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Shenlingbaizhu (SLBZ) formula, a classical traditional Chinese medicinal (TCM) formula, has been widely used for treating antibiotic-associated diarrhea (AAD). However, the underlying pharmacological mechanisms have not yet been investigated thoroughly. To explore the remission mechanism of SLBZ in the treatment of AAD, we conducted network pharmacological analysis and experimental validation in vitro and in vivo. In this study, the main compounds of SLBZ were identified by ultra-high-performance liquid chromatography-mass spectroscopy (UHPLC-MS) and online databases. The targets of the active components and AAD-related targets were predicted by network pharmacology, and the potential targets of SLBZ against AAD were obtained. Then the core targets were recognized after Protein-Protein Interaction (PPI) analysis. Based on these, gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) signaling pathway analyses were conducted, and the key pathway was screened. Subsequently, molecular docking was performed using Auto Dock Vina to find the key components that played a crucial role in that pathway. Molecular dynamics simulation was performed by Gromacs software to detect the binding mode. Finally, the results were confirmed by in vitro and in vivo experiments. A total of 66 active ingredients of SLBZ were detected by UHPLC-MS, and 128 active ingredients were screened out by network pharmacological analysis. Additionally, 935 drug targets and 1686 AAD-related targets were obtained. Seventy-eight intersected genes were selected as potential therapeutic targets and 19 genes were excavated as core targets. Enrichment analysis revealed PI3K-AKT signaling pathway was the key pathway in SLBZ against AAD. Topological analysis further revealed that JAK2, MTOR, TLR4, and SYK were the key targets affected by SLBZ on the PI3K-AKT pathway, and 52 components of SLBZ were associated with them. Molecular docking and dynamics simulation revealed strong binding affinities between MTOR and diosgenin. Subsequently, after SLBZ treatment, the expression levels of JAK2, MTOR, TLR4, and SYK were found significantly upregulated in the AAD model rats (p < 0.05). The cell experiment further validated the good binding ability between MTOR and diosgenin. We demonstrate that the therapeutic effect of SLBZ on AAD was achieved in part by inhibiting the PI3K-AKT pathway. [Display omitted] • Ultra-high-performance liquid chromatography-mass spectroscopy was used to identify the main compounds of Shenlingbaizhu (SLBZ) formula. • Network pharmacology and in vivo experiments showed that the therapeutic effect of SLBZ on antibiotic-associated diarrhea was achieved in part by inhibiting the PI3K/AKT pathway. • This study revealed for the first time that diosgenin in SLBZ was the main ingredient that inhibited the PI3K/AKT pathway. [ABSTRACT FROM AUTHOR]

WeiChang'An Pill (WCAP) is used in Traditional Chinese Medicine (TCM) to clinically treat diarrhoea-predominant irritable bowel syndrome (IBS-D); however, the underlying pharmacological mechanisms are unclear to date. To explore the mechanism underlying the therapeutic action of WCAP in IBS-D using a network pharmacology approach and in vivo experiments. The active compounds of WCAP were selected from the TCM Systems Pharmacology Database and TCM Integrated Database, and the potential targets were identified using the Swiss Target Prediction and Similarity Ensemble Approach (SEA) databases. The targets related to IBS-D were mined from the Therapeutic Target Database (TTD), National Center for Biotechnology Information Search database (NCBI), DrugBank database, and DisGeNET database. The intersecting protein-protein interactions (PPIs) of the drug-disease crossover genes were analysed, and the central PPI network was constructed using the String database, version 11.0, and Cytoscape version 3.7.2. Following Gene Ontology and Kyoto Encyclopaedia of Genes and Genomes pathway analyses, the gene-pathway network was constructed for identifying the key target genes and pathways. Based on the results and existing evidence, it was selected the cyclic adenosine monophosphate (cAMP) signalling pathway for further validation using in vivo experiments. A total of 872 targets were identified from the 77 active compounds in WCAP, which shared 78 targets that were predicted to be related to IBS-D. Twenty-one core targets were identified from the PPI network, which was constructed from the common targets. The