羧甲基茯苓多糖对溃疡性结肠炎大鼠的研究

Research of carboxymethyl pachymaran on ulcerative colitis rats

目的研究羧甲基茯苓多糖通过调控Toll样受体4/髓样分化因子88/核转录因子-κB(TLR4/MyD88/NF-κB)信号通路对溃疡性结肠炎大鼠的改善作用。方法从40只SPF级大鼠中随机分出8只大鼠作为正常组,其余大鼠用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模型,造模成功后大鼠随机分为模型组、对照组和低、高剂量实验组。低、高剂量实验组分别给予0.5%羧甲基茯苓多糖混悬液2 mL和1.5%CMP混悬液2 mL,灌胃;对照组给予0.5 g·kg^(-1)柳氮磺吡啶灌胃;正常组和模型组每日灌胃给予等量的生理盐水,连续给药14 d。末次给药结束后,测定大鼠体质量、脾质量、结肠长度和质量,用自动化血液学分析仪分析血液参数,用酶联免疫吸附(ELISA)法检测各组大鼠血清中肿瘤坏死因子-α(TNF-α)、白细胞介素-6(IL-6)和白细胞介素-12(IL^(-1)2)水平;用蛋白质印迹法检测TLR4/MyD88/NF-κB信号通路相关蛋白TLR4、MyD88和NF-κB p65的表达水平。结果正常组、模型组、对照组和低、高剂量实验组的肠质量分别为(1.98±0.15),(3.72±0.26),(2.13±0.25),(2.99±0.40)和(2.66±0.15)g,肠长分别为(21.63±2.13),(15.47±1.59),(20.76±2.42),(17.23±0.86)和(19.67±1.68)cm,TLR4蛋白表达水平分别为0.23±0.04,1.00±0.08,0.39±0.05,0.78±0.05和0.54±0.06;MyD88蛋白表达水平分别为0.38±0.02,0.97±0.08,0.50±0.05,0.66±0.08和0.58±0.04;NF-kB p65蛋白表达水平分别为0.39±0.04,1.05±0.11,0.57±0.08,0.71±0.05和0.62±0.09。以上指标,模型组与正常组比较,对照组和低、高剂量实验组与模型组比较,差异均有统计学意义(均P<0.05)。结论羧甲基茯苓多糖可以通过抑制TLR4/MyD88/NF-κB信号通路相关蛋白的表达,减轻炎症反应,从而对溃疡性结肠炎起到改善作用。

Objective To investigate the effect of carboxymethyl pachymaran on improving ulcerative colitis in rats by regulating the Toll-like receptor 4/myeloid differentiation factor 88/nuclear transcription factor-κB(TLR4/MyD88/NF-κB)signaling pathway.Methods Eight SPF rats were randomly selected from 40 rats as normal group,other rats were established ulcerative colitis model by 2,4,6-nitrobenzene sulfonic acid(TNBS)combined with ethanol modeling.After modeling,the rats were randomly divided into model group,control group and low,high dose experimental groups.Low,high dose experimental groups were given 0.5%suspension of carboxymethyl pachymaran 2 mL and 1.5%suspension of carboxymethyl pachymaran 2 mL,respectively.Control group was given 0.5 g·kg^(-1) sulfasalazine intragastric administration.Normal group and model group were given the same amount of normal saline by intragastric administration for 14 days.After the last administration,body weight,spleen weight,colon length and weight of rats were determined,blood parameters were analyzed by automatic hematology analyzer,and serum levels of tumor necrosis factor-α(TNF-α),interleukin-6(IL-6)and interleukin-12(IL^(-1)2)were determined by enzyme-linked immunosorbent assay(ELISA).Western blotting was used to detect the expression levels of TLR4/MyD88/NF-κB signaling pathway related proteins TLR4,MyD88 and NF-κB p65.Results The intestinal weight in normal group,model group,control group and low,high dose experimental groups were(1.98±0.15),(3.72±0.26),(2.13±0.25),(2.99±0.40)and(2.66±0.15)g;intestinal length were(21.63±2.13),(15.47±1.59),(20.76±2.42),(17.23±0.86)and(19.67±1.68)cm;TLR4 protein expression levels were 0.23±0.04,1.00±0.08,0.39±0.05,0.78±0.05 and 0.54±0.06;MyD88 protein expression levels were 0.38±0.02,0.97±0.08,0.50±0.05,0.66±0.08 and 0.58±0.04;NF-κB p65 protein levels were 0.39±0.04,1.05±0.11,0.57±0.08,0.71±0.05 and 0.62±0.09,respectively.There were statistically significant differences in the above indexes compared between model group and normal group,between control group,low dose experimental group,high dose experimental group and model group(all P<0.05).Conclusion Carboxymethyl pachyman can reduce the inflammatory response by inhibiting the expression of TLR4/MyD88/NF-κB signaling pathway-related proteins,thereby improving ulcerative colitis.