Nrf2/HO-1通路在甘草查尔酮A诱导的小鼠心肌损伤保护中的作用

Licochalcone A protects the heart against acute myocardial infarction through the activation of the NRF2/HO-1 signaling pathway

目的探讨甘草查尔酮A(Lico A)对急性心肌梗死(AMI)模型小鼠心肌损伤的保护作用及其机制。方法将雄性C57BL/6小鼠随机分为假手术组、心肌梗死(心梗)组和低、中、高剂量药物组以及核因子红系2-相关因子2(Nrf2)特异性抑制剂(ML385)组,每组5只。造模3 d测各组小鼠心功能后,取心肌组织进行苏木素伊红(HE)染色评估心肌组织坏死程度;免疫荧光法检测心肌组织活性氧(ROS)生成,酶标法检测心肌组织中谷胱甘肽(GSH)和丙二醛(MAD)含量,以及过氧化氢酶(CAT)活力;免疫印迹检测心肌组织中血红素加氧酶-1(HO-1)及Nrf2的表达,免疫组织化学方法进一步验证心肌组织中Nrf2的表达及定位情况。结果与假手术组相比,手术组小鼠心肌组织坏死显著,但给予Lico A治疗后,各剂量组小鼠心肌组织坏死程度及梗死面积明显减小。检测心肌局部氧化应激情况显示,Lico A治疗后,心肌组织ROS水平较心梗手术组显著降低,GSH含量升高、CAT活力增加同时MAD含量降低(P均<0.05)。免疫印迹分析显示,与假手术组相比,心梗组小鼠心肌组织总蛋白中HO-1和核蛋白中Nrf2表达轻度升高,而Lico A治疗组小鼠心肌组织总蛋白中HO-1及核蛋白中NRF2表达显著增高(P<0.05)。免疫组化结果同样证明Lico A可促进Nrf2核转移(P<0.05)。给予Nrf2特异性抑制剂ML385后,Lico A的上述保护作用被明显抑制。结论Lico A可通过激活Nrf2/HO-1通路来发挥抗氧化应激效应,从而有效地防止心梗后心肌的损伤。

Objective To investigate the protective effect and its mechanism of Licochalcone A(Lico A)on myocardial injury in mice with acute myocardial infarction(AMI).Methods Male C57BL/6 mice were randomly divided into Sham control group,acute myocardium infarction(AMI)group,three dose Lico A treatment group and Nrf2 inhibitor ML385 group(n=5).Cardiac function of each group was measured 3 days after modeling.The myocardial tissue was stained with hematoxylin eosin(HE)to evaluate the necrosis degree.Immunofluorescence assay was used to detect reactive oxygen species(ROS)production of myocardial tissue.The glutathione(GSH)and malondialdehyde(MAD)contents and catalase(CAT)activity in myocardium were detected by enzyme natation.The expressions of heme oxygenase-1(HO-1)and Nrf2 were detected by Western blot,and the expression and localization of Nrf2 in myocardial tissue were further verified by immunohistochemistry.Results Compared with sham group,myocardial tissue infarction was obvious in AMI group,and the myocardial infarction area and necrosis degree in the treatment group were significantly reduced due to Lico A(P<0.05).After Lico A treatment,ROS level of myocardial tissue was significantly lower than that of the AMI group(P<0.05),GSH expression increased,CAT activity increased and MAD expression decreased(all P<0.05).Western blot results showed that compared with the AMI group,the expression of HO-1 in total protein and Nrf2 in nuclear protein of mice in the Lico A treatment group was significantly increased(P<0.05).Immunohistochemical results showed that a large amount of Nrf2 expression was observed in the nucleus after Lico A treatment.The above protective effects of Lico A were inhibited by the administration of ML385.Conclusion Lico A can protect AMI mice from oxidative stress by activating Nrf2/HO-1 pathway.