Molecular Mechanism of Caulis Spatholobi in the Treatment of Chronic Myeloid Leukemia based on Network Pharmacology and Experimental Verification.

BACKGROUND: Caulis Spatholobi is one of the necessary Chinese herbal medicines for hematologists in the treatment of malignant tumors, but its potential targets and molecular mechanisms need further exploration. OBJECTIVE: This study aimed to predict the relevant targets of the treatment of chronic myeloid leukemia (CML) with Caulis Spatholobi by applying the network pharmacology method, and in vitro cell experiments were conducted to verify the mechanism of Caulis Spatholobi in the treatment of CML. METHODS: TCMSP, ETCM, Genecards, and GisGeNET databases were used to obtain relevant targets of Caulis Spatholobi in the treatment of CML. Go and KEGG analyses were performed using the David database. Using Cytoscape 3.7.2, the "active compounds-targets-pathways" network was constructed. Further validation was carried out by pharmacological experiments in vitro. The proliferation and apoptosis of K562 cells were observed by the MTT method and Hoechst 33242 fluorescence staining method. The predicted targets and their related signal pathways were verified by western blotting. RESULTS: In this study, 18 active compounds and 43 potential targets were obtained. The results of the MTT method showed that compared with the normal control group, 62.5-500 µg/mL alcohol extract of Caulis Spatholobi had an obvious inhibitory effect on K562 and the IC50 value was less than 100 µg/mL. The Hoechst 33242 fluorescence staining method showed that the alcohol extract of Caulis Spatholobi could promote apoptosis. The results of western blotting showed that compared with the normal control group, the expressions of Bax and Caspase-3 proteins in the 62.5 and 125 µg/mL alcohol extract of Caulis Spatholobi groups were significantly up-regulated (p < 0.05). The expression of Bcl-2 in the 125 µg/mL alcohol extract of the Caulis Spatholobi group was significantly down-regulated (p < 0.01), and the expression of Bcl-2 in the 62.5 and 31.25 µg/mL alcohol extract of Caulis Spatholobi groups was also significantly down-regulated (p < 0.05). It showed that the ethanol extract of Caulis Spatholobus could promote apoptosis by up-regulating the expression of Bax and caspase-3 and down-regulating the expression of the Bcl-2 protein. CONCLUSION: The treatment of Caulis Spatholobi for CML has the characteristics of multi-targets and multi-pathways. The results of in vitro pharmacological experiments demonstrated that its mechanism of action might be based on the expression of key target proteins, such as Caspase-3, Bcl-2, and Bax, thereby inhibiting cell proliferation and promoting cell apoptosis, which provides a scientific basis for the treatment of CML.

The Potential Mechanisms of Qufeng Zhitong Capsule against Rheumatoid Arthritis Based on Network Pharmacology and In Vitro Experiments.

Qufeng Zhitong capsule (QFZTC) is a traditional Chinese herbal formula with potential therapeutic efficacy in rheumatoid arthritis (RA). This study seeks to clarify the potential effects and mechanisms of QFZTC against RA. Active compounds and targets of QFZTC were retrieved from the Herbal Ingredients' Targets (HIT), Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP), and Traditional Chinese Medicine Integrated Database (TCMID) databases. RA-related targets were searched on GeneCards and DisGeNET databases. Protein-protein interaction (PPI) network was established using the STRING database. Gene ontology (GO) and Kyoto Encyclopedia of Gene and Genome (KEGG) enrichment analyses were performed on hub targets. Molecular docking was conducted on hub targets and active compounds. High-performance liquid chromatography (HPLC) was applied to characterize the active compounds in QFZTC. RA-fibroblast like synoviocytes (RA-FLSs) were cultured and treated by QFZTC-containing serum, in which proinflammatory cytokines and hub targets were detected. Cell viability was determined by cell counting kit-8 (CCK-8) assay. A total of 360 active compounds and 445 potential targets are identified for QFZTC against RA. Protein-protein interaction (PPI) network determined five hub targets, interleukin 6 (IL6), IL1B, VEGFA, JUN, and tumor necrosis factor (TNF). GO and KEGG analyses revealed that the MAPK pathway may be a critical signaling in QFZTC treating RA. Molecular docking showed that luteolin, kaempferol, and myricetin has good affinity with TNF, and they were identified by HPLC. In vitro experiments confirmed that QFZTC restrained the cell viability and inflammation in RA. This study revealed the active compounds and molecular targets for QFZTC treating RA. QFZTC is a promising drug and ameliorates RA by inhibiting inflammatory response.

Mechanism Study of Polydatin in Treating Spinal Cord Injury by Modulating Mitochondrial Membrane Potential Based on Network Pharmacology and Molecular Docking.

Spinal cord injury (SCI) is one of the most devastating central lesions, and mitochondrial function plays an important role in secondary injury after SCI. Polydatin (PD) is a natural glycosylated precursor of resveratrol, showing mitochondrial preservation effects in the central nervous system. This study aimed to identify the hub target genes of PD on mitochondrial membrane potential (MMP) in SCI. A comprehensive analysis was performed on SCI-related genes, MMP-related genes, and PD targets screening from public databases. Differential expression analysis was conducted to identify differentially expressed genes (DEGs) in SCI. Gene set enrichment analysis (GSEA) and gene set variation analysis (GSVA) were employed to assess pathway enrichment. Protein-protein interaction (PPI) network analysis and molecular docking were conducted to identify key genes and evaluate the binding affinity between PD and hub genes. A total of 16,958 SCI-related genes, 2,786 MMP-related genes, 318 PD-related target genes, and 7229 DEGs were identified. Intersection analysis revealed 46 genes common to all four categories. GSEA and GSVA analysis identified significant enrichment of pathways associated with suppressed and activated SCI biological processes. The PPI network analysis identified seven core hub genes: EGFR, SRC, VEGFA, STAT3, ERBB2, TP53, and RHOA. Molecular docking revealed strong binding affinities between PD and ERBB2, EGFR, and RHOA. The findings based on computational investigation from public databases suggest that PD may have therapeutic potential for SCI by modulating MMP. These results contribute to the understanding of SCI pathogenesis and the development of novel therapeutic strategies.

