Toxic effects and comparison of common amino antioxidants (AAOs) in the environment on zebrafish: A comprehensive analysis based on cells, embryos, and adult fish.

The ubiquity of amino antioxidants (AAOs) in the environment has attracted increasing attention, given their potential toxicity. This investigation represents a pioneering effort, systematically scrutinizing the toxicological effects of four distinct AAOs across the developmental spectrum of zebrafish, encompassing embryonic, larvae, and adult stages. The results indicate that four types of AAO exhibit varying degrees of cell proliferation toxicity. Although environmentally relevant concentrations of AAOs exhibit a comparatively circumscribed impact on zebrafish embryo development, heightened concentrations (300 µg/L) of N-(1,3-dimethylbutyl)-N'-phenyl-p-phenylenediamine (6PPD) and N-isopropyl-N'-phenyl-p-phenylenediamine (IPPD) distinctly evoke developmental toxicity. Behavioral analysis results indicate that at concentrations of 20 and 300 µg/L, the majority of AAOs significantly reduced the swimming speed and activity of larvae. Moreover, each AAO triggers the generation of reactive oxygen species (ROS) in larvae, instigating diverse levels of oxidative stress. The study delineates parallel toxicological patterns in zebrafish exposed to 300 µg/L of 6PPD and IPPD, thereby establishing a comparable toxicity profile. The comprehensive toxicity effects among the four AAOs is as follows: IPPD >6PPD > N-Phenyl-1-naphthylamine (PANA) > diphenylamine (DPA). These findings not only enrich our comprehension of the potential hazards associated with AAOs but also provide data support for structure-based toxicity prediction models.

Aerobic Oxidation of 5-Hydroxymethylfurfural via Hydrogen Bonds Reconstruction with Ternary Deep Eutectic Solvents.

Hydrogen bonding effect exists widely in various chemical and biochemical systems, primarily stabilizing the molecular structure as a positive factor. However, the presence of intermolecular hydrogen bonds among biomass molecules results in a formidable challenge for the efficient utilization of biomass resources. Here in, a novel strategy of "hydrogen bonds reconstruction" was developed by a series of ternary deep eutectic solvent (DESs) as molecular scissors, which disrupting the initial intermolecular hydrogen bonds and reconstructing the new ones to increase the reactivity of the biomass-based compound. The DESs played a crucial role in enhancing the reactivity of 5-hydroxymethylfurfural (HMF) and promoting its oxidation through reconstructing the hydrogen bonds interactions. Furthermore, DESs was also found to activate the Anderson-type catalyst Na5IMo6O24 (IMo6) through an electron-transfer mechanism, which facilitated the generation of oxygen vacancies and significantly enhances its ability to activate molecular oxygen. With this novel catalytic system, oxidation of HMF exhibited remarkable efficiency as HMF was almost entirely converted into FFCA with an impressive yield of 98 % under the optimized conditions. This finding offers novel insights into the utilization of biomass resources and endows the solvent with new functions in the chemical reaction.

[Therapeutic effect of ursodeoxycholic acid-berberine supramolecular nanoparticles on ulcerative colitis based on supramolecular system induced by weak bond].

Ulcerative colitis(UC) is a recurrent, intractable inflammatory bowel disease. Coptidis Rhizoma and Bovis Calculus, serving as heat-clearing and toxin-removing drugs, have long been used in the treatment of UC. Berberine(BBR) and ursodeoxycholic acid(UDCA), the main active components of Coptidis Rhizoma and Bovis Calculus, respectively, were employed to obtain UDCA-BBR supramolecular nanoparticles by stimulated co-decocting process for enhancing the therapeutic effect on UC. As revealed by the characterization of supramolecular nanoparticles by field emission scanning electron microscopy(FE-SEM) and dynamic light scattering(DLS), the supramolecular nanoparticles were tetrahedral nanoparticles with an average particle size of 180 nm. The molecular structure was described by ultraviolet spectroscopy, fluorescence spectroscopy, infrared spectroscopy, high-resolution mass spectrometry, and hydrogen-nuclear magnetic resonance(H-NMR) spectroscopy. The results showed that the formation of the supramolecular nano-particle was attributed to the mutual electrostatic attraction and hydrophobic interaction between BBR and UDCA. Additionally, supramolecular nanoparticles were also characterized by sustained release and pH sensitivity. The acute UC model was induced by dextran sulfate sodium(DSS) in mice. It was found that supramolecular nanoparticles could effectively improve body mass reduction and colon shortening in mice with UC(P<0.001) and decrease disease activity index(DAI)(P<0.01). There were statistically significant differences between the supramolecular nanoparticles group and the mechanical mixture group(P<0.001, P<0.05). Enzyme-linked immunosorbent assay(ELISA) was used to detect the serum levels of tumor necrosis factor-α(TNF-α) and interleukin-6(IL-6), and the results showed that supramolecular nanoparticles could reduce serum TNF-α and IL-6 levels(P<0.001) and exhibited an obvious difference with the mechanical mixture group(P<0.01, P<0.05). Flow cytometry indicated that supramolecular nanoparticles could reduce the recruitment of neutrophils in the lamina propria of the colon(P<0.05), which was significantly different from the mechanical mixture group(P<0.05). These findings suggested that as compared with the mechanical mixture, the supramolecular nanoparticles could effectively improve the symptoms of acute UC in mice. The study provides a new research idea for the poor absorption of small molecules and the unsatisfactory therapeutic effect of traditional Chinese medicine and lays a foundation for the research on the nano-drug delivery system of traditional Chinese medicine.

