基于TLR4/MyD88/NF-κB信号通路观察维生素D_(3)对大鼠炎症性肠病的改善作用

Effect of vitamin D_(3) on inflammatory bowel disease in rats based on TLR4/MyD88/NF-κB signaling pathway

目的基于Toll样受体4/髓样分化因子88/核转录因子κB(TLR4/MyD88/NF-κB)信号通路观察维生素D3(VD_(3))对大鼠炎症性肠病(IBD)的改善作用。方法选取32只大鼠随机分为对照组(Control组)、模型组(Model组)、低浓度VD_(3)组(VD_(3)low组)、高浓度VD_(3)组(VD_(3)high组)。采用3%右旋葡聚糖硫酸钠(DSS)建立大鼠IBD模型,分别给予不同浓度VD_(3)干预。造模结束后计算大鼠疾病活动指数(DAI)得分并测量结肠长度;行HE染色及结肠组织病理学评分;qRT-PCR、Western blot及免疫组化检测TLR4、MyD88、NF-κB p65 mRNA和蛋白表达水平;ELISA检测血清促炎因子肿瘤坏死因子-α(TNF-α)、白介素1β(IL-1β)、白介素6(IL-6)、抗炎因子白介素10(IL-10)含量。结果与Control组相比,Model组DAI评分、TLR4、MyD88、NF-κB p65 mRNA及蛋白表达水平,TNF-α、IL-1β、IL-6表达水平均显著升高,IL-10表达水平显著降低(P<0.05)。HE染色可见结肠组织黏膜及黏膜下层糜烂,形成溃疡、出血、隐窝脓肿,有大量炎症细胞、中性粒细胞浸润,杯状细胞减少。VD_(3)干预后,与Model组相比,VD_(3)low组、VD_(3)high组DAI评分、TLR4、MyD88、NF-κB p65 mRNA及蛋白表达水平,TNF-α、IL-1β、IL-6表达水平均显著下降,IL-10表达水平显著升高,结肠组织损伤有所缓解(P<0.05)。结论VD_(3)通过抑制TLR4/MyD88/NF-κB信号通路,降低下游促炎因子的表达,从而减轻IBD大鼠肠道损伤,改善结肠炎症反应,对IBD大鼠有很好的干预作用。

This study was designed to evaluate the ameliorating effect of vitamin D_(3)(VD_(3))on inflammatory bowel disease(IBD)in rats based on the Toll-like receptor 4/myeloid differentiation factor 88/nuclear transcription factorκB(TLR4/MyD88/NF-κB)signaling pathway.Total of 32 rats were recruited and randomly divided into control group,Model group,VD_(3) low group and VD3high group,while IBD model was established by using 3%dextran sulphate sodium(DSS).After modeling,disease activity index(DAI)was calculated and the length of the colon was measured;HE staining was performed for colonic histopathology score;qRT-PCR,Western blot and immunohistochemistry were used to detect the mRNA and protein expression levels of TLR4,MyD88,NF-κB p65;ELISA was used to detect the content of TNF-α,IL-1β,IL-6,and IL-10.Data showed that compared with the control group,the DAI score,the mRNA and protein expression levels of TLR4,MyD88,NF-κB p65,and the content of TNF-α,IL-1β,IL-6 in the vehicle group were significantly increased,while the expression level of IL-10 was significantly decreased(P<0.05).Rats of the vehicle group demonstrated were erosions in the mucosa and submucosa of the colon tissue,forming ulcers,hemorrhages,crypt abscesses,a large number ofinflammatory cells and neutrophils infiltrated,and lowlevel of goblet cells.After VD_(3) intervention,all these symptoms mentioned above were alleviated,and the damage of colon tissue was relieved(P<0.05).Taken together,VD_(3) inhibits the TLR4/My D88/NF-κB signaling pathway and reduces the expression of downstream pro-inflammatoryfactors,thereby reducing the intestinal injury of IBD rats and improving the inflammation of the colon.