Jiangu granules ameliorate postmenopausal osteoporosis via rectifying bone homeostasis imbalance: A network pharmacology analysis based on multi-omics validation.

BACKGROUND: Postmenopausal osteoporosis (PMOP) is a series of reactions to bone homeostasis dysregulation mediated by estrogen deficiency in elderly women. Jiangu granules, a traditional Chinese medicine formula, has been proven as an effective treatment approach for PMOP, which still needs more research iin its complex regulatory mechanisms. PURPOSE: Our study aimed to identify the putative targets and regulatory mechanisms of Jiangu granules in PMOP treating. METHODS: We utilized the NHANES database to compare the clinical information of normal population and PMOP patients. Associated with transcriptomics and proteomic data, we identified the PMOP-related genes, and further studied them with bioinformatic methods including and prognosis model. Network pharmacology was applied for confirming the action targets of Jiangu granules in PMOP. We verified the safety and effectiveness in PMOP treatments of Jiangu granules, and also demonstrated our hypothesis in rats. RESULTS: We discovered that the PMOP patients had higher monocytes than the normal women. Moreover, the transcriptomics and proteomic analysis suggested that the dysregulation of PMOP-related genes expression was associated with monocytes, and the Notch pathway were the critical targets representing bone homeostasis imbalance highly involved in the occurrence of PMOP. We also ascertained network pharmacology results further revealing that Jiangu granules might treat PMOP via recovering the bone homeostasis imbalance identified above. In vivo experiments, we confirmed the high efficacy which mainly resulted from function in mitigating the imbalance in bone homeostasis by recovering the normal expression of PMOP-related genes associated with monocytes, Notch, and steroid pathway in the rat models. CONCLUSION: Our finding underscored the clinical potential of Jiangu granules in treating PMOP, and enriched the comprehension of the related pathogenic and therapeutic mechanisms.

Study on the underlying mechanism of Poria in intervention of arrhythmia zebrafish by integrating metabolomics and network pharmacology.

BACKGROUND: Poria is an herb with both medicinal and dietary application. It has been used in various traditional Chinese patent medicines and medicinal decoctions for the treatment of arrhythmia. However, the specific mechanisms involved in the antiarrhythmic effects of Poria have, until now, remained unknown. PURPOSE: This present study sought to explore the potential compounds and mechanisms by which Poria ameliorates BaCl2-induced arrhythmia. METHOD: We initiated by using network pharmacology to predict probable components, targets, and associated signaling pathways before optimizing the extraction process of Poria. We then applied Poria extract to a zebrafish model of BaCl2-induced arrhythmia. We combined network pharmacology and untargeted metabolomic analysis to predict the likely signaling and metabolic pathways governed by Poria. Finally, we verified putative mRNA and metabolite targets of Poria involved in the intervention of arrhythmia by PCR, molecular docking, enzymatic inhibition and targeted metabolomics. RESULTS: We found that triterpenoids may be the main components of Poria responsible for its effects on arrhythmia, and that the optimal extraction process for its water extract is 9 volumes of water with the 7.5 h first extraction period, and the second extraction period of 1.5 h. Through experimentation, we have found that the water extract of Poria can interfere with BaCl2 induced arrhythmia in zebrafish by significantly increasing the heart rate, reducing the SV-BA distance, and pericardial area, and the degree of cardiomyocyte apoptosis in zebrafish. In addition, PCR validation revealed that Poria can regulate the calcium signaling pathway by upregulating the gene expression levels of ADRB1, HTR7, CALMB1, and PPP3CA. Meanwhile, through molecular docking and enzyme activity inhibition, it was found that the compounds in Poria can bind to ADRB1, HTR7, CALMB1, and PPP3CA, respectively. Targeted metabolism confirmed that Poria can downregulate the synthesis of cAMP in the calcium signaling pathway, as well as the synthesis of valine and isoleucine in valine, leucine, and isoleucine biosynthesis. CONCLUSION: Overall, our study indicates that Poria exerts its antiarrhythmic effect through regulating the calcium signaling pathway and valine, leucine, and isoleucine biosynthesis. Our findings not only establish a mechanistic framework for elucidating the antiarrhythmic effects of Chinese patent medicine containing Poria, but also provide a medicinal basis for the study of its dual use as medicine and food.

Qinzhizhudan formula dampens inflammation in microglia polarization of vascular dementia rats by blocking MyD88/NF-κB signaling pathway: Through integrating network pharmacology and experimental validation.

ETHNOPHARMACOLOGICAL RELEVANCE: Qinzhizhudan Formula (QZZD) is composed of Scutellaria baicalensis Georgi (Huang Qin) extract, Gardenia jasminoides (Zhizi) extract and Suis Fellis Pulvis (Zhudanfen) (ratio of 4:5:6). This formula is optimized from Qingkailing (QKL) injection. Regarding brain injury, QZZD is protective. However, the mechanism by which QZZD treats vascular dementia (VD) has not been elucidated. AIM OF THE STUDY: To ascertain QZZD's effect on the treatment of VD and further investigate the molecular mechanisms. MATERIALS AND METHODS: In this study, we screened the possible components and targets of QZZD against VD and microglia polarization using network pharmacology (NP), then an animal model of bilateral common carotid artery ligation method (2VO) was induced. Afterward, The Morris water maze was employed to evaluate cognitive ability, and pathological alterations in the CA1 area of the hippocampus were detected using HE and Nissl staining. To confirm the affect of QZZD on VD and its molecular mechanism, the contents of inflammatory factors IL-1ß, TNF-α, IL-4, and IL-10 were performed to detect by ELISA, the phenotype polarization of microglia cells was detected by immunofluorescence staining, and the expressions of MyD88, p-IκBα and p-NF-κB p65 in brain tissue were detected by western blot. RESULTS: A total of 112 active compounds and 363 common targets of QZZD, microglia polarization, and VD were identified, according to the NP analysis. 38 hub targets were screened out from the PPI network. GO analysis and KEGG pathway analysis showed that QZZD may regulate microglia polarization through anti-inflammatory mechanism such as Toll-like receptor signaling pathway and NF-κB signaling pathway. The further results showed that QZZD can alleviate the memory impairment induced by 2VO. QZZD profoundly rescued brain hippocampus neuronal damage and increased the number of neurons. These advantageous outcomes were linked to the control of microglia polarization. QZZD decreased M1 phenotypic marker expression while increasing M2 phenotypic marker expression. QZZD may controll the polarization of the M1 microglia by blocking the core part of Toll-like receptor signaling pathway, that is the MyD88/NF-κB signaling pathway, which reduced the neurotoxic effects of the microglia. CONCLUSION: Here, we explored the anti-VD microglial polarization characteristic of QZZD for the first time and clarified its mechanisms. These findings will provide valuable clues for the discovery of anti-VD agents.

