基于NLRP3信号通路探讨活血通脉颗粒对急性心肌梗死大鼠免疫炎症紊乱的干预作用  

Intervention effect of Huoxue Tongmai Granule on immune inflammation disorders in acute myocardial infarction rats based on NLRP3 signaling pathway

作　　者：韩玉泽[1] 田峥[2] 钟雷[3] HAN Yu-ze;TIAN Zheng;ZHONG Lei(Department of Cardiology, Dalian Friendship Hospital, Liaoning 116001, China;Department of Cardiology, Affiliated Zhongshan Hospital of Dalian University, Liaoning 116001, China;Department of Cardiology, The First Affiliated Hospital of Dalian Meidcal University, Liaoning 116001, China)

机构地区：[1]大连市友谊医院心内二科,辽宁116001 [2]大连大学附属中山医院心内科,辽宁116001 [3]大连医科大学附属第一医院心内科,辽宁116001

出　　处：《中国医药生物技术》2019年第5期436-444,共9页Chinese Medicinal Biotechnology

摘　　要：目的探讨活血通脉颗粒对急性心肌梗死(AMI)大鼠在免疫炎症紊乱的干预作用。方法大鼠 42 只随机选 6 只为空白对照组,其余 36 只大鼠 AMI 造模。建模后存活大鼠 30 只随机分成 5 组,阳性药物对照组,AMI 组,活血通脉颗粒低、中、高剂量组,每组 6 只。空白对照组、AMI 组每天灌胃生理盐水,阳性药物对照组 184 mg/(kg·d)灌胃辛伐他汀,活血通脉颗粒低、中、高剂量组按生药 2.7、5.4、10.8 g/(kg·d)灌胃,每日 1 次,连续 28 d。同时建立人 H9C2 心肌细胞缺氧模型,缺氧处理 21 h,加药处理。HE 染色观察心肌组织形态学变化;MTS 检测心肌细胞活力变化;ELISA 检测心肌组织和心肌细胞中 IL-1β、TNF-α水平;qRT-PCR 检测心肌组织中核苷酸结合寡聚化结构域样受体蛋白 3(NLRP3)、接头蛋白 ASC(ASC)、半胱氨酸天冬氨酸蛋白水解酶 1(caspase-1)mRNA 水平;蛋白免疫印迹检测心肌组织中 NLRP3、ASC、半胱氨酸天冬氨酸蛋白酶酶原(pro-caspase-1)、caspase-1 蛋白水平。结果大鼠和人心肌细胞模型中,AMI 组心肌细胞破坏、死亡严重,形态及结构损害严重。随着活血通脉颗粒剂量的增加,心肌细胞坏死现象减弱,逐渐恢复形态。大鼠模型中,与空白对照组相比,AMI 组心肌组织中 IL-1β、TNF-α、NLRP3、ASC、caspase-1 mRNA 和蛋白、pro-caspase-1 蛋白表达量升高(P < 0.05)。与 AMI 组相比,活血通脉颗粒低剂量组心肌组织中 ASC、pro-caspase-1、caspase-1 蛋白表达量降低;活血通脉颗粒中、高剂量组心肌组织中 IL-1β、NLRP3、ASC、caspase-1 mRNA 和蛋白、pro-caspase-1 蛋白表达量降低,并呈剂量依赖效应(P < 0.05)。人心肌细胞中加药 6 h 时 IL-1β、TNF-α的变化与大鼠模型中类似。结论活血通脉颗粒可能通过减弱 NLRP3 信号通路、抑制促炎因子表达,实现对 AMI 大鼠的保护作用。Objective To investigate the effect of Huoxue Tongmai Granule (HXTMG) on immune inflammation disorders in rats with acute myocardial infarction (AMI). Methods Six from forty-two rats were randomly selected as blank control group, and the rest thirty-six rats were made as models by AMI. Thirty surviving rats were randomly divided into five groups: positive drug control group, AMI group, low, middle and high-dose HXTMG groups, with six rats in each group. The blank control group and the AMI group were given normal saline daily for intragastric administration. The positive drug control group was given 184 mg/(kg·d) of simvastatin for intragastric administration, and the low, medium and high-dose HXTMG groups were given 2.7, 5.4, 10.8 g/(kg·d) of crude drug, respectively, once a day for 28 days. Human hypoxia model of H9C2 cardiomyocytes was established, and treated with hypoxia for 21 h, followed by treatment with drugs. HE staining was used to observe the histomorphological changes of myocardium. MTS was used to detect changes in myocardial cell viability. The levels of IL-1β and TNF-α in myocardium were detected by ELISA. Quantitative RT-PCR was used to detect the levels of nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3), apoptosis-associated speck-like proteincontain a caspase activation and recruitment domain (ASC), and cysteinyl aspartate specific proteinase 1 (caspase-1) mRNA in myocardium. The levels of NLRP3, ASC, cysteine-containing aspartate-specific-1 protease (pro-caspase-1) and caspase-1 proteins in myocardium were detected by Western blot. Results In the rat and cardiomyocyte models, the AMI group was severely damaged, with significant morphological and structural damages. With the increase of dose of HXTMG, the necrosis of myocardial cells is weakened and the morphology is gradually restored. In the rat model, expressions of IL-1β, TNF-α, NLRP3, ASC, caspase-1 mRNA and protein along with pro-caspase-1 protein in the myocardium of AMI group was increased as compared with

关 键 词：心肌梗死 活血通脉颗粒 免疫炎症 核苷酸结合寡聚化结构域样受体蛋白3信号通路 

分 类 号：R285.5[医药卫生—中药学] R-332[医药卫生—中医学]