In-silico prediction of drug targets, biological activities, signal pathways and regulating networks of dioscin based on bioinformatics.
ABSTRACT: Inverse docking technology has been a trend of drug discovery, and bioinformatics approaches have been used to predict target proteins, biological activities, signal pathways and molecular regulating networks affected by drugs for further pharmacodynamic and mechanism studies.In the present paper, inverse docking technology was applied to screen potential targets from potential drug target database (PDTD). Then, the corresponding gene information of the obtained drug-targets was applied to predict the related biological activities, signal pathways and processes networks of the compound by using MetaCore platform. After that, some most relevant regulating networks were considered, which included the nodes and relevant pathways of dioscin.71 potential targets of dioscin from humans, 7 from rats and 8 from mice were screened, and the prediction results showed that the most likely targets of dioscin were cyclin A2, calmodulin, hemoglobin subunit beta, DNA topoisomerase I, DNA polymerase lambda, nitric oxide synthase and UDP-N-acetylhexosamine pyrophosphorylase, etc. Many diseases including experimental autoimmune encephalomyelitis of human, temporal lobe epilepsy of rat and ankylosing spondylitis of mouse, may be inhibited by dioscin through regulating immune response alternative complement pathway, G-protein signaling RhoB regulation pathway and immune response antiviral actions of interferons, etc. The most relevant networks (5 from human, 3 from rat and 5 from mouse) indicated that dioscin may be a TOP1 inhibitor, which can treat cancer though the cell cycle- transition and termination of DNA replication pathway. Dioscin can down regulate EGFR and EGF to inhibit cancer, and also has anti-inflammation activity by regulating JNK signaling pathway.The predictions of the possible targets, biological activities, signal pathways and relevant regulating networks of dioscin provide valuable information to guide further investigation of dioscin on pharmacodynamics and molecular mechanisms, which also suggests a practical and effective method for studies on the mechanism of other chemicals.
Project description:We previously reported the promising effects of dioscin against liver injury, but its effect on liver fibrosis remains unknown. The present work investigated the activities of dioscin against liver fibrosis and the underlying molecular mechanisms. Dioscin effectively inhibited the cell viabilities of HSC-T6, LX-2 and primary rat hepatic stellate cells (HSCs), but not hepatocytes. Furthermore, dioscin markedly increased peroxisome proliferator activated receptor-γ (PPAR-γ) expression and significantly reduced a-smooth muscle actin (α-SMA), transforming growth factor-β1 (TGF-β1), collagen α1 (I) (COL1A1) and collagen α1 (III) (COL3A1) levels in vitro. Notably, dioscin inhibited HSCs activation and induced apoptosis in activated HSCs. In vivo, dioscin significantly improved body weight and hydroxylproline, laminin, α-SMA, TGF-β1, COL1A1 and COL3A1 levels, which were confirmed by histopathological assays. Dioscin facilitated matrix degradation, and exhibited hepatoprotective effects through the attenuation of oxidative stress and inflammation, in addition to exerting anti-fibrotic effects through the modulation of the TGF-β1/Smad, Wnt/β-catenin, mitogen-activated protein kinase (MAPK) and mitochondrial signaling pathways, which triggered the senescence of activated HSCs. In conclusion, dioscin exhibited potent effects against liver fibrosis through the modulation of multiple targets and signaling pathways and should be developed as a novel candidate for the treatment of liver fibrosis in the future.
Project description:Inhalation of crystalline silica particles leads to pulmonary fibrosis, eventually resulting in respiratory failure and death. There are few effective drugs that can delay the progression of this disease; thus, patients with silicosis are usually only offered supportive care. Dioscin, a steroidal saponin, exhibits many biological activities and health benefits including its protective effects against hepatic fibrosis. However, the effect of dioscin on silicosis is unknown.We employed experimental mouse mode of silicosis. Different doses of dioscin were gavaged to the animals 1 day after crystalline silica instillation to see the effect of dioscin on crystalline silica induced pulmonary fibrosis. Also, we used RAW264.7 and NIH-3T3 cell lines to explore dioscin effects on macrophages and fibroblasts. Dioscin was also oral treatment but 10 days after crystalline silica instillation to see its effect on established pulmonary fibrosis.Dioscin treatment reduced pro-inflammation and pro-fibrotic cytokine secretion by modulating innate and adaptive immune responses. It also reduced the recruitment of fibrocytes, protected epithelial cells from crystalline silica injury, inhibited transforming growth factor beta/Smad3 signaling and fibroblast activation. Together, these effects delayed the progression of crystalline silica-induced pulmonary fibrosis. The mechanism by which dioscin treatment alleviated CS-induced inflammation appeared to be via the reduction of macrophage, B lymphocyte, and T lymphocte infiltration into lung. Dioscin inhibits macrophages and fibroblasts from secreting pro-inflammatory cytokines and may also function as a modulator of T helper cells responses, concurrent with attenuated phosphorylation of the apoptosis signal-regulating kinase 1-p38/c-Jun N-terminal kinase pathway. Also, dioscin could block the phosphorylation of Smad3 in fibroblast. Oral treatment of dioscin could also effectively postpone the progression of established silicosis.Oral treatment dioscin delays crystalline silica-induced pulmonary fibrosis and exerts pulmonary protective effects in mice. Dioscin may be a novel and potent candidate for protection against crystalline silica-induced pulmonary fibrosis.
