Data mining-guided alleviation of hyperuricemia by Paeonia veitchii Lynch through inhibition of xanthine oxidase and regulation of renal urate transporters.

BACKGROUND: Hyperuricemia (HUA) is a metabolic disease characterized by a high level of uric acid (UA). The extensive historical application of traditional Chinese medicine (TCM) offers a range of herbs and prescriptions used for the treatment of HUA-related disorders. However, the core herbs in the prescriptions and their mechanisms have not been sufficiently explained. PURPOSE: Our current investigation aimed to estimate the anti-HUA effect and mechanisms of Paeonia veitchii Lynch, an herb with high use frequency identified from data mining of TCM prescriptions. METHODS: Prescriptions for HUA/gout treatment were statistically analyzed through a data mining approach to determine the common nature and use frequency of their composition herbs. The chemical constituents of Paeonia veitchii extract (PVE) were analyzed by UPLC-QTOF-MS/MS, while its UA-lowering effect was further evaluated in adenosine-induced liver cells and potassium oxonate (PO) and hypoxanthine (HX)-induced HUA mice. RESULTS: A total of 225 prescriptions involving 246 herbs were sorted out. The properties, flavors and meridians of the appearing herbs were mainly cold, bitter and liver, respectively, while their efficacy was primarily concentrated on clearing heat and dispelling wind. Further usage frequency analysis yielded the top 20 most commonly used herbs, in which PVE presented significant inhibitory activity (IC50 = 131.33 µg/ml) against xanthine oxidase (XOD), and its constituents showed strong binding with XOD in a molecular docking study and further were experimentally validated through XOD enzymatic inhibition and surface plasmon resonance (SPR). PVE (50 to 200 µg/ml) dose-dependently decreased UA levels by inhibiting XOD expression and activity in BRL 3A liver cells. In HUA mice, oral administration of PVE exhibited a significant UA-lowering effect, which was attributed to the reduction of UA production by inhibiting XOD activity and expression, as well as the enhancement of UA excretion by regulating renal urate transporters (URAT1, GLUT9, OAT1 and ABCG2). Noticeably, all doses of PVE treatment did not cause any liver injury, and displayed a renal protective effect. CONCLUSIONS: Our results first comprehensively clarified the therapeutic effect and mechanisms of PVE against HUA through suppressing UA production and promoting UA excretion with hepatic and renal protection, suggesting that PVE could be a promising UA-lowering candidate with a desirable safety profile for the treatment of HUA and prevention of gout.

Glytabastan B, a coumestan isolated from Glycine tabacina, alleviated synovial inflammation, osteoclastogenesis and collagen-induced arthritis through inhibiting MAPK and PI3K/AKT pathways.

The roots of Glycine tabacina are used to treat rheumatoid arthritis (RA) and joint infection in folk medicine. Glytabastan B (GlyB), a newly reported coumestan isolated from this species, was found to significantly attenuate IL-1ß-induced inflammation in SW982 human synovial cells at 3 and 6 µM, as evidenced by the decreased levels of pro-inflammatory mediators and matrix metalloproteinases (MMPs). GlyB also suppressed RANKL-induced osteoclastogenesis, decreased the expression of osteoclastogenic markers (NFATc1, CTSK, MMP-9) and osteoclast-mediated bone resorption. Further, GlyB administration (12.5 and 25 mg/kg) significantly inhibited inflammation, osteoclast formation and disease progression in collagen-induced arthritis (CIA) mice. Integration of network pharmacology, quantitative phosphoproteomic and experimental pharmacology results revealed that these beneficial actions were closely associated with the blockade of GlyB on the activation of MAPK, PI3K/AKT and their downstream signals including NF-κB and GSK3ß/NFATc1. Drug affinity responsive target stability (DARTS) assay, cellular thermal shift (CETSA) assay and molecular docking analysis confirmed that there were direct interactions between GlyB and its target proteins ERK2, JNK1 and class Ⅰ PI3K catalytic subunit p110 (α, ß, δ and γ), which significantly contributed to the inhibition of activation of MAPK and PI3K/AKT pathways. In conclusion, these results strongly suggest GlyB is a promising multiple-target candidate for the development of agents for the prevention and treatment of RA.

Investigation of the interaction between Chrysoeriol and xanthine oxidase using computational and in vitro approaches.