Integrative multi-omics unravels the amelioration effects of Zanthoxylum bungeanum Maxim. on non-alcoholic fatty liver disease.

BACKGROUND: The effect of Zanthoxylum bungeanum Maxim. (ZBM) on anti-obesity, lipid-lowering and liver protection has been identified, but the effect on the development of NAFLD induced by high-fat diet remains unclear. PURPOSE: To evaluate the alleviation effect of ZBM on NAFLD in vivo and explore the mechanisms by analyzing the liver transcriptome, microbiota and fecal metabolites. METHODS: NAFLD model was induced in C57BL/6J mice by feeding with high-fat diet (HFD). The potential mechanism of ZBM in improving NAFLD was studied by liver transcriptome analysis, real-time PCR, immunofluorescence, 16s rRNA sequencing and non-targeted metabonomics. RESULTS: ZBM has alleviation effects on HFD-induced NAFLD. The liver transcriptome, real-time PCR and immunofluorescence analysis showed that ZBM could efficiently regulate fatty acid and cholesterol metabolism. The 16S rRNA sequencing and LC-MS based metabonomic demonstrated that ZBM could rebalance gut microbiota dysbiosis and regulate metabolic profiles in HFD-induced NAFLD mice. Spearman correlation analysis revealed a strong correlation between gut microbiota and biochemical, pathological indexes and differential metabolic biomarkers. CONCLUSION: ZBM ameliorates HFD-induced NAFLD by regulating fatty acid and cholesterol metabolism, gut microbiota and metabolic profile.

Qu-Du-San-Jie decoction induces growth inhibition and vascular normalization in NF2-associated vestibular schwannoma.

Background: Neurofibromatosis type 2 (NF2) is a rare genetic syndrome that predisposes individuals to develop bilateral vestibular schwannomas (VSs) causing a high risk of life-threatening neurological complications. Traditional treatment options for NF2-associated VS usually cause neurological damage, and to date, there are no FDA-approved pharmacotherapies for NF2. The aim of this study was to evaluate the antitumor efficacy of Qu-Du-San-Jie (QDSJ) decoction, a traditional Chinese medicine formula, on NF2-associated VS and to investigate the potential underlying mechanisms. Methods: Ultra high-performance liquid chromatography-mass spectroscopy (UHPLC-MS) analysis was performed to identify the components of QDSJ and their targets. To determine the relationships between the putative targets of QDSJ and the differential genes of NF2-associated VS, the drug-disease crossover genes were screened using the UHPLC-MS data combined with our previous gene expression profiling data. The differentially expressed genes were imported into the STRING database to generate a PPI network. Differentially expressed gene targets and pathways were identified using GO and KEGG pathway enrichment analyses. The in vitro and in vivo drug efficacy of QDSJ decoction was tested using a patient-derived schwannoma cell line and a patient-derived xenograft mouse model, respectively. H&E staining, immunochemistry, and immunofluorescence staining were used to evaluate the cell proliferation and tumor vessels. Results: A total of 133 compounds were identified in QDSJ decoction using UHPLC-MS analysis. Network pharmacology showed that the regulation of necroptosis, apoptosis, cell cycle, angiogenesis, adherens junction, and neuroactive ligand-receptor interaction could be associated with the efficacy of QDSJ in treating NF2-associated VS. Treatment with QDSJ induced necrotic cell death and apoptosis of schwannoma cells in vitro and suppressed the tumor growth in vivo. Histopathological analysis revealed areas of cell necrosis and enlarged tumor blood vessels in the QDSJ-treated tumors. The numbers of cells positive for Cyclin D1 and Ki-67 were significantly reduced in QDSJ-treated tumors compared to control tumors. Immunofluorescence staining of CD31 and αSMA showed a decreased number and density of tumor vessels and normalized vessel structure in QDSJ-treated tumors. Conclusion: Our study demonstrates that QDSJ decoction shows significant antitumor activity against NF2-associated schwannoma and is a possible candidate for future clinical trials.

