红景天苷调控线粒体自噬抵抗冠状动脉内皮细胞缺血再灌注损伤

Salidroside protects coronary endothelial cells against ischemia-reperfusion injury by regulating mitophagy

目的探讨红景天苷(Sal)对心肌梗死后冠状动脉内皮细胞(CoECs)线粒体自噬的调控机制。方法CoECs分为正常对照组(Control组)、阴性对照组(PBS组)、红景天苷组(Sal组)、红景天苷+氯喹组(Sal+CQ组),其中Control组用常氧处理,其余3组进行氧糖剥夺及恢复处理(OGD/R);CCK-8法、荧光探针法、Western Blot检测细胞活性、凋亡、线粒体溶酶体共定位、线粒体膜电位(MMP)、ROS及自噬相关蛋白表达。18只C57/BL6小鼠随机分为假手术(Sham组)、缺血再灌注组(MIRI组)和缺血再灌注+红景天苷组(MIRI+Sal组),其中MIRI组、MIRI+Sal组在LAD结扎手术前后28 d分别腹腔注射生理盐水、50 mg/(kg·d)Sal;利用小动物超声成像系统、Masson染色和Western Blot检测射血分数(EF)、左室短轴缩短率(FS)、心肌纤维化和小鼠梗死冠状动脉中自噬相关蛋白表达。结果与PBS组比较,Sal组细胞活力、MMP、线粒体溶酶体共定位和PINK1、Beclin1、Parkin表达增加,而细胞凋亡率、ROS含量和Mtfr1、P62、LC3Ⅱ/LC3Ⅰ表达下降(P<0.05);与Sal组比较,Sal+CQ组细胞活力、线粒体溶酶体共定位和PINK1、Beclin1、Parkin表达下降,而细胞凋亡率、ROS含量和Mtfr1、P62、LC3Ⅱ/LC3Ⅰ表达增加(P<0.05)。与MIRI组比较,Sal+MIRI组FS、EF和梗死区域冠状动脉PINK1、Parkin和Beclin1表达增加,而心肌纤维化和Mtfr1、P62/SQSTM1和LC3Ⅱ/LC3Ⅰ表达下降(P<0.05)。结论Sal对OGD/R的CoECs和MIRI小鼠梗死区域冠状动脉内皮具有抑制氧化应激和促进线粒体自噬的保护作用。

Objective To investigate the modulation of salidroside(Sal)on mitophagy in coronary endothelial cells(CoECs)after myocardial infarction.Methods CoECs were divided into normal control,negative control(PBS),Sal,and Sal+Chloroquine(Sal+CQ)groups;Control group was treated with normoxic condition,while the other groups with oxygen/glucose deprivation and reperfusion(OGD/R).Cell viability,apoptosis,mitochondrial-lysosomal colocalization(MLCL),mitochondrial membrane potential(MMP),ROS,and autophagy-related protein levels were detected using CCK-8,fluorescent probe,and Western blot.Eighteen C57/BL6 mice were randomized to Sham,ischemia-reperfusion(MIRI),and MIRI+Sal groups.MIRI or MIRI+Sal group was intraperitoneally injected with normal saline or 50 mg/(kg·d)Sal 28 days before and after LAD ligation.Ejection fraction(EF),left ventricular fractional shortening(FS),myocardial fibrosis,and expression of autophagy-related proteins in infarcted coronary arteries were determined by small animal ultrasound imaging system,Masson stain and Western blot.Results Compared with PBS group,cell viability,MMP,MLCL,and expression of PINK1,Beclin1,and Parkin were increased(P<0.05),while apoptosis,ROS content,and expression of Mtfr1,P62,and LC3 Ⅱ/Ⅰ decreased in Sal Group(P<0.05).Compared with Sal group,cell viability,MLCL and expression of PINK1,Beclin1,and Parkin were reduced(P<0.05),while apoptosis,ROS content and expression of Mtfr1,P62,and LC3 Ⅱ/Ⅰ elevated(P<0.05)in Sal+CQ group.Compared with MIRI group,FS,EF and expression of PINK1,Parkin and Beclin1 were increased in infarcted coronary arteries,while myocardial fibrosis and expression of Mtfr1,P62/SQSTM1 and LC3 Ⅱ/Ⅰ decreased(P<0.05)in Sal+MIRI group.Conclusion Sal is protective for OGD/R-induced CoECs and infarcted coronary arteries in MIRI mice by inhibiting oxidative stress and promoting mitophagy.