results of enrichment analysis revealed that HRT2B, ADRA1A, ADRA1D, and CHRM2 could be the key targets of WCAP in IBS-D, and 11 signalling pathways, including the neuroactive ligand-receptor interaction, calcium signalling, and cAMP signalling pathways, were identified as crucial for the therapeutic activity of WCAP in IBS-D. We also identified the possibility of several interactions and crosstalk between the different pathways. Subsequent molecular biology experiments revealed that the expression levels of cAMP, phospho-(Ser/Thr) protein kinase A substrates (p -PKA), 5-hydroxytryptamine, and proteins in the cAMP signalling pathway, including G protein-coupled receptor (GPCR), adenylyl cyclase 5 (AC5), and cAMP-response element binding protein (CREB), were significantly upregulated in rat models of IBS-D following treatment with WCAP (P < 0.05). However, a reverse trend was observed in the expression of nuclear factor kappa-B (NF-κB) (P < 0.05), which could be attributed to the low-grade inflammation that occurs in IBS-D. We demonstrated that WCAP may alleviate the symptoms of diarrhoea and visceral sensitivity in IBS-D by regulating the cAMP signalling pathway. [Display omitted] [ABSTRACT FROM AUTHOR]

Standard therapy for asthma, a highly heterogeneous disease, is primarily based on bronchodilators and immunosuppressive drugs, which confer short-term symptomatic relief but not a cure. It is difficult to discover novel bronchodilators, although potential new targets are emerging. Traditional Chinese Medicine (TCM) formulas have been used to treat asthma for more than 2000 years, forming the basis for representative asthma treatments. Based on the efficacy of TCM formulas, anti-asthmatic herbal compounds bind proteins are potential targets for asthma therapy. This analysis will provide new drug targets and discovery strategies for asthma therapy. A list of candidate herbs for asthma was selected from the classical formulas (CFs) of TCM for the treatment of wheezing or dyspnea recorded in Treatise on Cold Damage and Miscellaneous Diseases (TCDMD) and from modern herbal formulas identified in the SAPHRON TCM Database using the keywords "wheezing" or "dyspnea". Compounds in the selected herbs and compounds that directly bind target proteins were acquired by searching the Herbal Ingredients' Targets Database (HITD), TCM Data Bank (TCMDB) and TCM Integrated Database (TCMID). Therapeutic targets of conventional medicine (CM) for asthma were collected by searching Therapeutic Target Database (TTD), DrugBank and PubMed as supplements. Finally, the enriched gene ontology (GO) terms of the targets were obtained using the Database for Annotation Visualization and Integrated Discovery (DAVID) and protein-protein interactions (PPI) networks were constructed using Search Tool for the Retrieval of Interacting Genes/Proteins (STRING). The effects of two selected TCM compounds, kaempferol and ginkgolide A, on cellular resistance in human airway smooth muscle cells (ASMCs) and pulmonary resistance in a mouse model were investigated. The list of 32 candidate herbs for asthma was selected from 10 CFs for the treatment of wheezing or dyspnea recorded in TCDMD and 1037 modern herbal formulas obtained from the SAPHRON TCM Database. A total of 130 compounds from the 32 selected herbs and 68 herbal compounds directly bind target proteins were acquired from HITD and TCMDB. Eighty-eight therapeutic targets of CM for asthma were collected by searching TTD and PubMed as supplements. DAVID and STRING analyses showed targets of TCM formulas are primarily related to cytochrome P450 (CYP) family, transient receptor potential (TRP) channels, matrix metalloproteinases (MMPs) and ribosomal protein. Both TCM formulas and CM act on the same types of targets or signaling pathways, such as G protein-coupled receptors (GPCRs), steroid hormone receptors (SHRs), and JAK-STAT signaling pathway. The proteins directly targeted by herbal compounds, TRPM8, TRPA1, TRPV3, CYP1B1, CYP2B6, CYP1A2, CYP3A4, CYP1A1, PPARA, PPARD, NR1I2, MMP1, MMP2, ESR1, ESR2, RPLP0, RPLP1 and RPLP2, are potential targets for asthma therapy. In vitro results showed kaempferol (1 × 10−2 mM) and ginkgolide A (1 × 10−5 mM) significantly increased the cell index (P < 0.05 vs. histamine, n = 3) and therefore relaxed human ASMCs. In vivo results showed kaempferol (145 μg/kg) and ginkgolide A (205 μg/kg) significantly reduced pulmonary resistance (P < 0.05 vs. methacholine, n = 6). Potential target discovery for asthma treatment based on the clinical effectiveness of TCM is a feasible strategy. Image 1 • 10 classical formulas for asthma and sovereign medicinal herbs of the classical formulas are discussed. • 130 herb compounds of 32 herbs and 68 targets for asthma are discussed. • 88 modern medicine targets for asthma from TTD and PubMed are discussed. • 18 proteins directly targeted by herbal compounds are potential targets for asthma therapy. [ABSTRACT FROM AUTHOR]

