Network Pharmacology-based Analysis Research Articles

Exploring Mechanism of Actions for Eugenol and Beta-Caryophyllene to Combat Colorectal Cancer Chemotherapy Using Network Pharmacology

This study explores the potential of essential oils, Eugenol (EUG), and Beta-Caryophyllene (BCP) in enhancing the efficacy of the chemotherapeutic drug 5-fluorouracil (5-FU) in treating metastatic colorectal cancer (CRC). Pharmacokinetic assessment through ADMET analysis indicates that EUG and BCP adhere to the rule of five with good bioavailability, ensuring their drug-likeness properties. The study employs a multitarget strategy to reduce drug dosage and enhance effectiveness, testing the compounds on the HCT116 human colorectal cancer cell line. MTT assay revealed in-vitro cytotoxic effects of EUG, BCP, and 5-FU, with a noteworthy reduction in IC50 values observed when combining the compounds, indicating synergistic effects (CI < 1) as depicted in the Fa-CI plot. Network pharmacology-based analysis of the compound-disease-target (C-D-T) network identifies 58, 24, and 49 target proteins for EUG, BCP, and 5-FU, respectively, in metastatic CRC. Venn diagram intersection reveals 11 common target proteins, and the merged C-D-T network highlights 84 target proteins, with 16 selected based on edge count, including HSP90AA1, IGF-1R, ESR1, and CASP3. Molecular docking studies indicate that EUG, BCP, and 5-FU effectively inhibit the core target protein HSP90AA1 within the C-D-T network, suggesting their potential as modulators for CRC metastasis. These findings propose a promising approach for developing drugs targeting specific proteins to mitigate metastasis in colorectal cancer.

Network pharmacology-based analysis on the key mechanisms of Yiguanjian acting on chronic hepatitis

Chronic hepatitis (CH) encompasses a prevalent array of liver conditions that significantly contribute to global morbidity and mortality. Yiguanjian (YGJ) is a classical traditional Chinese medicine with a long history of medicinal as a treatment for CH. Although it has been reported that YGJ can reduce liver inflammation, the intricate mechanism requires further elucidation. We used network pharmacology approaches in this work, such as gene ontology (GO) analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, and network-based analysis of protein-protein interactions (PPIs), to clarify the pharmacological constituents, potential therapeutic targets, and YGJ signaling pathways associated with CH. Employing the random walk restart (RWR) algorithm, we identified GNAS, GNB1, CYP2E1, SFTPC, F2, MAPK3, PLG, SRC, HDAC1, and STAT3 as pivotal targets within the PPI network of YGJ-CH. YGJ attenuated liver inflammation and inhibited GNAS/STAT3 signaling in vivo. In vitro, we overexpressed the GNAS gene further to verify the critical role of GNAS in YGJ treatment. Our findings highlight GNAS/STAT3 as a promising therapeutic target for CH, providing a basis and direction for future investigations.

NADES extraction, UHPLC-ELSD-based quantification, and network pharmacology-guided target identification of fourteen specialised metabolites from Trillium govanianum Wall. ex D.Don.

Trillium govanianum Wall. ex D.Don is a folk medicinal herb rich in structurally diverse steroidal saponins. The annual demand for this herb in India is about 200-500 metric tons, highlighting the need for a thorough quality assessment. The objective of this study is to develop an easy and reliable ultrahigh-performance liquid chromatography-evaporative light scattering detector (UHPLC-ELSD)-based quality assessment method with 14 specialised metabolites of T. govanianum and identify the potential targets of this herb using network pharmacology. A UHPLC-ELSD method was developed and validated with 14 markers of T. govanianum. The developed method and natural deep eutectic solvent (NADES)-assisted extraction were utilised for the recovery enhancement study of targeted specialised metabolites from rhizome samples (collected from five geographically distinct areas). In addition, the network pharmacology approach was performed for these 14 markers to predict the plausible biological targets of T. govanianum. The developed method showed good linearity (r2: 0.940-0.998), limit of detection (LOD) (2.4-9.0μg), limit of quantification (LOQ) (7.92-29.7μg), precision (intra-day relative standard deviations [RSDs] 0.77%-1.96% and inter-day RSDs 2.19-4.97%), and accuracy (83.24%-118.90%). NADES sample TG-1* showed the highest recovery (yield: 167.66 ± 4.39 mg/g of dry weight) of total saponin content (TSC) as compared to its hydroethanolic extract (yield: 103.95 ± 5.36 mg/g of dry weight). Sample TG-1* was the most favourable (yield: 167.66 ± 4.39 mg/g) in terms of TSC as compared to other analysed samples (32.68 ± 1.04-88.22 ± 6.79 mg/g). Govanoside D (yield: 3.43-28.06 mg/g), 22β-hydroxyprotodioscin (yield: 3.22-114.79 mg/g), and dioscin (yield: 1.07-20.82 mg/g) were quantified as the major metabolites. Furthermore, network pharmacology analysis of targeted 14 markers indicated that these molecules could be possible therapeutic agents for managing neuralgia, diabetes mellitus, and hyperalgesia. The current study represents the first report for the simultaneous quantification and a network pharmacology-based analysis of 14 chemical marker compounds isolated from T. govanianum.