Qinlian hongqu decoction ameliorates hyperlipidemia via the IRE1-α/IKKB-ß/NF-κb signaling pathway: Network pharmacology and experimental validation.

ETHNOPHARMACOLOGICAL RELEVANCE: Qinlian Hongqu decoction (QLHQD) is a traditional Chinese medicine (TCM) formula. It has previously been found to mitigate hyperlipidemia, although its mechanism requires further clarification. AIM OF THE STUDY: This study explored QLHQD's mechanism in treating hyperlipidemia based on network pharmacology and experimental validation. MATERIALS AND METHODS: The components of QLHQD were analyzed by means of ultrahigh performanceliquid chromatography-quadrupole/orbitrapmass spectrometry (UHPLC-Q-Orbitrap-HRMS) and the targets of hyperlipidemia were predicted using the Swiss ADME, GeneCards, OMIM, DrugBank, TTD, and PharmGKB databases. A drug-component-target-disease network was constructed using Cytoscape v3.7.1. Moreover, Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) enrichment analyses were performed using the Bioinformatics platform. Based on the KEGG results, the non-alcoholic fatty liver disease signaling pathways were selected for experimental validation in an animal model. RESULTS: We identified 34 components of QLHQD, 94 targets of hyperlipidemia, and 18 lipid metabolism-related pathways from the KEGG analysis. The results of the animal experiment revealed that QLHQD alleviated lipid metabolism disorders, obesity, insulin resistance, and inflammation in rats with hyperlipidemia induced by high-fat diets. Additionally, it reduced the expression of IRE1-α, TRAF2, IKKB-ß, and NF-κB proteins in the liver of hyperlipidemic rats. CONCLUSION: QLHQD is able to significantly mitigate hyperlipidemia induced via high-fat diets in rats. The mechanism of action in this regard might involve regulating the IRE1-α/IKKB-ß/NF-κB signaling pathway in the liver, thereby attenuating inflammatory responses and insulin resistance.

Integration of network pharmacology and proteomics to elucidate the mechanism and targets of traditional Chinese medicine Biyuan Tongqiao granule against allergic rhinitis in an ovalbumin-induced mice model.

ETHNOPHARMACOLOGICAL RELEVANCE: Biyuan Tongqiao granule (BYTQ) is a traditional Chinese medicine that has been used in China to clinically treat patients with allergic rhinitis (AR), yet its underlying mechanism and targets remains unclear. AIM OF THE STUDY: The study aimed to investigate the potential mechanism of BYTQ against AR using the ovalbumin (OVA) -induced AR mice model. Integrating network pharmacology and proteomics to investigate possible targets of BYTQ for AR. MATERIALS AND METHODS: The compounds in BYTQ were analyzed using UHPLC-ESI-QE-Orbitrap-MS. The OVA/Al(OH)3 were used to induce the AR mice model. The nasal symptoms, histopathology, immune subsets, inflammatory factors, and differentially expressed proteins were examined. Proteomics analysis elucidated the potential mechanisms of BYTQ to improve AR, which was further validated by Western blot (WB) assay. The compounds and potential targets of BYTQ were systematically elucidated by integrating network pharmacology and proteomics analysis to explore the mechanism. The binding affinity between key potential targets and corresponding compounds was then validated using molecular docking. Molecular docking results were verified by a western blotting and cellular thermal shift assay (CETSA). RESULTS: A total of 58 compounds were identified from BYTQ. BYTQ significantly suppressed AR symptoms by inhibiting the release of OVA-specific immunoglobulin E (IgE) and histamine, improving the pathological injury of nasal mucosal tissue, and regulating the proportions of lymphocytes to maintain immune balance. Proteomics analysis showed that the cell adhesion factors and focal adhesion pathway might be potential mechanism of BYTQ against AR. The levels of E-selectin, vascular endothelial cell adhesion molecule-1 (VCAM-1), and intercellular adhesion molecule-1 (ICAM-1) proteins in the nasal mucosal tissue were significantly downregulated in the BYTQ-H group compared to the AR group. Integrating network pharmacology and proteomics analysis identified that SRC, PIK3R1, HSP90AA1, GRB2, AKT1, MAPK3, MAPK1, TP53, PIK3CA, and STAT3 may be potential protein targets for BYTQ to treat AR. Molecular docking analysis indicated that the active compounds of BYTQ could bind tightly to these key targets. In addition, BYTQ could inhibit OVA-induced phosphorylation levels of PI3K, AKT1, STAT3 and ERK1/2. The CETSA data suggested that BYTQ could improve the heat stability of PI3K, AKT1, STAT3 and ERK1/2. CONCLUSIONS: BYTQ suppresses E-selectin and VCAM-1 and ICAM1 expression by regulating PI3K/AKT and STAT3/MAPK signaling pathways, thus alleviating inflammation in AR mice. BYTQ is the aggressive treatment for AR.

Network pharmacology and transcriptomics to determine Danggui Yifei Decoction mechanism of action for the treatment of chronic lung injury.