The Chinese herb Qianghuo is an antiphlogistic herb with many effects and complex components. In this study, the chemical composition of Qianghuo and its components in rat plasma after oral administration were investigated using ultra-high-performance liquid chromatography coupled with quadrupole-time-of-flight tandem mass spectrometry (UPLC-Q-TOF-MS/MS). The extracts, blank plasma, and plasma containing the drug were analyzed by mass spectrometry, and data collected in both positive and negative ion modes were analyzed using Masslynx software, and the structures were confirmed by combining the compound fragment ions and mass spectrometry cleavage pathways. A total of 62 in vitro chemical components were identified, including 27 coumarins, 18 organic acids, 5 amino acids, 5 glycosides, 2 flavonoids, 4 nucleotides, and 1 ester, which were summarized from the obtained compounds in terms of their possible cleavage patterns. Among the identified 31 compounds in rat plasma, 21 were prototypes, mostly coumarins, organic acids, and flavonoids, and 10 were metabolites, which were mainly generated via hydroxylation and methylation pathways. Based on these, this study provides a theoretical foundation for quality control and basic research on Qianghuo medicinal substances. [Display omitted] [ABSTRACT FROM AUTHOR]

Background: Elderly rheumatoid arthritis (ERA) population faces multiple treatment dilemma. Here we aim to investigate if Gancao Nourishing-Yin decoction (GCNY) added to methotrexate (MTX) exhibit better effects in an ERA mice model. Methods: ERA mice model was established by adding D-galactose (Dgal) to collagen-induced arthritis (CIA) mice. The model was then assigned into control group (CIA + Dgal), MTX treatment group (MTX), GCNY treatment group (GCNY), and integrative treatment group (MTX + GCNY). Pathological scoring was performed to evaluate the severity between the groups. Proteomic analysis was applied to investigate the secretory phenotype of the ERA mouse model and the underlying mechanism of GCNY, MTX and their combination. Representative cytokines related to proteomic results were further validated by ELISAs. Results: CIA + Dgal mice showed more aggressive joints damage than the CIA mice. Besides changes in the inflammatory pathway such as Pi3k-Akt signaling pathway in both model, differential expressed proteins (DEPs) indicated metabolism-related pathways were more obvious in CIA + Dgal mice. Low-dose MTX failed to show pathological improvement in CIA + Dgal mice, while GCNY improved joints damage significantly. Besides down-regulated inflammation-related targets, GCNY-regulated DEPs (such as Apoc1 ~ 3, Grk2 and Creb3l3) were broadly enriched in metabolism-related pathways. MTX + GCNY showed the best therapeutic effect, and the DEPs enriched in a variety of inflammatory,metabolism and osteoclast differentiation signaling pathway. Notably, MTX + GCNY treatment up-regulated Dhfr, Cbr1, Shmt1 involved in folic acid biosynthesis and anti-folate resistance pathways indicated a coincidence synergic action. ELISAs confirmed CPR and Akt that elevated in CIA + Dgal mice were significantly ameliorated by treatments, and adding on GCNY elevated folic acid levels and its regulator Dhfr. Conclusion: Aging aggravated joints damage in CIA, which probably due to metabolic changes rather than more severe inflammation. GCNY showed significant effects in the ERA mice model especially when integrated with MTX to obtain a synergic action. [ABSTRACT FROM AUTHOR]

