基于网络药理学和16S rDNA测序的当归六黄汤合煎与单煎治疗阴虚甲亢作用的差异研究

Difference between all-herb decoction and single-herb decoction of Danggui Liuhuang Decoction in treating yin-deficiency-hyperthyroidism based on network pharmacology and 16S rDNA sequencing

目的探讨当归六黄汤的抗阴虚甲亢作用,阐释传统汤剂合煎和单煎的差异。方法运用网络药理学方法,筛选当归六黄汤的主要活性成分和治疗阴虚甲亢疾病的相关靶点,并对交集靶点进行蛋白质-蛋白质相互作用(protein-protein interaction,PPI)网络分析、基因本体(gene ontology,GO)功能和京都基因与基因组百科全书(Kyoto encyclopedia of genes and genomes,KEGG)通路富集分析,构建药材-成分-靶点-通路网络;进一步进行实验验证,构建阴虚甲亢大鼠模型,评价当归六黄汤合煎和单煎对阴虚甲亢大鼠体质量、粪便含水率及相关血清学指标的影响;利用16S rDNA测序技术,揭示合煎和单煎对大鼠肠道微生物群落多样性的影响。结果筛选获得当归六黄汤方中102个活性成分,检索出与阴虚甲亢有关靶点116个。富集分析获得871条GO条目和158条KEGG相关条目(P<0.05),其中生物过程(biological process,BP)675条、细胞组分(cellular component,CC)77条、分子功能(molecular function,MF)119条,与当归六黄汤相关度较高的通路主要包括炎症、感染、肠病、癌症等通路。抗阴虚甲亢药效作用结果表明,与模型组比较,合煎高剂量组大鼠的体质量净增值和粪便含水率均显著增加(P<0.01),三碘甲状腺原氨酸(three iodine thyroid,T3)、甲状腺素(tetraiodothyronine,T4)等激素水平显著降低(P<0.01),且作用接近阳性药甲巯咪唑组。大鼠肠道菌群多样性结果显示,合煎高剂量组大鼠肠道微生物的丰富度和多样性接近对照组,Lachnospiraceae_NK4A136_group在模型组大鼠肠道显著富集,合煎给药后阿克曼氏菌属Akkermansia相对丰度明显增加,而单煎给药后普雷沃氏菌属Prevotella相对丰度明显增加。结论当归六黄汤合煎抗阴虚甲亢药效作用更好;阴虚甲亢等内分泌紊乱性疾病极有可能致肠道微生物群紊乱,当归六黄汤合煎可能通过调节肠道微生物群的方式治疗阴虚甲亢。

Objective To investigate the anti-yin-deficiency-hyperthyroidism effect of Danggui Liuhuang Decoction(当归六黄汤),and explain the difference between all-herb decoction and single-herb decoction.Methods The network pharmacology method was used to screen the main active components of Danggui Liuhuang Decoction and related targets for the treatment of yin-deficiencyhyperthyroidism.The protein-protein interaction(PPI)network analysis,gene ontology(GO)function and Kyoto encyclopedia of genes and genomes(KEGG)pathway enrichment analysis were performed on the intersection targets to construct the herb-componenttarget-pathway network.Further experimental verification was performed,yin-deficiency-hyperthyroidism rat model was constructed,effect of all-herb decoction and single-herb decoction of Danggui Liuhuang Decoction on body weight,fecal water content and related serological indicators of yin-deficiency-hyperthyroidism rats were evaluated.The effects of all-herb decoction and single-herb decoction on the diversity of intestinal microbial community in rats were revealed by 16S rDNA sequencing.Results A total of 94 active components in Danggui Liuhuang Decoction were screened and 116 targets related to yin-deficiency-hyperthyroidism were found.Enrichment analysis obtained 871 GO items and 158 KEGG related items(P<0.05),including 675 biological processes(BP),77 cellular components(CC),and 119 molecular functions(MF).The pathways with high correlation with Danggui Liuhuang Decoction mainly included inflammation,infection,intestinal disease,cancer and other pathways.The results of anti-yin-deficiencyhyperthyroidism efficacy showed that compared with model group,the net weight gain and fecal moisture content of rats in all-herb decoction high-dose group were significantly increased(P<0.01),levels of triiodothyronine(T3)and thyroxine(T4)were significantly decreased(P<0.01),and the effect was close to the positive drug methimazole group.The results of intestinal flora diversity in rats showed that the richness and diversity of intestinal microorganisms in all-herb decoction high-dose group were close to those in control group.Lachnospiraceae_NK4A136_group was significantly enriched in the intestinal tract of model rats.Akkermansia was significantly increased after administration of all-herb decoction,while Prevotella was significantly increased after administration of single-herb decoction.Conclusion The effect of Danggui Liuhuang Decoction for anti-yin-deficiency-hyperthyroidism is better.Endocrine disorders such as yin-deficiency-hyperthyroidism is likely to cause intestinal microflora disorders,Danggui Liuhuang Decoction may be through the regulation of intestinal microflora in the treatment of yin-deficiency-hyperthyroidism.