溃结康对溃疡性结肠炎小鼠肠粘膜炎症因子及肠屏障功能相关蛋白的影响

Study on the influence of Kuijiekang Decoction on the inflammatory cytokines and proteins related to intestinal barrier function in mice with ulcerative colitis

目的:探讨溃结康对溃疡性结肠炎小鼠肠粘膜炎症因子及肠屏障功能的影响。方法:建立葡聚糖硫酸钠诱导小鼠实验性溃疡性结肠炎模型,实验分为正常对照组、模型组、柳氮磺胺吡啶组(0.45g/kg)、溃结康组(12.8g/kg、6.4g/kg、3.2g/kg)。造模7天内,每天观察UC小鼠体质量及疾病活动指数变化,第8天实验结束时,采集结肠组织,量取各组小鼠结肠长度,HE染色观察小鼠结肠病理变化,酶联免疫法检测结肠组织IL-1β、TNF-α、IL-4、IL-10含量变化,实时荧光定量PCR检测结肠ITF、NF-κB、Occludin、Claudin 1mRNA表达,Western-blot法检测结肠ITF、NF-κB、Occludin、Claudin 1蛋白表达。结果:模型组小鼠体质量较正常对照明显减轻,疾病活动指数显著升高,结肠长度明显缩短,结肠组织损伤明显,病理学评分显著增加,结肠IL-1β、TNF-α含量显著增加,IL-4、IL-10含量显著降低,NF-κBmRNA及蛋白表达均明显增加,而ITF、Occludin、Claudin 1mRNA及蛋白表达均显著降低;溃结康组(12.8g/kg、6.4g/kg)及柳氮磺胺吡啶组(0.5g/kg)小鼠体质量及结肠长度较模型组增加,疾病活动指数降低,结肠病理损伤较模型组减轻,病理学评分降低,TNF-α含量显著降低,NF-κB mRNA及蛋白表达明显减少,而occludin mRNA水平明显增加;溃结康组(12.8g/kg)还可明显增加IL-4含量,上调ITF mRNA、蛋白及occludin蛋白表达,溃结康组(6.4g/kg)可降低IL-1β含量,增加IL-10含量,上调ITF mRNA水平。溃结康组(3.2g/kg)除可上调ITF mRNA水平外,对各指标均无明显影响。结论:溃结康可能通过降低结肠组织致炎因子含量、增加抑炎因子含量,同时促进肠粘膜修复,保护肠粘膜屏障功能而治疗溃疡性结肠炎。

Objective： To investigate the therapeutic effect of Kuijiekang Decoction（ KJKD） on inflammatory cytokines and the intestinal mucosal barrier in mice with ulcerative colitis. Methods： The animal model of ulcerative colitis induced by dextran sodium sulfate（ DSS） was established. Experimental animals were divided into the control group,the model group,SASP group（ 0. 45 g/kg） and KJKD groups（ 12. 8 g/kg,6.4 g/kg,3. 2 g/kg）. The body weight and disease activity index（ DAI） were evaluated everday. On the eighth day after the treatment,mice were sacrificed to collect colonic mucosa,the therapeutic effect was evaluated with the length of colon,Pathological changes of colon were observed under light microscope by HE staining,contents of IL-1β,TNF-α,IL-4,IL-10 were observed by ELISA method,expressions of ITF,NF-κB,Occludin,Claudin 1 mRNA were detected by realtime polymerase chain reaction（ RT-PCR）,And western blotting was used to detect the expressions of ITF,NF-κB,Occludin,Claudin. Results： Compared with the control group,the body weight and colonic length were reduced（ P〈0. 01）,the injury of colonic tissue was serious（ P〈0. 01）,DAI scores as well as contents of IL-1β,TNF-α,expressions of NF-κB mRNA and protein were significantly increased（ P〈0. 01）,whereas levels of IL-4 and IL-10 were decreased（ P〈0. 05 or P〈0. 01）. Compared with the model group,the body weight,length of colonic as well as the expression of occludin mRNA were improved（ P〈0. 05 or P〈0. 01）,whereas DAI and the colonic histological scores,the content of TNF-α as well as expressions of NF-κB mRNA and protein in KJKD groups（ 12. 8 g/kg、6. 4 g/kg） and SASP group（ 0. 5 g/kg） were decreased（ P〈0. 05 or P〈0. 01）. Meanwhile 12. 8 g/kg KJKD also increased the content of IL-4,enhanced the gene expression of ITF,protein levels of ITF and occludin.（ P〈0. 05）. The level of IL-1β was lower,levels of IL-10 and ITF mRNA were higher in KJKD group（ 6. 4 g/kg） than those in the model group（ P〈0. 05）. 3. 2 g/kg KJKD had no significant effects except upregulating the expression of ITF mRNA. Conclusion： KJKD ameliorates DSS-induced ulcerative colitis in mice might be through regulating levels of inflammatory cytokines and promoting the repair of intestinal mucosa.