铁死亡在支气管哮喘气道重塑中的作用

doi:10.6040/j.issn.1671-7554.0.2024.0467

摘要/Abstract

摘要： 目的通过建立卵清蛋白(ovalbumin, OVA)诱导的哮喘小鼠模型,研究铁死亡在支气管哮喘气道重塑发病中的作用,为治疗哮喘气道重塑提供新的靶点。方法将24只雌性C57BL/6小鼠随机分为4个组:对照组、哮喘模型组(OVA组)、干预组(Fer-1组)及Fer-1联合哮喘组(Fer-1+OVA组),每组6只,通过建立OVA诱导哮喘小鼠模型联合公共数据库筛选的对照组和OVA诱导的OVA组小鼠肺组织差异表达谱,探索铁死亡与哮喘的联系。应用铁离子检测试剂盒和普鲁士蓝铁染色法对小鼠肺组织中的铁表达进行测定,通过免疫印记及免疫组化评估肺组织谷胱甘肽过氧化物酶4(glutathione peroxidase 4, GPX4)蛋白的表达,检测小鼠肺组织脂质过氧化代谢物丙二醛(malondialdehyde, MDA)的表达,透射电子显微镜观察小鼠气道上皮超微结构改变,应用MASSON组织染色法对小鼠气道上皮下胶原沉积进行观察。为了进一步验证铁死亡抑制剂对哮喘气道重塑的影响,给予哮喘模型小鼠铁死亡抑制剂干预后,对气道周围胶原沉积情况及肺组织GPX4蛋白表达进行评估,并分析气道重塑指标的改变。结果生物信息学分析显示,哮喘小鼠肺组织中铁死亡通路较对照组明显富集,OVA组小鼠气道上皮中GPX4表达较对照组显著降低,表明铁死亡可能参与哮喘的发病过程。OVA组小鼠肺组织中铁含量增加,气道上皮GPX4表达降低,肺组织MDA表达升高,气道上皮下胶原沉积增加,伴有小鼠气道上皮特征性超微结构改变,表明铁死亡与哮喘气道重塑密切相关。铁死亡抑制剂干预哮喘小鼠后,气道上皮GPX4表达升高,气道上皮下胶原沉积减轻,气道重塑标志蛋白表达降低,上皮标志蛋白表达升高。结论铁死亡参与支气管哮喘气道重塑的发病过程,铁死亡抑制剂Fer-1可减轻哮喘气道重塑的结构改变,为治疗支气管哮喘气道重塑提供新靶点。

关键词: 哮喘, 气道重塑, 铁死亡, Ferrostatin-1, 治疗靶点

Abstract: Objective To explore the contribution of ferroptosis to airway remodeling in asthma by establishing a ovalbumin(OVA)-induced asthma mice model, and to provide new directions in the treatment of airway remodeling of asthma. Methods Twenty-four female C57BL/6 mice were stochastically divided into four groups: the control group(n=6), the OVA group(n=6), the ferrostatin(Fer)-1 group(n=6), and the Fer-1+OVA group(n=6). The association between ferroptosis and asthma was clarified by establishing the differentially expressed genes of lung tissues in the OVA-induced asthma mice and controls screened from public databases. The iron concentration was measured by iron detection kit and Perls stain, and the expression of glutathione peroxidase 4(GPX4)protein was evaluated by Western blotting and immunohistochemical staining. The levels of the lipid peroxidation product malondialdehyde(MDA)in lung tissue were assayed and the ultra-structural changes in the airway epithelium were observed using transmission electron microscopy(TEM). Lung tissue slices were stained with Masson staining to evaluate collagen deposition. To further verify the effect of ferroptosis inhibitor on the airway remodeling in asthma, the mice were intervened with Fer-1, and then collagen deposition was evaluated by MASSON staining, the expression of GPX4 protein was detected by immunofluorescence and Western blotting. Results The results of bioinformatics analysis showed that the ferroptosis signaling pathway was significantly enriched in OVA group, suggesting that ferroptosis played a certain role in the pathogenesis of asthma. Meanwhile, in this gene set, GPX4 was significantly decreased in OVA group. In asthma mice model, collagen deposition was accompanied by a series of ferroptosis features such as increased iron concentration, decreased GPX4 expression, MDA accumulation, increased collagen deposition, and characteristic ultra-structural changes in the airway epithelium, suggesting that ferroptosis was intimately linked with asthma airway remodeling. After treated with Fer-1, for the asthma model mice, the GPX4 expression was elevated, the collagen deposition was reducted, the expressions of airway remodeling indicators were suppressed, and epithelial cell markers were increased. Conclusion Ferroptosis participates in the pathogenesis of asthma airway remodeling, ferroptosis inhibitor Fer-1 attenuates airway remodeling in asthma, and the application of Fer-1 provides a new therapeutic target for asthmatic airway remodeling.

Key words: Asthma, Airway remodeling, Ferroptosis, Ferrostatin-1, Therapeutic target

中图分类号:

R562.2+5

孙丛丛,崔文静,张锦涛,张东,刘晓菲,潘云,亓倩,徐嘉蔚,曾荣,郭红喜,董亮. 铁死亡在支气管哮喘气道重塑中的作用[J]. 山东大学学报 (医学版), 2024, 62(7): 1-9.

SUN Congcong, CUI Wenjing, ZHANG Jintao, ZHANG Dong, LIU Xiaofei, PAN Yun, QI Qian, XU Jiawei, ZENG Rong, GUO Hongxi, DONG Liang. Roles of ferroptosis in asthmatic airway remodeling[J]. Journal of Shandong University (Health Sciences), 2024, 62(7): 1-9.

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