摘要目的 探讨酪醇对糖尿病大鼠心肌缺血再灌注损伤的影响及沉默调节蛋白1(SIRT1)/腺苷酸活化蛋白激酶(AMPK)/内皮型一氧化氮合酶(eNOS)信号通路在其中的作用.方法 SPF级健康成年雄性SD大鼠,体重200～220 g,采用腹腔注射1％链脲佐菌素60 mg/kg制备糖尿病模型.取糖尿病模型制备成功的大鼠56只,采用随机数字表法分为4组(n=14):假手术组(S组)、心肌缺血再灌注组(I/R组)、心肌缺血再灌注+酪醇组(I/R+T组)和心肌缺血再灌注+酪醇+SIRT1抑制剂EX527组(I/R+T+E组).I/R+T组和I/R+T+E组灌胃法连续45 d给予酪醇20 mg·kg-1·d-1,其他2组给予等体积生理盐水.I/R+T+E组于缺血前连续3d腹腔注射EX527 5 mg·kg-1·d-1,并于再灌注前20 min时腹腔注射EX527 5 mg/kg.采用结扎左冠状动脉前降支30 min,再灌注2h的方法制备心肌缺血再灌注损伤模型.采用TTC法测量心肌梗死体积,ELISA法检测血清CK-MB、LDH、15-F2t-isoprostane和心肌组织SOD水平,Western blot法检测心肌组织SIRT1、AMPK、磷酸化AMPK(p-AMPK)、eNOS和磷酸化eNOS(p-eNOS)的表达水平.结果 与S组比较,I/R组血清CK-MB、LDH和15-F2t-Isoprostane的水平升高,心肌梗死体积增加,心肌组织SOD活性和SIRT1表达水平降低,I/R+T组和I/R+T+E组血清CK-MB、LDH和15-F2t-Isoprostane的水平升高,心肌梗死体积增加,心肌组织SOD活性和SIRT1表达水平降低,p-AMPK和p-eNOS表达上调(P＜0.05);与I/R组比较,I/R+T组血清CK-MB、LDH和15-F2t-Isoprostane的水平降低,心肌梗死体积减少,心肌组织SOD活性和SIRT1、p-AMPK、p-eNOS的表达水平升高(P＜0.05),I/R+T+E组上述指标差异无统计学意义(P＞0.05);与I/R+T组比较,I/R+T+E组血清CK-MB、LDH和15-F2t-Isoprostane的水平升高,心肌梗死体积增加,心肌组织SOD活性和SIRT1、p-AMPK、p-eNOS表达水平降低(P＜0.05).结论 酪醇可减轻糖尿病大鼠心肌缺血再灌注损伤,其机制与激活SIRT1/AMPK/eNOS信号通路,抑制氧化应激反应有关.

abstractsObjective To evaluate the effect of tyrosol on myocardial ischemia-reperfusion (I/R) injury in diabetic rats and the role of silent mating-type information regulation 1 (SIRT1)/adenosine monophosphate-activated protein kinase (AMPK)/endothelial nitric oxide synthase (eNOS) signaling pathway.Methods SPF healthy adult male Sprague-Dawley rats,weighing 200-220 g,were intraperitoneally injected with streptozotocin 60 mg/kg to establish the model of diabetes mellitus.Fifty-six diabetic rats were divided into 4 groups (n =14 each) using a random number table method:sham operation group (S group),myocardial I/R group (I/R group),myocardial I/R plus tyrosol group (I/R+T group),and myocardial I/R plus tyrosol plus SIRT1 inhibitor EX527 group (I/R+T+E group).In I/R+T and I/R+T+E groups,tyrosol 20 mg · kg-1 · d-1 was given by gavage for 45 consecutive days,and the equal volume of normal saline was given in the other two groups.In I/R+T+E group,EX527 5 mg · kg-1 · d-1 was intraperitoneally injected for 3 consecutive days before ischemia,and EX527 5 mg/kg was intraperitoneally injected at 20 min before repeffusion.Myocardial I/R was induced by ligation of the left anterior descending branch of coronary artery for 30 min followed by 2-h reperfusion.The myocardial infarct volume was measured by TTC staining.The levels of creatine kinase-MB (CK-MB),lactic dehydrogenase (LDH) and 5-F2t-isoprostane in serum and superoxide dismutase (SOD) in myocardial tissues were detected by enzyme-linked immunosorbent assay.The expression of SIRT1,AMPK,phosphorylated AMPK (p-AMPK),eNOS and p-eNOS was detected by Western blot.Results Compared with S group,the levels of serum CK-MB,LDH and 15-F2t-Isoprostane and myocardial infarction volume were significantly increased,the SOD activity was decreased,and the SIRT1 expression was down-regulated in I/R group,and the levels of serum CKMB,LDH and 15-F2t-Isoprostane and myocardial infarction volume were significantly increased,the SOD activity was decreased,the SIRT1 expression was down-regulated,and the expression of p-AMPK and peNOS was up-regulated in I/R+T and I/R+T+E groups (P＜0.05).Compared with I/R group,the levels of serum CK-MB,LDH and 15-F2t-Isoprostane and myocardial infarction volume were significantly decreased,the SOD activity was increased,and the expression of SIRT1,p-AMPK and p-eNOS was up-regulated in I/R+T group (P＜0.05),and no significant change was found in the parameters mentioned above in I/R+ T+E group (P＞0.05).Compared with I/R+T group,the levels of CK-MB,LDH and 15-F2t-isoprostane in serum and myocardial infarct volume were significantly increased,the SOD activity was increased,and the expression of SIRT1,p-AMPK and p-eNOS was down-regulated in I/R+T+E group (P＜0.05).Conclusion Tyrosol can mitigate myocardial I/R injury,and the mechanism may be related to activating SIRT1/AMPK/eNOS signaling pathway and inhibiting oxidative stress response in diabetic rats.