As a common malignant tumor, the morbidity and mortality of lung cancer have been rising in recent years. The concept of "premetastatic niche" may lead to a revolutionary change in antitumor metastasis therapeutic strategies. Traditional Chinese medicine with multitargets and lower poisonous agents may be a potentially effective means to intervene in the "premetastatic niche (PMN)" to prevent and treat tumor metastasis. Astragalus polysaccharide (APS) is a substance with strong immune activity in Astragalus membranaceus that has excellent biological activities such as immunomodulation, anti-inflammatory, and antitumor. In this study, we constructed a tumor lung metastasis animal model to explore the intervention mechanism of APS on the premetastatic niche. We found that APS inhibited the formation of the lung premetastatic niche and inhibited the recruitment of myeloid-derived suppressor cells (MDSCs) in the lung. Mechanistically, we showed that the proteins and gene expression of S1PR1, STAT3, and p-STAT3 in the S1PR1/STAT3 signaling pathway were suppressed by APS. In line with the above findings, our results confirmed that APS may inhibit the accumulation of MDSCs in the premetastatic niche through the intervention of the S1PR1-STAT3 signaling pathway to achieve the antitumor effect. [ABSTRACT FROM AUTHOR]

Perioperative or postoperative adjuvant chemotherapy based on 5-fluorouracil (5-FU) is a common first-line adjuvant therapy for gastric cancer (GC). However, drug resistance and the side effects of 5-FU have reduced its efficacy. Among these side effects, gastrointestinal (GI) toxicity is one of the most common. Xianglian Pill (XLP) is a Chinese patent medicine that is commonly used for the treatment of diarrhoea. It can reduce inflammation and has a protective effect on the intestinal mucosa. Recent studies have shown that many components of XLP can inhibite tumor cell growth. However, the therapeutic effect of XLP in combination with 5-FU on GC is unclear. To investigate whether the combination of XLP and 5-FU can enhance anti-GC activity while reducing GI toxicity. XLP was administered orally during intraperitoneal injection of 5-FU in GC mice model. Mice were continuously monitored for diarrhea and xenograft tumor growth. After 2 weeks, the mice were sacrificed and serum was collected to determine interleukin-6 levels. Pathological changes, the expression of pro-inflammatory factors and p38 mitogen-activated protein kinase (MAPK) in GI tissue were determined by Western blot analysis. Pathological changes, apoptosis levels and p38 MAPK expression levels in xenograft tissues were also determined. The results showed that XLP could alleviate GI mucosal injury caused by 5-FU, alleviated diarrhea, and inhibited the expression of nuclear factor (NF)-κB and myeloid differentiation primary response-88. Besides, XLP could promote the 5-FU-induced apoptosis of GC cells and enhance the inhibitory effect of 5-FU on tumor xenografts. Further study showed that XLP administration could regulate the expression of p38 MAPK. XLP in combination with 5-FU could alleviate its GI side effects and enhance its inhibitory effect on xenograft tumor. Moreover, these effects were found to be related to the regulation of the p38 MAPK/NF-κB pathway. [Display omitted] • XLP combined with 5-FU could alleviate the diarrhea and reduce the inflammation of gastrointestinal mucosa in GC mice model. • XLP combined with 5-FU could inhibit the growth of xenogaft and promote tumor cell apoptosis in GC mice model. • Those effects were related to the regulation of p38MAPK/NF-κB signaling pathway by XLP. [ABSTRACT FROM AUTHOR]

Postcontrast acute kidney injury (PC-AKI) is directly caused by the use of contrast, indicating a clear causal relationship between the contrast and the injury. Salvianolic acid B (Sal B), a water-soluble compound of Salvia miltiorrhiza, has a potent anti-inflammatory effect. We conducted a study to explore whether the protective effect of Sal B on iopromide-induced injury in human proximal tubular epithelial cells (HK-2 cells) is related to inhibition of the TLR4/NF-κB/NLRP3 signal pathway. The results showed that 100 μmol/L Sal B counteracted the decrease in cell viability, the increase of ROS and the number of apoptotic cells, and the decrease of mitochondrial membrane potential (ΔΨm) induced by iopromide. Molecular docking analysis showed that Sal B binds TLR4 and NLRP3 proteins. Moreover, 100 μmol/L Sal B also decreased the expression of TLR4, NLRP3, ASC, Caspase-1, IL-18, IL-1β, TNF-α, p-NF-κB, cleaved caspase-3, and the ratio of Bax/Bcl-2 induced by iopromide. TAK-242, a TLR4 antagonist, was added to further explore the mechanism of Sal B. However, the cotreatment group with TAK-242 and Sal B had no significant difference in cell viability and apoptosis rate compared to the treatment group with TAK-242 or Sal B alone. These results indicated that Sal B can inhibit the TLR4/NF-κB/NLRP3 signal pathway, resulting in the alleviation of iopromide-induced HK-2 cell injury. [ABSTRACT FROM AUTHOR]

