壳寡糖的制备及其对三硝基苯磺酸诱导的大鼠结肠炎模型的治疗作用

Preparation and therapeutic effects of chitosan oligosaccharide on trinitrobenzene sulfonic acid-induced rat model of inflammatory bowel disease

目的探究壳寡糖对结肠炎大鼠模型的治疗作用及分子机制。方法以脱乙酰度为87%、分子量为140 ku壳聚糖为原料,经酶解制备壳寡糖(COS)。将大鼠随机分为正常对照组(CON)、结肠炎模型组(TNBS)、5-氨基水杨酸(5-ASA)治疗组(TNBS+5-ASA)、COS治疗组(TNBS+COS)。以50%乙醇溶液为溶剂,用100 mg/kg 2,4,6-三硝基苯磺酸(TNBS)诱导大鼠结肠炎模型。实验过程中记录大鼠体质量并进行疾病活动指数(DAI)评估;于第2周处死大鼠,实验结束后取出结肠组织进行病理染色并评分;通过ELISA法测定大鼠血清中白细胞介素1β(IL-1β)、肿瘤坏死因子α(TNF-α)和白细胞介素6(IL-6)的浓度;利用血细胞分析仪检测大鼠血液中血红蛋白和红细胞含量;免疫组化方法检测结肠组织Ⅰ型胶原蛋白(collagen Ⅰ)、α平滑肌肌动蛋白(α-SMA)和核因子kappa-B(NF-κB)表达水平。结果与TNBS组比较,TNBS+COS组大鼠的DAI、组织学评分、血清中TNF-α及IL-6等炎症指标显著降低(P<0.01);杯状细胞损伤减少,数量显著恢复(P<0.01);collagenⅠ和α-SMA表达减少(P<0.01),纤维化明显改善。结论COS对实验性结肠炎具有较好的治疗作用,能够缓解结肠组织炎症反应,保护肠道黏膜屏障,抑制肠道纤维化进程。

Objective To investigate the therapeutic effect of chitosan oligosaccharide(COS)on the rat model of inflammatory bowel disease(IBD)and the underlying mechanisms.Methods COS was prepared from chitosan with a deacetylation degree of 87%and a molecular weight of 140 ku by enzymatic hydrolysis.The rats were randomly divided into four groups:normal control(CON),IBD model group(TNBS),5-aminosalicylic acid treated group(TNBS+5-ASA),and chitosan oligosaccharide treated group(TNBS+COS).IBD rat model was established by intracolonic injection of 100 mg/kg TNBS in 50% ethanol.Bodyweight was daily recorded,and fecal status was scored using the disease activity index(DAI).The rats were sacrificed at 2 weeks.The colon tissues were removed for histopathological staining and scoring.Serum concentrations of interleukin 1 beta(IL-1β),tumor necrosis factor-α(TNF-α),interleukin 6(IL-6)were detected by commercialenzyme linked immunosorbent assay(ELISA)kits.The contents of hemoglobin and red blood cells in rat blood were determined by the blood cell analyzer.The expression levels of typeⅠcollagen(collagen Ⅰ),alpha-smooth muscle actin(α-SMA),and nuclear factor kappa-B(NF-κB)were detected by immunohistochemistry.Results Compared with the TNBS group,DAI,histological score,serum TNF-α,and IL-6 expression significantly decreased by COS treatment(P<0.01).Goblet cell injury decreased and the number of goblet cells recovered significantly(P<0.01).Compared with the TNBS group,the expression of collagenⅠandα-SMA decreased in the TNBS+COS group(P<0.01).Conclusion COS has therapeutic effects on experimental IBD,which alleviates the inflammatory response of colon tissue,protecting the intestinal mucosal barrier,and inhibiting the progression of intestinal fibrosis.