Deciphering the multi-functional role of Indian propolis for the management of Alzheimer's disease by integrating LC-MS/MS, network pharmacology, molecular docking, and in-vitro studies.

The conventional one-drug-one-disease theory has lost its sheen in multigenic diseases such as Alzheimer's disease (AD). Propolis, a honeybee-derived product has ethnopharmacological evidence of antioxidant, anti-inflammatory, antimicrobial and neuroprotective properties. However, the chemical composition is complex and highly variable geographically. So, to leverage the potential of propolis as an effective treatment modality, it is essential to understand the role of each phytochemical in the AD pathophysiology. Therefore, the present study was aimed at investigating the anti-Alzheimer effect of bioactive in Indian propolis (IP) by combining LC-MS/MS fingerprinting, with network-based analysis and experimental validation. First, phytoconstituents in IP extract were identified using an in-house LC-MS/MS method. The drug likeness and toxicity were assessed, followed by identification of AD targets. The constituent-target-gene network was then constructed along with protein-protein interactions, gene pathway, ontology, and enrichment analysis. LC-MS/MS analysis identified 16 known metabolites with druggable properties except for luteolin-5-methyl ether. The network pharmacology-based analysis revealed that the hit propolis constituents were majorly flavonoids, whereas the main AD-associated targets were MAOB, ESR1, BACE1, AChE, CDK5, GSK3β, and PTGS2. A total of 18 gene pathways were identified to be associated, with the pathways related to AD among the topmost enriched. Molecular docking analysis against top AD targets resulted in suitable binding interactions at the active site of target proteins. Further, the protective role of IP in AD was confirmed with cell-line studies on PC-12, in situ AChE inhibition, and antioxidant assays.

Experimental and pharmacoinformatic approaches unveil the neuropharmacological and analgesic potential of chloroform fraction of Roktoshirinchi (Achyranthes ferruginea Roxb.)

Ethnopharmacological relevanceAchyranthes ferruginea (A. ferruginea) Roxb. is a common plant used in traditional medicine in Asia and Africa. It has a variety of local names, including “Gulmanci” in Nigeria, “Dangar” in Pakistan, “Thola” in Ethiopia, and “Roktoshirinchi” in Bangladesh. It is edible and has several ethnomedical uses for a wide range of illnesses, including hysteria, dropsy, constipation, piles, boils, asthma, and shigellosis. However, the neuropharmacological and analgesic potential of A. ferruginea remains uninvestigated. Aim of the studyTo assess the neuropharmacological and analgesic potential of A. ferruginea through a multifaceted approach encompassing both experimental and computational models. Materials and methodsMethanol was used to extract the leaves of A. ferruginea. It was then fractionated with low to high polar solvents (n-hexane, chloroform, ethyl acetate, and water) to get different fractions, including chloroform fraction (CLF). The study selected CLF at different doses and conducted advanced chemical element and proximate analyses, as well as phytochemical profiling using GC-MS. Toxicological studies were done at 300 μg per rat per day for 14 days. Cholinesterase inhibitory potential was checked using an in-vitro colorimetric assay. Acetic acid-induced writhing (AAWT) and formalin-induced licking tests (FILT) were used to assess anti-nociceptive effects. The forced swim test (FST), tail suspension test (TST), elevated plus maze (EPM), hole board test (HBT), and light and dark box test (LDB) were among the behavioral tests used to assess depression and anxiolytic activity. Network pharmacology-based analysis was performed on selected compounds using the search tool for interacting chemicals-5 (STITCH 5), Swiss target prediction tool, and search tool for the retrieval of interacting genes and proteins (STRING) database to link their role with genes involved in neurological disorders through gene ontology and reactome analysis. ResultsQualitative chemical element analysis revealed the presence of 15 elements, including Na, K, Ca, Mg, P, and Zn. The moisture content, ash value, and organic matter were found to be 11.12, 11.03, and 88.97%, respectively. GC-MS data revealed that the CLF possesses 25 phytoconstituents. Toxicological studies suggested the CLF has no effects on normal growth, hematological and biochemical parameters, or cellular organs after 14 days at 300 μg per rat. The CLF markedly reduced the activity of both acetylcholinesterase and butyrylcholinesterase (IC50: 56.22 and 13.22 μg/mL, respectively). Promising dose-dependent analgesic activity (p < 0.05) was observed in chemically-induced pain models. The TST and FST showed a dose-dependent substantial reduction in immobility time due to the CLF. Treatment with CLF notably increased the number of open arm entries and time spent in the EPM test at doses of 200 and 400 mg/kg b.w. The CLF showed significant anxiolytic activity at 200 mg/kg b.w. in the HBT test, whereas a similar activity was observed at 400 mg/kg b.w. in the EPM test. A notable increase in the amount of time spent in the light compartment was observed in the LDB test by mice treated with CLF, suggesting an anxiolytic effect. A network pharmacology study demonstrated the relationship between the phytochemicals and a number of targets, such as PPARA, PPARG, CHRM1, and HTR2, which are connected to the shown bioactivities. ConclusionsThis study demonstrated the safety of A. ferruginea and its efficacy in attenuating cholinesterase inhibitory activity, central and peripheral pain, anxiety, and depression, warranting further exploration of its therapeutic potential.