ETHNOPHARMACOLOGICAL RELEVANCE: Several children with pneumonia (especially severe cases) have symptoms of cough and expectoration during the recovery stage after standard symptomatic treatment, which eventually results in chronic lung injury. Danggui yifei Decoction (DGYFD), a traditional Chinese formula, has shown clinical promise for the treatment of chronic lung injury during the recovery stage of pneumonia, however, its mechanism of action is yet to be deciphered. AIM OF THIS STUDY: To investigate the therapeutic mechanism of DGYFD for the treatment of chronic lung injury by integrating network pharmacology and transcriptomics. MATERIALS AND METHODS: BALB/c mice were used to establish the chronic lung injury mouse model by intratracheal instillation of lipopolysaccharide (LPS). Pathological analysis of lung tissue, lung injury histological score, lung index, protein levels in bronchoalveolar lavage fluid (BALF), immunohistochemical staining, blood rheology, inflammatory cytokines, and oxidative stress levels were used to evaluate the pharmacological effects of DGYFD. Chemical components of DGYFD were identified using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Integrated network pharmacology together with transcriptomics was used to predict potential biological targets. Western blot analysis was used to verify the results. RESULTS: In this study, we demonstrated that DGYFD could improve lung injury pathological changes, decreases lung index, down-regulate NO and IL-6 levels, and regulate blood rheology. In addition, DGYFD was able to reduce the protein levels in BALF, up-regulate the expression levels of occludin and ZO-1, improve the ultrastructure of lung tissues, and reverse the imbalance of AT I and AT II cells to repair the alveolar-capillary permeability barrier. Twenty-nine active ingredients of DGYFD and 389 potential targets were identified by UPLC-MS/MS and network pharmacology, and 64 differentially expressed genes (DEGs) were identified using transcriptomics. GO and KEGG analysis revealed that the MAPK pathway may be the molecular target. Further, we found that DGYFD inhibits phosphorylation levels of p38 MAPK and JNK in chronic lung injury mouse models. CONCLUSIONS: DGYFD could regulate the imbalance between the excessive release of inflammatory cytokines and oxidative stress, repair the alveolar-capillary permeability barrier and improve the pathological changes during chronic lung injury by regulating the MAPK signaling pathway.

Symptom-oriented network pharmacology revealed the mechanism of HuangQi-DanShen herb pair against cerebral ischemia coupled with comprehensive chemical characterization.

ETHNOPHARMACOLOGICAL RELEVANCE: In the clinical practice of traditional Chinese medicine, HuangQi-DanShen (HD) is an important drug pair for the treatment of cerebral ischemia (CI). AIM OF THE STUDY: Elucidate the mechanism of HD against CI based on symptom-oriented network pharmacology coupled with comprehensive chemical characterization. MATERIALS AND METHODS: UHPLC-Q-Exactive Orbitrap-MS technology was firstly used to obtain the chemical profile of HD constituents. A comprehensive strategy combining in-house library, diagnostic ions, Compound Discover software and network databases was then established to identify its chemical constitutes. Symptomatic treatment is a treatment aimed at relieving or eliminating symptoms which is often characterized as a stop-gap measure due to its inability to cure the disease fundamentally. Nevertheless, symptomatic treatment is an indispensable part of clinical practice and has an important place in medical therapeutics. Therefore, network pharmacology technique were used to elucidate molecular mechanisms from the symptoms of CI. Finally, some literatures were further mined to support our conclusions. RESULTS: A total of 190 ingredients were identified in HD. Symptom-oriented network pharmacology analysis indicated that compounds of HD relieved "blood" through the regulation of ADORA2A, ADORA1, PTPN11, MMP9 and EGFR, relieved "qi" via the regulation of ADORA2A, EGFR, MMP9 and CA2. The therapeutic effect of HD on "faint" was linked to PTPN11 and MMP9, while the regulation of "dyskinesia" was related to ADORA2A and EGFR, and ADORA1, PTPN11 and MMP9 were associated withe its effect on "speech disorder". ADORA1, ADORA2A and MMP9 were key to the HD component in treating "visual disturbance". CONCLUSION: The approach of symptom-oriented network pharmacology coupled with comprehensive chemical characterization proposed a further orientation for exploring the mechanisms of HD against CI.

Network pharmacology, molecular docking and experimental verification of the mechanism of huangqi-jixuecao herb pair in treatment of peritoneal fibrosis.

ETHNOPHARMACOLOGICAL RELEVANCE: The Huangqi-Jixuecao herb pair (HQJXCHP) is a traditional herbal formula composed of two widely applied TCM prescriptions, Huangqi (Astragalus membranaceus (Fisch.) Bunge) and Jixuecao (Centella asiatica (L.) Urb.), used for hundreds of years to replenish qi and clear away heat. However, the therapeutic effects of HQJXCHP against peritoneal fibrosis (PF) and potential targets are currently unclear. AIMS OF THE STUDY: The main objective of this study was preliminary prediction and validation of the effects and molecular mechanisms of action of HQJXCHP against PF based on network pharmacology analysis and experimental verification. MATERIALS AND METHODS: The ingredients of HQJXCHP were analyzed via HPLC-Q-TOF/MS. Bioactive compounds of HQJXCHP used for network pharmacology analysis were obtained from the TCMSP database. HQJXCHP-related therapeutic targets in PF were obtained from the GeneCards, OMIM, Therapeutic Targets and PharmGkb databases. Therapeutic target-related signaling pathways were predicted via GO and KEGG pathway enrichment analyses. The targets of HQJXCHO were further validated in a PDS-induced PF mouse model in vivo and PMCs MMT model in vitro. RESULTS: A total of 23 bioactive compounds of HQJXCHP related 188 target genes were retrieved. The HQJXCHP compound-target and PF-related target networks identified 131 common target genes. Subsequent protein-protein interaction (PPI) network analysis results disclosed Akt1, TP53, TNF, VEGFA and CASP3 as the top five key targets of HQJXCHP. Further molecular docking data revealed strong affinity of the two key compounds of HQJXCHP, quercetin and kaempferol, for these key targets. GO and KEGG pathway enrichment analyses further showed that PI3K/Akt, IL-17, TNF and TLR pathways contribute to the therapeutic effects of HQJXCHP on PF. An in vivo PDS-induced PF mouse model and in vitro PMCs mesothelial-to-mesenchymal transition (MMT) model with or without HQJXCHP intervention were used to confirm the effects and mechanisms of action of HQJXCHP. Western blot and qRT-PCR results showed that HQ, JXC and HQJXCHP reduced PDS-induced inflammatory cell aggregation and peritoneal thickening through suppressing the MMT process, among which HQJXCHP exerted the greatest therapeutic effect. Moreover, HQJXCHP inhibited activation of the PI3K/Akt, IL-17, TNF and TLR signaling pathways induced by PDS. CONCLUSIONS: This is the first study to employ network pharmacology and molecular docking analyses to predict the targets of HQJXCHP with therapeutic effects on PDS-related PF. Data from in vivo and in vitro validation experiments collectively showed that HQJXCHP delays the PF process through inhibiting PI3K/Akt, IL-17, TNF and TLR signaling pathways. Overall, our findings highlight the successful application of network pharmacology theory to provide a scientific basis for clinical utility of HQJXCHP against PF.

