黄芩素激活Nrf2/GPX4通路保护大鼠脑出血铁死亡损伤实验研究

Baicalein reduces ferroptosis after intracerebral hemorrhage in rats by activating Nrf2/GPX4 pathway

关键词:  脑出血  铁死亡  谷胱甘肽过氧化物酶4  脂质过氧化物

Key words:  intracerebral hemorrhage  ferroptosis  glutathione peroxidase 4  lipid hydroperoxide

摘要:

目的 基于核因子E2相关因子(Nrf2)/谷胱甘肽过氧化物酶4(GPX4)途径探讨黄芩素对大鼠脑出血后铁死亡损伤的保护作用。方法 50只SD大鼠随机分为5组：假手术组,模型组,黄芩素组,黄芩素+RSL3组和黄芩素+ML385组,每组10只。基底节注入60μl自体动脉血建立脑出血(ICH)模型。腹腔注射黄芩素(100mg/kg)﹑RSL3(10mg/kg)和ML385(10mg/kg)进行干预6d。采用神经功能缺损评分(mNSS)评价神经功能,ELISA法检测脂质过氧化物(LPO)含量,酶标法检测谷光甘酞(GSH)含量,RT-PCR法检测GPX4和Nrf2 mRNA表达,Western-blot法检测Nrf2和GPX4蛋白表达,电镜观察线粒体变化。结果 与假手术组比较,模型组mNSS评分﹑LPO﹑神经元线粒体损伤﹑GPX4mRNA﹑Nrf2mRNA和Nrf2蛋白明显增加,GSH和GPX4蛋白明显减少 (P <0.05)；与模型组比较,黄芩素组GSH﹑GPX4mRNA﹑GPX4蛋白﹑Nrf2mRNA和Nrf2蛋白明显增加,mNSS评分﹑LPO﹑神经元线粒体损伤明显减少 (P <0.05)；与黄芩素组比较,黄芩素+RSL3组mNSS评分﹑LPO和神经元线粒体损伤明显增加,GSH和GPX4蛋白明显减少 (P <0.05),黄芩素+ML385组mNSS评分﹑LPO和神经元线粒体损伤明显增加,GSH﹑GPX4mRNA﹑GPX4蛋白和Nrf2蛋白明显减少 (P <0.05)。结论 黄芩素可减轻脑出血后铁死亡损伤,其机制可能与激活Nrf2/GPX4通路有关。

Abstract:

ABSTRACT: OBJECTIVE To explore the protective effect of baicalein on ferroptosis after intracerebral hemorrhage in rats based on the nuclear factor E2 related factor (Nrf2)/glutathione peroxidase 4 (GPX4) pathway. METHODS Fifty SD rats were randomly divided into five groups by digital table: sham operation group, model group, baicalein group, baicalein + RSL3 group and baicalein + ML385 group. Basal ganglia injection 60 μL autologous arterial blood was used to establish ICH model. Baicalein (100mg/kg), RSL3 (10mg/kg) and ML385 (10mg/kg) were injected intraperitoneally for 6 days. The neurological function was evaluated by modified neurological severity score（mNSS）, the content of LPO was detected by ELISA, the content of glutathione (GSH) was detected by enzyme labeling, the expression of GPX4 and Nrf2 mRNA was detected by RT-PCR, the expression of Nrf2 and GPX4 protein was detected by Western-blot, and the changes of mitochondria were observed by electron microscope. RESULTS Compared with the sham-operated group, the mNSS score, LPO, neuronal mitochondrial damage, GPX4 mRNA and Nrf2 mRNA in the model group increased significantly, while GSH, GPX4 protein and Nrf2 protein decreased significantly (P<0.05); Compared with the model group, GSH, GPX4 mRNA, GPX4 protein, Nrf2 mRNA and Nrf2 protein in baicalein group increased significantly, while mNSS score, LPO and neuronal mitochondrial damage decreased significantly (P<0.05); Compared with the baicalein group, the mNSS score, LPO and neuronal mitochondrial damage in the baicalein+RSL3 group increased significantly, GSH and GPX4 protein decreased significantly (P<0.05), the mNSS score, LPO and neuronal mitochondrial damage in the baicalein+ML385 group increased significantly, GSH, GPX4 mRNA, GPX4 protein and Nrf2 protein decreased significantly (P<0.05). CONCLUSION Baicalein can reduce ferroptosis after intracerebral hemorrhage, and its mechanism may be related to the activation of Nrf2/GPX4 pathway.