当归散改善克罗米芬诱导子宫内膜血管发育不良的生物机制探讨

Biological mechanism of Danggui Powder in improving endometrial vascular dysplasia induced by clomiphene citrate

关键词:  当归散  克罗米芬  血管内皮生长因子  血管生成素  子宫内膜血管发育  网络药理学

Key words:  Danggui Powder  clomiphene citrate  vascular endothelial growth factor（VEGF）  angiogenin  endometrial angiogenesis  network pharmacology

摘要:

[目的] 基于网络药理学和动物实验，探究当归散及其拆方对克罗米芬诱导子宫内膜血管发育不良的作用效果。[方法] 通过中药系统药理学数据库与分析平台（TCMSP）、Swiss Target Predicition数据库获取当归散成分及靶点，利用人类基因数据库（Gene Cards）、人类孟德尔遗传（OMIM）数据库检索克罗米芬导致子宫内膜血管发育不良相关的疾病靶点，两者取交集后，构建“中药-成分-靶点”网络图、蛋白互作（PPI）网络，进行GO功能、京都基因和基因组百科全书（KEGG）通路富集分析。模型组和各给药组大鼠于动情期后第1天起，连续5 d灌胃克罗米芬混悬液造模；各给药组每日分别予阳性对照药物阿司匹林、当归散、川芎-当归-芍药、黄芩-白术供试液，连续8 d。免疫组化法测定大鼠子宫内膜微血管密度、血管内皮生长因子（VEGFA）及其受体血管内皮生长因子受体1（FLT1）、血管内皮细胞生长因子受体2（KDR）、血管生成素（Ang）1、Ang2及其受体内皮细胞TEK酪氨酸激酶（Tie2）的表达。[结果] 网络药理学得到VEGFA、表皮生长因子受体（EGFR）、成纤维细胞生长因子2（FGF2）、KDR、原癌基因酪氨酸蛋白激酶（SRC）、FLT1等当归散核心作用靶点24个，GO富集分析涉及生物过程1 524个条目、细胞组分36个条目和分子功能94个条目，KEGG通路富集得到磷脂酰肌醇-3-激酶-蛋白激酶（PI3K-Akt）信号通路、Rap1信号通路、松弛素信号通路等118条通路。与正常组相比，模型组子宫内膜微血管密度显著降低，VEGFA及其受体FLT1、KDR，Ang1、Ang2及其受体Tie2表达水平显著下降（P＜0.01）。与模型组相比，各药物组微血管密度提高，KDR、FLT1、Ang1、Ang2、Tie2表达升高（P＜0.05或P＜0.01）；除黄芩白术组，其余各用药组大鼠子宫内膜VEGFA的表达水平均显著升高（P＜0.01）。[结论] 当归散及其拆方可以不同程度地改善克罗米芬造成的大鼠子宫内膜微血管密度下降，恢复VEGFA及其受体FLT1、KDR，Ang1、Ang2及其受体Tie2表达水平，提高子宫内膜容受性。

Abstract:

[Objective] To study the effect of Danggui Powder and its disassembled prescriptions on dysplasia endometrial vascular caused by clomiphene citrate，based on network pharmacology and experimental verification. [Methods] TCMSP and SwissTargetPredicition databases were used to obtain the active components and potential drug targets of Danggui Powder. GeneCards and OMIM databases were used to search the targets of dysplasia endometrial vascular caused by clomiphene citrate. The intersection of drugs and dysplasia endometrial vascular related targets was selected. The network of “Chinese medicine-core component-target” and the protein-protein interactions（PPI） network were conducted. The enrichment analysis of GO and KEGG were carried out. On the first day after estrus，the model group and each treatment group were given clomiphene suspension for 5 consecutive days. Meanwhile，the treatment groups were given aspirin，Danggui Powder，Chuanxiong-Danggui-Shaoyao，Huangqin-Baizhu solution for 8 consecutive days. The rat endometrial microvessel density，the expression of vascular endothelial growth factor VEGFA，FLT1，KDR，Ang1，Ang2 and Tie2 were determined by Immunohistochemistry. [Results] A total of 24 potential targets of Danggui Powder were obtained by network pharmacology，such as VEGFA，EGFR，FGF2，KDR，SRC，FLT1，etc. A total of 1524 items of biological processes，36 items of cellular components and 94 items of molecular functions were involved in the enrichment analysis of GO. And 118 pathways were involved in the enrichment analysis of KEGG，such as PI3K-Akt signaling pathway，Rap1 signaling pathway，Relaxin signaling pathway，etc. Compared with the normal group，the rat endometrial microvessel density，VEGFA，FLT1，KDR，Ang1，Ang2 and Tie2 in the model group significantly decreased（P＜0.01）. Compared with the model group，the rat endometrial microvessel density，KDR，FLT1，Ang1，Ang2 and Tie2 in each treatment group increased（P＜0.05 or P＜0.01）. In addition to the Huangqin-Baizhu group，VEGFA in the other treatment groups significantly increased（P＜0.01）. [Conclusion] Danggui Powder and its disassembled prescription can reduce the damage of clomiphene citrate for endometrial to varying degrees，restore the expression of VEGFA，FLT1，KDR，Ang1，Ang2 and Tie2，and improve the endometrial receptivity.