汉黄芩素调节AMPK/NLRP3信号通路对H/R诱导的心肌细胞凋亡的影响  

Effect of Wogonin on H/R Induced Cardiomyocyte Apoptosis by Regulating the AMPK/NLRP3 Signaling Pathway

作　　者：李均[1] 徐艺[1] 冉阳 徐刚[1] 王均生[1] LI Jun;XU Yi;RAN Yang;XU Gang;WANG Jun-sheng(Department of Cardiovascular Medicine,Chongqing Emergency Medical Center(Chongqing Fourth People's Hospital/Chongqing University Central Hospital),Chongqing,400014,China)

机构地区：[1]重庆市急救医疗中心(重庆市第四人民医院/重庆大学附属中心医院)心血管内科,重庆400014

出　　处：《现代生物医学进展》2024年第17期3211-3216,3225,共7页Progress in Modern Biomedicine

基　　金：重庆市自然科学基金面上项目(cstc2021jcyj-msxmX1061)。

摘　　要：目的:探讨汉黄芩素(WOG)调节单磷酸腺苷活化蛋白激酶(AMPK)/NOD样受体蛋白3(NLRP3)信号通路对缺氧/再氧合(H/R)诱导的心肌细胞凋亡的影响。方法:将H9C2细胞分为Control组(正常培养)、H/R组(H/R诱导)、L(低剂量)-WOG组、M(中剂量)-WOG组、H(高剂量)-WOG组(在H/R诱导的基础上分别加入40、80、120μmol/L的WOG)、WOG+Compound C组(在H-WOG组基础上加入10μmol/L AMPK抑制剂Compound C)。噻唑蓝(MTT)法和5-乙炔基-2′脱氧尿嘧啶核苷(EdU)染色检测WOG对H9C2细胞增殖的影响;流式细胞术检测WOG对H9C2细胞凋亡的影响;酶联免疫吸附试验(ELISA)检测H9C2细胞血清氧化应激指标[丙二醛(MDA)、活性氧类物质(ROS)、超氧化物歧化酶(SOD)]和炎症因子[白介素(IL)-1β、IL-18]水平;蛋白免疫印迹(WB)法检测H9C2细胞AMPK、NLRP3、B细胞淋巴瘤-2(Bcl-2)、Bcl-2相关X蛋白(Bax)蛋白表达。结果:H/R组H9C2细胞的光密度值(OD490)、EdU阳性细胞率、SOD、AMPK、Bcl-2低于Control组,细胞凋亡率、IL-1β、IL-18、ROS、MDA、NLRP3、Bax高于Control组(P<0.05);与H/R组比较,L-WOG组、M-WOG组、H-WOG组OD_(490)、EdU阳性细胞率、SOD、AMPK、Bcl-2表达升高,细胞凋亡率、IL-1β、IL-18、ROS、MDA、NLRP3、Bax降低(P<0.05);WOG+Compound C组OD490、EdU阳性细胞率、SOD、AMPK、Bcl-2低于H-WOG组,细胞凋亡率、IL-1β、IL-18、ROS、MDA、NLRP3、Bax表达高于H-WOG组(P<0.05)。结论:WOG可以抑制H/R诱导的心肌细胞凋亡,其机制可能是通过介导AMPK/NLRP3信号通路有关。Objective:To investigate the effect of wogonin(WOG)on hypoxia/reoxygenation(H/R)-induced cardiomyocyte apoptosis by regulating adenosine monophosphate-activated protein kinase(AMPK)/NOD-like receptor protein 3(NLRP3)signaling pathway.Methods:H9C2 cells were divided into Control group(normal culture),H/R group(H/R induction),L(low dose)-WOG group,M(medium dose)-WOG group,H(high dose)-WOG group(40,80,120μmol/L WOG were added on the basis of H/R induction),WOG+Compound C group(10μmol/L AMPK inhibitor Compound C was added on the basis of H-WOG group).The effect of WOG on the proliferation of H9C2 cells was detected by methyl thiazolyl tetrazolium(MTT)assay and 5-ethynyl-2'-deoxyuridine(EdU)staining.The effect of WOG on apoptosis of H9C2 cells was detected by flow cytometry.The levels of serum oxidative stress indexes[malondialdehyde(MDA),reactive oxygen species(ROS),superoxide dismutase(SOD)]and inflammatory factors[interleukin(IL)-1β,IL-18]in H9C2 cells were detected by enzyme-linked immunosorbent assay(ELISA).The expression of AMPK,NLRP3,B cell lymphoma-2(Bcl-2)and Bcl-2 associated X protein(Bax)in H9C2 cells was detected by Western blot(WB).Results:The optical density(OD490),EdU positive cell rate,SOD,AMPK and Bcl-2 of H9C2 cells in H/R group were lower than those in Control group,and the apoptosis rate,IL-1β,IL-18,ROS,MDA,NLRP3 and Bax were higher than those in Control group(P<0.05).Compared with H/R group,OD_(490),EdU positive cell rate,SOD,AMPK and Bcl-2 expression in L-WOG group,M-WOG group and H-WOG group increased,while apoptosis rate,IL-1β,IL-18,ROS,MDA,NLRP3 and Bax decreased(P<0.05).The OD490,EdU positive cell rate,SOD,AMPK and Bcl-2 in WOG+Compound C group were lower than those in H-WOG group,and the apoptosis rate,IL-1β,IL-18,ROS,MDA,NLRP3 and Bax expression were higher than those in H-WOG group(P<0.05).Conclusion:WOG can inhibit H/R-induced cardiomyocyte apoptosis,which may be relate to the AMPK/NLRP3 signaling pathway.

关 键 词：黄芩素 AMPK/NLRP3信号通路 心肌细胞 细胞凋亡 

分 类 号：R-33[医药卫生] R542.22