七叶皂苷影响急性心肌梗死心肌损伤的机制研究

Protective effect of escin on acute myocardial infarction in rats and its mechanism

目的探讨七叶皂苷影响急性心肌梗死(AMI)心肌损伤的机制。方法60只雄性SD大鼠随机分为4组,假手术组、AMI组、AMI+低剂量七叶皂苷组(LE组)和AMI+高剂量七叶皂苷组(HE组),最终每组6只。建立AMI大鼠模型,其中假手术组只开胸不结扎。LE组和HE组在造模前分别2.10 mg/kg七叶皂苷灌胃7 d。建立大鼠心肌细胞H9C2株低氧损伤模型,设立对照组、低氧组、七叶皂苷组和模拟物组。除对照组外,其余各组均低氧(1%O_(2))培养,七叶皂苷组加入10μmol/L七叶皂苷,模拟物组加入10μmol/L七叶皂苷和20 nmol/L R-140-5P模拟物。通过超声检测各组大鼠心功能,采用TTC染色法检测心脏梗死面积,采用Western blot法检测大鼠心脏组织和H9C2细胞中B淋巴细胞瘤-2(Bcl-2)和B淋巴细胞瘤-2相关X蛋白(Bax)表达水平,采用qRT-PCR法检测miR-140-5p的mRNA表达水平。采用细胞增殖和毒性检测(CCK-8)法检测H9C2细胞活力,采用原位末端标记法(TUNEL)检测细胞凋亡水平。结果与假手术组比较,AMI组大鼠心脏左心室舒张末期内径(LVEDd)和左心室收缩末期内径(LVESd)均显著增大(均P<0.05),射血分数(EF)和左室短轴缩短率(FS)均显著降低(均P<0.05),同时Bax表达水平显著升高,Bcl-2表达水平均显著降低,心脏梗死面积增大,miR-140-5p表达水平显著增加(均P<0.05);与AMI组比较,LE组和HE组LVEDd和LVESd显著减小,EF和FS显著增加,Bax表达水平显著降低,Bcl-2表达水平显著升高,心脏梗死面积减小,miR-140-5p表达水平显著降低(均P<0.05),且HE组变化更显著(均P<0.05)。与对照组比较,低氧组miR-140-5p表达水平显著升高,细胞活力显著降低,Bax表达水平显著升高,Bcl-2表达水平显著降低,同时细胞凋亡水平升高(均P<0.05);与低氧组比较,七叶皂苷组细胞损伤得到明显改善(P<0.05);然而,miR-140-5p模拟物显著逆转了七叶皂苷对低氧心肌细胞损伤的改善作用(P<0.05)。结论七叶皂苷通过调控miR-140-5p表达改善AMI心肌损伤。

Objective To investigate the effect of escin on acute myocardial infarction(AMI)and its mechanism in rats.Methods Sixty male SD rats were randomly divided into AMI model group,sham group,AMI+low-dose escin(LE)group and AMI+high-dose escin(HE)group,each group ended up with six animals.AMI model was constructed in rats.The sham group only opened the chest without ligation,LE group and HE group were given 2 and 10 mg/kg escin 7 d by intragastric administration before modeling,respectively.The cardiac function of rats in each group was detected by ultrasonography,the infarct size was detected by TTC staining,the levels of B-cell lymphoblastoma-2(Bcl-2)and Bcl-2-associated X(Bax)protein were detected by Western blot,and the expression levels of miR-140-5p were detected by qRT-PCR.Rat cardiomyocyte H9C2 cells were divided into control group,hypoxic group,escin group and simulated group.In addition to the control group,the other 3 groups were all hypoxic(1%oxygen)culture,while 10μmol/L escin was added to the escin group,and 10μmol/L escin and 20 nmol/L miR-140-5p were added to the simulated group.The expression level of miR-140-5p were detected by qRT-PCR,cell viability was detected by CCK-8,the expression levels of Bcl-2 and Bax were detected by Western blot,and apoptosis was detected by TUNEL staining.Results In animal experiments,compared with sham group,the left ventricular end diastolic diameter(LVEDd)and left ventricular endsystolic diameter(LVESd)in AMI group were significantly increased(all P<0.05),ejection fraction(EF)and left ventricular short-axis shortening rate(FS)were significantly decreased(all P<0.05);the expression level of Bax was significantly increased,expression level of Bcl-2 was significantly decreased(P<0.05),heart infarction area was increased,miR-140-5p expression level was significantly increased(P<0.05).Compared with AMI group,LVEDd and LVESd in LE group were significantly decreased,EF and FS were significantly increased,Bax expression level was significantly decreased,Bcl-2 expression level was significantly increased,heart infarction area was decreased,and miR-140-5p expression level was significantly decreased(all P<0.05);the change of above indicators in HE group was more significant(all P<0.05).In cell experiments,compared with the control group,the hypoxia group showed a significant increase in miR-140-5p expression level,a significant decrease in cell viability,a significant increase in Bax expression level,a significant decrease in Bcl-2 expression level,and an increase in cell apoptosis level;compared with hypoxia group,the cell damage in hypoxia+escin group was significantly attenuated(P<0.05).However,miR-140-5p mimics significantly reversed the ameliorative effect of escin on hypoxic cardiomyocyte injury(P<0.05).Conclusion Escin can improve myocardial injury in rat AMI model,which is associated with the down-regulation of miR-140-5p expression in cardiomyocytes.