基于TLR4/NF-κB通路探讨黄芪甲苷对炎症性贫血的改善作用

Effects of Astragaloside Ⅳ on Inflammatory Anemia by Inhibiting TLR4/NF-κB Pathway

目的:研究黄芪甲苷(Astragaloside IV,ASIV)通过TLR4/NF-κB信号通路对脂多糖(lipopolysaccharide,LPS)诱导的炎症性贫血(anemia of inflammation,AI)的影响并探讨其可能机制。方法:40只C57BL/6J小鼠随机分为4组:正常对照组、LPS(10 mg/kg)组、LPS+ASIV(40 mg/kg)、LPS+ASIV(80 mg/kg)组。ASIV给药组提前给予小鼠黄芪甲苷灌胃14 d后,除正常对照外,给予小鼠10 mg/kgLPS一次性腹腔注射,8 h后取肝脏组织和血清置于-80℃冰箱保存。全自动生化分析仪检测小鼠全血中红细胞计数(RBC)、血红蛋白(HB)、红细胞压积(HCT)、平均红细胞体积(MCV)、平均红细胞血红蛋白浓度(MCHC)、红细胞生成素(EPO)及血清和肝组织铁含量。酶联免疫吸附法测定(ELISA)小鼠血清TNF-α(肿瘤坏死因子)、IL-18(白介素-18)和IL-1β(白介素-1β)的表达水平。Western Blot检测肝组织铁调素、TLR 4(TOLL样受体4)、p-p65(磷酸化核因子-κBp65)及转铁蛋白的表达。结果:与正常对照组相比,LPS组RBC、HB、HCT、MCV、MCHC、EPO明显降低;血清TNF-α、IL-18和IL-1β含量升高;血清铁含量降低,肝组织铁含量升高;肝组织转铁蛋白的表达降低,铁调素、TLR 4和p-p 65的表达增高。与LPS组相比,ASIV能增加全血中血常规的相关指标,降低血清中炎症因子水平,增加血清铁含量,降低肝组织铁含量;同时ASIV还可以增加转铁蛋白的表达,抑制铁调素、TLR 4和p-p 65表达的增高。结论:ASIV可以通过抑制TLR4/NF-κB信号通路,改善LPS引起的铁代谢障碍,减轻炎症性贫血。

Objective:To study the effects of Astragaloside IV(ASIV)on anemia of inflammation(AI)induced by lipopolysaccharide(LPS)by TLR4/NF-κB signal transduction pathway and explore its possible mechanism.Methods:Forty C57BL/6J mice were randomly divided into four groups:normal control group,LPS(10 mg/kg)group,LPS+ASIV(40 mg/kg)group,LPS+ASIV(80 mg/kg)group.ASIV administration groups were given ASIV 14 days before modeling,and then given 10 mg·kg^-1 of LPS intraperitoneal injection except the control group.After 8 hours,liver tissues and serum were taken and placed in a refrigerator at-80℃.Red blood cell count(RBC),hemoglobin(HB),hematocrit(HCT),mean corpuscular volume(MCV),mean corpuscular hemoglobin concentration(MCHC),erythropoietin(EPO)and iron content in whole blood and liver tissue were measured.The levels of TNF-α,IL-18 and IL-β in serum were detected by ELISA.The protein levels of hepcidin,TLR4,p-p65 and transferrin were detected by Western Blot.Results:Compared with the normal control group,RBC,HB,HCT,MCV,MCHC and EPO in LPS group were significantly lower;TNF-α,IL-18 and IL-β in serum were increased;iron content in serum was decreased,iron content in liver tissue was increased;transferrin expression in liver tissue was decreased,and the expressions of hepcidin,TLR4 and P-P65 were increased.Compared with LPS group,ASIV could increase the blood routine related indicators,reduce the level of inflammatory factors in serum,increase the serum iron content,and reduce the liver iron content.At the same time,the application of ASIV can also increase the expression of transferrin and inhibit the expression of hepcidin,TLR 4 and P-P 65.Conclusion:ASIV can alleviate the occurrence of inflammatory anemia by inhibiting TLR4/NF-κB signaling pathway and improving iron metabolism disorder caused by LPS.