Based on network pharmacology and experiments to explore the underlying mechanism of Mahonia bealei (Fortune) Carrière for treating alcoholic hepatocellular carcinoma.

ETHNOPHARMACOLOGICAL RELEVANCE: Mahonia bealei (Fortune) Carrière (M. bealei) is a traditional medicine widely used by the Hmong community in Guizhou. It possesses diverse biological activities and shows promise in cancer treatment; however, contemporary pharmacological research in this area is lacking. AIMS OF THE STUDY: This study aimed to investigate the effects and underlying mechanisms of M. bealei on alcoholic hepatocellular carcinoma (HCC). MATERIALS AND METHODS: We initially employed the LC-MS/MS method to identify the compounds present in M. bealei serum. Subsequently, its potential targets were predicted using public databases. Bioinformatics and network pharmacology approaches, such as univariate Cox regression and random forest (RF) algorithms, were utilized to identify differentially expressed genes (DEGs) associated with the prognosis of alcoholic HCC. Survival curve and receiver operating characteristic (ROC) analyses were conducted using alcoholic HCC-related data from TCGA and GEO to determine the diagnostic value of the identified DEGs. Molecular docking using the CDOCKER approach based on CHARMm was performed to validate the affinity between the predictive compounds and targets. Additionally, we evaluated the impact of M. bealei on cell proliferation, migration, and conducted western blot assays. RESULTS: The LC-MS/MS approach identified 17 therapeutic components and predicted 483 component-related targets, of which 63 overlapped with alcoholic HCC targets and were considered potential therapeutic targets. GO and KEGG pathway analysis revealed significant associations between the 63 overlapping targets and alcoholic HCC progression. Through various approaches in the Cytoscape 3.9.0 software, we confirmed 9 hub genes (CDK1, CXCR4, DNMT1, ESR1, KIT, PDGFRB, SERPINE1, TOP2A, and TYMS) as core targets. TOP2A and CDK1 genes were identified as advantageous for diagnosing alcoholic HCC using univariate Cox regression, RF, survival curve, and ROC analysis. Molecular docking analysis demonstrated strong binding affinity between key bioactive components cyclamic acid, perfluoroalkyl carboxylic acid, perfluorosulfonic acid, alpha-linolenic acid, adenosine receptor antagonist (CGS 15943), and Prodigiosin and TOP2A and CDK1. In vitro experiments confirmed that M. bealei significantly suppressed cell proliferation and migration of HepG2 cells, while downregulating TOP2A and CDK1 expression. CONCLUSION: This study highlights the potential of M. bealei as a natural medicine for the treatment of alcoholic HCC. Six compounds (cyclamic acid, perfluoroalkylic carboxylic acids, perfluorosulfonic acid, alpha-linolenic acid, adenosine receptor antagonist (CGS 15943), and Prodigiosin) present in M. bealei serum may exhibit therapeutic effects against alcoholic HCC by downregulating CDK1 and TOP2A expression levels in vitro.

Quality control assessment, toxicity profiling, and experimental validation of network pharmacology-predicted anti-inflammatory potential of Natsiatum herpeticum Buch.-Ham. Ex Arn.

ETHNOPHARMACOLOGICAL RELEVANCE: Natsiatum herpticum Buch.-Ham. Ex Arn., a least-explored plant, is being considered a wild edible plant by the Bankariya community of Nepal and the Mishing, Sonowal Kachari, and several ethnic groups in the north-east region of India. It is also used as a traditional remedy for the treatment of pain and inflammation-associated conditions like cuts and wounds, stomach ache, backache, and headache as a practice of a folkloristic system of medicine. In spite of several previous publications suggesting its use by different tribes, no documentation or scientific approaches have been made hitherto to validate its ethnopharmacological claims. AIM OF STUDY: The study aimed at the botanical quality control assessment, toxicity profiling, and network pharmacology-assisted experimental validation of the anti-inflammatory potential of the aqueous extract of N. herpeticum to fill the lacunae in the current knowledge. MATERIAL AND METHOD: Plant material was authenticated using a classical taxonomical approach and DNA barcoding. The quality control methods, acute toxicity study, and repeated dose 28-day oral toxicity study were performed as per standard guidelines. QToF-MS analysis, drug-likeness properties, network pharmacology-based anti-inflammatory prediction, and in vitro assays were carried out. RESULTS: Quality control assessment was done for the plant. Toxicity studies revealed the aqueous extract to be non-toxic when consumed for short periods at low doses. Alterations in food and water intake, biochemical parameters, and alterations in liver histology (n = 2 female rats) implicate repeated exposure to high doses (2000 mg/kg) that may possess deleterious effects, particularly in hepatic tissues. 21 representative compounds (14 drug-like molecules) were detected by QToF-MS analysis and then subjected to network pharmacology to predict anti-inflammatory effects. It was found that an anti-inflammatory effect may be exerted by modulating inflammatory pathways involving genes such as TNF, PTGS2, EGFR, STAT3, PPARG, PTGER4, PPARA, NOS2, TRPV1, and JAK2. Further, in vitro studies demonstrated plant extract to possess a good anti-inflammatory effect with IC50 values of 98.76, 85.73, and 96.16 µg/ml in protein denaturation, proteinase inhibition, and haemolysis inhibition assays, respectively. CONCLUSION: The plant extract was found to be safer at acute dose but may cause potential liver toxicity on prolonged use. The anti-inflammatory property predicted by network pharmacology was further supported by the positive results of in vitro experiments. In summary, to further establish the toxicity profile of this edible plant and its anti-inflammatory properties, chronic toxicity study and in vivo experiments are required.