Xanthine oxidase (XO) plays a vital role in inducing hyperuricemia and increasing the level of superoxide free radicals in blood, and is proved as an important target for gout. Chrysoeriol (CHE) is a natural flavone with potent XO inhibitory activity (IC50 = 2.487 ± 0.213 µM), however, the mechanism of interaction is still unclear. Therefore, a comprehensive analysis of the interaction between CHE and XO was accomplished by enzyme kinetics, isothermal titration calorimetry (ITC), multi-spectroscopic methods, molecular simulation and ADMET. The results showed that CHE acted as a rapid reversible and competitive-type XO inhibitor and its binding to XO was driven by hydrogen bonding and hydrophobic interaction. Moreover, CHE exhibited a strong fluorescence quenching effect through a static quenching procedure and induced conformational changes of XO. Its binding pattern with XO was revealed by docking study and the binding affinity to XO was enhanced by the interactions with key amino acid residues in the active pocket of XO. Further, CHE showed good stability and pharmacokinetic behavior properties in molecule dynamic simulation and ADMET prediction. Overall, this study shed some light on the mechanism of interaction between CHE and XO, also provided some valuable information concerning the future therapeutic application of CHE as natural XO inhibitor.

Naturally occurring coumestans from plants, their biological activities and therapeutic effects on human diseases.

Naturally occurring coumestans are known as a collection of plant-derived polycyclic aromatic secondary metabolites which are characterized by the presence of an oxygen heterocyclic four-ring system comprising a coumarin moiety and a benzofuran moiety sharing a CË­C bond. Recently, there is an increasing attention in excavating the medicinal potential of coumestans, particularly coumestrol, wedelolactone, psoralidin and glycyrol, in a variety of diseases. This review is a comprehensive inventory of the chemical structures of coumestans isolated from various plant sources during the period of 1956-2020, together with their reported biological activities. 120 molecules were collected and further classified as coumestans containing core skeleton, dimethylpyranocoumestans, furanocoumestans, O-glycosylated coumestans and others, which showed a wide range of pharmacological activities including estrogenic, anti-cancer, anti-inflammatory, anti-osteoporotic, organ protective, neuroprotective, anti-diabetic and anti-obesity, antimicrobial, immunosuppressive, antioxidant and skin-protective activities. Furthermore, this review focuses on the counteraction of coumestans against bone diseases and organ damages, and the involved molecular mechanisms, which could provide important information to better understand the medicinal values of these compounds. This review is intended to be instructive for the rational design and development of less toxic and more effective drugs with a coumestan scaffold.

Low-Dose Berberine Attenuates the Anti-Breast Cancer Activity of Chemotherapeutic Agents via Induction of Autophagy and Antioxidation.

Berberine (BBR), a major active component of Rhizoma coptidis, is one of the most promising agents for breast cancer adjuvant therapy. It is well accepted that BBR could exhibit remarkable anticancer efficacy with few side effects, and when treated with chemotherapeutic agents in combination, BBR could enhance the chemosensitivity of cancer cells. Our previous study reported that low-dose BBR (LDB) induced hormetic effect and attenuated the anticancer activity of chemotherapeutic agents. However, the underlying mechanisms are still unclear. In this study, we confirmed that LDB could promote cancer cell proliferation and antagonize the anti-breast cancer activities of chemotherapeutic agents. And the mechanisms were proved to be induction of autophagy and antioxidation by LDB. Our results showed that LDB could mildly induce reactive oxygen species, raise the level of autophagy by promoting the phosphorylation of adenosine monophosphate-activated protein kinase, and promote antioxidant enzymes expression through activating nuclear factor erythroid 2-related factor 2 in breast cancer cells. These findings revealed a potential negative impact of BBR on its adjuvant anti-breast cancer therapy, providing guidance for a safe and effective use of naturally originated medicines in the clinic.

Isolation and Identification of Antiarthritic Constituents from Glycine tabacina and Network Pharmacology-Based Prediction of Their Protective Mechanisms against Rheumatoid Arthritis.

