基于TAK1/MKK3/p38 MAPK通路探讨养心定悸胶囊防治室性心律失常的作用机制  

Mechanism of Yangxin Dingji Capsules in Preventing Ventricular Arrhythmia Based on TAK1/MKK3/p38 MAPK Pathway

作　　者：李棉 张征[1] 李欣玥 田雪 郑文璐 吴晋苇 刘刚[1] 梁文杰 LI Mian;ZHANG Zheng;LI Xinyue;TIAN Xue;ZHENG Wenlu;WU Jinwei;LIU Gang;LIANG Wenjie(The First Hospital of Hebei Medical University,Shijiazhuang 050031,China;The Third Hospital of Hebei Medical University,Shijiazhuang 050051,China;Hebei University of Chinese Medicine,Shijiazhuang 050200,China)

机构地区：[1]河北医科大学第一医院,石家庄050031 [2]河北医科大学第三医院,石家庄050051 [3]河北中医药大学,石家庄050200

出　　处：《中国实验方剂学杂志》2024年第20期86-95,共10页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：河北省重点研发计划-卫生健康领域面上项目(213777550)。

摘　　要：目的:基于转化生长因子β激活激酶1(TAK1)/丝裂原活化蛋白激酶激酶3(MKK3)/p38丝裂原活化蛋白激酶(p38 MAPK)信号通路,探讨养心定悸胶囊对异丙肾上腺素(ISO)诱导的室性心律失常大鼠心肌细胞的保护作用及机制。方法:50只SPF级雄性SD大鼠随机分为正常组、模型组、普萘洛尔组、养心定悸胶囊(中药)低、高剂量组。采用ISO“6+1”方式构建室性心律失常模型。普萘洛尔组给予普萘洛尔0.015 g·kg^(-1)·d^(-1),中药低、高剂量组给予养心定悸胶囊0.5、2 g·kg^(-1)·d^(-1),正常组和模型组给予等体积生理盐水溶液。BL-420F生物机能实验系统记录大鼠心电图变化;苏木素-伊红(HE)染色观察大鼠心脏病理形态学变化;酶联免疫吸附测定法(ELISA)检测血清肌钙蛋白(cTnI)、肌酸激酶同工酶(CK-MB)、白细胞介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)、转化生长因子-β1(TGF-β1)水平;实时荧光定量聚合酶链式反应(Real-time PCR)检测IL-1β、TNF-α的mRNA表达;免疫荧光检测活性氧(ROS)表达;蛋白免疫印迹法(Western blot)或免疫组化检测TAK1、磷酸化(p)-TAK1、MKK3、p-MKK3、p38MAPK、p-p38MAPK、核转录因子-κB(NF-κB)、p-NF-κB、IL-1β、TNF-α蛋白的表达。结果:与正常组比较,模型组大鼠心电图发生了明显室性心律失常,心律失常评分增加(P<0.01),心肌组织病理损伤明显,血清cTnI、CK-MB、IL-1β、TNF-α、TGF-β1水平升高(P<0.01);心肌组织IL-1β、TNF-α的mRNA及蛋白表达升高(P<0.01);心肌组织ROS水平增加,p-TAK1、p-MKK3、p-p38 MAPK、p-NF-κB蛋白表达升高(P<0.01);与模型组比较,普萘洛尔组和中药高剂量组大鼠持续性室性心动过速(SVT)发生减少,心律失常评分降低(P<0.05,P<0.01),心肌细胞病理损伤减轻,心肌损伤相关指标降低,血清及心肌组织炎症因子表达降低,心肌组织ROS水平降低(P<0.01),p38 MAPK通路中各分子表达均降低(P<0.05,P<0.01)。结论:p38 MAPK通路中各分�Objective:To explore the protective effect and mechanism of Yangxin Dingji capsules on isoproterenol(ISO)-induced ventricular arrhythmia in SD rat cardiomyocytes based on the transforming growth factor-βactivated kinase 1(TAK1)-mitogen-activated protein kinase kinase 3(MKK3)-p38 mitogen-activated protein kinase(p38 MAPK)signaling pathway.Method:Fifty male SPF-grade SD rats were randomly divided into a normal group,a model group,a propranolol group,a low-dose Chinese medicine group,and a high-dose Chinese medicine group.The ventricular arrhythmia model was constructed using the ISO"6+1"method.The propranolol group received propranolol at 0.015 g·kg^(-1)·d^(-1).The Chinese medicine groups received Yangxin Dingji capsules at doses of 0.5、2 g·kg^(-1)·d^(-1),respectively.The normal and model groups were given an equal volume of 0.9%NaCl solution.Electrocardiogram(ECG)changes in SD rats were recorded using the BL-420F biological function experimental system.Hematoxylin-eosin(HE)staining was used to observe the pathological changes in the heart.Serum levels of cardiac troponin I(cTnI),creatine kinase-MB(CK-MB),interleukin-1β(IL-1β),tumor necrosis factor-α(TNF-α),and transforming growth factor-β1(TGF-β1)were measured using enzyme-linked immunosorbent assay(ELISA).The mRNA expression of IL-1βand TNF-αwas assessed using real-time fluorescence quantitative polymerase chain reaction(Real-time PCR).Reactive oxygen species(ROS)expression was detected using immunofluorescence.Protein expression levels of TAK1,phosphorylated TAK1(p-TAK1),MKK3,phosphorylated MKK3(p-MKK3),p38 MAPK,phosphorylated p38 MAPK(p-p38 MAPK),nuclear factor-κB(NF-κB),phosphorylated NF-κB(p-NF-κB),IL-1β,and TNF-αwere measured using Western blot or immunohistochemistry.Result:Compared with normal group,the model group showed significant ventricular arrhythmia in ECG,with an increased arrhythmia score(P<0.01).Pathological damage to myocardial tissue was evident,and serum levels of cTnI,CK-MB,IL-1β,TNF-α,and TGF-β1 were elevated(P<0.01).

关 键 词：室性心律失常 养心定悸胶囊 转化生长因子β激活激酶1 丝裂原活化蛋白激酶激酶3 P38丝裂原活化蛋白激酶 炎症 

分 类 号：R2-0[医药卫生—中医学] R33R289R541