摘要
目的探讨白藜芦醇(Res)对低氧复氧(H/R)损伤心肌细胞的保护作用。方法采用三气培养箱法制备乳大鼠心肌细胞H/R损伤模型(H12 h/R12 h)。乳大鼠心肌细胞分为5组:细胞对照组、H/R模型组、模型+Res 5和20μmol·L^-1组(Res预处理细胞12 h,然后H12 h/R12 h)和模型+Res 20μmol·L^-1+氯喹(CQ)10μmol·L^-1组(Res预处理细胞12 h,然后含CQ的培养基H12 h/R12 h);CCK-8法检测心肌细胞存活率,比色法测定心肌细胞乳酸脱氢酶(LDH)漏出率,AnnexinⅤ/PI双染法检测细胞凋亡,免疫荧光法检测心肌细胞内微管相关蛋白1轻链3(LC3)阳性细胞率,Western印迹法检测心肌细胞Beclin 1,LC3Ⅱ和P62蛋白表达水平。结果与细胞对照组比较,H/R组心肌细胞存活率下降(P<0.01),细胞凋亡率和LDH漏出率上升(P<0.01),LC3阳性细胞率和Beclin 1表达水平无明显变化,LC3Ⅱ和P62表达水平下降(P<0.05)。与H/R组比较,Res 5和20μmol·L^-1可升高心肌细胞存活率(P<0.05),降低凋亡率和LDH漏出率(P<0.01),LC3阳性细胞率增高(P<0.05),Beclin 1,LC3Ⅱ和P62蛋白表达水平增加(P<0.05)。与模型+Res 20μmol·L^-1组比较,加用CQ组心肌细胞存活率下降(P<0.05),凋亡率升高(P<0.05),LDH漏出率增高(P<0.01),Beclin 1,LC3Ⅱ和P62蛋白水平和LC3阳性细胞率无明显变化。结论Res对H/R损伤心肌细胞具有保护作用,其机制可能与活化自噬有关。
OBJECTIVE To explore the protective effects of resveratrol(Res)against hypoxia/reox⁃ygenation injury(H/R)of myocardial cells.METHODS A newborn rat cardiomyocyte H/R model was established using the tri-gas incubator method(H for 12 h,R for 12 h).Myocardial cells of newborn rats were divided into 5 groups:cell control group,H/R model group,model+Res 5 and 20μmol·L^-1 group(cardiomyocytes were pretreated with Res for 12 h,and then processed with H12 h/R12 h)and model+Res 20μmol·L^-1+chloroquine(CQ)10μmol·L^-1 group(myocardial cells were pretreated with Res for 12 h,and then processed with H12 h/R12 h with medium containing CQ).The cell viability was measured by CCK-8 assay.The leakage rate of lactate dehydrogenase(LDH)was determined by LDH kit.The apop⁃tosis was detected by AnnexinⅤ/PI double staining.The rate of intracellular microtubule associated protein 1 light chain 3(LC3)positive cells was observed by immunofluorescence.The expressions of autophagy-related proteins such as Beclin 1,LC3Ⅱand P62 were evaluated by Western blotting.RESULTS Compared with cell control group,the cell viability in H/R group was significantly decreased(P<0.01),the apoptosis rate and leakage rate of LDH were obviously increased(P<0.01),the rate of LC3 positive cells and the protein expression of Beclin 1 did not change significantly,but LC3Ⅱand P62 protein expressions were decreased(P<0.05).Compared with H/R group,the cell viability of Res 5 and 20μmol·L^-1 group was increased(P<0.05),the apoptosis rate and leakage rate of LDH were all de⁃creased(P<0.01),the rate of LC3 positive cells was increased(P<0.05),and the protein expressions of Beclin 1,LC3Ⅱand P62 were increased(P<0.05).Compared with Res 20μmol·L^-1 group,the cell viability of Res 20μmol·L^-1+CQ 10μmol·L^-1 group was decreased(P<0.05),the apoptosis rate and leakage rate of LDH were increased(P<0.05),but there was no significant difference in protein expres⁃sions of Beclin 1,LC3Ⅱand P62 or in the rate of LC3 positive cells.CONCLUSION Res could pr
作者
张金芳
刘梦思
孙秀玉
刘立亚
李志勇
黄秀兰
ZHANG Jin-fang;LIU Meng-si;SUN Xiu-yu;LIU Li-ya;LI Zhi-yong;HUANG Xiu-lan(School of Pharmacy,Minzu University of China,Beijing 100081,China;College of Life and Environmental Sciences,Minzu University of China,Beijing 100081,China)
出处
《中国药理学与毒理学杂志》
CAS
北大核心
2020年第8期584-590,共7页
Chinese Journal of Pharmacology and Toxicology
基金
国家自然科学基金(81173656)
国家自然科学基金(81470182)。
关键词
白藜芦醇
心肌细胞
低氧复氧损伤
自噬
resveratrol
myocardial cells
hypoxia/reoxygenation injury
autophagy