Glycine tabacina (Labill.) Benth is an edible medicinal herb for rheumatoid arthritis (RA) treatment in folk medicine. Current phytochemical research on this dried herb led to the isolation of eight new coumestans, named glytabastan A-H (1-8), and twenty-three known compounds 9-31. Their structures were elucidated using spectroscopic methods. The antiarthritic activities of all isolates were evaluated, and the results showed that coumestans 1-6 and 8-10 could inhibit arthritic inflammation in vitro, while coumestans 1, 2, 9, and 10 significantly blocked the osteoclastogenesis induced by receptor activator of nuclear factor (NF) κB ligand (RANKL). Moreover, network pharmacological analysis revealed that the anti-RA effect of G. tabacina involved multitargets, multipathways such as PI3K/Akt and MAPK signaling pathways, and various biological processes such as inflammatory response and cytokine-mediated signaling pathways. These results suggested that this species and its novel coumestans could serve as potential antiarthritic agents for functional food or medicinal use.

Dolichosin A, a coumestan isolated from Glycine tabacina, inhibits IL-1ß-induced inflammation in SW982 human synovial cells and suppresses RANKL-induced osteoclastogenesis: From network pharmacology to experimental pharmacology.

ETHNOPHARMACOLOGICAL RELEVANCE: Glycine tabacina (Labill.) Benth has been used as a traditional Chinese herbal medicine for the treatment of rheumatoid arthritis (RA) and joint infection. It is also one of the sources of the renowned native herbal medicine 'I-Tiao-Gung' in Taiwan. AIM OF THE STUDY: This study aimed to investigate anti-arthritic effects and underlying mechanisms of dolichosin A (DoA), a coumestan compound isolated from G. tabacina, by the integration of network pharmacology and experimental pharmacology. MATERIALS AND METHODS: Putative therapeutic targets and potential pharmacological mechanisms of DoA for RA treatment were predicted by network pharmacology approach. The regulated network of DoA acting on RA was constructed using Cytoscape 3.7.1. Anti-arthritic effects of DoA and predicted mechanisms were further validated using IL-1ß-induced SW982 human synovial cell model and RANKL-induced osteoclastogenesis model. RESULTS: A regulatory network of DoA-targets-pathways-RA was successfully constructed using network pharmacology approach. In this network, 65 candidate targets of DoA related to its therapeutic effect on RA were identified and the functional enrichment analysis revealed that these candidate targets were significantly involved in 12 central signaling pathways such as PI3K/AKT pathway, MAPK pathway and osteoclast differentiation. Furthermore, we found that DoA could significantly inhibit IL-1ß-induced inflammation in SW982 human synovial cells, as evidenced by the decreased levels of pro-inflammatory mediators (TNF-α, IL-6 and COX-2) and MMP-3. DoA also suppressed RANKL-induced osteoclastogenesis in vitro, as evidenced by decreased number of TRAP-positive multinucleated osteoclasts and reduced TRAP activity. Further experimental mechanism evidence confirmed the predicted results of network pharmacology that the blockade of PI3K/AKT and MAPK pathways activation was closely associated with these regulated processes of DoA. CONCLUSIONS: Our results demonstrated that DoA exhibited strong anti-arthritic activity through suppressing PI3K/AKT and MAPK pathways activation in activated synovial cells and osteoclasts, suggesting its potential as a hopeful candidate for the development of novel agents for the prevention and treatment of RA.

A new isoflavone with anti-inflammatory effect from the seeds of Millettia pachycarpa.

A new isoflavone, milletenol A (1), along with four known flavonoids (2-5) were isolated from the seeds of Millettia pachycarpa. The structure of 1 was established by extensive spectroscopic methods while known compounds were identified by comparisons with literature data. Compound 1 and 2 showed significant anti-inflammatory activities against nitric oxide production in LPS-induced RAW264.7 macrophages. The state of CuSO4-stimulated inflammation was effectively alleviated by compound 1 in zebrafish. However, no significant cytotoxicity against human breast cancer cells was observed among all isolates.

Flavonoids from Rhynchosia minima root exerts anti-inflammatory activity in lipopolysaccharide-stimulated RAW 264.7 cells via MAPK/NF-κB signaling pathway.

