基于铁死亡探讨补肾益气方治疗哮喘小鼠的作用机制

Mechanism of action of Bushen Yiqi formula in treatment of asthma based on ferroptosis in mice

中文关键词: 支气管哮喘    铁死亡    氧化性应激    炎症    卵清蛋白

英文关键词: bronchial asthmaferroptosisoxidative stressinflammationovalbumin

中文摘要:

      目的 探讨补肾益气方（BSYQF）对支气管哮喘小鼠模型的治疗效果及其作用机制。方法 通过卵清蛋白（OVA）致敏与激发建立Balb/c小鼠哮喘模型，将40只小鼠随机分为正常对照组（NC组）、哮喘模型组（A组）、BSYQF低剂量组（BSYQF/L组）、BSYQF高剂量组（BSYQF/H组）及铁死亡抑制剂组（Fer-1组），每组8只。采用肺功能仪检测各组小鼠的肺力学功能，包括气道阻力和肺动态顺应性;通过苏木精-伊红（HE）和过碘酸希夫（PAS）染色观察各组肺组织中炎症细胞浸润、杯状细胞化生及黏液分泌情况;通过血细胞分类计数仪计数各组支气管肺泡灌洗液（BALF）中炎症细胞的数量;通过生化法及普鲁士蓝-二氨基联苯胺（Perls-DAB）染色法分别检测肺组织中Fe²⁺铁含量及Fe³⁺铁沉积情况;采用酶联免疫吸附试验检测肺组织中丙二醛（MDA）与谷胱甘肽（GSH）的含量。采用单因素方差分析或重复测量双因素方差分析进行数据统计，组间比较使用Tukey检验。结果 A组小鼠气道阻力较NC组明显增加（P<0.01），肺动态顺应性大幅降低（P<0.01）;经BSYQF与Fer-1干预后各组小鼠气道阻力下降（P<0.05），肺动态顺应性提升（P<0.05）; HE及PAS染色提示A组小鼠肺组织炎症细胞浸润较明显（P<0.001）,BSYQF与Fer-1干预后各组小鼠炎症细胞浸润改善（P<0.01），杯状细胞化生及黏液高分泌状态减轻; A组小鼠组BALF中各组炎症细胞（淋巴细胞、嗜酸性粒细胞及单核细胞）数量均明显增多（均P<0.01）,BSYQF与Fer-1干预后各组小鼠炎症细胞数量均有所减少（P<0.05）; Fe²⁺含量检测及Perls-DAB染色提示A组小鼠肺组织存在Fe²⁺铁过载（P<0.01）与Fe³⁺铁沉积现象，BSYQF/H与Fer-1可有效降低Fe²⁺铁含量（P<0.05）并减轻Fe³⁺沉积; A组小鼠MDA水平较NC组明显升高（P<0.01）,GSH水平降低（P<0.001）,BSYQF/H与Fer-1干预后MDA水平明显降低（P<0.05 ）; BSYQF与Fer-1组GSH的表达提升（P<0.01）。结论 BSYQF与Fer-1对哮喘小鼠模型展现出了良好的治疗效果，可能通过调节铁代谢与抑制脂质过氧化来抑制铁死亡，从而减轻气道炎症与上皮损伤，在调控炎症方面显示出了多靶点协同作用的潜力。BSYQF/H组有效降低肺组织Fe²⁺含量、减轻Fe³⁺沉积，并调节氧化应激指标（降低MDA、提升GSH），提示其具有抑制铁死亡的作用。

英文摘要:

      Objective To investigate the therapeutic effects and mechanism of action of Bushen Yiqi formula (BSYQF) in a mouse model of bronchial asthma. Methods An allergic asthma model was established in Balb/c mice through ovalbumin (OVA)sensitization and challenge. 40 mice were randomly divided into five groups (n=8 per group):normal control group (NC group),model group (group A),BSYQF low-dose group (BSYQF/L group),BSYQF high-dose group (BSYQF/H group),and ferroptosis inhibitor group (Fer-1 group). Pulmonary mechanical function,including airway resistance and lung dynamic compliance,were measured using a pulmonary function apparatus. Hematoxylin and eosin (HE)and periodic acid-schiff (PAS)staining were used to observe inflammatory cell infiltration,goblet cell metaplasia,and mucus secretion in lung tissue.The number of inflammatory cells in bronchoalveolar lavage fluid(BALF)was counted with an automated hematology analyzer.The Fe2+content and Fe3+deposition in lung tissue were detected by biochemical assay and Perls-DAB staining,respectively.The levels of malondialdehyde(MDA)and glutathione(GSH)in lung tissue were measured by enzyme-linked immunosorbent assay.Data were analyzed by one-way ANOVA or two-way repeated-measures ANOVA,and the Tukey's test was used for inter-group comparison.Results Compared with the NC group,the group A showed a significant increase in airway resistance(P＜0.01)and a marked decrease in lung dynamic compliance(P＜0.01).After BSYQF and Fer-1 intervention,airway resistance decreased(P＜0.05)and lung dynamic compliance improved(P＜0.05)in all treatment groups.HE and PAS staining revealed pronounced inflammatory infiltration in the lung tissue of the group A(P＜0.001),which was alleviated by BSYQF and Fer-1 treatment(P＜0.01),accompanied by reduced goblet cell metaplasia and mucus hypersecretion.The numbers of inflammatory cells(lymphocytes,eosinophils,and monocytes)in BALF were significantly elevated in the group A(all P＜0.01)and were reduced after BSYQF and Fer-1 intervention(P＜0.05).Fe2+content measurement and Perls-DAB staining indicated the presence of Fe2+overload(P＜0.01)and Fe3+deposition in the lung tissue of the group A.Both BSYQF/H and Fer-1 effectively lowered the Fe2+content(P＜0.05)and reduced Fe3+deposition.MDA level in the group A was significantly higher than that in the NC group(P＜0.01),while GSH level was lower(P＜0.001).After BSYQF/H and Fer-1 intervention,MDA level decreased significantly(P＜0.05),and GSH expression increased(P＜0.01).Conclusion Both BSYQF and Fer-1 demonstrated favorable therapeutic effects in the mouse asthma model.It may inhibit ferroptosis by regulating iron metabolism and suppressing lipid peroxidation,thereby reducing airway inflammation and epithelial damage.This demonstrates the potential for multi-target synergy in regulating inflammation.The BSYQF/H group effectively reduce lung-tissue Fe2+content,attenuate Fe3+deposition,and regulate oxidative-stress markers(decreasing MDA and increasing GSH),suggesting that it has Objective To investigate the therapeutic effects and mechanism of action of Bushen Yiqi formula (BSYQF) in a mouse model of bronchial asthma. Methods An allergic asthma model was established in Balb/c mice through ovalbumin (OVA)sensitization and challenge. 40 mice were randomly divided into five groups (n=8 per group):normal control group (NC group),model group (group A),BSYQF low-dose group (BSYQF/L group),BSYQF high-dose group (BSYQF/H group),and ferroptosis inhibitor group (Fer-1 group). Pulmonary mechanical function,including airway resistance and lung dynamic compliance,were measured using a pulmonary function apparatus. Hematoxylin and eosin (HE)and periodic acid-schiff the effect of inhibiting ferroptosis.