摘要
目的研究黄芪多糖(APS)对缺氧/复氧(H/R)所致乳鼠心肌细胞损伤的保护作用及其机制。方法分离并体外培养乳鼠心肌细胞3 d后随机分为五组,正常组、H/R组、APS低、中、高剂量(20、40、80μmol/mL)组;给药干预30 min后,H/R组和APS各剂量组给予缺氧2.5 h后复氧处理。复氧2 h后,CCK-8法检测细胞增殖抑制率,Annexin V-FITC/PI染色法检测细胞凋亡水平,生化分析法检测培养液中乳酸脱氢酶(LDH)、肌酸激酶(CK)、肌酸激酶同工酶(CK-MB)水平,酶联免疫吸附(ELISA)法检测培养液中N末端脑钠肽前体(NT-proBNP)、肿瘤坏死因子-α(TNF-α)、白细胞介素-1β(IL-1β)水平,Western blot法检测糖调节蛋白78(GRP78)、CCAAT/增强子结合蛋白同源蛋白(CHOP)、激活型半胱氨酸蛋白酶-12(cleved caspase-12)、B细胞淋巴瘤基因-2(bcl-2)、bcl-2相关X蛋白(Bax)、核因子-κB(NF-κB)蛋白表达。结果与正常组比较,H/R组细胞增殖抑制率和凋亡率升高(P<0.01),培养液中LDH、CK、CK-MB水平和NT-proBNP、TNF-α、IL-1β水平升高(P<0.01);GRP78、CHOP、Cleved Caspase-12、Bax、NF-κB蛋白表达上调而bcl-2表达下调(P<0.01),bcl-2/Bax比值降低(P<0.01)。与H/R组比较,APS中、高剂量细胞增殖抑制率和凋亡率降低(P<0.01),培养液中LDH、CK、CK-MB水平和NT-proBNP、TNF-α、IL-1β水平降低(P<0.05);GRP78、CHOP、Cleved Caspase-12、Bax、NF-κB蛋白表达下调而bcl-2表达上调(P<0.01),bcl-2/Bax比值升高(P<0.01)。结论APS可以减轻H/R诱导的乳鼠心肌细胞损伤,可能与APS抑制内质网应激通路介导的细胞凋亡和炎症反应有关。
Objective To investigate the protective effect of astragalus polysaccharide(APS)on hypoxia/reoxygenation(H/R)induced cardiomyocyte damage in neonatal rats and its mechanism.Methods The neonatal rat cardiomyocytes were isolated and cultured in vitro for 3 days and randomly divided into 5 groups,normal group,H/R group and APS low-,medium-and high-dose(20,40,80μmol/mL)group.After the drugs were given for 30min,the cell in H/R group and the APS groups were given hypoxia for 2.5 hours and then reoxygenation.2h after reoxygenation,the cell proliferation inhibition rate was detected by CCK-8 method,the cell apoptosis level was detected by Annexin V-FITC/PI staining;the content of lactate dehydrogenase(LDH),phosphokinase(CK)and phosphokinase isoenzyme(CK-MB)in culture fluid were detected by biochemical analysis;the level of N-terminal brain natriuretic peptide(NT-proBNP),tumor necrosis factor-α(TNF-α)and interleukin-1β(IL-1β)in culture medium were detected by enzyme-linked immunosorbent assay(ELISA)method;the expression of glycoregulated protein 78(GRP78),CCAAT/enhancer binding protein homologous protein(CHOP),activated caspase-12(Cleved Caspase-12),B cell lymphoma gene-2(bcl-2),bcl-2 related X protein(Bax),nuclear factor-κB(NF-κB)were detected by Western blot.Results Compared with normal group,the proliferation inhibition rate and apoptosis rate in H/R group increased(P<0.01);the content of LDH,CK,CK-MB and the level of NT-proBNP,TNF-α,IL-1βin culture medium increased(P<0.01);the expression of GRP78,CHOP,Cleved Caspase-12,Bax,NF-κB up-regulated and the expression of bcl-2 down-regulated(P<0.01),and the ratio of bcl-2/Bax decreased(P<0.01).Compared with H/R group,the proliferation inhibition rate and apoptosis rate in APS medium-and high-dose groups decreased(P<0.01);the content of LDH,CK,CK-MB and the level of NT-proBNP,TNF-α,IL-1βin culture medium decreased(P<0.05);the expression of GRP78,CHOP,Cleved Caspase-12,Bax,NF-κB down-regulated and the expression of bcl-2 up-regulated(P<0.01),and the ratio of bcl-2/Bax increased(P<0.01).Conclusion APS can alleviate H/R-induced cardiomyocyte damage in neonatal rats,which may be related to the inhibition of endoplasmic reticulum stress pathway-mediated apoptosis and inflammation.
作者
沈玉珏
Shen Yujue(Second Department of Cardiology,Handan Central Hospital,Handan 056000,China)
出处
《心脑血管病防治》
2021年第5期441-444,479,共5页
CARDIO-CEREBROVASCULAR DISEASE PREVENTION AND TREATMENT
基金
邯郸市科学技术研究与发展计划项目(1623208070ZC)。
关键词
黄芪多糖
心肌细胞
缺氧/复氧
内质网应激
凋亡
Astraglus polysaccharides
Myocardial cells
Hypoxia/reoxygenation
Endoplasmic reticulum stress
Apoptosis