Manpixiao Decoction Halted the Malignant Transformation of Precancerous Lesions of Gastric Cancer: From Network Prediction to In-Vivo Verification.

Manpixiao decoction (MPX), a traditional Chinese medicine formula, is mainly used to improve the gastric mucosal pathology and stomach discomfort in patients with gastric precancerous lesions. Precancerous lesion of gastric cancer (PLGC) refers to intestinal metaplasia and/or dysplasia based on gastric mucosal atrophy. Effective prevention and treatment of PLGC is of great significance to reduce the incidence of gastric cancer. Because of the complexity of the etiology and pathogenesis of PLGC, there is no unified and effective treatment plan in western medicine. In recent years, traditional Chinese medicine has shown obvious advantages in the treatment of PLGC and the prevention of its further progression to gastric cancer, relying on its multi-approach and multi-target comprehensive intervention characteristics. This study is designed to examine the protective effect of MPX against PLGC and further to reveal the engaged mechanism via integrating network pharmacology and in vivo experimental evidence. Network pharmacology results demonstrated that inflammation, immune responses, and angiogenesis might be associated with the efficacy of MPX in the treatment of PLGC, in which the PI3K-Akt, cellular senescence, P53 and protein processing in endoplasmic reticulum were involved. Then, we established a rat model of PLGC using a combination of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), sodium salicylate, irregular fasting, and ranitidine, and observed the effects after MPX treatment. Our result showed that MPX improved the pathological condition of gastric mucosa in PLGC rats and reduced the incidence of gastric cancer. Next, the analysis of serum inflammatory cytokines showed that MPX reduced the inflammation-related cytokines (such as IL-1α, IL-7, CSF-1, and CSF-3) in the serum. Additionally, MPX also had a regulation effect on the "protein/protein phosphorylation-signaling pathway" network in the core region of the PLGC rats. It is showed that MPX can inhibit the phosphorylation of PI3K-AKT, and downregulates the EGFR, ß-catenin, and N-cadherin protein levels. These results indicate that MPX halted the PLGC progression through inhibiting EGFR-PI3K-AKT related epithelial-mesenchymal transition process.

Gut Microbial Dysbiosis and Changes in Fecal Metabolic Phenotype in Precancerous Lesions of Gastric Cancer Induced With N-Methyl-N'-Nitro-N-Nitrosoguanidine, Sodium Salicylate, Ranitidine, and Irregular Diet.

Background and Aims: Precancerous lesions of gastric cancer (PLGC) are the most important pathological phase with increased risk of gastric cancer (GC) and encompass the key stage in which the occurrence of GC can be prevented. In this study, we found that the gut microbiome changed significantly during the process of malignant transformation from chronic gastritis to GC in N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) multiple factors-induced rat model. Accumulating evidence has shown that alterations in gut microbiota and metabolism are potentially linked to chronic inflammation and cancer of the gastrointestinal tract. However, the correlation of gut microbiota and metabolites, inflammatory factors, and the potential mechanism in the formation of PLGC have not yet been revealed. Methods: In this study, multiple factors including MNNG, sodium salicylate drinking, ranitidine feed, and irregular diet were used to establish a PLGC rat model. The pathological state of the gastric mucosa of rats was identified through HE staining and the main inflammatory cytokine levels in the serum were detected by the Luminex liquid suspension chip (Wayen Biotechnologies, Shanghai, China). The microbial composition and metabolites in the stool samples were tested by using 16S ribosomal RNA (rRNA) gene sequencing and non-targeted metabolomics. The correlation analysis of gut microbiota and inflammatory cytokines in the serum and gut microbiota and differential metabolites in feces was performed to clarify their biological function. Results: The results showed that compared to the control group, the gastric mucosa of the model rats had obvious morphological and pathological malignant changes and the serum levels of inflammatory cytokines including interleukin-1ß (IL-1ß), interleukin-4 (IL-4), interleukin-6 (IL-6), interleukin-10 (IL-10), interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), and macrophage colony-stimulating factor (M-CSF) increased significantly, while the level of chemokine (C-X-C motif) ligand 1 (CXCL1) in serum reduced significantly. There were significant differences in the composition of the gut microbiota and fecal metabolic profiles between the model and control rats. Among them, Lactobacillus and Bifidobacterium increased significantly, while Turicibacter, Romboutsia, Ruminococcaceae_UCG-014, Ruminococcaceae_UCG-005, and Ruminococcus_1 reduced significantly in the model rats compared to the control rats. The metabolites related to the lipid metabolism and peroxisome proliferator-activated receptor (PPAR) signaling pathway have also undergone significant changes. In addition, there was a significant correlation between the changes of the differential inflammatory cytokines in the serum, fecal metabolic phenotypes, and gut microbial dysbiosis in model rats. Conclusion: The activation of the inflammatory response, disturbance of the gut microbiota, and changes in the fecal metabolic phenotype could be closely related to the occurrence of PLGC. This study provides a new idea to reveal the mechanism of risk factors of chronic gastritis and GC from the perspective of inflammation-immune homeostasis, gut microbiota, and metabolic function balance.

