基于“通路-疾病”交互网络的葛根芩连汤及其指标成分组合物干预小鼠溃疡性结肠炎相关结肠癌的作用

Effect of Gegen Qinlian Decoction and composition of index components on ulcerative colitis related colon cancer in mice based on “pathway-disease” interaction network

目的建立溃疡性结肠炎相关结肠癌(UCRCC)与NF-κB信号通路的网络模型,预测UCRCC的关键炎性靶标,明确葛根芩连汤及其指标成分组合物对这些靶标的影响。方法采用HPLC法,流动相为乙腈-0.2%磷酸水溶液,梯度洗脱,测定10种指标成分;检索Pubmed、ScienceDirect等数据库,分别查阅UCRCC疾病、NF-κB通路的上下游蛋白,采用Cytoscape软件,预测UCRCC的关键炎性靶标;在AOM/DSS诱导的UCRCC小鼠模型中,通过免疫组化、ELISA方法明确葛根芩连汤及其指标成分组合物对关键炎性靶标的影响。结果葛根芩连汤中葛根素、大豆苷、甘草苷、药根碱、黄芩苷、巴马汀、小檗碱、黄芩素、甘草酸、汉黄芩素的含量分别为1.677、0.154、0.159、0.045、0.448、0.035、0.095、0.013、0.111、0.006μg/g。交互网络中综合排名较高的前8位炎症因子分别是NF-κBp65、i NOS、COX-2、Bcl-2、TNF-α、IL-1β、ICAM-1和VCAM-1。与模型组比较,葛根芩连汤低、中、高剂量组及其指标成分组合物组、美沙拉秦肠溶片(5-ASA)组均显著下调UCRCC模型小鼠结肠组织中NF-κBp65、i NOS、Bcl-2、COX-2蛋白表达(P<0.01),并显著降低血清中TNF-α、IL-1β、ICAM-1、VCAM-1水平(P<0.05、0.01)。结论葛根芩连汤及其指标成分组合物能够改善UCRCC小鼠结肠组织病理情况,并下调炎性因子的表达,对UCRCC具有一定的干预作用,为确定葛根芩连汤质量标志物(Q-marker)奠定了基础。

Objective A network model of ulcerative colitis-associated colon cancer(UCRCC) and NF-κB signaling pathway was established, in order to predict the key inflammatory targets of UCRCC and identify the effect of GQD and composition of index components on these targets. Methods HPLC method was used, the mobile phase was acetonitrile-0.2% phosphoric acid aqueous solution, and the gradient elution was carried out. The 10 indicators were determined. The literature of databases such as Pubmed and ScienceDirect were searched, the terms of the upstream and downstream proteins of UCRCC disease and NF-κB pathway were examined. The Cytoscape software was used to predict the key inflammatory targets of UCRCC. The UCRCC model was established by AOM/DSS method, and the effect of GQD and composition of index components on key inflammatory targets were identified by immunohistochemistry and ELISA. Results The contents of puerarin, daidzin, glycyrrhizin, jatrorrhizine, baicalin, palmatine, berberine, baicalein, glycyrrhizic acid and wogonin in GQD were 1.677, 0.154, 0.159, 0.045, 0.448, 0.035, 0.095, 0.013, 0.111 and 0.006 μg/g, respectively. The top eight inflammatory factors of the key protein in the interaction network were NF-κBp65, i NOS, COX-2, Bcl-2, TNF-α, IL-1β, ICAM-1 and VCAM-1. Compared with the model group, the high-, medium-and low-dose groups of GQD, the composition of index components group and the 5-ASA group were able to down-regulate the relative expression levels of NF-κBp65, iNOS, Bcl-2, COX-2 in colonic tissues of UCRCC mice(P < 0.01), and down-regulate the expression levels of TNF-α, IL-1β, ICAM-1, VCAM-1 in serum of UCRCC mice(P < 0.05, 0.01). Conclusion GQD and composition of index components can improve the pathological condition of colon tissue in UCRCC mice, down-regulate the expression of inflammatory factors in colon tissue of UCRCC mice, and have a certain intervention effect on UCRCC, which laid the foundation for determining the quality marker(Q-marker) of GQD.