Unraveling the mechanisms of Fenugreek seed for managing different gynecological disorders: steroidal saponins and isoflavones revealed as key bioactive metabolites

This study employed network pharmacology-based analysis and reverse molecular docking to investigate the molecular targets and pathways associated with gynecological disorders, particularly those related to steroidal hormones and their receptors, and the potential therapeutic effects of fenugreek (Trigonella foenum-graecum L.) constituents. The STITCH 5.0 database was utilized to identify potential molecular targets, and a compound-target network was constructed. The main targets associated with gynecological disorders included estrogen receptor beta (ESR2), estrogen-related receptor gamma (GPER1), oxytocin receptor (OXTR), progesterone receptor (PGR), prolactin receptor (PRLR), and several enzymes involved in sex hormone biosynthesis. Additionally, network topological analysis revealed that specific compounds, such as quercetin, luteolin, genistein, and vitexin, had significant interactions with the identified targets. Reverse molecular docking analysis confirmed the interactions between the identified compounds and target proteins where quercetin, luteolin, genistein, 4'-methylgenistein, trigoneoside IIB, diosgenin, and vitexin possessed the highest combined docking scores, indicating their multi-target nature. The results highlighted the potential of steroidal saponins, isoflavones, and flavones as active constituents of fenugreek with implications for lactation, reproductive processes, and estrogenic activity. The chemical profiling of saponin-enriched and flavonoid-enriched fractions using UPLC/MS/MS further supported the presence of these bioactive compounds. In an animal model study, the steroidal saponins-enriched fraction of fenugreek seed exhibited a significant increase in the body weight of lactating female rats and serum prolactin levels while the flavonoids-enriched fraction showed an increase in serum estradiol levels and improved the histological structure of ovaries.

Network pharmacology-based analysis of Jin-Si-Wei on the treatment of Alzheimer's disease

Ethnopharmacological relevanceJin-Si-Wei (JSW), a traditional Chinese medicine (TCM) formula, have cognitive enhancing effect and delay the memory decline in an animal model of AD， which has been reported. However, the therapeutic mechanism of JSW in the treatment of AD remains unclear. Aim of the studyThis study aimed to verify the pharmacodynamics of JSW in the treatment of AD, and to explore its potential mechanism based on network pharmacology, molecular docking and experimental validation both in vitro and in vivo. Materials and methodsIn this study, the underlying mechanism of JSW against AD was investigated by the integration of network pharmacology. Then, the core pathways and biological process of JSW were verified by experiment, including behavioral test and pathological and biochemical assays with 6-month-old APPswe/PS1ΔE9 transgenic (APP/PS1) mice in vivo and verified with Aβ1-42-stimulated SH-SY5Y cells in vitro. At last, molecular docking was used to show the binding activity of each active ingredient to the core genes of JSW treatment in AD. ResultsA Drug-Ingredient-Target network was established, which included 363 ingredients and 116 targets related to the JSW treatment of AD. The main metabolic pathway of JSW treatment for AD is neuroactive ligand-receptor interaction pathway, and biological processes are mainly involved in Aβ metabolic process. In vivo experiments, compared with APP/PS1 mice, the cognitive and memory ability of mice was significantly improved after JSW administration. In brain tissue of APP/PS1 mice, JSW could increase the contents of low-density lipoprotein receptor-related protein 1 (LRP-1), enkephalinase (NEP) and Acetyl choline (ACh), and decrease the contents of Aβ1-42, amyloid precursor protein (APP) and receptor for advanced glycation end products (RAGE), decrease the vitality of cholinesterase (AChE) and choline acetyltransferase (ChAT). Besides, JSW could increase α-secretase expression and decrease β/γ-secretase expression, and improve the number and morphology of synapses in CA1 region of the hippocampus of APP/PS1 mice. In vitro experiments, Drug-Containing Serum (JSW-serum) has a neuroprotective effect by reducing the apoptosis on Aβ1-42-stimulated SH-SY5Y cells. Molecular docking results showed that 2-Isopropyl-8-methylphenanthrene-3,4-dione had strong binding activity with PTGS2, which maybe a potential ingredient for the treatment of AD. ConclusionsJSW improves AD in APP/PS1 mice, and this therapeutic effect may be achieved in part by altering the neuroactive ligand-receptor interaction pathway.