Erianin, the main active ingredient of Dendrobium chrysotoxum Lindl, inhibits precancerous lesions of gastric cancer (PLGC) through suppression of the HRAS-PI3K-AKT signaling pathway as revealed by network pharmacology and in vitro experimental verification.

ETHNOPHARMACOLOGICAL RELEVANCE: Dendrobium chrysotoxum Lindl, a well-known traditional Chinese medicinal herb used in the treatment of gastric disease, is distinguished as the first of the "nine immortal grasses". Dendrobium chrysotoxum Lindl and the traditional Chinese medicine prescriptions containing Dendrobium chrysotoxum Lindl are often prescribed clinically to treat chronic gastritis and precancerous lesions of gastric cancer (PLGC), showing favorable clinical effects and medicinal value in the prevention of gastric cancer. However, the effective ingredients and pharmacological mechanisms through which Dendrobium chrysotoxum Lindl prevents and treats PLGC have not been adequately identified or interpreted. AIM OF THE STUDY: The present study aimed to evaluate the effective ingredients and pharmacological mechanisms of Dendrobium chrysotoxum Lindl in the prevention and treatment of PLGC using network pharmacology. In addition, in vitro verification was performed to evaluate the mechanism of action of Erianin, the main active ingredient in Dendrobium chrysotoxum Lindl, providing experimental evidence for the clinical use of Dendrobium chrysotoxum Lindl in the treatment of PLGC. MATERIALS AND METHODS: Using network pharmacology methods, the main ingredients in Dendrobium chrysotoxum Lindl were screened from the ETCM, BATMAN-TCM, and TCMID databases, and their potential targets were predicted using the Swiss Target Prediction platform. The targets related to PLGC were retrieved through the GeneCard database, and the targets common to the main ingredients of Dendrobium chrysotoxum Lindl and PLGC were analyzed. The protein-protein interaction (PPI) network was obtained via the STRING database and analyzed visually using Cytoscape 3.7.2. The underlying mechanisms of the common targets identified through gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were analyzed using DAVID online. The "component-target-pathway" networks of Dendrobium chrysotoxum Lindl and Erianin were visually constructed by Cytoscape 3.7.2. The biological activity evaluation of Erianin's effect on PLGC was carried out using MC cell lines, the PLGC cell model established using MNNG to induce damage in normal gastric mucosal epithelial cell (GES-1). After the intervention of different concentrations of Erianin, MC cell viability was explored using the MTT assays, cell migration was determined by wound healing assays, the cell cycle and apoptosis were analyzed using flow cytometry, and the expression levels of related proteins and their phosphorylation in the HRAS-PI3K-AKT signaling pathway were detected by Western blot. RESULTS: The "component-target-pathway" network constructed in this study showed 37 active ingredients from Dendrobium chrysotoxum Lindl and 142 overlapping targets related to both Dendrobium chrysotoxum Lindl and PLGC. The targets were associated with a variety of cancer-related signaling pathways, including Pathways in cancer, PI3K-Akt signaling pathway, Rap1 signaling pathway, Focal adhesion, Ras signaling pathway, and MAPK signaling pathway. Notably, the network showed that Erianin, the primary active ingredient from Dendrobium chrysotoxum Lindl and the component associated with the most targets, could regulate Pathways in cancer, PI3K-AKT signaling pathway, Focal adhesion, Rap1 signaling pathway, cell cycle, and RAS signaling pathway in the treatment of PLGC. Verification through in vitro experiments found that Erianin can significantly inhibit MC cell viability, inhibit cell migration, block the cell cycle in the G2/M phase, and induce cell apoptosis in a dose-dependent manner. The results of the Western blot experiment further showed that Erianin can significantly decrease the protein expression levels of HRAS, AKT, p-AKT, MDM2, Cyclin D1, and p-Gsk3ß, and increase the protein expression level of p21, which suggests that Erianin can treat PLGC by regulating the HRAS-PI3K-AKT signaling pathway. CONCLUSION: This study explained the positive characteristics of multi-component, multi-target, and multi-approach intervention with Dendrobium chrysotoxum Lindl in the treatment of PLGC. Our results suggest that Erianin may be a promising candidate in the development of prevention and treatment methods for PLGC. This study provided experimental evidence for the clinical use of Dendrobium chrysotoxum Lindl to treat PLGC and prevent gastric cancer.

