松果菊苷延缓人脐静脉内皮细胞衰老的机制  

Mechanism by which echinacoside delays the senescence of human umbilical vein endothelial cells

作　　者：田甜[1] 欧阳·菊艳 李瑜[1] 米叶赛·艾尼瓦尔 何娟丽 李振华 王红[1] Tian Tian;Ouyang·Juyan;Li Yu;Miyesai·Ainiwaer;He Juanli;Li Zhenhua;Wang Hong(Second Department of Comprehensive Internal Medicine,The First Affiliated Hospital of Xinjiang Medical University,Urumqi 830000,Xinjiang UygurAutonomous Region,China)

机构地区：[1]新疆医科大学第一附属医院综合内二科,新疆维吾尔自治区乌鲁木齐市830000

出　　处：《中国组织工程研究》2022年第31期5014-5019,共6页Chinese Journal of Tissue Engineering Research

基　　金：克拉玛依市科技计划项目(2017RC001A-16),项目负责人:王红;新疆医科大学研究生创新创业项目(CXCY2021008),项目负责人:田甜。

摘　　要：背景:前期工作发现同型半胱氨酸通过活性氧加速脐静脉内皮细胞衰老,此次实验初步研究影响内皮细胞衰老的相关机制。目的:探讨松果菊苷对人脐静脉内皮细胞中CPEB1、SIRT1、P53、P21基因表达的影响。方法:体外培养人脐静脉内皮细胞,加入10 g/L D-半乳糖构建细胞衰老模型,CCK-8法分析松果菊苷(0,10,20,50,100,200,300,400,500 mg/L)预处理12,24 h后的细胞增殖活性,筛选松果菊苷最佳质量浓度及干预时间;然后将人脐静脉内皮细胞分为5组:正常对照组,D-半乳糖组,松果菊苷100,200,300 mg/L组。β-半乳糖苷酶染色法检测松果菊苷预处理12 h后细胞染色阳性率,实时荧光定量-聚合酶链反应、蛋白免疫印迹法检测松果菊苷预处理12 h后人脐静脉内皮细胞中CPEB1、Sirt1、P53、P21 mRNA的表达和CPEB1、Sirt1、P21蛋白的表达。结果与结论:①CCK-8检测结果表明:松果菊苷预处理12 h的细胞存活率明显高于预处理24 h,与D-半乳糖组相比,松果菊苷100,200,300 mg/L组均不同程度提高细胞活力(P<0.0001),3组间对比差异有显著性意义(P<0.01);②与正常对照组相比,D-半乳糖组β-半乳糖苷酶染色阳性率最高(P<0.0001),100,200,300 mg/L松果菊苷预处理12 h后,可一定程度降低β-半乳糖苷酶染色阳性率(P<0.0001);③与正常对照组相比,D-半乳糖组CPEB1、P53、P21 mRNA表达水平均升高(P<0.0001),Sirt1 mRNA表达水平降低(P<0.0001);与D-半乳糖组相比,100,200,300 mg/L松果菊苷预处理12 h后,CPEB1、P53、P21 mRNA表达水平均降低(P<0.0001),Sirt1 mRNA表达水平均升高(P<0.0001);蛋白免疫印迹分析结果与实时荧光定量-聚合酶链反应分析结果一致;④结果表明,松果菊苷可能通过下调CPEB1、P53、P21表达水平,上调Sirt1表达水平,来延缓D-半乳糖诱导的人脐静脉内皮细胞衰老。BACKGROUND:Preliminary work has found that homocysteine accele rates the aging of umbilical vein endothelial cells through reactive oxygen species.This expe riment preliminarily studies the relevant mechanisms affecting endothelial cell aging.OBJECTIVE:To investigate the effects of echinacoside on the expression of cytoplasmic polyadenylate binding protein 1(CPEB1),sirtuinl(SIRT1),P53 and P21 genes in human umbilical vein endothelial cells.METHODS:Human umbilical vein endothelial cells were cultured in vitro.D-galactose 10 g/L was used to construct cell aging model.The prolife ration viability of cells treated with echinacoside(0,10,20,50,100,200,300,400,500 mg/L) for 12 and 24 hours was analyzed by CCK-8 assay,and the optimal mass concentration and inte rvention time of echinacoside were screened.Human umbilical vein endothelial cells were divided into normal control group,D-galactose group and echinacoside groups(100,200,and 300 mg/L).β-Gala ctosidase staining was used to detect the positive staining rate of cells after echinacoside pre-intervention for 12 hours.Real time RT-PCR and western-blot assay were used to detect the mRNA expression levels of CPEB1,Sirt1,P53 and P21 and the protein expression levels of CPEB1,Sirt1 and P21 in human umbilical vein endothelial cells after echinacoside pretreatment for 12 hours.RESULTS AND CONCLUSION:(1) The results of CCK-8 assay showed that the survival rate of echinacoside group at 12 hours was significantly higher than that at 24 hours.Compared with the D-galactose group,cell viability increased in the echinacoside 100,200,and 300 mg/L groups to varying degrees(P <0.0001),and there was a significant difference among the three groups(P <0.01).(2) Compared with the normal control group,the D-galactose group had the highest positive rate of β-galactosidase staining(P<0.0001).100,200,and 300 mg/L echinacoside pretreatment for 12 hours could reduce the positive rate ofβ-gala ctosidase staining to a certain extent(P <0.000 1).(3) Compared with the normal control group,the

关 键 词：松果菊苷 人脐静脉内皮细胞 衰老 CPEB1 SIRT1 P53 P21 

分 类 号：R446[医药卫生—诊断学] R318[医药卫生—临床医学]