Xiaoqinglong decoction suppresses childhood cough variant asthma and inhibited the body inflammatory response by regulating IL-6/STAT3 signalling pathway.

Xiaoqinglong decoction (XQLD) is widely used clinically in the treatment of childhood cough variant asthma (CVA). However, its potential mechanism is still unknown. In the present study, the authors investigate the biological network and signalling pathway of XQLD in treatment of childhood CVA using network pharmacology-based analysis and experimental validation. By using the Bioinformatics Analysis Tool Molecular Mechanism of Traditional Chinese Medicine (BATMAN-TCM) database, the authors confirmed the correlation between XQLD and asthma, and the authors screened 1338 potential target genes of Mahuang and Guizhi, the most active herbs in XQLD. By overlapping "Childhood asthma-related genes" of DisGeNET database, the authors identified 58 intersecting genes of Childhood asthma and 1338 target genes of Mahuang and Guizhi. The intersecting genes were used to construct the protein-to-protein interaction and performed Gene Ontology (GO) functional and the Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses. Gene Ontology enrichment analysis demonstrated 359 Biological Process terms, 16 Cellular Component terms, and 26 Molecular Function terms. Meantime, 75 terms of Kyoto Encyclopedia of Genes and Genomes signalling pathway were involved in enrichment analysis. These candidates showed a significant correlation with inflammatory response and positive regulation of tyrosine phosphorylation of STAT protein. In addition, XQLD treatment significantly upregulated serum interferon-γ expression, and downregulated serum interlukin-6 expression of CVA mice. XQLD treatment significantly inhibited phosphorylation of STAT3 in bronchial-lung tissues. Our data suggest that XQLD effectively alleviated bronchial-lung tissue damage in CVA mice and inhibited the body inflammatory response by regulating interlukin-6/STAT3 signalling pathway.

Anti-atherosclerotic effects of naringenin and quercetin from Folium Artemisiae argyi by attenuating Interleukin-1 beta (IL-1β)/ matrix metalloproteinase 9 (MMP9): network pharmacology-based analysis and validation

Effective components and related target genes of Folium Artemisiae argyi were screened from Traditional Chinese Medicines for Systems Pharmacology Database and Analysis Platform. The therapeutic targets of atherosclerosis were searched in the MalaCards and OMIM databases. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were performed in WebGestalt online and verified according to ClueGo and Pedia apps in Cytoscape. Then, the protein-protein interaction network was analyzed using the STRING database and constructed using Cytoscape. Differential expression of target genes was identified in GSE9128 and GSE71226 by GEO2R. And then, molecular docking was performed using the Molecular Operating Environment. Finally, we validated the protein expression of Interleukin-6 (IL-6)/IL-1β /MMP9 by qRT-PCR and Western blot in Raw264.7 which was induced by LPS. A total of 232 potential target genes and 8 ingredients of Folium Artemisiae argyi were identified. Quercetin and naringenin are potential candidate bioactive agents in treating atherosclerosis. Vascular endothelial growth factor (VEGFA), MMP9 and IL-1β could be potential target genes. KEGG analysis demonstrated that the fluid shear stress and atherosclerosis pathway play a crucial role in the anti-atherosclerosis effect of Folium Artemisiae argyi. Gene Expression Omnibus (GEO) validation demonstrated that VEGFA was downregulated, while MMP9 and IL-1β were upregulated in patients with atherosclerosis. Molecular docking suggested that only MMP9 had a good combination with quercetin. The cell experiment results suggested that naringenin and quercetin have strong anti-inflammation effects, and significantly inhibit the expression of MMP9.Practical ApplicationsArtemisiae argyi is a traditional Chinese herbal medicine that has been widely used for its antibacterial and anti-inflammatory effects. This research demonstrated the bioactive ingredients, potential targets, and molecular mechanism of Folium Artemisiae argyi in treating atherosclerosis. It also suggests a reliable approach in investigating the therapeutic effect of traditional Chinese herbal medicine in treating Atherosclerotic cardiovascular disease (ASCVD).