The mechanisms of action of WeiChang'An Pill (WCAP) treat diarrhoea-predominant irritable bowel syndrome (IBS-D) using network pharmacology approach and in vivo studies.

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

Potential effect of Maxing Shigan decoction against coronavirus disease 2019 (COVID-19) revealed by network pharmacology and experimental verification.

ETHNOPHARMACOLOGICAL RELEVANCE: Since the occurrence of coronavirus disease 2019 (COVID-19) in Wuhan, China in December 2019, COVID-19 has been quickly spreading out to other provinces and countries. Considering that traditional Chinese medicine (TCM) played an important role during outbreak of SARS and H1N1, finding potential alternative approaches for COVID-19 treatment is necessary before vaccines are developed. According to previous studies, Maxing Shigan decoction (MXSGD) present a prominent antivirus effect and is often used to treat pulmonary diseases. Furthermore, we collected 115 open prescriptions for COVID-19 therapy from the National Health Commission, State Administration of TCM and other organizations, MXSGD was identified as the key formula. However, the underlying molecular mechanism of MXSGD against COVID-19 is still unknown. AIM OF THE STUDY: The present study aimed to evaluate the therapeutic mechanism of MXSGD against COVID-19 by network pharmacology and in vitro experiment verification, and screen the potential components which could bind to key targets of COVID-19 via molecular docking method. MATERIALS AND METHODS: Multiple open-source databases related to TCM or compounds were employed to screen active ingredients and potential targets of MXSGD. Network pharmacology analysis methods were used to initially predict the antivirus and anti-inflammatory effects of MXSGD against COVID-19. IL-6 induced rat lung epithelial type Ⅱ cells (RLE-6TN) damage was established to explore the anti-inflammatory damage activity of MXSGD. After MXSGD intervention, the expression level of related proteins and their phosphorylation in the IL-6 mediated JAK-STAT signaling pathway were detected by Western blot. Molecular docking technique was used to further identify the potential substances which could bind to three key targets (ACE2, Mpro and RdRp) of COVID-19. RESULTS: In this study, 105 active ingredients and 1025 candidate targets were selected for MXSGD, 83 overlapping targets related to MXSGD and COVID-19 were identified, and the protein-protein interaction (PPI) network of MXSGD against COVID-19 was constructed. According to the results of biological enrichment analysis, 63 significant KEGG pathways were enriched, and most of them were related to signal transduction, immune system and virus infection. Furthermore, according the relationship between signal pathways, we confirmed MXSGD could effectively inhibit IL-6 mediated JAK-STAT signal pathway related protein expression level, decreased the protein expression levels of p-JAK2, p-STAT3, Bax and Caspase 3, and increased the protein expression level of Bcl-2, thereby inhibiting RLE-6TN cells damage. In addition, according to the LibDock scores screening results, the components with strong potential affinity (Top 10) with ACE2, Mpro and RdRp are mainly from glycyrrhiza uralensis (Chinese name: Gancao) and semen armeniacae amarum (Chinese name: Kuxingren). Among them, amygdalin was selected as the optimal candidate component bind to all three key targets, and euchrenone, glycyrrhizin, and glycyrol also exhibited superior affinity interactions with ACE2, Mpro and RdRp, respectively. CONCLUSION: This work explained the positive characteristics of multi-component, multi-target, and multi-approach intervention with MXSGD in combating COVID-19, and preliminary revealed the antiviral and anti-inflammatory pharmacodynamic substances and mechanism of MXSGD, which might provide insights into the vital role of TCM in the prevention and treatment of COVID-19.

Integrated strategy of LC-MS and network pharmacology for predicting active constituents and pharmacological mechanisms of Ranunculus japonicus Thunb. for treating rheumatoid arthritis.

