摘要
目的探讨槲皮素(QCT)通过抑制Toll样受体4(TLR4)/核苷酸结合寡聚化结构域样受体3(NLRP3)信号通路对缺血再灌注诱发的大鼠心肌细胞凋亡的影响。方法60只SD雄性大鼠,随机选择15只作为sham假手术组,剩余45只大鼠构建心肌缺血再灌注损伤模型,造模后大鼠随机分为I/R模型组、QCT治疗组(50 mg/kg QCT)、QCT+BMS⁃986299(NLRP3激动剂)组(50 mg/kg QCT+10 mg/kg BMS⁃986299),每组15只大鼠。取材后将心肌组织固定切片,TTC染色直接观察梗死区域大小;固定心肌组织后进行HE及Masson染色观察心肌病理结构改变;收集大鼠血清进行酶联免疫吸附试验(ELISA)检测血清心肌损伤标志物(CK⁃MB、cTnI)及促炎因子(TNF⁃α、IL⁃6)水平;心肌组织切片进行TUNEL染色观察心肌细胞凋亡情况;心肌组织切片进行NLRP3的免疫组化染色观察NLRP3表达;Western blot(WB)法测定大鼠心肌组织凋亡及TLR4/NLRP3通路蛋白表达。结果与sham组比较,缺血再灌注心肌损伤组大鼠心肌组织可观察到大片梗死区域,血清中CK⁃MB、cTnI、TNF⁃α、IL⁃6水平更高,心肌组织中TUNEL+凋亡细胞比例增高同时伴随NLRP3表达增高,WB检测发现心肌组织Bax、NLRP3、ASC、Cleaved⁃caspase 1、TLR4表达增高而Bcl⁃2表达降低(P<0.05)。给药QCT治疗后,治疗组大鼠心肌梗死区域面积减小,血清中CK⁃MB、cTnI、TNF⁃α、IL⁃6水平下降,心肌组织中凋亡细胞比例及NLRP3表达均下降,心肌组织中Bax、NLRP3、ASC、Cleaved⁃caspase 1、TLR4表达下降而Bcl⁃2表达增高(P<0.05)。但给药QCT的同时给药BMS⁃986299(NLRP3激动剂)会部分抵消QCT对于I/R大鼠心肌的保护作用,使大鼠心肌梗死面积扩大并加重心肌损伤和炎症反应。结论槲皮素可能通过抑制TLR4/NLRP3通路对I/R大鼠发挥心肌保护作用。
Objective To investigate the influence of quercetin(QCT)on ischemia/reperfusion(I/R)⁃in⁃duced myocardial apoptosis in rats by the toll⁃like receptor 4(TL4)/NOD⁃like receptor protein 3(NLRP3)path⁃way.Methods Sixty SD male rats were randomly divided into sham group,I/R model group,QCT group(50 mg/kg QCT),and QCT+BMS⁃986299(NLRP3 agonist)group(50 mg/kg QCT+10 mg/kg BMS⁃986299)with 15 rats in each group.Except for the sham group,rats in other groups received temporary coronary ligation to estab⁃lish myocardial ischemia/reperfusion(I/R)model.After cardiac tissue being fixed and sliced,TCC staining was performed to observe myocardial infarction area,while HE and Masson staining were performed to evaluate myo⁃cardial pathological changes.Enzyme⁃linked immunosorbent(ELISA)assay was used to detect the levels of CK⁃MB,cTnI,TNF⁃α,IL⁃6 in rat serum.TUNEL staining was performed to assess myocardial cell apoptosis ratio,and immunohistochemical staining of NLRP3 was performed to assess NLRP3 expression in vivo.Western blot was utilized to determine the protein expression of apoptosis⁃specific and TLR4/NLRP3 pathway in rat myocardial tissue.Results Compared with the sham group,I/R rats exhibited larger myocardial infarction area,elevated se⁃rum CK⁃MB,cTnI,TNF⁃α,IL⁃6 levels,increased TUNEL+apoptosis cell ratio and enhanced NLRP3 expression in vivo;meanwhile,the protein expression of Bax,NLRP3,ASC,Cleaved⁃caspase 1,TLR4 increased,but the expression of Bcl⁃2 decreased in myocardial tissues(P<0.05).Notably,QCT treatment significantly ameliorated I/R⁃induced myocardial injury,as evidenced by decreased myocardial infarction area,reduced serum CK⁃MB,cT⁃nI,TNF⁃α,IL⁃6 levels,decreased TUNEL+apoptosis cell ratio and NLRP3 expression in vivo;compared with the I/R group,the protein expression of Bax,NLRP3,ASC,Cleaved⁃caspase 1,TLR4 decreased,and the ex⁃pression of Bcl⁃2 increased in the QCT group(P<0.05).However,BMS⁃986299(NLRP3 agonist)administration partially abrogated the prote
作者
陶真茂
杨振
郭春峰
刘秀红
TAO Zhen-mao;YANG Zhen;GUO Chun-feng;LIU Xiu-hong(Department of Cardiology,Zaoyang First Peoples Hospital,Zaoyang,Hubei Province 441200,China)
出处
《解剖学研究》
CAS
2023年第4期307-313,321,共8页
Anatomy Research
基金
湖北省卫生健康委员会科研项目(20190732)。