Network Pharmacology-based Analysis to Investigate Bioactive from the Root of Astragalus membranaceus against Immunomodulatory Protein and their Role in Autoimmune Thyroiditis (AIT)

Background The thyroid is the prime organ of infection during the onset of autoimmune disease. The disease is known as Autoimmune thyroiditis (AIT). In recent times, AIT has been alarming worldwide, with growing attention towards pharmaceutical companies for finding new drugs for therapy. The present investigation aims to use a multi-targeting approach to identify the best compound from the root of Astragalus membranaceus for synergistic therapy of AIT. Materials and Methods To explore the mechanism of action of the compounds, the present study uses network pharmacology-based methods. Totally, 48 genes involved in AIT were collected from the Kyoto Encyclopedia of Genes and Genomes pathway (KEGG). Gene ontology (GO) annotation, disease ontology, and pathway enrichment analysis were conducted. The chemical structure of the compounds from A. membranaceus was retrieved from the PubChem database. To study the toxic and pharmacological efficiency of the chemical structure, absorption, distribution, metabolism, excretion, and toxicity (ADMET) screening was performed. Results The ononin and astroside were identified as suitable with all the pharmacokinetic properties can be employed to identify the disease targets. The intersection between the compound target (C-T-N) and the AIT-related genes was identified. Totally 16 genes among the C-T networks of ononin and astroside have shown interaction with thyroid disease-related pathways and signaling pathways. Consequently, the overlapping genes between the C-T network and AIT-related genes were determined as IL2. However, from the pathway enrichment, AIT-related genes have been involved in several viral infection pathways. The genes involved in infection pathways are also targeted by ononin and astroside. Conclusion The underlying mechanism of action of the compounds on AIT-related pathways was also determined. Therefore, a network pharmacology-based strategy will provide new insight into AIT therapy. However, further studies are required to validate the investigation that can prove bioactive as a promising candidate for the treatment of AIT.

Network pharmacology-based analysis of potential mechanisms of myocardial ischemia-reperfusion injury by total salvianolic acid injection.

In this review, we investigated the potential mechanism of Total Salvianolic Acid Injection (TSI) in protecting against myocardial ischemia reperfusion injury (MI/RI). To achieve this, we predicted the component targets of TSI using Pharmmapper and identified the disease targets of MI/RI through GeneCards, DisGenNET, and OMIM databases. We constructed protein-protein interaction networks by analyzing the overlapping targets and performed functional enrichment analyses using Gene Ontology and Kyoto Encyclopedia of Genes and Genomes. Our analysis yielded 90 targets, which were implicated in the potential therapeutic effects of TSI on MI/RI. Seven critical signaling pathways significantly contributed to TSI's protective effects, namely, PI3K signaling, JAK-STAT signaling, Calcium signaling, HIF-1 signaling, Nuclear receptor signaling, Cell Cycle, and Apoptosis. Subsequently, we conducted a comprehensive literature review of these seven key signaling pathways to gain further insights into their role in the TSI-mediated treatment of MI/RI. By establishing these connections, our study lays a solid foundation for future research endeavours to elucidate the molecular mechanisms through which TSI exerts its beneficial effects on MI/RI.

Forecasting of potential anti-inflammatory targets of some immunomodulatory plants and their constituents using in vitro, molecular docking and network pharmacology-based analysis

Most synthetic immunomodulatory medications are extremely expensive, have many disadvantages and suffer from a lot of side effects. So that, introducing immunomodulatory reagents from natural sources will have great impact on drug discovery. Therefore, this study aimed to comprehend the mechanism of the immunomodulatory activity of some natural plants via network pharmacology together with molecular docking and in vitro testing. Apigenin, luteolin, diallyl trisulfide, silibinin and allicin had the highest percentage of C-T interactions while, AKT1, CASP3, PTGS2, NOS3, TP53 and MMP9 were found to be the most enriched genes. Moreover, the most enriched pathways were pathways in cancer, fluid shear stress and atherosclerosis, relaxin signaling pathway, IL-17 signaling pathway and FoxO signaling pathway. Additionally, Curcuma longa, Allium sativum, Oleu europea, Salvia officinalis, Glycyrrhiza glabra and Silybum marianum had the highest number of P-C-T-P interactions. Furthermore, molecular docking analysis of the top hit compounds against the most enriched genes revealed that silibinin had the most stabilized interactions with AKT1, CASP3 and TP53, whereas luteolin and apigenin exhibited the most stabilized interactions with AKT1, PTGS2 and TP53. In vitro anti-inflammatory and cytotoxicity testing of the highest scoring plants exhibited equivalent outcomes to those of piroxicam.

