黄芪多糖对溃疡性结肠炎大鼠的研究

Research of Astragalus polysaccharide on ulcerative colitis in rats

目的探讨黄芪多糖(APS)通过Toll样受体4(TLR4)/核转录因子κB(NF-κB)信号通路减轻氧化应激和炎症反应对溃疡性结肠炎大鼠的保护作用。方法用2,4,6-三硝基苯磺酸(TNBS)联合乙醇造模法建立大鼠溃疡性结肠炎(UC)模型,将SD大鼠随机分为对照组、模型组、柳氮磺砒啶组(0.5 g·kg^(-1) SASP,SASP组)和低、中、高剂量实验组(0.3,0.6,1.2 g·kg^(-1)·d^(-1)黄芪多糖)。用苏木精-伊红染色法分析黄芪多糖对溃疡性结肠炎大鼠的影响;用生化法检测黄芪多糖对溃疡性结肠炎大鼠氧化应激的影响;用酶联免疫吸附(ELISA)法测定黄芪多糖对溃疡性结肠炎大鼠炎症和促炎细胞因子的影响;以蛋白质印迹(Western blot)法检测黄芪多糖对溃疡性结肠炎大鼠炎症TLR4/NF-κB信号通路相关蛋白表达的影响;以免疫组化法检测各组大鼠TLR4的表达。结果对照组、模型组、SASP组和低、中、高剂量实验组的结肠黏膜损伤指数分别为0.31±0.04,2.68±0.23,1.48±0.15,1.65±0.18,1.86±0.16和2.43±0.17,TLR4蛋白表达水平分别为0.28±0.05,1.09±0.13,0.34±0.03,0.75±0.05,0.57±0.06,0.40±0.03;NF-κB p65表达水平分别为0.25±0.04,0.98±0.07,0.37±0.05,0.71±0.07,0.61±0.06,0.38±0.05。与对照组比较,模型组大鼠结肠组织中超氧化物歧化酶(SOD)、过氧化氢酶(CAT)及谷胱甘肽S-转移酶(GST)活性降低(均P<0.05),大鼠血清中肿瘤坏死因子-α(TNF-α)、白细胞介素^(-1)β(IL^(-1)β)、白细胞介素-6(IL-6)、髓过氧化物酶(MPO)及一氧化氮(NO)水平升高(均P<0.05);与模型组比较,SASP组和低、中、高剂量实验组SOD、CAT及GST活性均升高(均P<0.05),细胞因子TNF-α、IL^(-1)β、IL-6、MPO及NO水平均降低(均P<0.05),且呈现剂量依赖性。结论黄芪多糖通过调节TLR4/NF-κB信号通路,能够明显减轻氧化应激和炎症反应,对溃疡性结肠炎大鼠具有保护作用。

Objective To investigate the protective effect of Astragalus Polysaccharide(APS)through toll-like receptor 4(TLR4)/nuclear factor-kappa B(NF-κB)signal pathway on rats with ulcerative colitis by reducing oxidative stress and inflammatory response.Methods Rat model of ulcerative colitis(UC)was established by modeling 2,4,6-trinitrobenzene sulfonic acid(TNBS)with ethanol.The SD rats were randomly divided into control group,model group,salazosulfapyridine(0.5 g·kg^(-1) SASP,SASP group)and low,medium,high dose experimental groups(0.3,0.6,1.2 g·kg^(-1)·d^(-1) APS).The effects of APS on rats with ulcerative colitis were analyzed by hematoxylin eosin staining;the effects of APS on oxidative stress in rats with ulcerative colitis were detected by biochemical method;the effects of APS on inflammation and pro-inflammatory cytokines in rats with ulcerative colitis were measured by enzyme linked immunosobent assay(ELISA);the effects of APS on inflammation TLR4/NF-κB in rats with ulcerative colitis were detected by Western blot;the expression of TLR4 was detected by immunohistochemistry.Results The damage indices of colonic mucosa in control group,model group,SASP group and low,medium,high dose experimental groups were 0.31±0.04,2.68±0.23,1.48±0.15,1.65±0.18,1.86±0.16 and 2.43±0.17,the expression levels of TLR4 protein in each group were 0.28±0.05,1.09±0.13,0.34±0.03,0.75±0.05,0.57±0.06,0.40±0.03,the expression levels of NF-κB p65 were 0.25±0.04,0.98±0.07,0.37±0.05,0.71±0.07,0.61±0.06,0.38±0.05,respectively.Compared with control group,the activities of superoxide dismutase(SOD),catalase(CAT)and glutathione S-transferase(GST)in the colon tissue of rats in model group were decreased(all P<0.05),and the levels of tumor necrosis factor-α(TNF-α),interleukin 1β(IL^(-1)β),interleukin 6(IL-6),myeloperoxidase(MPO)and nitric oxide(NO)in the serum of rats were increased(all P<0.05).Compared with model group,the activities of SOD,CAT and GST in the SASP group and low,medium,high dose experimental groups were increased(all P<0.05),while the levels of cytokines TNF-α,IL^(-1)β,IL-6,MPO and NO were decreased(all P<0.05).In addition,the effect of Astragalus polysaccharide was dose-dependent.Conclusion Astragalus polysaccharides regulate TLR4/NF-κB signaling pathway can significantly reduce oxidative stress and inflammatory response,and has a protective effect on rats with ulcerative colitis.