ETHNOPHARMACOLOGICAL RELEVANCE: Ranunculus japonicus Thunb. (short for R. japonicus) is a topically applied herb with the activities of removing jaundice, nebula and edema, preventing malaria, stopping asthma, promoting diuresis and relieving pain. It was firstly recorded in Zhouhou Beiji Fang and has been used for the treatment of malaria, ulcers, carbuncle, jaundice, migraine, stomachache, toothache and arthritis for over 1800 years. AIM OF THE STUDY: This study aimed to uncover the potentially effective components of R. japonicus and the pharmacological mechanisms against rheumatoid arthritis (RA) by combing LC-MS and network pharmacology. MATERIALS AND METHODS: Firstly, the chemical constituents of R. japonicus were qualitatively identified by UPLC-ESI-LTQ-Orbitrap MS. Then we performed target prediction by PharmMapper, protein-protein interaction (PPI) analysis via String, GO and KEGG pathway enrichment analysis by DAVID and constructed the compound-target-pathway network using Cytoscape. Thirdly, crucial compounds in the network were quantitatively analyzed to achieve quality control of R. japonicus. Finally, the pharmacological activities of R. japonicus and two potentially bioactive ingredients were validated in RA-FLSs (Rheumatoid Arthritis Fibroblast-like Synoviocytes) in vitro. RESULTS: Overall fifty-four ingredients of R. japonicus were identified and forty-five components were firstly discovered in R. japonicus. Among them, twenty-seven validated compounds were predicted to act on twenty-five RA-related targets and they might exhibit therapeutic effects against RA via positive regulation of cell migration, etc. Nine potentially bioactive components of R. japonicus which played important roles in the compound-target-pathway network were simultaneously quantified by an optimized UPLC-ESI-Triple Quad method. In vitro, compared to control group, R. japonicus extract, berberine and yangonin significantly inhibited the migration capacity of RA-FLSs after 24 h treatment. CONCLUSION: This study clarified that R. japonicus and the bioactive ingredients berberine and yangonin might exert therapeutic actions for RA via suppressing the aggressive phenotypes of RA-FLSs through combined LC-MS technology and network pharmacology tools for the first time. The present research provided deeper understanding into the chemical profiling, pharmacological activities and quality control of R. japonicus and offered reference for further scientific research and clinical use of R. japonicus in treating RA.

Synergistic Effect of Berberine-Based Chinese Medicine Assembled Nanostructures on Diarrhea-Predominant Irritable Bowel Syndrome In Vivo.

Diarrhea-predominant irritable bowel syndrome (IBS-D) is one common chronic functional disease of the digestive system with limited treatments. The microbiota-gut-brain axis (MGBA) has a central function in the pathogeny of IBS-D, which includes the participation of many various factors, such as brain-gut peptides (BGPs), immune inflammation, and intestinal flora. Inspired by the drug combination in traditional Chinese medicine (TCM), our previous study discovered that berberine (BBR) and baicalin (BA) could form natural self-assemblies as BA-BBR nanoparticles (BA-BBR NPs) and showed synergistic effects against IBS-D. Here, we investigated the synergistic effects of BA-BBR NPs on IBS-D model mice induced by chronic restraint stress plus Senna alexandrina Mill decoction with the influence on MGBA. BA-BBR NPs showed the best therapeutic effect on improving visceral hypersensitivity and diarrhea on IBS-D model mice, compared with BBR, BA, and BA/BBR mixture. Furthermore, BA-BBR NPs significantly (P<0.05) reduced the levels of 5-hydroxytryptamine (5-HT), vasoactive intestinal polypeptide (VIP) and choline acety transferase (CHAT) in colon tissues or of serum from BGPs; it lowered the expressions of the nuclear factor kappa-B (NF-κB) in colon tissues and changed the levels of basophil granulocyte (BASO) and leukomonocyte (LYMPH) in whole blood from immune inflammation; it altered the intestinal flora of Bacteroidia, Deferribacteres, Verrucomicrobia, Candidatus_Saccharibacteria, and Cyanobacteria from intestinal flora. In conclusion, BA-BBR NPs, after forming the natural self-assembly between BBR and BA, promoted the synergistic effect on IBS-D mice than the sum of BBR and BA effects, based to the formation of self-assemblies rather than the simple mixing. It further proved that synergistic effect of BA-BBR NPs on IBS-D mice might be related to BGPs, immune inflammation, and intestinal flora from three important interrelated components of MGBA. This study will provide a novel idea for the interpretation of TCM compatibility theory and provide the basis for BA-BBR NPs as a medicinal plant-derived natural and efficient nanomaterial for clinical use.

Correction: Cui, H., et al. BA-12 Inhibits Angiogenesis via Glutathione Metabolism Activation. Int. J. Mol. Sci. 2019, 20, 4062.

The authors wish to make the following corrections to this paper [...].

Celastrol inhibits ezrin-mediated migration of hepatocellular carcinoma cells.