Jingfang granule alleviates Pseudomonas aeruginosa-induced acute lung inflammation through suppression of STAT3/IL-17/NF-κB pathway based on network pharmacology analysis and experimental validation.

ETHNOPHARMACOLOGICAL RELEVANCE: Pseudomonas aeruginosa is an opportunistic bacterial pathogen which is the second leading cause of hospital-acquired pneumonia. Jingfang granule (JFG) is an herbal formula of Traditional Chinese medicine (TCM) widely used in treatment of acute respiratory tract infections in China. However, the molecular mechanisms of JFG in treatment of P. aeruginosa-induced acute pneumonia are not clear. AIM OF STUDY: This study aimed to investigate the mechanisms underlying the effects of JFG on P. aeruginosa-induced acute inflammation using a mouse model of bacterial acute pneumonia. MATERIALS AND METHODS: The chemical components and targets of JFG were retrieved from Traditional Chinese Medicine Systems Pharmacology (TCMSP) database, and the P. aeruginosa pneumonia-related targets were obtained from the disease databases, including Online Mendelian Inheritance in Man (OMIM), GeneCards and DisGeNet. The protein-protein interaction (PPI) network was constructed using STRING database. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were performed using the Database for Annotation, Visualization and Integrated Discovery (DAVID). Molecular docking was performed using AutoDockTools 1.5.6. Further in vivo experiments employed a mouse model of P. aeruginosa acute pneumonia to verify the target proteins and signaling pathways affected by JFG, which were predicted by the network pharmacology analysis. RESULTS: A total of 218 active components and 257 targets of JFG were retrieved from TCMSP database. Moreover, 99 intersectant targets were obtained between the 257 JFG targets and 694 disease targets. Among the intersectant targets, STAT3, IL-6, AKT1, TNF, MAPK1, MAPK3 and EGFR were identified to be the key therapeutic targets through PPI network analysis, and STAT3 was in the center of the network, which is a key regulator of IL-17 expression. KEGG pathway enrichment analysis suggested that IL-17 signaling pathway was one of the crucial inflammatory pathways affected by JFG in treatment of P. aeruginosa pneumonia. Furthermore, the in vivo experiments demonstrated that the JFG-treated mice displayed reduced proinflammatory cytokine production (IL-17, IL-1ß, IL-6 and TNF), diminished neutrophil infiltration and decreased mortality, compared with the non-drug-treated mice during P. aeruginosa lung infection. Moreover, the expression or phosphorylation levels of the key regulators in STAT3/IL-17/NF-κB axis including STAT3, ERK1/2 (MAPK3/1), AKT, NF-κB p65 and RORγt were significantly reduced in the lung tissues of the JFG-treated mice. CONCLUSION: JFG was effective in treatment of P. aeruginosa acute lung infection, which reduced inflammatory responses through suppressing STAT3/IL-17/NF-κB pathway.

Prediction of the Molecular Mechanism of Corni Fructus-Epimedii Folium- Rehmanniae Radix Praeparata in the Treatment of Postmenopausal Osteoporosis based on Network Pharmacology and Molecular Docking.

INTRODUCTION: In this study, core drugs of clinical postmenopausal osteoporosis were retrieved using data mining, the drug molecular action target was predicted through network pharmacology, the key nodes of interaction were identified by combining postmenopausal osteoporosis-related targets, and the pharmacological mechanism of Traditional Chinese Medicine (TCM) against postmenopausal osteoporosis and other action mechanisms was explored. METHODS: TCMISS V2.5 was used to collect TCM prescriptions of postmenopausal osteoporosis from databases, including Zhiwang, Wanfang, PubMed, etc., for selecting the highest confidence drugs. TCMSP and SwissTargetPrediction databases were selected to screen the main active ingredients of the highest confidence drugs and their targets. Relevant targets for postmenopausal osteoporosis were retrieved from GeneCards and GEO databases, PPI network diagrams construction and selection of core nodes in the network, GO and KEGG enrichment analysis, and molecular docking validation. RESULTS: Correlation analysis identified core drug pairs as 'Corni Fructus-Epimedii Folium- Rehmanniae Radix Praeparata' (SZY-YYH-SDH). After TCMSP co-screening and de-weighting, 36 major active ingredients and 305 potential targets were selected. PPI network graph was built from the 153 disease targets and 24 TCM disease intersection targets obtained. GO, KEGG enrichment results showed that the intersectional targets were enriched in the PI3K-Akt signalling pathway, etc. The target organs were mainly distributed in the thyroid, liver, CD33+_Myeloid, etc. Molecular docking results showed that the core active ingredients of the 'SZY-YYH-SDH' were able to bind to the pair core nodes and PTEN and EGFR. CONCLUSION: The results showed that 'SZY-YYH-SDH' can provide the basis for clinical application and treat postmenopausal osteoporosis through multi-component, multi-pathway, and multitarget effects.