Background. Cerebral infarction (CI) is a common brain disease in clinical practice, which is mainly due to the pathological environment of ischemia and hypoxia caused by difficult cerebral circulation perfusion function, resulting in ischemic necrosis of local brain tissue and neurological impairment. In traditional Chinese medicine (TCM) theory, CI is mainly due to blood stasis in the brain. Therefore, blood-activating and stasis-dissipating drugs are often used to treat CI in clinical practice. Salvia miltiorrhiza Bunge (SMB) is a kind of traditional Chinese medicine with good efficacy in promoting blood circulation and removing blood stasis, and treatment of CI with it is a feasible strategy. Based on the above analysis, we chose network pharmacology to investigate the feasibility of SMB in the treatment of CI and to study the possible molecular mechanisms by providing some reference for the treatment of CI with TCM. Methods. The active ingredients and related targets of SMB were obtained through the Traditional Chinese Medicine Systems Pharmacology (TCMSP) database, and CI-related targets were obtained from the GeneCards and DisGeNET databases. The target of SMB for the treatment of CI was obtained using Cytoscape software and visualized. GO and KEGG enrichment analysis was performed based on "clusterProfiler" within R, and the prediction results were validated by molecular docking technique. Results. By constructing a compound-target (C-T) network, it was found that the active components in SMB mainly treated CI by regulating key proteins such as AKT1, IL-6, and EGFR. These key proteins mainly involve in pathways such as immune regulation, cancer and lipid metabolism, such as lipid and atherosclerosis, chemical carcinogenesis-receptor activation pathways, and IL-17 signaling pathway. In the GO term, it mainly regulates the response to steroid hormones, membrane rafts, and G protein-amine receptor coupled activity. Eventually, we verified that the luteolin and tanshinone IIA components in SMB have a good possibility of action with AKT1 and IL-6 by in silico techniques, indicating that SMB has some scientificity in the treatment of CI. Conclusion. SMB mainly treats CI by regulating 94 proteins involved in lipid and atherosclerosis, chemical carcinogenesis-receptor activation, and IL-17 signaling pathway. Our research strategy provided a template for the drug development of TCM for the treatment of CI. [ABSTRACT FROM AUTHOR]

Moluodan concentrated pill (MLD) is a traditional herbal formula used in China for the treatment of chronic atrophic gastritis (CAG). However, its pharmacological mechanism of action remains unclear. The aim of this study was to investigate the therapeutic effect and mechanism of action of MLD in the treatment of CAG using network pharmacology and in vivo experiments. The active compounds of MLD were determined using network pharmacology, utilizing various Chinese medicine databases such as the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform, Traditional Chinese Medicine Integrated Database, Integrative Pharmacology-based Research Platform of Traditional Chinese Medicine, and a comprehensive database of Traditional Chinese Medicine on Immuno-Oncology. The compounds found in the root of Anemone altaica Fisch. were extracted from the China National Knowledge Infrastructure literature database. Additionally, the Swiss Target Prediction database and Similarity Ensemble Approach were employed to identify the potential targets of these components. CAG-related targets were gathered from the GeneCards and DisGeNET databases. Protein-protein interactions (PPIs) of the genes associated with the drug-disease crossover were examined, and a core PPI network was constructed using the STRING database (version 11.5) and Cytoscape (version 3.7.2). A gene-pathway network was established to identify significant target genes and pathways through Gene Ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. Finally, based on these findings and existing data, the tumor necrosis factor (TNF) signaling pathway was selected for further validation through in vivo experiments. A total of 724 active molecules in MLD yielded 961 identified target genes, of which 179 were found to be potentially associated with CAG. From the common targets, a PPI network revealed ten core targets. Enrichment analysis suggested that MLD may primarily target TNF and AKT in the treatment of CAG. Essential signaling pathways, such as the PI3K-AKT and TNF pathways, were