人参皂苷Rb1改善自噬流抗离体大鼠心脏心肌缺血再灌注损伤  

Ginsenoside Rb1 improving autophagic flux against myocardial ischemia reperfusion injury in isolated-heart of rat

作　　者：李洋 姜永良 陆地 董川书 普俞维 杨萍 孙林[2] LI Yang;JIANG Yong-liang;LU Di;DONG Chuan-shu;PU Yu-wei;YANG Ping;SUN Lin(Department of Anatomy and Histology,Kunming Medical University College of Basic Medicine,Kunming 650500,China;Department of Cardiology,the Second Affiliated Hospital of Kunming Medical University,Kunming 650101,China;Kunming Medical University Biomedical Engineering Research Center,Kunming 650500,China)

机构地区：[1]昆明医科大学基础医学院人体解剖学与组织学胚胎学系,昆明650500 [2]昆明医科大学第二附属医院心内科,昆明650101 [3]昆明医科大学生物医学工程研究中心,昆明650500

出　　处：《解剖学报》2020年第2期265-272,共8页Acta Anatomica Sinica

基　　金：国家自然科学基金(81760087,81560050);云南省应用基础研究重点资助项目[2017FA035,2017FE467(-008)];云南省应用基础研究(昆医联合专项)[2017FE468(-182)]。

摘　　要：目的通过Langendorff系统构建离体大鼠心肌缺血/再灌注(I/R)损伤模型,探讨人参皂苷Rb1对心肌I/R损伤的保护作用及机制。方法选取健康成年雄性SD大鼠60只,随机分为假手术组、I/R组和人参皂苷Rb1(1、5、10和20μmol/L)预处理组,每组10只。大鼠开胸、结扎主动脉后,取出心脏置于Langendorff系统进行灌流,按照分组情况构建相应模型。采用Lab Chart电生理系统检测心率(HR)、左心室发展压(LVDP)、左室压上升/下降最大速率(±dp/dtmax)等相关心功能指标;采用TTC染色法检测心肌梗死面积;采用Western blotting检测Beclin 1、LC3、p62和Lamp 2表达情况;采用免疫组织化学法检测Beclin 1表达情况。结果人参皂苷Rb1(1、5、10和20μmol/L)改善由I/R损伤导致的LVDP和±dp/dtmax的下降,减少心肌梗死面积,其中10μmol/L浓度人参皂苷Rb1对心功能保护作用最显著(P<0.05)。Beclin 1在I/R组心肌细胞胞质中阳性表达率显著增高,加入人参皂苷Rb1(10μmol/L)后表达量减少(P<0.05)。与sham组相比,在I/R组发现自噬流受损:自噬相关蛋白Beclin 1、LC3和p62表达水平升高,Lamp 2表达水平下调(P<0.05)。加入人参皂苷Rb1(10μmol/L)后上述蛋白的调控作用被反转,自噬流通畅(P<0.05)。结论人参皂苷Rb1预处理通过改善自噬流受损发挥抗大鼠离体心肌I/R损伤的保护作用,其中以10μmol/L浓度的人参皂苷Rb1保护作用最好。Objective To explore the protective effect of ginsenoside Rb1 on the myocardial ischemia/reperfusion(I/R)injury in rats in vitro.Methods Totally 60 adult male SD rats were randomly divided into 6 groups:sham group,I/R group,ginsenosde Rb1 pretreatment groups(at the doses of 1μmol/L,5μmol/L,10μmol/L and 20μmol/L,respectively),10 in each group.The Langendorff perfusion system was used to establish I/R model.The Lab Chart electrophysiological system was used to monitor real-time heart function by monitoring heart rate(HR),left ventricular development pressure(LVDP)and left ventricular development pressure(±dp/dtmax).TTC staining method was used to measure myocardial infarct size.The Western blotting were used to assay Beclin 1,LC3,p62 and Lamp 2 expression,respectively.The immunohistochemistry were used to assay Beclin 1 expression.Results Ginsenoside Rbl of all the four different concent rations improved the decrease of LVDP and±dp/dtmax arising from myocardial I/R injury.Meanwhile,ginsenoside Rbl significantly decreased the area of cardial infarction.Ginsenoside Rb1(10μmol/L)precondition group protected the heart most significantly(P<0.05).The expression of Beclin 1 with I/R increased significantly in the cytoplasm of cardiomyocytes.Moreover,Beclin 1 expression decreased after addition pretreatment with ginsenoside Rb1(10μmol/L)(P<0.05).Compared with sham group,we found that the autophagic flux was impaired in I/R group which the expression of Beclin 1,LC3 and p62 increased significantly,as well as the expression of Lamp 2 decreased significantly.On the other hand,pretreatment with ginsenoside Rb1(10μmol/L)could reverse impaired autophagic flux(P<0.05).Conclusion Ginsenoside Rbl demonstrates pharmacological preconditioning effect and protects against myocardial I/R injury by improving damaged-autophagy flux,the dose of 10μmol/L precondition protectes the heart most significantly.

关 键 词：离体心脏 心肌缺血再灌注损伤 自噬流 人参皂苷RB1 Langendorff系统 免疫印迹法 大鼠 

分 类 号：R541.4[医药卫生—心血管疾病]