南方医科大学学报

南方医科大学学报 ›› 2022, Vol. 42 ›› Issue (2): 256-262.doi: 10.12122/j.issn.1673-4254.2022.02.13

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抑制铁死亡可减轻脓毒症小鼠的心肌损伤：脂钙蛋白-2的作用

黄毓慧，张共鹏，梁 欢，曹珍珍，叶红伟，高 琴   

  1. 蚌埠医学院生理学教研室，蚌埠医学院临床医学院，蚌埠医学院第一附属医院呼吸与危重症医学科，安徽 蚌埠 233000
  • 出版日期:2022-02-20 发布日期:2022-03-16

Inhibiting ferroptosis attenuates myocardial injury in septic mice: the role of lipocalin-2

HUANG Yuhui, ZHANG Gongpeng, LIANG Huan, CAO Zhenzhen, YE Hongwei, GAO Qin   

  1. Department of Physiology, Department of Clinical Medicine, Bengbu Medical College, Bengbu 233000, China; Department of Respiratory and Critical Care Medicine, First Affiliated Hospital of Bengbu Medical College, Bengbu 233000, China
  • Online:2022-02-20 Published:2022-03-16

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摘要： 目的 观察铁死亡在盲肠结扎与穿孔（CLP）法诱导的脓毒症小鼠心肌损伤中的作用，并探讨脂钙蛋白-2（Lcn2）在铁死亡中的可能作用。方法 选取8周龄雄性C57BL/6小鼠，采用CLP法诱导脓毒症心肌损伤模型。小鼠随机分为3组（10只/组）：假手术组、脓毒症组（CLP，小鼠接受CLP手术）、脓毒症+铁死亡抑制剂Ferrostain-1组（CLP+Fer-1，小鼠腹腔注射浓度为5 mg/mL的Fer-1 5 mg/kg，1 h后接受CLP手术）。各组小鼠术后24 h通过超声心动图检测小鼠心功能。H&E染色观察心肌损伤，透射电镜观察心肌纤维细微结构和线粒体的变化；ELISA法测定血清中炎症因子肿瘤坏死因子-α（TNF-α）水平；组织铁试剂盒测定心肌组织铁含量的变化；Western blot法检测心肌组织中Lcn2蛋白和铁死亡相关蛋白谷胱甘肽过氧化物酶4（GPX4）和铁死亡抑制蛋白1（FSP1）的表达变化。结果 与假手术组相比，CLP术后24 h，脓毒症小鼠心脏收缩与舒张功能减弱，左心室射血分数（LVEF%）、左心室缩短分数（LVFS%）和左心室舒张末期内径（LVIDd）降低（P<0.05）；左心室收缩期末期内径（LVIDs）升高（P<0.05）。与CLP组相比，CLP+铁死亡抑制剂Fer-1组LVEF%、LVFS%和LVIDd升高（P<0.05）；LVIDs降低（P<0.05）。光镜下观察到CLP小鼠心肌纤维排列不整齐，部分变性，有炎症细胞浸润，间质水肿，横纹模糊，红细胞渗出。透射电镜观察到部分线粒体嵴减少，外膜破裂，部分线粒体变小，膜密度增高。CLP+Fer-1组小鼠心肌形态学有改善，线粒体损伤减轻。与假手术组相比，CLP组血清TNF-α水平、心肌组织铁含量、Lcn2蛋白表达升高（P<0.01），GPX4、FSP1蛋白表达降低（P<0.01）。与CLP组相比，CLP+Fer-1组血清TNF-α水平、心肌组织铁含量、和Lcn2蛋白表达降低（P<0.05）；GPX4、FSP1蛋白表达升高（P<0.05）。结论 来源于GPX4、FSP1不同途径的铁死亡参与CLP引起的脓毒症性心肌损伤的发生，抑制铁死亡可减轻脓毒症心肌损伤，Lcn2可能参与其中。

关键词: 脓毒症；心肌损伤；铁死亡；ferrostatin-1；脂钙蛋白-2

Abstract: Objective To explore the contribution of ferroptosis to myocardial injury in mouse models of sepsis and the role lipocalin-2 (Lcn2) in ferroptosis. Methods Adult male C57BL/6 mice were randomized equally into sham-operated group, cecal ligation and puncture (CLP)-induced sepsis group, and CLP + Fer-1 group where the mice received intraperitoneal injection of 5 mg/mL Fer-1 (5 mg/kg) 1 h before CLP. The left ventricular functions (including LVEF%, LVFS%, LVIDd and LVIDs) of the mice were assessed by echocardiography at 24 h after CLP. Myocardial injury in the mice was observed with HE staining, and the changes of myocardial ultrastructure and mitochondria were observed using transmission electron microscopy (TEM). Serum TNF-α level was measured with ELISA, and the changes of myocardial iron content were detected using tissue iron kit. The protein expressions of myocardial Lcn2, glutathione peroxidase 4 (GPX4) and ferroptosis suppressor protein 1 (FSP1) were determined with Western blotting. Results The septic mice showed significantly decreased LVEF%, LVFS% and LVIDd and increased LVIDs at 24 h after CLP (P<0.05), and these changes were significantly improved by Fer-1 treatment. Sepsis caused obvious myocardial pathologies and changes in myocardial ultrastructure and mitochondria, which were significantly improved by Fer-1 treatment. Fer-1 treatment also significantly ameliorated sepsis-induced elevations of serum TNF-α level, myocardial tissue iron content, and Lcn2 protein expression and the reduction of GPX4 and FSP1 protein expression levels (P<0.05). Conclusion GPX4- and FSP1-mediated ferroptosis are involved in myocardial injury in mice with CLP-induced sepsis, and inhibition of ferroptosis can attenuate septic myocardial injury, in which Lcn2 may play a role.

Key words: sepsis; myocardial injury; ferroptosis; ferrostatin-1; lipocalin-2