found to be crucial for the therapeutic effects of MLD on CAG. Furthermore, potential interactions and crosstalk between these pathways were identified. Moreover, we confirmed that MLD effectively improved gastric mucosa atrophy and cellular ultrastructural damage, while increasing pepsinogen secretion and decreasing gastrin, somatostatin, and motilin levels. Subsequent molecular biology studies in rat models of CAG demonstrated that MLD treatment significantly reduced the expression levels of TNF-α, phosphatidylinositol 3′-kinase (PI3K), and phosphorylated Akt (P < 0.05). Notably, the expression of nuclear factor kappa-B (NF-κB) exhibited a contrasting trend (P < 0.05), potentially associated with the crucial tumor suppressor role of NF-κB p105. This study provides evidence that MLD effectively alleviates stomach mucosal atrophy through modulation of the TNF/PI3K/AKT signaling pathway. These findings establish a solid theoretical foundation for the practical management of CAG. [Display omitted] • A rat model of chronic atrophic gastritis was established by the MNNG multifactorial modeling method. • The efficacy and mechanism of Moluodan for chronic atrophic gastritis were revealed in vivo experiments for the first time. • Moluodan exerts gastric mucosal protective effects via the TNF/PI3K/AKT signaling pathway. [ABSTRACT FROM AUTHOR]

Background. Qixuehe capsule (QXH), a Chinese patent medicine, has been demonstrated to be effective in the treatment of menstrual disorders. In traditional Chinese medicine (TCM) theory, qi stagnation and blood stasis syndrome (QS-BSS) is the main syndrome type of menstrual disorders. However, the pharmacodynamic effect of QXH in treating QS-BSS is not clear, and the main active compounds and underlying mechanisms remain unknown. Methods. A rat model of QS-BSS was established to evaluate the pharmacodynamic effect of QXH. Thereafter, a network pharmacology approach was performed to decipher the active compounds and underlying mechanisms of QXH. Results. QXH could significantly reduce the rising whole blood viscosity (WBV) and plasma viscosity (PV) but also normalize prothrombin time (PT), activated partial thromboplastin time (APTT), thrombin time (TT), and fibrinogen (FIB) content in QS-BSS rats. Based on partial least-squares-discriminant analysis (PLS-DA), the low-dose QXH-intervened (QXH-L) and the high-dose QXH-intervened (QXH-H) groups seemed the most effective by calculating the relative distance to normality. Through network pharmacology, QXH may improve hemorheological abnormality mainly via 185 compounds-51 targets-28 pathways, whereas 184 compounds-68 targets-28 pathways were associated with QXH in improving coagulopathy. Subsequently, 25 active compounds of QXH were verified by UPLC-Q/TOF-MS. Furthermore, 174 active compounds of QXH were shared in improving hemorheological abnormality and coagulopathy in QS-BSS, each of which can act on multiple targets to be mainly involved in complement and coagulation cascades, leukocyte transendothelial migration, PPAR signaling pathway, VEGF signaling pathway, and arachidonic acid metabolism. The attribution of active compounds indicated that Angelicae Sinensis Radix (DG), Paeoniae Radix Rubra (CS), Carthami Flos (HH), Persicae Semen (TR), and Corydalis Rhizoma (YHS) were the vital herbs of QXH in treating QS-BSS. Conclusion. QXH can improve the hemorheology abnormality and coagulopathy of QS-BSS, which may result from the synergy of multiple compounds, targets, and pathways. [ABSTRACT FROM AUTHOR]

Abstract Ethnopharmacological relevance Danggui buxue Decoction (DBD) has been frequently used to treat with blood deficiency, which consisted of Danggui (DG) and Huangqi (HQ) at a ratio of 1:5. Accumulating evidence showed that blood deficiency in traditional Chinese medicine (TCM) was similar to anemia in modern medicine. Aim of the study The purpose of this study was to explore its therapeutic mechanism of with network pharmacology approach. Materials and methods We explored the chemical compounds of DBD and used compound ADME screening to identify the potential compounds. Targets for the therapeutic actions of DBD were obtained from the PharmMapper, Swiss, SEA and STITCH. GO analysis and pathway enrichment analysis was performed using the DAVID webserver. Cytoscape was used to visualize the compound-target-pathway network for DBD. The pharmacodynamics and crucial targets were also validated. Results Thirty-six potential active components in DBD and 49 targets which the active components acted on were identified. 