苦参素对实验性自身免疫性脑脊髓炎小鼠的改善作用及机制

Improvement effect and mechanism of matrine on experimental autoimmune encephalomyelitis mice

目的探讨苦参素能否通过调控铁死亡途径发挥对实验性自身免疫性脑脊髓炎(EAE)小鼠的改善作用。方法将30只雌性C57BL/6小鼠随机分为正常组、模型组、苦参素组,每组10只。模型组和苦参素组小鼠以含灭活结核分枝杆菌和MOG35-55的抗原乳剂诱导复制EAE模型。苦参素组小鼠自免疫后第7天开始腹腔注射苦参素注射液(50 mg/kg),正常组和模型组小鼠腹腔注射等体积生理盐水,每天1次至免疫后第18天。记录各组小鼠的神经功能学评分,以苏木精-伊红染色法和勒克斯光蓝染色法分别观察脊髓组织炎症细胞浸润和髓鞘脱失情况,以定量逆转录聚合酶链反应法和Western blot法分别检测其转铁蛋白受体1(TFR1)、核受体共激活因子4(NCOA4)、膜铁转运辅助蛋白(Heph)m RNA和胱氨酸-谷氨酸反向转运体(xCT)、谷胱甘肽过氧化物酶4(GPx4)蛋白的表达情况。结果与正常组比较,模型组小鼠的累计神经功能学评分显著升高(P<0.01),脊髓组织内炎症细胞浸润和髓鞘脱失现象明显,相关评分均显著升高(P<0.01),髓鞘组织中TFR1、NCOA4 mRNA的表达均显著上调,Heph mRNA和xCT、GPx4蛋白的表达均显著下调(P<0.05或P<0.01)。与模型组比较,苦参素组小鼠上述指标均显著改善(P<0.05或P<0.01)。结论苦参素对EAE小鼠有一定的改善作用,且这种作用可能与调节铁死亡中的铁代谢途径和xCT/GPx4通路有关。

OBJECTIVE To investigate whether matrine exerts improvement effect on experimental autoimmune encephalomyelitis(EAE)mice by regulating ferroptosis pathway.METHODS Totally 30 female C57BL/6 mice were randomly assigned into normal group,model group and matrine group,with 10 mice in each group.Model group and matrine group were given antigen emulsion containing inactivated Mycobacterium tuberculosis and MOG35-55 to induce EAE model.Matrine group was injected with Matrine injection(50 mg/kg)intraperitoneally since the 7th day after immunization;normal group and model group were given constant volume of normal saline intraperitoneally,once a day,since 18th day after immunization.The neurofunctional score of mice was recorded,and hematoxylin and eosin staining and Luxol fast blue staining were used to observe inflammatory cell infiltration and demyelination in spinal cord tissue.The quantitative reverse transcription PCR and Western blot assay were performed to determine the mRNA expressions of transferrin receptor 1(TFR1),nuclear receptor coactivator 4(NCOA4)and hephaestin(Heph),and the protein expressions of system Xc-(xCT)and glutathione peroxidase 4(GPx4).RESULTS Compared with normal group,accumulative neurofunctional score was significantly increased in model group(P<0.01);inflammatory cell infiltration and demyelination were obvious in spinal cord tissue,and related scores were increased significantly(P<0.01).The mRNA expressions of TFR1 and NCOA4 in myelin tissue were up-regulated significantly,while the mRNA expression of Heph and the protein expressions of xCT and GPx4 were down-regulated significantly(P<0.05 or P<0.01).Compared with model group,above indexes of matrine group were all improved significantly(P<0.05 or P<0.01).CONCLUSIONS Matrine can improve EAE mice,the mechanism of which may be associated with regulating iron metabolism pathway and xCT/GPx4 pathway in ferroptosis.