Identification and verification of effective components of Huanghuai for dysfunctional uterine bleed

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Objective:The Huanghuai (HH),which is made from the dried roots ofScutellaria baicalensis (Huangqin in Chinese) and the dried flowers and buds of Sophora japonica (Huaihua in Chinese),is a traditional Chinese formula used to treat dysfunctional uterine bleeding (DUB) (Benglou in Chinese) and proven to treat hemostasis effectively in our previous study.Network pharmacology and molecule docking were per-formed to study the underlying mechanism of Huanghuai (HH),and pharmacodynamic experiments were conducted to verify its curative effect.Methods:TCMSP,UniProt,GeneCards,STRING,DAVID databases,and Cytoscape 3.7.2 were utilized for the construction of a compound-target-pathway network.Docking.the potential effective components with potential targets.The HPLC analysis of the potential effective components was performed.In vivo,the hot plate test model was used to study the analgesic activity,the egg white was used to study the swollen reaction in the sole in mice,and the hemostasis effect was studied by the capillary method,tail-breaking method and abortion uterus test.Results:The results showed that six compounds (acacetin,beta-sitosterol,wogonin,baicalein,kaemp-ferol and quercetin) and four potential targets (PTGS2,AKT1,TP53 and TNF) in the compound-target-pathway network were the potential material basis for HH to treat DUB.It can be seen that the binding energy of the acacetin,wogonin,baicalein,beta-sitosterol,kaempferol and quercetin in HH docked with the receptor proteins PTGS2,AKT1,TP53,and TNF were far less than-5.0 kJ/mol,which means the mole-cules have low conformational energy,stable structure and high binding activity.And the result of HPLC analysis showed that acacetin,wogonin,baicalein,kaempferol and quercetin were the potential effective components of the hemostasis mechanism of HH,beta-sitosterol was removed due to low content.In vivo testing of the potential effective components,it revealed that the group of potential effective components identified by HPLC could increase the pain threshold,inhibit the swelling hind paws of mice induced by egg white,reduce the bleeding time and clotting time,reduce uterine bleeding,decrease the uterine weight,increase the content of Ca and ET-1,and reduce the content of NO in uterine homogenate tissue,and decrease of E2 and P content in uterine serum in aborted rats,whose efficacy was equal to HH.Conclusion:The results indicated that HH and potential active ingredient groups obtained from network pharmacology can treat DUB and play a hemostatic effect.The results obtained by network pharmacology have certain reliability.This study provides new indications for further mechanism research of HH on DUB and the development of HH or its components as an alternative therapy for patients with DUB.At the same time,the application of network pharmacology strategy may provide a powerful tool for explor-ing the mechanism of traditional Chinese medicine and discovering new biologically active ingredients.
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