Therapeutic mechanism of Guizhi Gancao Decoction for heart failure: a network pharmacology-based analysis

To predict the targets and pathways in the therapeutic mechanism of Guizhi Gancao Decoction (GZGCD) against heart failure (HF) based on network pharmacology. The chemical components of GZGCD were analyzed using the databases including TCMSP, TCMID and TCM@Taiwan, and the potential targets of GZGCD were predicted using the SwissTargetPrediction database. The targets of HF were obtained using the databases including DisGeNET, Drugbank and TTD. The intersection targets of GZGCD and HF were identified using VENNY. Uniport database was used to convert the information, and the components-targets-disease network was constructed using Cytoscape software. The Bisogene plug-in, Merge plug-in, and CytoNCA plug-in in Cytoscape software were used for protein-protein interaction (PPI) analysis to acquire the core targets. Metascape database was used for GO and KEGG analysis. The results of network pharmacology analysis were verified with Western blot analysis. Three factors (PKCα, ERK1/2 and BCL2) were screened according to the degree value of network pharmacology results and the degree of correlation with heart failure process. The pentobarbtal sodium was dissolvein H9C2 cells treated with serum-free high glucose medium to simulate the ischemic anoxic environment of heart failure. The total proteins of myocardial cells were extracted. The protein contents of PKCα, ERK1/2 and BCL2 were determined. We identified a total of 190 intersection targets between GZGCD and HF using Venny database, involving mainly the circulatory system process, cellular response to nitrogen compounds, cation homeostasis, and regulation of the MAPK cascade. These potential targets were also involved in 38 pathways, including the regulatory pathways in cancer, calcium signal pathway, cGMP-PKG signal pathway, and cAMP signal pathway. Western blot analysis showed that in an in vitro H9C2 cell model of HF, treatment with GZGCD downregulated PKCα and ERK1/2 expressions and upregulated BCL2 expression. The therapeutic mechanism of GZGCD for HF involves multiple targets including PRKCA, PRKCB, MAPK1, MAPK3, and MAPK8 and multiple pathways including the regulatory pathway in cancer and the calcium signaling pathway.

Network pharmacology-based analysis on geniposide, a component of gardenia jasminoides, beneficial effects to alleviate LPS-induced immune stress in piglets.

Geniposide is the main medicinal component of Gardenia jasminoides, and its content is approximately 3-8% depending on its origin. Geniposide is a class of cyclic enol ether terpene glucoside compounds with strong antioxidant, free radical quenching and cancer-inhibiting activities. Many studies have reported that geniposide has hepatoprotective, cholestatic, neuroprotective, blood sugar and blood lipid regulation, soft tissue damage treatment, antithrombotic, antitumor and other effects. As a traditional Chinese medicine, gardenia, whether used as gardenia alone, as the monomer geniposide or as the effective part of cyclic either terpenoids, has been reported to have anti-inflammatory effects when used in the right amounts. Recent studies have found that geniposide has important roles in pharmacological activities such as anti-inflammation activity, inhibition of the NF-κB/IκB pathway, and cell adhesion molecule production. In this study, we predicted the anti-inflammatory and antioxidant effects of geniposide in piglets through network pharmacology based on the LPS-induced inflammatory response-regulated signaling pathway. The effects of geniposide on changes in inflammatory pathways and cytokine levels in the lymphocytes of inflammation-stressed piglets were investigated using in vivo and in vitro models of piglet lipopolysaccharide-induced oxidative stress. Network pharmacology identified 23 target genes, of which the main pathways of action were lipid and atherosclerosis, fluid shear stress and atherosclerosis, and Yersinia infection. The main relevant target genes were VEGFA, ROCK2, NOS3, and CCL2. Validation experiments showed that the interventional effects of geniposide reduced the relative expression of NF-κB pathway proteins and genes, restored the expression of COX-2 genes to normal levels, and increased the relative expression of tight junction proteins and genes in IPEC-J2 cells. This indicates that the addition of geniposide can alleviate inflammation and improve the level of cellular tight junctions.

Network Pharmacology Prediction and Experimental Verification for Anti-Ferroptosis of Edaravone After Experimental Intracerebral Hemorrhage

