慈姑多糖调控TLR4/MyD88/NF-κB抑制细胞炎症和氧化损伤对多种重金属诱导L02肝细胞凋亡的影响  

Effect of Sagittaria Sagittifolia Polysaccharide on Regulating TLR4/MyD88/NF-κB to Inhibit Inflammation and Oxidative Damage in L02 Hepatocyte Apoptosis Injury by Multiple Heavy Metals

作　　者：周漫钰 刘红双 刘冰清 高鑫 梁垚 廖艳 ZHOU Manyu;LIU Hongshuang;LIU Bingqing;GAO Xin;LIANG Yao;LIAO Yan(School of Traditional Chinese Medicine,Beijing University of Chinese Medicine,Beijing 102446,China)

机构地区：[1]北京中医药大学中医学院,北京102446

出　　处：《中华中医药学刊》2025年第1期117-121,I0014,I0015,共7页Chinese Archives of Traditional Chinese Medicine

基　　金：国家重点研发计划项目(2018YFC1706800)。

摘　　要：目的探索慈姑多糖(Sagittaria Sagittifolia polysaccharide,SSP)是否通过介导Toll样受体4(Toll-like receptors 4,TLR4)/髓样分化因子88(myeloid differentiation factor 88,MyD88)/核转录因子-κB(nuclear transcription factor-kappa B,NF-κB)信号通路对6种重金属联合所致L02肝细胞损伤发挥保护作用。方法首先根据北京市售大米6种重金属膳食摄入比例,构建6种重金属混合染毒液所致L02细胞损伤体外模型。应用MTT法检测不同质量浓度慈姑多糖(0.25、0.5、1、2 mg/mL)提前作用24 h对L02细胞存活率的影响;Annexin V-FITC/碘化丙啶(PI)双染流式结合Hoechst 33342荧光检测细胞凋亡;DCFH-DA荧光探针检测细胞中活性氧(reactive oxygen species,ROS)含量变化;Western blot法检测细胞中多聚ADP核糖聚合酶1(poly-ADP ribose polymerase1,PARP1)、动力相关蛋白1(dynamin-related protein 1,Drp1)、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白介素(interleukin,IL)-1β、IL-17、TLR4、MyD88、NF-κB p65蛋白表达情况。结果MTT结果显示,SSP在1 mg/mL作用24 h保护效果最佳;与模型组相比,SSP治疗组中ROS含量及细胞凋亡率均明显下降(P<0.05),Drp1、TNF-α、IL-1β、IL-17和TLR4、MyD88、NF-κB p65蛋白表达均显著降低(P<0.05),PARP1蛋白表达显著升高(P<0.01)。结论SSP可有效改善6种重金属联用诱导的L02细胞凋亡,可能与调控TLR4/MyD88/NF-κB信号通路抑制氧化损伤和炎症反应相关。Objective To explore whether Sagittaria Sagittifolia polysaccharide(SSP)plays a protective role in L02 hepatocyte injured by six heavy metals mixture by mediating the Toll-like receptors 4(TLR4)/myeloid differentiation factor 88(MyD88)/nuclear transcription factor-κB(NF-κB)signaling pathway.Methods Firstly,according to the dietary intake of six heavy metals in rice sold in Beijing,a heavy metal mixture was prepared for inducing L02 cell injury.MTT assay was used to detect the effect of SSP(0.25 mg/mL,0.5 mg/mL,1 mg/mL,2 mg/mL)on the L02 cells survival rate with 24 h pre-treatment.Annexinv FITC/propidium iodide(PI)double staining combined with flow cytometry and Hoechst 33342 staining were used for detecting the cell apoptosis.The content of reactive oxygen species(ROS)in cells was detected by using DCFH-DA fluorescence probe.The protein expressions of poly-ADP ribose polymerase1(PARP1),dynamic related protein 1(Drp1),tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β),interleukin-17(IL-17),TLR4,MyD88 and NF-κB p65 were detected by Western blot.Results MTT assay showed that SSP(1 mg/mL,24 h)had the best protective effect.Compared with those of the model group,the ROS content and apoptosis rate in SSP treatment group were significantly decreased(P<0.05),and the protein expressions of Drp1,TNF-α,IL-1β,IL-17,TLR4,MyD88 and NF-κB p65 decreased significantly(P<0.05)and the protein expression of PARP1 increased significantly(P<0.01).Conclusion SSP can effectively improve the apoptosis of L02 cells induced by six heavy metals mixture,which may be related to the regulation of TLR4/MyD88/NF-κB signaling pathway to inhibit the oxidative damage and inflammatory response.

关 键 词：慈姑多糖 多种重金属 TLR4/MyD88/NF-κB通路 氧化应激 炎症反应 L02细胞 

分 类 号：R282.71[医药卫生—中药学]