Progression of hepatocellular carcinoma involves multiple genetic and epigenetic alterations that promote cancer invasion and metastasis. Our recent study revealed that hyperphosphorylation of ezrin promotes intrahepatic metastasis in vivo and cell migration in vitro. Celastrol is a natural product from traditional Chinese medicine which has been used in treating liver cancer. However, the mechanism of action underlying celastrol treatment was less clear. Here we show that ROCK2 is a novel target of celastrol and inhibition of ROCK2 suppresses elicited ezrin activation and liver cancer cell migration. Using cell monolayer wound healing, we carried out a phenotype-based screen of natural products and discovered the efficacy of celastrol in inhibiting cell migration. The molecular target of celastrol was identified as ROCK2 using celastrol affinity pull-down assay. Our molecular docking analyses indicated celastrol binds to the active site of ROCK2 kinase. Mechanistically, celastrol inhibits the ROCK2-mediated phosphorylation of ezrin at Thr567 which harnesses liver cancer cell migration. Our findings suggest that targeting ROCK2-ezrin signaling is a potential therapeutic niche for celastrol-based intervention of cancer progression in hepatocellular carcinoma.

Meta-Analysis of Paclitaxel-Based Chemotherapy Combined With Traditional Chinese Medicines for Gastric Cancer Treatment.

This study aimed to compare the efficacy and safety of traditional Chinese medicines (TCMs) combined with paclitaxel-based chemotherapy and paclitaxel-based chemotherapy alone for gastric cancer treatment. Literature searches (up to September 25, 2019) were performed using the Cochrane Library, EMBASE, PubMed, Chinese Science and Technology Journals (CQVIP), Wanfang, and China Academic Journals (CNKI) databases. Data from 14 randomized controlled trials (RCTs), with 1,109 participants, were included. The results indicated that, compared with paclitaxel-based chemotherapy alone, the combination of TCMs and paclitaxel-based chemotherapy significantly improved the tumor response rate (TRR; RR: 1.39; 95% CI: 1.24-1.57; p < 0.001, I 2 = 12%), increased the quality of life based on the Karnofsky Performance Scale score (RR: 1.53; 95% CI: 1.19-1.96; p < 0.001, I 2 = 0%), and reduced the side effects, such as neutropenia (RR: 0.68; 95% CI: 0.55-0.84; p < 0.001, I 2 = 44%), leukopenia (RR: 0.69; 95% CI: 0.54-0.90; p < 0.01, I 2 = 40%), thrombocytopenia (RR: 0.66; 95% CI: 0.46-0.96; p < 0.05, I 2 = 32%), and nausea and vomiting (RR: 0.50; 95% CI: 0.32-0.80; p < 0.01, I 2 = 85%). Hepatic dysfunction (RR: 0.63; 95% CI: 0.33-1.20; p = 0.16, I 2 = 0%), neurotoxicity (RR: 0.64; 95% CI: 0.26-1.55; p = 0.32, I 2 = 0%), and anemia (RR: 0.65; 95% CI: 0.40-1.04; p = 0.07, I 2 = 0%) were similar between the two groups. Evidence from the meta-analysis suggested that compared with paclitaxel-based chemotherapy alone, the combination of TCMs and paclitaxel-based chemotherapy may increase the TRR, improve quality of life, and reduce multiple chemotherapy-related side effects in gastric cancer patients. Additional rigorously designed large RCTs are required to confirm the efficacy and safety of this treatment.

Dysbiosis of gut microbiota is associated with gastric carcinogenesis in rats.

OBJECTIVES: Although many studies have examined changes in gut microbiota composition in gastric carcinogenesis to clarify the mechanism of action of anticancer drugs, it is unclear whether animal models of gastric carcinogenesis adequately reflect the disease in humans. METHODS: To address this issue, the present study investigated changes in the gut microbiome profile of a rat model of gastric carcinogenesis established using a combination of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), sodium salicylate, irregular fasting, and ranitidine. The rats were divided into control (Normal), chronic non-atrophic gastritis (CNAG), chronic atrophic gastritis (CAG), precancerous lesion of gastric cancer (PLGC), and gastric cancer (GC) groups according to histopathological features. Gut microbiome in gastric carcinogenesis profiling was performed by 16S rRNA gene sequencing of rat feces samples. RESULTS: We found that gut bacterial species richness increased whereas species diversity decreased during gastric carcinogenesis, with the most significant changes detected in the PLGC group. Gut microbiota community composition differed across groups, with the greatest similarities observed between CNAG and CAG groups and between PLGC and GC groups. There were significant differences in taxonomic representation at the phylum level: the PLGC group had the highest ratio of Firmicutes/Bacteroidetes whereas the GC group had the highest abundance of Proteobacteria and Actinobacteria. CONCLUSIONS: These results indicate that changes in the gut microbiome in a rat model of MNNG-induced gastric carcinogenesis are similar to those observed in humans, thus providing a useful tool for evaluating the efficacy and mechanism of action of novel monotherapies or drug combinations for the treatment of gastric carcinogenesis.