Neuronal ferroptosis plays an important role in secondary brain injuries after intracerebral hemorrhage (ICH). Edaravone (Eda) is a promising free radical scavenger that inhibits ferroptosis in neurological diseases. However, its protective effects and underlying mechanisms in ameliorating post-ICH ferroptosis remain unclear. We employed a network pharmacology approach to determine the core targets of Eda against ICH. Forty-two rats were subjected to successful striatal autologous whole blood injection (n=28) or sham operation (n=14). The 28 blood-injected rats were randomly assigned to either the Eda or vehicle group (n=14) for immediate administration and then for 3 consecutive days. Hemin-induced HT22 cells were used for in vitro studies. The effects of Eda in ICH on ferroptosis and the MEK/ERK pathway were investigated in vivo and in vitro. Network pharmacology-based analysis revealed that candidate targets of Eda-treated ICH might be related to ferroptosis; among which prostaglandin G/H synthase 2 (PTGS2) was a ferroptosis marker. In vivo experiments showed that Eda alleviated sensorimotor deficits and decreased PTGS2 expression (all p<0.05) after ICH. Eda rescued neuron pathological changes after ICH (increased NeuN+ cells and decreased FJC+ cells, all p<0.01). In vitro experiments showed that Eda reduced intracellular reactive oxygen species and reversed mitochondria damage. Eda repressed ferroptosis by decreasing malondialdehyde and iron deposition and by influencing ferroptosis-related protein expression (all p<0.05) in ICH rats and hemin-induced HT22 cells. Mechanically, Eda significantly suppressed phosphorylated-MEK and phosphorylated-ERK1/2 expression. These results indicate that Eda has protective effects on ICH injury through ferroptosis and MEK/ERK pathway suppression.

The inhibitory effect and mechanism of Resina Draconis on the proliferation of MCF-7 breast cancer cells: a network pharmacology-based analysis

Resina Draconis (RD) is known as the "holy medicine for promoting blood circulation" and possesses antitumor properties against various types of cancer, including breast cancer (BC); however, the underlying mechanism is not well understood. To explore the potential mechanism of RD against BC using network pharmacology and experimental validation, data on bioactive compounds, potential targets of RD, and related genes of BC were obtained from multiple public databases. Gene Ontology (GO) and KEGG pathway analyses were performed via the DAVID database. Protein interactions were downloaded from the STRING database. The mRNA and protein expression levels and survival analysis of the hub targets were analyzed using the UALCAN, HPA, Kaplan‒Meier mapper, and cBioPortal databases. Subsequently, molecular docking was used to verify the selected key ingredients and hub targets. Finally, the predicted results of network pharmacology methods were verified by cell experiments. In total, 160 active ingredients were obtained, and 148 RD target genes for the treatment of BC were identified. KEGG pathway analysis indicated that RD exerted its therapeutic effects on BC by regulating multiple pathways. Of these, the PI3K-AKT pathway was indicated to play an important role. In addition, RD treatment of BC seemed to involve the regulation of hub targets that were identified based on PPI interaction network analysis. Validation in different databases showed that AKT1, ESR1, HSP90AA1, CASP3, SRC and MDM2 may be involved in the carcinogenesis and progression of BC and that ESR1, IGF1 and HSP90AA1 were correlated with worse overall survival (OS) in BC patients. Molecular docking results showed that 103 active compounds have good binding activity with the hub targets, among which flavonoid compounds were the most important active components. Therefore, the sanguis draconis flavones (SDF) were selected for subsequent cell experiments. The experimental results showed that SDF significantly inhibited the cell cycle and cell proliferation of MCF-7 cells through the PI3K/AKT pathway and induced MCF-7 cell apoptosis. This study has preliminarily reported on the active ingredients, potential targets, and molecular mechanism of RD against BC, and RD was shown to exert its therapeutic effects on BC by regulating the PI3K/AKT pathway and related gene targets. Importantly, our work could provide a theoretical basis for further study of the complex anti-BC mechanism of RD.

Insights into the mechanisms of triptolide nephrotoxicity through network pharmacology-based analysis and RNA-seq.

Triptolide (TPL) is a promising plant-derived compound for clinical therapy of multiple human diseases; however, its application was limited considering its toxicity. To explore the underlying molecular mechanism of TPL nephrotoxicity, a network pharmacology based approach was utilized to predict candidate targets related with TPL toxicity, followed by deep RNA-seq analysis to characterize the features of three transcriptional elements include protein coding genes (PCGs), long noncoding RNAs (lncRNAs) and circular RNAs (circRNAs) as well as their associations with nephrotoxicity in rats with TPL treatment. Although the deeper mechanisms of TPL nephrotoxcity remain further exploration, our results suggested that c-Jun is a potential target of TPL and Per1 related circadian rhythm signaling is involved in TPL induced renal toxicity.

Antiviral and anti-inflammatory activity of natural compounds against Japanese encephalitis virus via inhibition of NS5 protein and regulation of key immune and inflammatory signaling pathways.