Project description:The purpose of present study was to investigate the effect of dioscin on activity of adriamycin (ADR) in ADR-sensitive (MCF-7) and ADR-resistant (MCF-7/ADR) human breast cancer cells and to clarify the molecular mechanisms involved. Antiproliferation effect of ADR was enhanced by dioscin in MCF-7 and MCF-7/ADR cells. Dioscin significantly inhibited MDR1 mRNA and protein expression and MDR1 promoter and nuclear factor κ-B (NF-κB) activity in MCF-7/ADR cells. Additionally, inhibitor κB-α (IκB-α) degradation was inhibited by dioscin. Moreover, dioscin induced the formation of vacuoles in the cytoplasm and protein level of LC3-II in MCF-7 and MCF-7/ADR cells. Autophagy inhibitor 3-MA abolished the effect of dioscin on ADR cytotoxicity. Dioscin inhibited phosphorylation of PI3K and Akt, resulting in upregulation of LC3-II expression. In conclusion, dioscin increased ADR chemosensitivity by down-regulating MDR1 expression through NF-κB signaling inhibition in MCF-7/ADR cells. Autophagy was induced by dioscin to ameliorate the cytotoxicity of ADR via inhibition of the PI3K/AKT pathways in MCF-7 and MCF-7/ADR cells. These findings provide evidence in support of further investigation into the clinical application of dioscin as a chemotherapy adjuvant.
Project description:Hepatic stellate cells (HSCs) migration, an important bioprocess, contributes to the development of liver fibrosis. Our previous studies have found the potent activity of dioscin against liver fibrosis by inhibiting HSCs proliferation, triggering the senescence and inducing apoptosis of activated HSCs, but the molecular mechanisms associated with cell migration were not clarified. In this work, iTRAQ (isobaric tags for relative and absolution quantitation)-based quantitative proteomics study was carried out, and a total of 1566 differentially expressed proteins with fold change ?2.0 and p < 0.05 were identified in HSC-T6 cells treated by dioscin (5.0 ?g/mL). Based on Gene Ontology classification, String and KEGG pathway assays, the effects of dioscin to inhibit cell migration via regulating SDC-4 were carried out. The results of wound-healing, cell migration and western blotting assays indicated that dioscin significantly inhibit HSC-T6 cell migration through SDC-4-dependent signal pathway by affecting the expression levels of Fn, PKC?, Src, FAK, and ERK1/2. Specific SDC-4 knockdown by shRNA also blocked HSC-T6 cell migration, and dioscin slightly enhanced the inhibiting effect. Taken together, the present work showed that SDC-4 played a crucial role on HSC-T6 cell adhesion and migration of dioscin against liver fibrosis, which may be one potent therapeutic target for fibrotic diseases.
Project description:The present work aimed to investigate the activities and underlying mechanisms of dioscin against alcoholic liver fibrosis (ALF). In vivo liver fibrosis in mice was induced by an alcoholic liquid diet, and in vitro studies were performed on activated HSC-T6 and LX2 cells treated with lipopolysaccharide. Our results showed that dioscin significantly attenuated hepatic stellate cells (HSCs) activation, improved collagen accumulation, and attenuated inflammation through down-regulating the levels of myeloid differentiation factor 88 (MyD88), nuclear factor ?B (NF-?B), interleukin (IL)-1, IL-6 and tumour necrosis factor-? by decreasing Toll-like receptor (TLR)4 expression both in vivo and in vitro. TLR4 overexpression was also decreased by dioscin, leading to the markedly down-regulated levels of MyD88, NF-?B, transforming growth factor-?1 (TGF-?1), ?-smooth muscle actin (?-SMA) and type I collagen (COL1A1) in cultured HSCs. Suppression of cellular MyD88 by ST2825 or abrogation of NF-?B by pyrrolidine dithiocarbamate eliminated the inhibitory effects of dioscin on the levels of TGF-?1, ?-SMA and COL1A1. In a word, dioscin exhibited potent effects against ALF via altering TLR4/MyD88/NF-?B signaling pathway, which provided novel insights into the mechanisms of this compound as an antifibrogenic candidate for the treatment of ALF in the future.