BA-12 Inhibits Angiogenesis via Glutathione Metabolism Activation.

There is a need for an efficient and low-cost leading compound discovery mode. However, drug development remains slow, expensive, and risky. Here, this manuscript proposes a leading compound discovery strategy based on a combination of traditional Chinese medicine (TCM) formulae and pharmacochemistry, using a ligustrazine-betulinic acid derivative (BA-12) in the treatment of angiogenesis as an example. Blocking angiogenesis to inhibit the growth and metastasis of solid tumors is currently one recognized therapy for cancer in the clinic. Firstly, based on a traditional Prunella vulgaris plaster, BA-12 was synthesized according to our previous study, as it exhibited better antitumor activities than other derivatives on human bladder carcinoma cells (T24); it was then uploaded for target prediction. Secondly, the efficacy and biotoxicity of BA-12 on angiogenesis were evaluated using human umbilical vein endothelial cells (HUVECs), a quail chick chorioallantoic membrane, and Caenorhabditis elegans. According to the prediction results, the main mechanisms of BA-12 were metabolic pathways. Thus, multiple metabolomics approaches were applied to reveal the mechanisms of BA-12. Finally, the predictive mechanisms of BA-12 on glutathione metabolism and glycerophospholipid metabolism activation were validated using targeted metabolomics and pharmacological assays. This strategy may provide a reference for highly efficient drug discovery, with the aim of sharing TCM wisdom for unmet clinical needs.

Identifying the Antiproliferative Effect of Astragalus Polysaccharides on Breast Cancer: Coupling Network Pharmacology With Targetable Screening From the Cancer Genome Atlas.

Background: Astragalus polysaccharides (APS), natural plant compounds, have recently emerged as a promising strategy for cancer treatment, but little is known concerning their effects on breast cancer (BC) tumorigenesis. Methods: We obtained breast cancer genetic data from The Cancer Genome Atlas (TCGA) database, network pharmacology to further clarify its biological properties. Survival analysis and molecular docking techniques were implemented for the final screening to obtain key target information. Our experiments focused on the detection of intervention effects of APS on BC cells (MCF-7 and MDA-MB-231), and quantitative RT-PCR (qRT-PCR) was used to assess the expression of key targets. Results: A total of 1,439 differentially expressed genes (DEGs) were identified by TCGA and used to build disease networks. Module analysis, gene ontology and pathway analysis revealed characteristic of the DEGs network. Topological properties were used to identify key targets, survival analysis and molecular docking finally found that the targets of APS regulation of BC cells may be CCNB1, CDC6, and p53. Through cell viability, migration and invasion assays, we found that APS interferes with the development of breast cancer in MCF7 and MDA-MB-231 cells in a dose-dependent manner. Furthermore, qRT-PCR verification suggested that the expression of CCNB1 and CDC6 in breast cancer cells was significantly downregulated in response to APS, while expression of the tumor suppressor gene P53 was significantly increased. Conclusion: Results of this study suggest therapeutic potential for APS in BC treatment, possibly through interventions with CCNB1, CDC6, and P53. Furthermore, these findings illustrate the feasibility of using network pharmacology to connect large-scale target data as a way to discover the mechanism of natural products interfering with disease.

Oleanolic Acid-amino Acids Derivatives: Design, Synthesis, and Hepatoprotective Evaluation In Vitro and In Vivo.

Activated hepatic stellate cells (HSCs) are the main extracellular matrix (ECM)-producing cells in the injured liver and the key mediators of liver fibrosis; they also promote the progression of hepatocellular carcinoma (HCC). In the acidic extracellular microenvironment of HCC, HSCs are activated to promote the migration of HCC cells. It is worth attempting to alter the weak acidic microenvironment to promote activated HSC apoptosis to treat liver fibrosis and liver cancer. In the present study, a series of novel OA-amino acids analogues were designed and synthesized to introduce different amino acids in the 3-hydroxyl of OA using the ester condensation reaction to enhance hydrophilicity, alkalinity, and biological activity. We found that OA-lysine derivative (3g) could improve the hydrophilic of OA and induce HSCs apoptosis via inducing MMP depolarization and increasing intracellular Ca2+ levels. Additionally, 3g displayed a better hepatoprotective effect than OA (20 mg/kg, intragastric administration) against the acute liver injury induced by carbon tetrachloride (CCl4) in mice. The results suggested that basic amino acids (lysine) could effectively enhance OA's hydrophilicity, alkalinity, and hepatoprotective activity in vitro and in vivo, which might be likely associated with increasing bioavailability and altering an extracellular weak acidic microenvironment with further verification. Therefore, the OA-lysine derivative (3g) has the potential to be developed as an agent with hepatoprotective activity.