Japanese encephalitis virus (JEV) is the foremost cause of viral encephalitis in Southeast Asia and Australia leading to approximately 68 000 clinical cases and about 13 600-20 400 deaths annually. Vaccination is not completely sure and safe. Despite this, no specific antiviral has been available or approved for JEV infection yet and treatment is generally symptomatic. Therefore, this study aims to examine the antiviral activity of natural compounds against JEV proteins. The antiviral activity of natural compounds was investigated via molecular docking, cytopathic effect (CPE) inhibition assay, western blotting, and indirect immunofluorescence assay. Physiochemical, pharmacokinetics, and toxicity analysis were evaluated for the safety and efficacy of natural compounds. Network pharmacology-based approaches have been used to study the molecular mechanisms of drug-target interactions. Molecular docking results suggested that the NS5 protein of JEV is the major target for natural compounds. Network pharmacology-based analysis revealed that these drugs majorly target IL6, AKT1, tumor necrosis factor (TNF), and PTGS2 to regulate key immune and inflammatory pathways such as nuclear factor kappa B, PI3K-Akt, and TNF signaling, during JEV infection. Our in vitro results show that among the natural compounds, curcumin provides the highest protection against JEV infection via reducing the JEV-induced CPE (IC50 = 5.90 ± 0.44 µM/mL), and reduces the expression of NS5 protein, IL6, AKT1, TNF-α, and PTGS2. However, other natural compounds also provide protection to some extent but their efficacy is lower compared to curcumin. Therefore, this study shows that natural compounds, mainly curcumin, may offer novel therapeutic avenues for the treatment of JEV via inhibiting key viral proteins and regulating crucial host pathways involved in JEV replication.

Network pharmacology-based analysis of Resinacein S against non-alcoholic fatty liver disease by modulating lipid metabolism.

Ganoderma lucidum is reportedly the best source of traditional natural bioactive constituents. Ganoderma triterpenoids (GTs) have been verified as an alternative adjuvant for treating leukemia, cancer, hepatitis and diabetes. One of the major triterpenoids, Resinacein S, has been found to regulate lipid metabolism and mitochondrial biogenesis. Nonalcoholic fatty liver disease (NAFLD) is a common chronic liver disease that has become a major public health problem. Given the regulatory effects on lipid metabolism of Resinacein S, we sought to explore potential protective effects against NAFLD. Resinacein S was extracted and isolated from G. lucidum. And mice were fed with high fat diet with or without Resinacein S to detect hepatic steatosis. According to Network Pharmacology and RNA-seq, we analyzed the hub genes of Resinacein S against NAFLD disease. Our results can be summarized as follows: (1) The structure of Resinacein S was elucidated using NMR and MS methods. (2) Resinacein S treatment could significantly attenuate high-fat diet (HFD)-induced hepatic steatosis and hepatic lipid accumulation in mouse. (3) GO terms, KEGG pathways and the PPI network of Resinacein S induced Differentially Expressed Genes (DEGs) demonstrated the key target genes of Resinacein S against NAFLD. (4) The hub proteins in PPI network analysis could be used for NAFLD diagnosis and treatment as drug targets. Resinacein S can significantly change the lipid metabolism in liver cells and yield a protective effect against steatosis and liver injury. Intersected proteins between NAFLD related genes and Resinacein S-induced DEGs, especially the hub protein in PPI network analysis, can be used to characterize targets of Resinacein S against NAFLD.

Usnic acid as potential inhibitors of BCL2 and P13K protein through network pharmacology-based analysis, molecular docking and molecular dynamic simulation

Usnic acid (UA) lately piqued the interest of researchers for its extraordinary biological characteristics, including anticancer activity. Here, the mechanism was clarified through network pharmacology,molecular docking and molecular dynamic simulation. Sixteen proteins were selected through network pharmacology study as they are probable to interact with UA. Out of these proteins, 13 were filtered from PPI network analysis based on their significance of interactions (p < 0.05). KEGG pathway analysis has also aided us in determining the three most significant protein targets for UA, which are BCL2, PI3KCA and PI3KCG. Therefore molecular docking and molecular dynamic (MD) simulations throughout 100 ns were performed for usnic acid onto the three proteins mentioned. However, UA's docking score in all proteins is lower than their co-crystalised ligand, especially for BCL2 (−36.5158 kcal/mol) and PI3KCA (−44.5995 kcal/mol) proteins. The only exception is PI3KCG which has comparable results with the co-crystallised ligand with (−41.9351 kcal/mol). Furthermore, MD simulation has also revealed that usnic acid does not stay fit in the protein throughout the simulation trajectory for PI3KCA protein evident from RMSF and RMSD plots. Nevertheless, it still poses good ability in inhibiting BCL2 and PI3KCG protein in MD simulation. In the end, usnic acid has exhibited good potential in the inhibition of PI3KCG proteins, rather than the other proteins mentioned. Thus further study on structural modification of usnic acid could enhance the ability of usnic acid in the inhibition of PI3KCG as anti-colorectal and anti-small cell lung cancer drug candidate.Communicated by Ramaswamy H. Sarma