyin granules were administered 0.5 h before reperfusion and 20 h after reperfusion. 24 h after reperfusion, cardiac function, myocardial infarct index, serum myocardial enzyme and myocardial inflammatory factors were measured. The activity of superoxide dismutase(SOD), the content of malondialdehyde(MDA) and total antioxidant capacity(T-AOC) were detected. The content of reactive oxygen species(ROS) was observed in the frozen section, and the activation level of NLRP3 inflammasome was analyzed by Western blot. Results: Compared with the control group, the LVESD was significantly increased in the model group, the levels of EF and FS were decreased, the infarct area was significantly increased, the serum levels of AST、LDH、CK-MB and MDA were significantly increased(P<0.05 or P<0.01), the activity of SOD and T-AOC was decreased, the contents of TNF-α、IL-1β and IL-6 in cardiac tissues were increased, the ROS level was increased, the protein expressions of NLRP3、ASC、cleaved Caspase-1 and cleaved IL-1β were significantly up regulated(P<0.05 or P<0.01). In 3 g/kg and 9 g/kg Danshenyin granules groups, the cardiac function was improved, the myocardial infarct area was reduced, the level of myocardial enzyme, myocardial inflammatory factors and ROS level was significantly decreased, the serum level of MDA was decreased, while the levels of SOD and T-AOC were increased. Danshenyin granules also significantly reduced the protein levels of NLRP3, apoptosis associated speck like protein containing a C-terminal caspase recruitment domain(ASC), cleaved caspase-1 and cleaved IL-1β. Conclusion: Danshenyin granules have certain protective effect against MIRI in rats, which may be related to suppressing oxidative stress, alleviating the activation of NLRP3 inflammasome and its anti-inflammation effect.