黄连碱衍生物Q3通过调节TLR4/NF-κB信号通路和肠道微生物改善葡聚糖硫酸钠诱导的小鼠溃疡性结肠炎研究  

Coptisine Derivative Q3 Ameliorates DSS-induced Ulcerative Colitis in Mice by Modulating TLR4/NF-κB Signaling Pathway and Intestinal Microbiota

作　　者：迟笑倩 柴常伟 张海婧 吴练秋 CHI Xiaoqian;CHAI Changwei;ZHANG Haijing;WU Lianqiu(State Key Laboratory of Digestive Health,Institute of Materia Medica,Chinese Academy of Medical Sciences and Peking Union Medical College,Beijing 100050,China)

机构地区：[1]中国医学科学院、北京协和医学院药物研究所消化健康全国重点实验室,北京100050

出　　处：《中国药学杂志》2024年第17期1581-1589,共9页Chinese Pharmaceutical Journal

基　　金：中国医学科学院医学与健康科技创新工程资助(2022-I2M-1-014)。

摘　　要：目的从炎症信号通路Toll样受体4(TLR4)/核因子κB(NF-κB)以及肠道微生物群组成改变的角度探究黄连碱衍生物Q3抗葡聚糖硫酸钠(dextran sodium sulfate,DSS)诱导小鼠溃疡性结肠炎(ulcerative colitis,UC)的作用机制。方法将50只小鼠随机分为正常对照组、模型组、阳性药柳氮磺吡啶(SASP)组、Q3-低剂量组和Q3-高剂量组,每组10只,除正常对照组外其余组小鼠以2.5%DSS溶液喂饮建立UC模型。SASP组灌胃给予700 mg·kg^(-1)·d^(-1)SAPA,Q3低与高两个剂量组分别灌胃给予50和100 mg·kg^(-1)·d^(-1)Q3,其余组给予等量蒸馏水。给药6 d后取小鼠结肠组织进行疾病活动指数(disease activity index,DAI)评分和长度测量,苏木精-伊红染色(hematoxylin-eosin staining,HE)检测各组病理损伤程度,16S rRNA高通量测序检测小鼠肠内容物中肠道菌群的变化,免疫组织化学(IHC)检测结肠组织TLR4和p-p65的表达,Western blot法检测结肠组织中TLR4、p-p65和p-IκBα的蛋白表达,体外实验,利用脂多糖(lipopolysaccharide,LPS)诱导炎症反应,免疫荧光法检测IEC6细胞和RAW264.7细胞中NF-κB p65的核转位。结果体内:在DSS诱导的小鼠UC模型中,与模型组相比,Q3能够显著改善UC小鼠体重降低、结肠挛缩和DAI评分升高等情况,HE染色结果显示Q3明显改善UC小鼠肠道上皮破坏、隐窝结构紊乱和杯状细胞减少等病理损伤;16S rRNA结果表明Q3可以增加DSS作用后肠道菌群的生物多样性,调节肠道菌群组成。免疫组化和Western blot结果显示Q3可以显著降低模型组小鼠结肠组织中TLR4和p-p65的表达;体外:免疫荧光和Western blot结果显示Q3能够抑制IEC6和RAW264.7两种细胞中NF-κB p65的核转位。结论Q3能够通过抑制TLR4/NF-κB通路以及调节肠道菌群组成改善肠道炎症,发挥抗UC作用,有望成为治疗UC的候选化合物。OBJECTIVE To explore the mechanism of action of coptisine derivative Q3 against dextran sulfate sodium(DSS)-induced ulcerative colitis(UC)in mice from the perspective of changes in the inflammatory signaling pathway TLR4/NF-κB and compositions of the intestinal microbiota.METHODS Fifty mice were randomly divided into normal control group,model group,sulfasalazine(SASP)group,Q3-low dose group,Q3-high dose group,with 10 in each group.Mice in the groups except the control group were orally administered with 2.5%DSS solution to induce UC model.The SASP was given 700 mg·kg^(-1)·d^(-1)of sulfasalazine by intragastric administration,and the Q3 low and high dose groups were given 50 and 100 mg·kg^(-1)·d^(-1)of Q3,respectively.The other groups were given an equal amount of distilled water.After 6 days of administration,the mouse colon tissues were taken for DAI score and length measurement.HE staining was used to detect the degree of pathological damage in each group.16S rRNA high-throughput sequencing was used to detect changes in intestinal flora in the intestinal contents of mice.Immunohistochemistry and Western blot were used to detect the expression of TLR4 and p-p65 in colon tissue.The protein expressions of TLR4,p-p65 and p-IκBαand the nuclear translocation of NF-κB p65 in IEC6 cells and RAW264.7 cells were detected by Western blot and immunofluorescence.RESULTS In the DSS-induced mouse ulcerative colitis model,compared with the model group,in vivo Q3 could significantly improve the weight loss,colon length,and increase in DAI scores of UC mice.HE staining results showed that Q3 significantly improved the intestinal pathological damages such as tract epithelial damage,crypt structure disorder and goblet cell reduction;immunohistochemistry and Western blot results showed that Q3 could significantly reduce the expression of TLR4 and p-p65 in the colon tissue of mice in the model group.The results of 16S rRNA showed that Q3 could increase the biodiversity of intestinal microbiota and regulate the composition o

关 键 词：溃疡性结肠炎 黄连碱衍生物 肠道菌群 TOLL样受体4 NF-ΚB 

分 类 号：R966[医药卫生—药理学]