47 KEGG pathways which DBD acted on were also come to light. And then, according to KEGG pathway annotation analysis, only 16 pathways seemed to be related to the blood nourishing effect of DBD, such as PI3K-AKT pathway, and so on. Only 32 targets participated in these 16 pathways and they were acted on by 29 of the 36 active compounds. Whole pharmacodynamic experiments showed that DBD had significant effects to blood loss rats. Furthermore, DBD could promote the up-regulation of hematopoietic and immune related targets and the down-regulation of inflammatory related targets. Significantly, with the results of effective rate, molecular docking and experimental validation, we predicted astragaloside IV in HQ, senkyunolide A and senkyunolide K in DG might be the major contributing compounds to DBD's blood enriching effect. Conclusion In this study, a systematical network pharmacology approach was built. Our results provided a basis for the future study of senkyunolide A and senkyunolide K as the blood enriching compounds in DBD. Furthermore, combined network pharmacology with validation experimental results, the nourishing blood effect of DBD might be manifested by the dual mechanism of enhancing immunity and promoting hematopoiesis. Graphical abstract fx1 [ABSTRACT FROM AUTHOR]

Guizhi Fuling Capsule (GFC) is a famous traditional Chinese medicine (TCM) formula recorded in Synopsis of the Golden Chamber, which has achieved obvious effects in the treatment of uterine fibroids (UFs). Mediator complex subunit 12 (Med12) mutations were closely related to UFs in 85% of fibroid cases. The Wnt/β-Catenin signaling pathway plays an important role in the occurrence and development of UFs. This study aims to explore the pharmacological mechanism of GFC against UFs in which the Med12-mediated Wnt/β-Catenin pathway is involved. Med12 was silenced in uterine fibroid cells (UFCs) using a lentivirus-based Med12 gene-specific RNA interference (RNAi) strategy. Cell proliferation was performed by CCK-8 assay, cell apoptosis and cell cycle were measured by flow cytometry. The rat model of UFs was established by injecting estradiol benzoate and progesterone. Forty-eight rats were divided into six groups, the low-dose GFC (L-GFC) group, the medium-dose GFC (M-GFC) group and the high-dose GFC (H-GFC) group were intragastrically treated with GFC solution at 0.25 g/kg, 0.50 g/kg and 1.00 g/kg per day for 8 weeks, the positive control (PC) group was administrated with mifepristone (2.70 mg/kg/day), the normal control (NC) group and the model control (MC) group were given equal volume of normal saline once a day for 8 weeks. The histopathological changes of uterine tissues were evaluated by H&E staining. The expression of Med12 in uterine tissues were detected by immunohistochemistry. The protein and mRNA levels of associated genes were evaluated by western bolt and real time-PCR, respectively. Related indicators involved in Wnt/β-Catenin pathway, such as Wnt1, β-Catenin, Cyclin D1, TCF1/TCF7 and C-myc, were compared among different groups. The Wnt/β-Catenin signaling pathway was inhibited after Med12 gene was knocked out in UFCs. GFC-containing serum could induce cell apoptosis, make the cell cycle stagnated in G0/G1 phase to inhibiting the proliferation and reduce the expression of Wnt1, β-Catenin, Cyclin D1, TCF1/TCF7, and C-myc in control-shRNA cells, while had no significant effect on Med12-shRNA cells. Compared with the MC group, the weight, endometrial thickness, and pathological structure of the uterus in the GFC treated groups were significantly improved. The expression of Med12, Wnt1, β-Catenin, Cyclin D1, TCF1/TCF7, and C-myc that related to Wnt/β-Catenin pathway in the GFC treated groups were decreased with the increase of dosage administration. GFC inhibited UFs growth, which was directly associated with Med12 modulated Wnt/β-Catenin signaling pathway. This study provided new perspective to understand the therapeutic mechanism of UFs. [Display omitted] • Guizhi Fuling Capsule (GFC) inhibited the proliferation of uterine fibroid cells by disrupting the normal Med12 expression. • GFC induced apoptosis and made the cell cycle stagnated in G0/G1 phase of uterine fibroid cells. • GFC improved the weight, endometrial thickness, and pathological structure of the uterus in uterine fibroids model rats. • GFC regulated the Med12-mediated Wnt/β-Catenin pathway to treating uterine fibroids in model rats. [ABSTRACT FROM AUTHOR]