摘要
目的基于Nrf2/GPX4介导的铁死亡途径探讨柚皮素(Naringenin,Nar)对胆红素脑病新生小鼠海马神经元损伤的保护作用。方法将新生小鼠随机分成对照(Control)组、模型(Model)组、柚皮素低剂量(Nar-L)组、柚皮素高剂量(Nar-H)组和高剂量柚皮素联合Nrf2抑制剂ML385(Nar-H+ML385)组,每组15只。除Control组外,其他各组小鼠均采用经小脑延髓池注射胆红素溶液(20μg/g)建立新生小鼠胆红素脑病模型,造模后经腹腔注射柚皮素(25、100 mg/kg)或ML385(30 mg/kg)干预,每天1次,连续7天。干预结束后,观察各组小鼠神经行为学变化;水迷宫实验检测各组小鼠远期学习与记忆能力;Nissl染色观察小鼠海马神经元损伤情况;化学法检测小鼠海马组织中丙二醛(Malondialdehyde,MDA)、4-羟基壬烯醛(4-Hydroxynonenal,4-HNE)、超氧化物歧化酶(Superoxidedismutase,SOD)、谷胱甘肽过氧化物酶(Glutathione peroxidase,GSH-Px)水平;比色法检测小鼠海马组织中Fe^(2+)含量;Western blot法检测小鼠海马组织中核相关因子2(Nuclear associated factor 2,Nrf2)、溶质载体家族7成员11(Solute carrier family 7 member 11,SLC7A11)、谷胱甘肽过氧化酶4(Glutathione peroxidase 4,GPX4)蛋白表达水平。结果与Control组比较,Model组小鼠出现不同程度的神经异常行为,且远期学习与记忆能力显著降低(P<0.05),同时海马神经元损伤严重,海马组织中MDA、4-HNE以及Fe^(2+)含量显著升高(P<0.05),SOD、GSH-Px活性以及Nrf2、SLC7A11、GPX4蛋白表达水平显著降低(P<0.05)。与Model组比较,Nar-L、Nar-H组小鼠神经异常行为减少,且远期学习与记忆能力显著增强(P<0.05),同时海马神经元损伤明显改善,海马组织中MDA、4-HNE以及Fe^(2+)含量显著降低(P<0.05),而SOD、GSH-Px活性以及Nrf2、SLC7A11、GPX4蛋白表达水平显著升高(P<0.05)。然而,联合ML385干预能明显逆转Nar对胆红素脑病新生小鼠海马神经元损伤的改善作用。结论Nar可改善胆红素脑病新生小鼠海马神经元损伤,其作用途径可能是通过调控Nrf2/GPX4轴介导的铁死亡实现的。
Objective To investigate the effect of naringenin(Nar)on hippocampal neuronal injury in neonatal mice with bilirubin encephalopathy,focusing on the Nrf2/GPX4-mediated ferroptosis pathway.Methods Neonatal mice were randomly divided into Control group,Model group,Nar low(Nar-L),high dose group(Nar-H)and high dose naringenin combined with Nrf2 inhibitor ML385 group(Nar-H+ML385),with 15 mice in each group.With the exception of the Control group,mice in the other groups were injected with bilirubin solution(20μg/g)through the cerebellar bulbar cisterna to establish neonatal mouse bilirubin encephalopathy model.After the establishment of the model,intraperitoneal injection of naringin(25 or 100 mg/kg)or ML385(30 mg/kg)was administered once daily for a consecutive period of 7 days.After intervention,the neurobehavioral changes of each group of mice were observed.The water maze experiment was conducted to assess the long-term learning and memory abilities of the mice in each group.Nissl staining was performed to observe hippocampal neuron damage in mice.Chemical methods were used to measure the levels of malondialdehyde(MDA),4-hydroxynonenal(4-HNE),superoxide dismutase(SOD),and glutathione peroxidase(GSH-Px)in the hippocampal tissue of mice.Colorimetric analysis was employed to determine the content of Fe^(2+)in hippocampal tissue.Western blotting was utilized to detect protein expression levels of nuclear associated factor 2(Nrf2),solute carrier family 7 member 11(SLC7A11)and glutathione peroxidase 4(GPX4)protein in the hippocampal tissue of mice.Results Compared with the Control group,the Model group showed different degrees of abnormal neural behavior,and the long-term learning and memory ability were significantly reduced(P<0.05),at the same time,the hippocampal nerve was seriously damaged,and the contents of MDA,4-HNE and Fe^(2+)in hippocampal tissue were significantly increased(P<0.05),the activities of SOD and GSH-Px and the protein expression levels of Nrf2,SLC7A11 and GPX4 were significantly decreased(P<0.05).Compared with the Model group,the Nar-L and Nar-H groups had less abnormal behavior,and the long-term learning and memory ability were significantly enhanced(P<0.05),at the same time,the hippocampal nerve injury was significantly improved,and the contents of MDA,4-HNE and Fe^(2+)in hippocampal tissue were significantly decreased(P<0.05),the activities of SOD and GSH-Px and the protein expression levels of Nrf2,SLC7A11 and GPX4 were significantly increased(P<0.05).However,the combined intervention of ML385 significantly attenuates the beneficial effects of Nar on hippocampal neuron damage in neonatal mice with bilirubin encephalopathy.Conclusion This study suggested that Nar can ameliorate neuronal damage in hippocampus of neonatal mice with bilirubin encephalopathy,possibly through the regulation of iron death mediated by Nrf2/GPX4 axis.
作者
罗克踊
段淼
蒋亮
黄婷
陈宗礼
LUO Keyong;DUAN Miao;JIANG Liang;HUANG Ting;CHEN Zongli(The First People's Hospital of Zunyi,Zunyi 563000,China)
出处
《世界科学技术-中医药现代化》
北大核心
2025年第5期1477-1484,共8页
Modernization of Traditional Chinese Medicine and Materia Medica-World Science and Technology
基金
合肥市优秀青年科技创新人才培养项目(遵优青科【2018】10号):从肠道菌群的角度靶向干预新生儿坏死性小肠结肠炎,负责人:段淼。