Project description:BACKGROUND AND PURPOSE:Dioscin shows potent effects against cancers. We aimed to elucidate its pharmacological effects and mechanisms of action on hepatocellular carcinoma (HCC) in vivo and in vitro. EXPERIMENTAL APPROACH:Effects of dioscin were investigated in SMMC7721 and HepG2 cells, diethylnitrosamine-induced primary liver cancer in rats, and cell xenografts in nude mice. Isobaric tags for relative and absolution quantitation (iTRAQ)-based proteomics was used to find dioscin's targets and investigate its mechanism. KEY RESULTS:In SMMC7721 and HepG2 cells dioscin markedly inhibited cell proliferation and migration, induced apoptosis, autophagy, and DNA damage. It inhibited DEN-induced primary liver cancer in rats, markedly changed body weights and restored levels of ? fetoprotein, alanine transaminase, aspartate transaminase, ?-glutamyltransferase, alkaline phosphatase, and Ki67. It also inhibited growth of xenografts in mice. In SMMC7721 cells, 191 differentially expressed proteins were found after dioscin, based on iTRAQ-based assay. TP53-inducible glycolysis and apoptosis regulator (TIGAR) was identified as being significantly down-regulated by dioscin. Dioscin induced cell apoptosis, autophagy, and DNA damage via increasing expression levels of p53, cleaved PARP, Bax, cleaved caspase-3/9, Beclin-1, and LC3 and suppressing those of Bcl-2, p-Akt, p-mammalian target of rapamycin (mTOR), CDK5, p-ataxia telangiectasia-mutated gene (ATM). The transfection of TIGAR siRNA into SMMC7721 cells and xenografts in nude mice further confirmed that the potent activity of dioscin against HCC is evoked by adjusting TIGAR-mediated inhibition of p53, Akt/mTOR, and CDK5/ATM pathways. CONCLUSIONS AND IMPLICATIONS:The data suggest that dioscin has potential as a therapeutic, and TIGAR as a drug target for treating HCC.
Project description:TarFisDock is a web-based tool for automating the procedure of searching for small molecule-protein interactions over a large repertoire of protein structures. It offers PDTD (potential drug target database), a target database containing 698 protein structures covering 15 therapeutic areas and a reverse ligand-protein docking program. In contrast to conventional ligand-protein docking, reverse ligand-protein docking aims to seek potential protein targets by screening an appropriate protein database. The input file of this web server is the small molecule to be tested, in standard mol2 format; TarFisDock then searches for possible binding proteins for the given small molecule by use of a docking approach. The ligand-protein interaction energy terms of the program DOCK are adopted for ranking the proteins. To test the reliability of the TarFisDock server, we searched the PDTD for putative binding proteins for vitamin E and 4H-tamoxifen. The top 2 and 10% candidates of vitamin E binding proteins identified by TarFisDock respectively cover 30 and 50% of reported targets verified or implicated by experiments; and 30 and 50% of experimentally confirmed targets for 4H-tamoxifen appear amongst the top 2 and 5% of the TarFisDock predicted candidates, respectively. Therefore, TarFisDock may be a useful tool for target identification, mechanism study of old drugs and probes discovered from natural products. TarFisDock and PDTD are available at http://www.dddc.ac.cn/tarfisdock/.