Rhynchosia minima root, a folk herbal medicine in southern China, is used to relieve itch and swelling. In this study, we examined the anti-inflammatory property of an ethanol fraction (EEF6) from R. minima root on lipopolysaccharide (LPS)-induced RAW 264.7 cells, as well as its underlying mechanism. The compound composition of EEF6 was determined by high-performance liquid chromatography-mass spectrometry. The result showed that five flavonoids compounds, 2',4',5,7-tetrahydroxyisoflavone, genistein-8-C-glucopyranoside, tricin, genistein, and daidzein, were identified in EEF6. In addition, EEF6 exhibited potent anti-inflammatory ability against LPS-stimulated RAW 264.7 cells via MAPK/NF-κB signaling pathways by decreasing the secretion of nitric oxide (NO), interleukin (IL)-6, TNF-α, and monocyte chemotactic protein (MCP)-1, inhibiting the translocation of p65 from cytoplasm to nucleus, and suppressing the phosphorylation of ERK, JNK, and p38. These results indicated that EEF6 could be a promising ingredient for inflammation management.

Synergistic anti-breast cancer effect of pulsatilla saponin D and camptothecin through interrupting autophagic-lysosomal function and promoting p62-mediated ubiquitinated protein aggregation.

Autophagy is an evolutionarily conserved mechanism to protect the cells from unfavorable environmental conditions. Inhibition of autophagy has been contemplated as a novel strategy to enhance anticancer efficacy of existing chemotherapeutic agents. We previously reported that pulsatilla saponin D (PSD) was a potent autophagy inhibitor. However, its anticancer potential as adjuvant and underlying mechanisms are still unknown. In this study, we identified that PSD induced the formation of autophagosome in MCF-7 and MDA-MB-231 breast cancer cells. However, PSD alone and particularly co-treatment with camptothecin remarkably increased p62 protein levels, indicating that PSD strongly inhibited the autophagic cargo degradation. The mechanistic study indicated that PSD profoundly abolished the co-localization of EGFP-LC3 and lysosomal-specific probe LysoTracker Red, suggesting that the autophagosome-lysosome fusion was blocked by PSD, which is similar to the action of chloroquine. In addition, PSD significantly increased lysosomal pH and inhibited the activation of lysosomal cathepsins in both breast cancer cell lines. Furthermore, the accrued p62 resulted in accumulation of ubiquitinated proteins owing to the interaction with p62 and delivery to the malfunctioned autophagosome by PSD. Finally, we demonstrated that PSD synergistically enhanced the anticancer activity of camptothecin (CPT) in cultured breast cancer cells and in mouse xenograft tumor models. Our results indicated that PSD inhibited autophagic flux via blocking autophagosome-lysosome fusion and lysosomal acidification, which may confer a synergistic anti-breast cancer activity of PSD and CPT.

Highly selective carbamate-based butyrylcholinesterase inhibitors derived from a naturally occurring pyranoisoflavone.

This current study described the design and synthesis of a series of derivatives based on a natural pyranoisaflavone, which was obtained from the seeds of Millettia pachycarpa and displayed attractive BChE inhibition and high selectivity in our previous study. The inhibitory potential of all derivatives against two cholinesterases was evaluated. Only a few compounds demonstrated AChE inhibitory activity at the tested concentrations, while 26 compounds showed significant inhibition on BChE (the IC50 values varied from 9.34 µM to 0.093 µM), most of them presented promising selectivity to ward BChE. Prediction of ADME properties for 7 most active compounds was performed. Among them, 9g (IC50 = 222 nM) and 9h (IC50 = 93 nM) were found to be the most potent BChE inhibitors with excellent selectivity over AChE (SI ratio = 1339 and 836, respectively). The kinetic analysis demonstrated both of them acted as mixed-type BChE inhibitors, while the molecular docking results indicated that they interacted with both residues in the catalytic active site. A cytotoxicity test on PC12 cells showed that both 9g and 9h had a therapeutic safety range similar to tacrine. Overall, the results indicate that 9h could be a good candidate of BChE inhibitors.

In Vitro and In Vivo Anti-Inflammatory Effects of Polyphyllin VII through Downregulating MAPK and NF-κB Pathways.