Network pharmacology analysis of the active ingredients of Corydalis hendersonii Hemsl. and their effects on eliminating neuroinflammation and improving motor functions in MPTP-intoxicated mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Corydalis hendersonii Hemsl. (CH), is a traditional Tibetan medicine used in highland areas for the treatment of alpine polycythemia, ulcers and various inflammatory diseases. Its antioxidant and anti-inflammatory effects have been demonstrated in experimental mice. Loss of dopaminergic neurons due to oxidative damage is thought to be an important factor in the development of PD, the potential antioxidant, anti-inflammatory effects of CH could potentially be used for PD treatment. AIM OF THE STUDY: To identify potential targets of CH using network pharmacology and to investigate the neuroprotective effects in cultured cell models and in MPTP-intoxicated mice. MATERIALS AND METHODS: The main chemical components of CH were analyzed by UPLC-MS/MS and their potential targets of action or signaling pathways were analyzed using network pharmacology. MPP + or LPS was added to SH-SY5Y or BV2 cells, respectively, to establish cellular models. MPTP was administered to C57BL/6J mice to induce inflammation and dopaminergic neuron loss as well as dyskinesia, followed by behavioral analysis to determine the role of CH in eliminating inflammation, avoiding neuron loss, and improving dyskinesia. RESULTS: CH contains 241 alkaloids, 213 flavonoids, 177 terpenoids and 114 phenolic compounds. The targets crossover between CH and PD yielded 210 potential therapeutic targets, especially growth factors and inflammatory pathway-related genes, such as BDNF, NF-κB, as potential key targets. In cultured cells, CHE eliminated MPP + -induced impairment of cell viability as well as LPS-induced inflammation, respectively. In mice, CHE ameliorated MPTP-induced dyskinesia and rescued the loss of dopaminergic neurons in the substantia nigra and striatum. Mechanistically, CHE effectively maintained the activity of the BDNF-TrkB/Akt signaling pathway, accordingly, inhibited inflammatory signaling pathways such as HIF-1α/PKM2 and Notch/NF-kB. CONCLUSIONS: CH performed well in eliminating inflammation and improving locomotor deficits in mice, and its potent active ingredients are worthy of subsequent research and development.

Network pharmacology and experimental validation on yangjing zhongyu decoction against diminished ovarian reserve.

ETHNOPHARMACOLOGICAL RELEVANCE: Diminished ovarian reserve (DOR) was considered a refractory reproductive endocrine condition that negatively affected female reproductivity. Yangjing Zhongyu Decoction (YJZYD) had effects on treating infertility. However, there were few studies on the mechanisms of YJZYD preserving ovarian reserve. AIM OF THE STUDY: To explore the possible mechanisms of YJZYD against DOR by UPLC-ESI-MS/MS, network pharmacology, and experimental validation. METHODS: The chemicals of YJZYD were measured by UPLC-ESI-MS/MS. The correlating targets of YJZYD and DOR were identified by the ETCM database, GeneCards database, and PubMed database. The common targets were employed with the DAVID database and visualized with the PPI network. GO and KEGG enrichment analyses were carried out to explore biological progression and pathways. In vivo experiments, energy production was assessed by ATP, and apoptosis rate was analyzed by TUNEL. The serum FSH, AMH, and E2 levels were evaluated by ELISA. Western blotting and immunohistochemistry were used to measure the expression of SIRT1, PGC1α, NRF1, COX IV, FSHR, CYP19A1, PI3K, p-Akt, Akt, Bcl-2, and Bax. RESULTS: 132 components in YJZYD were identified by UPLC-ESI-MS/MS. 149 overlapped targets were extracted from YJZYD and DOR, and the top 20 common targets included AKT1 and CYP19A1. ATP binding was involved in GO analysis. In the KEGG enrichment analysis, the metabolic pathway was the top, and the PI3K-Akt signaling pathway was included. In vivo experiments, YJZYD improved ovarian index and histomorphology. After YJZYD treatment, serum FSH, E2, and AMH were well-modulated, and the content of ATP was up-regulated. Besides, the expression of Bax was suppressed in ovarian tissue, while the expressions of SIRT1, PGC1α, NRF1, COX IV, FSHR, CYP19A1, PI3K, Bcl-2, and p-Akt/Akt were enhanced. CONCLUSION: YJZYD could attenuate reproductive endocrine disturbance and ovarian lesions in vivo by mediating steroidogenesis, energy metabolism, and cell apoptosis. This study uncovered the mechanisms of YJZYD against DOR, providing a theoretical basis for further study.

An integrated RNA-Seq and network pharmacology approach for exploring the preventive effect of Corydalis bungeana Turcz. Extract and Acetylcorynoline on LPS-induced acute lung injury.

ETHNOPHARMACOLOGICAL RELEVANCE: Corydalis bungeana Turcz. (KDD) is a Chinese herbal medicine with anti-inflammatory, lung cleansing, detoxification and other functions. Clinically, it is commonly used to treat respiratory infections. This study uses ALI as the research model, which is consistent with the clinical use of KDD. Acetylcorynoline (AC) is the main alkaloid component of the KDD extracts, and network pharmacology studies suggest that it may be the main active ingredient in the prevention of ALI. AIM OF THE STUDY: The aim of this study is to explore the underlying mechanisms and to study the efficacy material basis of KDD in anti-ALI effect by LPS-induced mice and using a combination of RNA sequencing (RNA-Seq) technology and network pharmacology. MATERIALS AND METHODS: Establish a mouse model of ALI by intraperitoneal injection of LPS (5 mg/kg). The main active ingredients of KDD were identified and analyzed by high performance liquid chromatography with quadrupole time-of-flight mass spectrometry (HPLC-QTOF-MS) and network pharmacology. IL-18, IL-1ß, and IL-6 levels in serum and bronchoalveolar lavage fluid (BALF), lung histopathological changes, and lung myeloperoxidase (MPO) activity were assessed. We investigated the possible molecular mechanisms of KDD and AC in an LPS-induced mouse ALI models with RNA-Seq technology. In addition, the anti-inflammatory effect of AC was verified in vitro by establishing an LPS-stimulated RAW264.7 inflammation model. Molecular docking further validated AC as the efficacy material basis of KDD in anti-ALI. RESULTS: Based on HPLC-QTOF-MS technology and network pharmacology, KDD is more strongly associated with lung tissue, and that AC may be the main active ingredient of KDD. Subsequently, in vivo experiments results showed that KDD and AC reduced the levels of pro-inflammatory cytokines in serum and BALF, reduced MPO levels and reduced inflammatory damage in the lungs. To elucidate its underlying mechanism, based on RNA-Seq analysis techniques performed in lung tissue, enrichment analysis showed that KDD and AC intervened through the NLR signaling pathway, thereby mitigating LPS-induced ALI. Then, RT-qPCR, IF, WB and other technologies were used to verify the anti-ALI core difference genes of KDD and AC from the gene transcription and protein expression levels of the NLR signaling pathway, and confirmed the anti-ALI. In vitro experimental results also showed that AC has anti-inflammatory effects in RAW264.7. Finally, the biotransformation and molecular docking results also further indicated that AC is the active ingredient of KDD in anti-ALI. CONCLUSIONS: Studies have shown that KDD has a good therapeutic effect on ALI, and AC is the main pharmacodynamic material basis for its therapeutic effect in ALI.

