Ziyuglycoside II attenuated OVX mice bone loss via inflammatory responses and regulation of gut microbiota and SCFAs.

BACKGROUND AND PURPOSE: Osteoporosis (OP) is a frequent clinical problem for the elderly. Traditional Chinese Medicine (TCM) has achieved beneficial results in the treatment of OP. Ziyuglycoside II (ZGS II) is a major active compound of Sanguisorba officinalis L. that has shown anti-inflammation and antioxidation properties, but little information concerning its anti-OP potential is available. Our research aims to investigate the mechanism of ZGS II in ameliorating bone loss by inflammatory responses and regulation of gut microbiota and short chain fatty acids (SCFAs) in ovariectomized (OVX) mice. METHODS: We predicted the mode of ZGS II action on OP through network pharmacology and molecular docking, and an OVX mouse model was employed to validate its anti-OP efficacy. Then we analyzed its impact on bone microstructure, the levels of inflammatory cytokines and pain mediators in serum, inflammation in colon, intestinal barrier, gut microbiota composition and SCFAs in feces. RESULTS: Network pharmacology identified 55 intersecting targets of ZGS II related to OP. Of these, we predicted IGF1 may be the core target, which was successfully docked with ZGS II and showed excellent binding ability. Our in vivo results showed that ZGS II alleviated bone loss in OVX mice, attenuated systemic inflammation, enhanced intestinal barrier, reduced the pain threshold, modulated the abundance of gut microbiota involving norank_f__Muribaculaceae and Dubosiella, and increased the content of acetic acid and propanoic acid in SCFAs. CONCLUSIONS: Our data indicated that ZGS II attenuated bone loss in OVX mice by relieving inflammation and regulating gut microbiota and SCFAs.

Decoding the mechanism of Eleutheroside E in treating osteoporosis via network pharmacological analysis and molecular docking of osteoclast-related genes and gut microbiota.

Objective: Eleutheroside E (EE) is an anti-inflammatory natural compound derived from the edible medicinal herb Acanthopanax senticosus. This study aims to investigate the underlying mechanism of the anti-osteoporosis action of EE through network pharmacology, molecular docking and gut microbiota. Materials and methods: Network pharmacology was used to explore the potential core targets and main pathways mediated by EE in osteoporosis (OP) treatment. Molecular docking was exploited to investigate the interactions between the active anti-OP compounds in EE and the potential downstream targets. Following the multi-approach bioinformatics analysis, ovariectomy (OVX) model was also established to investigate the in vivo anti-OP effects of EE. Results: The top 10 core targets in PPI network were TP53, AKT1, JUN, CTNNB1, STAT3, HIF1A, EP300, CREB1, IL1B and ESR1. Molecular docking results that the binding energy of target proteins and the active compounds was approximately between -5.0 and -7.0 kcal/mol, which EE has the lowest docking binding energy with HIF1A. Enrichment analysis of GO and KEGG pathways of target proteins indicated that EE treatment could potentially alter numerous biological processes and cellular pathways. In vivo experiments demonstrated the protective effect of EE treatment against accelerated bone loss, where reduced serum levels of TRAP, CTX, TNF-α, LPS, and IL-6 and increased bone volume and serum levels of P1NP were observed in EE-treated mice. In addition, changes in gut microbiota were spotted by 16S rRNA gene sequencing, showing that EE treatment increased the relative abundance of Lactobacillus and decreased the relative abundance of Clostridiaceae. Conclusion: In summary, these findings suggested that the characteristics of multi-target and multi-pathway of EE against OP. In vivo, EE prevents the onset of OP by regulating gut microbiota and inflammatory response and is therefore a potential OP drug.

Unraveling the mechanism of ethyl acetate extract from Prismatomeris connata Y. Z. Ruan root in treating pulmonary fibrosis: insights from bioinformatics, network pharmacology, and experimental validation.

Introduction: Pulmonary fibrosis is a terminal lung disease characterized by fibroblast proliferation, extracellular matrix accumulation, inflammatory damage, and tissue structure destruction. The pathogenesis of this disease, particularly idiopathic pulmonary fibrosis (IPF), remains unknown. Macrophages play major roles in organ fibrosis diseases, including pulmonary fibrosis. The phenotype and polarization of macrophages are closely associated with pulmonary fibrosis. A new direction in research on anti-pulmonary fibrosis is focused on developing drugs that maintain the stability of the pulmonary microenvironment. Methods: We obtained gene sequencing data and clinical information for patients with IPF from the GEO datasets GSE110147, GSE15197, GSE24988, GSE31934, GSE32537, GSE35145, GSE53845, GSE49072, GSE70864, and GSE90010. We performed GO, KEGG enrichment analysis and GSEA analysis, and conducted weighted gene co-expression network analysis. In addition, we performed proteomic analysis of mouse lung tissue. To verify the results of bioinformatics analysis and proteomic analysis, mice were induced by intratracheal instillation of bleomycin (BLM), and gavaged for 14 days after modeling. Respiratory function of mice in different groups was measured. Lung tissues were retained for histopathological examination, Western Blot and real-time quantitative PCR, etc. In addition, lipopolysaccharide, interferon-γ and interleukin-4 were used to induce RAW264.7 cells for 12h in vitro to establish macrophage inflammation and polarization model. At the same time, HG2 intervention was given. The phenotype transformation and cytokine secretion of macrophages were investigated by Western Blot, RT-qPCR and flow cytometry, etc. Results: Through bioinformatics analysis and experiments involving bleomycin-induced pulmonary fibrosis in mice, we confirmed the importance of macrophage polarization in IPF. The analysis revealed that macrophage polarization in IPF involves a change in the phenotypic spectrum. Furthermore, experiments demonstrated high expression of M2-type macrophage-associated biomarkers and inducible nitric oxide synthase, thus indicating an imbalance in M1/M2 polarization of pulmonary macrophages in mice with pulmonary fibrosis. Discussion: Our investigation revealed that the ethyl acetate extract (HG2) obtained from the roots of Prismatomeris connata Y. Z. Ruan exhibits therapeutic efficacy against bleomycin-induced pulmonary fibrosis. HG2 modulates macrophage polarization, alterations in the TGF-ß/Smad pathway, and downstream protein expression in the context of pulmonary fibrosis. On the basis of our findings, we believe that HG2 has potential as a novel traditional Chinese medicine component for treating pulmonary fibrosis.

