黄芪甲苷从Nrf2/SLC7A11/GPX4通路抑制铁死亡改善大鼠脊髓缺血再灌注损伤的研究OA
Astragaloside Ⅳ alleviates spinal cord ischemia-reperfusion injury in rats by inhibiting ferrop-tosis via the Nrf2/SLC7A11/GPX4 pathway
目的 探讨黄芪甲苷改善大鼠脊髓缺血再灌注损伤(SCII)的作用及其机制.方法 60只SD大鼠随机分为假手术组,SCII组,黄芪甲苷高、低剂量组,甲基强的松龙(MP)组.假手术组仅行开腹后分离腹主动脉,其余各组均采用改良Zivin,s法造模.术后24 h进行BBB评分后,收集L2～L5节段脊髓组织.苏木精-伊红(HE)、尼氏染色法观测大鼠脊髓神经元数目及形态变化;生化试剂盒检测髓组织超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、丙二醛(MDA)、脂质过氧化物(LPO)、亚铁离子(Fe2+)、8-羟基脱氧乌苷(8-OHdG)水平;免疫组化法检测脊髓组织中长链酰基辅酶A合成酶4(ACSL4)、谷胱甘肽过氧化物酶4(GPX4)蛋白表达情况;蛋白质免疫印迹法(WB)和实时荧光定量聚合酶链式反应(RT-PCR)检测相应脊髓组织中Kelch样ECH相关蛋白1(Keap1)、核因子E2相关因子2(Nrf2)、血红素氧合酶-1(HO-1)、GPX4、溶质载体家族7成员11(SLC7A11)表达情况.结果 与假手术组比较,SCII组大鼠BBB评分显著降低,脊髓组织神经元数量减少,神经元周围可见空泡状坏死区域及大量瘢痕组织;SOD、CAT活性明显下降、MDA、LPO、Fe2+、及8-OHdG的含量则显著上升;铁死亡相关特征蛋白中,ACSL4的表达量大幅度增加,而GPX4的表达量明显减少;蛋白表达方面,Keap1和细胞质Nrf2的蛋白表达量显著提高,细胞核Nrf2、HO-1、SLC7A11、GPX4的蛋白表达量显著降低;mRNA水平方面,Nrf2、HO-1、SLC7A11、GPX4 mRNA表达水平也呈现显著低表达趋势,Keap1 mRNA表达水平显著升高(P<0.05,P<0.01).经黄芪甲苷和甲基强的松龙干预后,上述指标出现显著逆转情况.结论 黄芪甲苷可能通过Nrf2/SLC7A11/GPX4信号通路抑制铁死亡,改善大鼠SCII.
Objective To investigate the effect of Astragaloside Ⅳ(AS-Ⅳ)on spinal cord ischemia-reperfusion injury(SCII)in rats and its underlying mechanism.Methods Sixty SD rats were randomly divided into five groups:sham,SCII,AS-Ⅳ high/low-dose(AS-IVH/L),and methylprednisolone(MP).Except for the sham group,all other groups underwent SCII modeling using the modified Zivin's method.Corresponding drug interventions were administered intraperitoneally once at 2 h after surgery.The sham group underwent laparotomy and abdominal aortic isolation without occlusion.Neurological function was assessed using the Basso,Beattie,Bresnahan(BBB)locomotor rating scale at 24 h post-surgery,after which the L2-5 spinal cord segments were harvested.Spinal cord morphology and neuronal survival were observed by HE and Nissl staining.The activities of SOD and CAT,and the levels of MDA,LPO,ferrous ion(Fe2+),and 8-hydroxy-2'-deoxyguanosine(8-OHdG)were measured using commercial assay kits.The protein expression of acyl-CoA synthetase long-chain family member 4(ACSL4)and glutathione peroxidase 4(GPX4)was detected by immunohistochemistry(IHC).The protein and mRNA expression levels of Keap1,Nrf2,HO-1,GPX4,and SLC7A11 were determined by Western blot(WB)and quantitative real-time polymerase chain reaction(qRT-PCR),respectively.Results Compared with the sham group,rats in the SCII group exhibited significantly lower BBB scores,reduced neuronal count,and evident pathological damage including edema and necrosis in spinal cord tissue.The activities of SOD and CAT were markedly decreased,while the contents of MDA,LPO,Fe2+,and 8-OHdG were increased.The expression of the ferroptosis marker ACSL4 was up-regulated,whereas GPX4 was down-regulated at the pro-tein level.Furthermore,WB analysis showed that the protein expression of Keap1 and cytosolic Nrf2 was increased,while that of nuclear Nrf2,HO-1,SLC7A11,and GPX4 was decreased.Consistently,qRT-PCR revealed decreased mRNA levels of Nrf2,HO-1,SLC7A11,and GPX4,and an increased mRNA level of Keap1(all P<0.05 or P<0.01).Treatment with either AS-Ⅳ or MP signifi-cantly reversed these changes induced by SCII.Conclusion Astragaloside Ⅳ alleviates spinal cord ischemia-reperfusion injury in rats,likely by suppressing ferroptosis through activation of the Nrf2/SLC7A11/GPX4 signaling pathway.
陈元平;张翔;周伟;周立亚
湖北中医药大学附属襄阳医院,湖北 襄阳 441000北京中医药大学东直门医院洛阳医院,北京 471000湖北中医药大学附属襄阳医院,湖北 襄阳 441000广州中医药大学附属深圳平乐骨伤科医院,广东 广州 518100
医药卫生
脊髓缺血再灌注损伤黄芪甲苷铁死亡Nrf2/SLC7A11/GPX4
Spinal cord ischemia-reperfusion injuryAstragaloside ⅣFerroptosisNrf2/SLC7A11/GPX4 pathway
《时珍国医国药》 2026 (18)
3466-3473,8
湖北省卫健委中医药科研项目(ZY2021M050)
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