Altered metabolic profiles and targets relevant to the protective effect of acteoside on diabetic nephropathy in db/db mice based on metabolomics and network pharmacology studies.

ETHNOPHARMACOLOGICAL RELEVANCE: Diabetic nephropathy (DN) was a major cause of end-stage renal failure and a common microvascular complication in patients with diabetes mellitus (DM). Acteoside (ACT) was the main ingredient extracted from the leaves of Rehmannia glutinosa, which had the functions of entering the lung, moisturizing the skin and relieving itching, nourishing yin and tonifying the kidney, cooling blood, and stopping bleeding. ACT had attracted worldwide interest because of its therapeutic effects on DM and its complications. AIM OF THE STUDY: To clarify the metabolic profiles and targets of ACT in db/db mice based on metabolomics and network pharmacology studies. MATERIALS AND METHODS: Db/db mice were used to observe the biochemical indices and histopathological changes in the kidney to evaluate the pharmacological effects of ACT on DN. Untargeted metabolomics studies were performed to investigate by UHPLC-LTQ-Orbitrap MS on urine, serum, and kidney samples. The key targets and pathways were analyzed by network pharmacology. For the pathways enriched by untargeted metabolomics, targeted metabolomics by UHPLC-QQQ-MS/MS was performed in kidney samples for validation. Sensitive biomarkers in kidney samples were evaluated. The effect of ACT on the improvement of DN from the perspective of metabolism of small molecules in vivo was described. RESULTS: ACT could delay the progression of DN and improve the degree of histopathological damage to the kidney. The pathways were focused on amino acid metabolism by untargeted metabolomics. Through network pharmacology analysis, the effect pathways were related to signal transduction, carbohydrate, lipid, amino acid metabolism and mainly affected the endocrine and immune systems. Amino acid metabolism was disturbed in the kidney of db/db mice, which could be callback by ACT, such as tryptophan, glutamine, cysteine, leucine, threonine, proline, phenylalanine, histidine, serine, arginine, asparagine by targeted metabolomics. CONCLUSIONS: In conclusion, this study provided strong support for ACT on DN treatment in clinics. Meanwhile, the Rehmannia glutinosa was used fully to raise the income level of farmers economically, while achieving the social benefit of empowering rural revitalization.

Combining transcriptomics and network pharmacology to reveal the mechanism of Zuojin capsule improving spasmolytic polypeptide-expressing metaplasia.

ETHNOPHARMACOLOGICAL RELEVANCE: Spasmolytic polypeptide-expressing metaplasia (SPEM) is a gastric precancerous lesion (GPL). Zuojin capsule (ZJC), consisting of Coptis chinensis Franch. (Ranunculaceae, recorded in the Chinese Pharmacopoeia as Rhizoma Coptidis) and Tetradium ruticarpum (A.Juss.) T.G.Hartley (Rutaceae, recorded in the Chinese Pharmacopoeia as Fructus Evodiae), has long been used for various gastrointestinal diseases. However, the effect and mechanism of ZJC on SPEM remain unclear. AIM OF THE STUDY: To clarify the role of ZJC in improving SPEM and study its mechanism. MATERIALS AND METHODS: The study utilized SPEM mice induced by 250 mg/kg body weight of tamoxifen (TAM) to assess the effects of ZJC and investigate its possible mechanisms. A strategy of transcriptomics combined with network pharmacology was conducted to explore the targets and mechanisms of ZJC in improving SPEM. The "ingredients-target-pathway" network was constructed, and the possible connections were verified by RT-qPCR and Western blot assays. RESULTS: ZJC significantly attenuated the abnormal serological indices, destruction of the gastric mucosal structure, hyperplasia of gastric pits, increased gastric mucus, massive secretion of CD44 and TFF2, oxyntic atrophy and massive proliferation of stem/progenitor cells in TAM-induced SPEM mice. Combined transcriptomics and network pharmacology analysis, 50 core targets of ZJC related to SPEM improvement were obtained. KEGG results showed that the core targets were significantly enriched in the cell cycle, and PI3K-AKT signaling pathway. The top-ranked targets according to PPI network analysis were CDK1, CCNB1, and CCNA2, which are also associated with cell cycle. Combined experiments demonstrated that ZJC can induce G2/M phase cycle arrest and inhibit TAM-induced malignant proliferation by regulating abnormal activation of cell cycle-related proteins such as CDK1, CCNB1, CCNA2 and PI3K-AKT signaling pathways. CONCLUSIONS: ZJC may improve TAM-induced SPEM by inhibiting abnormal activation of cell cycle-related proteins (CDK1, CCNB1, CCNA2) and PI3K-AKT signaling pathway. This finding supports the use of ZJC, a famous traditional Chinese medicine compound, as a potential treatment for gastric precancerous lesions.