Project description:Recent researches have shown that estrogen receptor-β (ERβ) activator may be a potent anticancer agent for prostate cancer (PCa), and our previous study also indicated that dioscin can upregulate the expression of ERβ in MC3T3-E1 cell. In the present work, the activity and mechanism of dioscin, a natural product, against PCa were investigated. The results showed that dioscin markedly inhibited cell viability, colony formation, motility and induced apoptosis in PC3 cells. Moreover, dioscin disrupted the formation of PC3 cell-derived mammospheres and reduced aldehyde dehydrogenase (ALDH) level and the CD133+/CD44+ cells, indicating that dioscin had a potent inhibitory activity on prostate cancer stem cells (PCSCs). In vivo results also showed that dioscin significantly suppressed the tumor growth of PC3 cell xenografts in nude mice. Furthermore, mechanism investigation showed that dioscin markedly upregulated ERβ expression level, subsequently increased prolyl hydroxylase 2 level, decreased the levels of hypoxia-inducible factor-1α, vascular endothelial growth factor A and BMI-1, and thus induced cell apoptosis by regulating the expression levels of caspase-3 and Bcl-2 family proteins. In addition, transfection experiment of ERβ-siRNA further indicated that diosicn showed excellent activity against PCa in vitro and in vivo by increasing ERβ expression level. The co-immunoprecipitation (Co-IP) results further suggested that dioscin promoted the interaction of c-ABL and ERβ, but did not change c-ABL expression. Moreover, the molecular docking assay showed that dioscin processed powerful affinity toward to ERβ mainly through the strong hydrogen bonding and hydrophobic effects, and the actions of dioscin on ERβ activation and tumor cells inhibition were significantly weakened in the mutational (Phe-336, Phe-468) PC3 cells. Collectively, these findings proved that dioscin exerted efficient anti-PCa activity via activation of ERβ, which should be developed as an efficient candidate in clinical for treating this cancer in the future.
Project description:Intrahepatic cholestasis, a clinical syndrome, is caused by excessive accumulation of bile acids in body and liver. Proper regulation of bile acids in liver cells is critical for liver injury. We previously reported the effects of dioscin against ?-naphthylisothio- cyanate (ANIT)-induced cholestasis in rats. However, the pharmacological and mechanism data are limited. In our work, the animals of rats and mice, and Sandwich-cultured hepatocytes (SCHs) were caused by ANIT, and dioscin was used for the treatment. The results showed that dioscin markedly altered relative liver weights, restored ALT, AST, ALP, TBIL, GSH, GSH-Px, MDA, SOD levels, and rehabilitated ROS level and cell apoptosis. In mechanism study, dioscin not only significantly regulated the protein levels of Ntcp, OAT1, OCT1, Bsep and Mrp2 to accelerate bile acids excretion, but also regulated the expression levels of Bak, Bcl-xl, Bcl-2, Bax, Caspase 3 and Caspase 9 in vivo and in vitro to improve apoptosis. In addition, dioscin markedly inhibited PI3K/Akt pathway and up-regulated the levels of Nrf2, GCLc, GCLm, NQO1 and HO-1 against oxidative stress (OS) caused by bile acids. These results were further validated by inhibition of PI3K and Akt using the inhibitors of wortmannin and perifosine in SCHs. Our data showed that dioscin had good action against ANIT-caused intrahepatic cholestasis through regulating transporters, apoptosis and OS. This natural product can be considered as one active compound to treat intrahepatic cholestasis in the future.
Project description:Pathway analysis is widely used to gain mechanistic insights from high-throughput omics data. However, most existing methods do not consider signal integration represented by pathway topology, resulting in enrichment of convergent pathways when downstream genes are modulated. Incorporation of signal flow and integration in pathway analysis could rank the pathways based on modulation in key regulatory genes. This implementation can be facilitated for large-scale data by discrete state network modeling due to simplicity in parameterization. Here, we model cellular heterogeneity using discrete state dynamics and measure pathway activities in cross-sectional data. We introduce a new algorithm, Boolean Omics Network Invariant-Time Analysis (BONITA), for signal propagation, signal integration, and pathway analysis. Our signal propagation approach models heterogeneity in transcriptomic data as arising from intercellular heterogeneity rather than intracellular stochasticity, and propagates binary signals repeatedly across networks. Logic rules defining signal integration are inferred by genetic algorithm and are refined by local search. The rules determine the impact of each node in a pathway, which is used to score the probability of the pathway's modulation by chance. We have comprehensively tested BONITA for application to transcriptomics data from translational studies. Comparison with state-of-the-art pathway analysis methods shows that BONITA has higher sensitivity at lower levels of source node modulation and similar sensitivity at higher levels of source node modulation. Application of BONITA pathway analysis to previously validated RNA-sequencing studies identifies additional relevant pathways in in-vitro human cell line experiments and in-vivo infant studies. Additionally, BONITA successfully detected modulation of disease specific pathways when comparing relevant RNA-sequencing data with healthy controls. Most interestingly, the two highest impact score nodes identified by BONITA included known drug targets. Thus, BONITA is a powerful approach to prioritize not only pathways but also specific mechanistic role of genes compared to existing methods. BONITA is available at: https://github.com/thakar-lab/BONITA.