白藜芦醇抑制大鼠高原缺氧心肌损伤的作用及机制研究

The effects and mechanism of resveratrol on myocardial injury induced by plateau hypoxia

目的探究白藜芦醇抑制大鼠高原缺氧心肌损伤作用及机制。方法选用8周龄雄性Wistar大鼠,将其随机分为常氧对照组(0.5%羧甲基纤维素钠,n=15)、低氧暴露组(0.5%羧甲基纤维素钠,n=15)、白藜芦醇干预组(低氧暴露+400 mg/kg白藜芦醇,n=15)。各组动物预先灌胃15 d,低氧暴露组和白藜芦醇干预组转至低压氧舱进行低氧干预(固定海拔高度6000 m,20 h/d,30 d)。小动物超声仪测定超声心动图。干预完成后,腹主动脉取血,分离心脏用于后续检测。TUNEL染色检测心肌细胞凋亡情况;DHE染色检测心脏组织活性氧(ROS)水平;TBA法和比色法检测血清超氧化物歧化酶(SOD),谷胱甘肽过氧化物酶(GSH-Px)水平。酶联免疫吸附法测定血清中白介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)水平。免疫荧光染色测定自噬标志蛋白LC3和线粒体外膜蛋白TOMM20。结果与常氧对照组相比,低氧暴露组大鼠心脏指数显著升高(P<0.05),心室壁增厚,心腔容积减少,射血分数和左心室缩短分数降低(P<0.05);与低氧暴露组相比,白藜芦醇干预组心脏指数显著下降(P<0.05),室壁增厚现象有所改善,心腔容积显著扩大,射血分数和左心室缩短分数显著回升(P<0.05)。与常氧对照组相比,低氧暴露组细胞凋亡率升高;与低氧暴露组相比,白藜芦醇干预组细胞凋亡显著减少,差异均有统计学意义(P<0.05)。与常氧对照组相比,低氧暴露组ROS水平显著升高;与低氧暴露组相比,白藜芦醇干预组ROS水平显著降低,差异均有统计学意义(P<0.05)。与常氧对照组相比,低氧暴露组大鼠血清中SOD,GSH-Px含量显著降低;与低氧暴露组相比,白藜芦醇干预组大鼠血清中抗氧化酶SOD,GSH-Px含量显著升高,差异均有统计学意义(P<0.05)。与常氧对照组相比,低氧暴露组大鼠血清中炎症因子IL-6、TNF-α水平显著升高;与低氧暴露组比,白藜芦醇干预组血清中IL-6、TNF-α水平显著降低,差异均有统计学意义(P<0.05)。与常氧对照组相比,低氧暴露组LC3表达量显著升高,TOMM20表达量显著降低,自噬及线粒体自噬水平显著增强;与低氧暴露组相比,白藜芦醇干预组LC3表达量降低,TOMM20表达量升高,自噬及线粒体自噬水平减弱,差异均有统计学意义(P<0.05)。结论白藜芦醇可显著缓解长期低氧暴露引发的心肌损伤,推测白藜芦醇可通过调节氧化应激介导的自噬及线粒体自噬发挥对缺氧心肌细胞的保护作用。

Objective To explore the inhibiting effects and mechanism of resveratrol on myocardial injury induced by plateau hypoxia in rats.Methods Wistar rats(8 weeks old)were randomly divided into normal oxygen control group(0.5%sodium carboxymethylcellulose,n=15),hypoxia group(0.5%sodium carboxymethylcellulose,n=15),and resveratrol administration group(hypoxia exposure,400 mg/kg resveratrol,n=15).After lavage for 15 days,the animals in each group were transferred to lowpressure oxygen chamber for hypoxia intervention(fixed altitude 6000 m,20 h/d,30 d).Echocardiography was determined by small animal ultrasonic apparatus.After the intervention,blood was taken from the abdominal aorta and the heart was separated for subsequent detection.TUNEL staining was used to detect myocardial cell apoptosis.ROS levels in heart tissue were detected by DHE staining.Serum superoxide dismutase(SOD)and glutathione peroxidase(GSH-Px)levels were detected by TBA and colorimetry.Serum levels of interleukin-6(IL-6)and tumor necrosis factor-α(TNF-α)were determined by enzyme-linked immunosorbent assay.Autophagy marker protein LC3 and mitochondrial outer membrane protein TOMM20 were determined by immunofluorescence.Results Compared with the normal oxygen control group,the cardiac index of the hypoxia exposure group significantly increased(P<0.05),the ventricular wall was thickened,the volume of the heart cavity significantly decreased,the ejection fraction and fractional shortening significantly decreased(P<0.05);compared with the hypoxic exposure group,the heart index in the resveratrol administration group significantly decreased(P<0.05),the ventricular wall thickening was improved,the cardiac volume significantly enlarged,and the ejection fraction and fractional shortening significantly increased(P<0.05).Compared with the normal oxygen control group,the apoptosis rate of the hypoxic exposure group significantly increased;compared with the hypoxia group,the apoptosis rate of the resveratrol administration group significantly reduced(P<0.05).Compared with the normal oxygen control group,ROS levels in the hypoxia group significantly increased(P<0.05);compared with the hypoxia group,ROS levels in the resveratrol administration group significantly decreased(P<0.05).Compared with the normal oxygen control group,the serum SOD,GSH-Px levels of the hypoxic exposure group significantly decreased;compared with the hypoxic exposure group,the serum SOD,GSH-Px levels of the resveratrol administration group significantly increased(P<0.05).Compared with the normal oxygen control group,the serum levels of inflammatory cytokines IL-6 and TNF-αin the low oxygen exposure group significantly increased(P<0.05);compared with the hypoxic exposure group,the serum IL-6 and TNF-αlevels of the resveratrol administration group significantly decreased(P<0.05).Compared with the normal oxygen control group,LC3 expression level significantly increased,TOMM20 expression levels significantly decreased,autophagy and mitophagy levels significantly enhanced in the hypoxia exposure group;compared with the hypoxia exposure group,LC3 expression level significantly decreased,TOMM20 expression level significantly increased,autophagy and mitochondrial autophagy levels significantly weakened in the resveratrol administration group(P<0.05).Conclusion Resveratrol can significantly relieve myocardial injury caused by long-term hypoxic exposure,and it is speculated that resveratrol plays a protective role in hypoxic cardiomyocytes by regulating autophagy and mitophagy mediated by oxidative stress.