Integrated spatially resolved metabolomics and network toxicology to investigate the hepatotoxicity mechanisms of component D of Polygonum multiflorum Thunb.

ETHNOPHARMACOLOGICAL RELEVANCE: The liver toxicity of Reynoutria multiflora (Thunb.) Moldenke. (Polygonaceae) (Polygonum multiflorum Thunb, PM) has always attracted much attention, but the related toxicity materials and mechanisms have not been elucidated due to multi-component and multi-target characteristics. In previous hepatotoxicity screening, different components of PM were first evaluated and the hepatotoxicity of component D [95% ethanol (EtOH) elution] in a 70% EtOH extract of PM (PM-D) showed the highest hepatotoxicity. Furthermore, the main components of PM-D were identified and their hepatotoxicity was evaluated based on a zebrafish embryo model. However, the hepatotoxicity mechanism of PM-D is unknown. AIM OF THE STUDY: This work is to explore the hepatotoxicity mechanisms of PM-D by integrating network toxicology and spatially resolved metabolomics strategy. MATERIALS AND METHODS: A hepatotoxicity interaction network of PM-D was constructed based on toxicity target prediction for eight key toxic ingredients and a hepatotoxicity target collection. Then the key signaling pathways were enriched, and molecular docking verification was implemented to evaluate the ability of toxic ingredients to bind to the core targets. The pathological changes of liver tissues and serum biochemical assays of mice were used to evaluate the liver injury effect of mice with oral administration of PM-D. Furthermore, spatially resolved metabolomics was used to visualize significant differences in metabolic profiles in mice after drug administration, to screen hepatotoxicity-related biomarkers and analyze metabolic pathways. RESULTS: The contents of four key toxic compounds in PM-D were detected. Network toxicology identified 30 potential targets of liver toxicity of PM-D. GO and KEGG enrichment analyses indicated that the hepatotoxicity of PM-D involved multiple biological activities, including cellular response to endogenous stimulus, organonitrogen compound metabolic process, regulation of the apoptotic process, regulation of kinase, regulation of reactive oxygen species metabolic process and signaling pathways including PI3K-Akt, AMPK, MAPK, mTOR, Ras and HIF-1. The molecular docking confirmed the high binding activity of 8 key toxic ingredients with 10 core targets, including mTOR, PIK3CA, AKT1, and EGFR. The high distribution of metabolites of PM-D in the liver of administrated mice was recognized by mass spectrometry imaging. Spatially resolved metabolomics results revealed significant changes in metabolic profiles after PM-D administration, and metabolites such as taurine, taurocholic acid, adenosine, and acyl-carnitines were associated with PM-D-induced liver injury. Enrichment analyses of metabolic pathways revealed tht linolenic acid and linoleic acid metabolism, carnitine synthesis, oxidation of branched-chain fatty acids, and six other metabolic pathways were significantly changed. Comprehensive analysis revealed that the hepatotoxicity caused by PM-D was closely related to cholestasis, mitochondrial damage, oxidative stress and energy metabolism, and lipid metabolism disorders. CONCLUSIONS: In this study, the hepatotoxicity mechanisms of PM-D were comprehensively identified through an integrated spatially resolved metabolomics and network toxicology strategy, providing a theoretical foundation for the toxicity mechanisms of PM and its safe clinical application.

Molecularly imprinted polymers combined with membrane-protected solid-phase extraction to detect triazines in tea samples.

Spherical molecularly imprinted polymers (MIPs) were prepared by emulsion polymerization. The isothermal adsorption and selective adsorption indicated that the MIPs obtained exhibit excellent specific recognition for the template (atrazine) and its analogues. The MIPs were encapsulated in a polypropylene microporous membrane to fabricate MIP adsorption packages for the direct extraction of triazines in uncentrifuged and unfiltered tea extracts. The extraction conditions affecting the extraction efficiency, including the type and volume of extraction solvent, the number of MIP adsorption packages, the surface area of the MIP adsorption packages, the mass of MIPs in the MIP adsorption packages, the extraction time, the eluting solvent, and the eluting volume, were optimized. Under the optimal extraction and high-performance liquid chromatography-tandem mass spectrometry conditions, the method exhibited excellent linearity in the range from 0.5 to 250 ng g-1, with R2 ≥ 0.9992. The detection limit of the method was 0.09-0.18 ng g-1. The intraday and interday relative standard deviations ranged from 3.1% to 7.5% and from 3.1% to 7.9%, respectively. The method was successfully used to detect triazines in five tea samples. At a spiking concentration of 2 ng g-1, satisfactory recoveries ranging from (81 ± 3)% to (104 ± 7)% were obtained. The membrane-protected solid-phase extraction method based on molecularly imprinted material is expected to be widely used to enrich triazines in complex samples. Graphical Abstract Schematic illustration of the MIPs combined with membrane-protected solid-phase extraction of triazines in tea sample.