二甲双胍对缺血再灌注损伤大鼠心肌的保护作用及其机制

Protective effect of metformin on myocardial ischemia-reperfusion injury in rats and its mechanism

目的探讨二甲双胍对缺血再灌注损伤大鼠心肌的保护作用及其机制。方法30只缺血再灌注损伤SD大鼠随机数字表法分为假手术组、模型组和二甲双胍组,模型组和二甲双胍组大鼠采用结扎大鼠冠状动脉左前降支再灌注构建心肌缺血再灌注损伤模型。二甲双胍组大鼠给予饮用水喂食二甲双胍10 mg/(kg·d),假手术组和模型组给予饮用水,连续治疗1周后,采用苏木精-伊红(HE)染色分析心肌病理变化;采用放射免疫法分析血清乳酸脱氢酶酶(LDH)和肌酸激酶(CK)的水平;酶联免疫吸附试验(ELISA)法检测心肌白细胞介素-6(IL-6)和肿瘤坏死因子-α(TNF-α)水平,采用蛋白质印迹法(Western blot)分析3组大鼠心肌腺苷酸活化蛋白激酶(AMPK)和沉默信息调节因子2相关酶1(SIRT1)表达水平。组间比较采用单因素方差分析。结果二甲双胍组大鼠LDH和CK水平[(1150.20±128.69)、(185.16±12.11)U/L]明显低于模型组大鼠[(784.21±69.58)、(35.84±4.02)U/L],差异有统计学意义(t=14.530、22.330,P<0.05)。二甲双胍组大鼠血清TNF-α和IL-6水平[(36.70±5.92)、(68.96±11.34)pg/ml]明显低于模型组大鼠,差异有统计学意义(t=13.000、12.021,P<0.05)。二甲双胍组大鼠心肌细胞凋亡比例[(13.96±2.94)%]明显低于模型组大鼠[(39.46±3.14)%],差异有统计学意义(t=19.680,P<0.05)。二甲双胍组大鼠心肌组织磷酸化AMPK(p-AMPK)水平(1.51±0.26)明显高于模型组和对照组(0.76±0.06、0.67±0.07),差异有统计学意义(t=3.005、7.331,P>0.05)。二甲双胍组大鼠心肌组织中SIRT1表达水平(2.38±0.32)明显高于模型组(1.04±0.09、0.98±0.09),差异有统计学意义(t=3.005、7.331,P>0.05)。结论二甲双胍通过AMPK/SIRT1通路抑制大鼠心肌缺血再灌注损伤,减轻炎性反应并发挥心肌保护作用。

Objective To investigate the protective effect of metformin on myocardial ischemia-reperfusion injury in rats and its mechanism.Methods Totally,30 SD rats with ischemia-reperfusion injury were randomly divided into sham operation group,model group and metformin group by a random number table.Rats in model group and metformin group were subjected to myocardial ischemia-reperfusion injury model by ligating the left anterior descending coronary artery of rats.The rats in metformin group were given drinking water and metformin 10 mg/(kg·d),and the sham operation group and model group were given drinking water only.After continuous treatment for 1 week,the hematoxylin-eosin(HE)staining was used to analyze the pathological changes of myocardium.The levels of serum lactate dehydrogenase(LDH)and creatine kinase(CK)were analyzed by radioimmunoassay.The myocardial interleukin-6(IL-6)and tumor necrosis factor-α(TNF-α)were analyzed by enzyme linked immunosorbent assay(ELISA).The expression levels of adenosine monophosphate-actived protein kinase(AMPK)and silent information regulator 2 homolog 1(SIRT1)in the myocardium of the three groups were analyzed by Western blotting.One way ANOVA was used for comparison between groups.Results The levels of LDH and CK in metformin group[(1150.20±128.69),(185.16±12.11)U/L]were significantly lower than those in model group[(784.21±69.58),(35.84±4.02)U/L,t=14.530,22.330,P<0.05].Serum TNF-αand IL-6 levels in metformin group[(36.70±5.92),(68.96±11.34)pg/ml]were significantly lower than those in the model group(t=13.000,12.021,P<0.05).The apoptosis rate of cardiomyocytes in metformin group[(13.96±2.94)%]was significantly lower than that in model group[(39.46±3.14)%,t=19.680,P<0.05].The level of phosphorylated AMPK(p-AMPK)in myocardial tissue of metformin group(1.51±0.26)was significantly higher than that in model group and control group(0.76±0.06,0.67±0.07,t=3.005,7.331,P>0.05).The expression level of SIRT1 in myocardial tissue of metformin group(2.38±0.32)was significantly higher than that in model group(1.04±0.09,0.98±0.09,t=3.005,7.331,P>0.05).Conclusion Metformin inhibits myocardial ischemia-reperfusion injury in rats through AMPK/SIRT1 pathway,reduces inflammatory response and plays a myocardial protective role.