Background: Polyphyllin VII (PP7), a steroidal saponin from Paris polyphylla, has been found to exert strong anticancer activity. Little is known about the anti-inflammatory property of PP7. In this study, the anti-inflammatory activity and its underlying mechanisms of PP7 were evaluated in lipopolysaccharide (LPS)-stimulated RAW264.7 cells and in multiple animal models. Methods: The content of nitric oxide (NO) was determined by spectrophotometry. The levels of prostaglandin E2 (PGE2) and cytokines were measured by enzyme-linked immunosorbent assay (ELISA) assay. The mRNA expression of pro-inflammatory genes was determined by qPCR. The total and phosphorylated protein levels were examined by Western blotting. The in vivo anti-inflammatory activities were evaluated by using mouse and zebrafish models. Results: PP7 reduced the production of NO and PGE2 and the protein and mRNA expressions of pro-inflammatory cytokines (TNF-α, IL-1ß, and IL-6) and enzymes (inducible NO synthase [iNOS], cyclooxygenase-2 [COX-2], and Matrix metalloproteinase-9 [MMP-9]) in LPS-induced RAW264.7 cells by suppressing the NF-κB and MAPKs pathways. Notably, PP7 markedly inhibited xylene-induced ear edema and cotton pellet-induced granuloma formation in mice and suppressed LPS and CuSO4-induced inflammation and toxicity in zebrafish embryos. Conclusion: This study demonstrates that PP7 exerts strong anti-inflammatory activities in multiple in vitro and in vivo models and suggests that PP7 is a potential novel therapeutic agent for inflammatory diseases.

Glycine tabacina ethanol extract ameliorates collagen-induced arthritis in rats via inhibiting pro-inflammatory cytokines and oxidation.

ETHNOPHARMACOLOGICAL RELEVANCE: The whole plant of Glycine tabacina (Labill.) Benth has been used as a traditional herbal medicine to treat rheumatism, ostealgia and nephritis in China. It is also one of the sources of the renowned native herbal medicine 'I-Tiao-Gung' in Taiwan. AIM OF THE STUDY: This study aimed to investigate the anti-arthritic effect of ethanol extract of G. tabacina (GTE) in a collagen-induced arthritis (CIA) rat model. MATERIALS AND METHODS: The chemical profile of GTE was analyzed by HPLC-UV. The CIA was induced in male Wistar rats by intradermal injection of bovine type II collagen at tail root, back and ankle joints. The rats were orally administrated daily with GTE (1.11, 2.22 and 4.44 g dry weight of herb powder per kg body weight) from day 0 and continued for 30 days. Swelling volume and thickness of paw, arthritis index, X-radiographs and histopathological changes were examined to assess the severity of arthritis. Furthermore, the levels of pro-inflammatory cytokines, such as interleukin1ß (IL-1ß), IL-6 and tumor necrosis factor α (TNF-α), total superoxide dismutase (T-SOD) activity and malonaldehyde (MDA) level were measured to preliminarily explore the possible mechanisms. RESULTS: Oral administration of GTE significantly ameliorated the arthritic symptoms in CIA rat model, as indicated by the effects on paws swelling and arthritis index. X-radiographic analysis and histopathological examinations demonstrated that GTE effectively protected the bone and cartilage of joints from erosion, lesion and deformation. The efficacy of GTE treatment on CIA was comparable to that of indomethacin (positive drug). Besides, the overproduction of IL-1ß, IL-6 and TNF-α was remarkably inhibited in the serum of all GTE treatment groups. The restoration of serum T-SOD activity and MDA level proved that GTE administration alleviated the oxidative stress in CIA rats. CONCLUSIONS: GTE exhibited strong anti-CIA activity through inhibiting pro-inflammatory cytokines and oxidation in rats, suggesting its potential preventive and therapeutic effects on rheumatoid arthritis (RA).

Two new stilbenoids from Bletilla striata.

Two new stilbenoids bletilol D (1) and bletilol E (2), together with five known compounds were isolated from Bletilla striata. Three of them (3, 4, and 7) were obtained from this genus for the first time. Their structures were elucidated by spectroscopic methods and comparing with data reported in literatures. The cytotoxic activities of compounds 1-7 against MCF-7 (human breast cancer) and A549 (human lung carcinoma) cell lines were evaluated by MTT assay. Compound 2 showed weak cytotoxicity against MCF-7 and A549 cell lines with IC50 values of 36.32 ± 1.17 and 36.48 ± 1.12 µM, respectively, and 5 exhibited weak cytotoxicity against MCF-7 cell line with IC50 value of 57.09 ± 2.03 µM.

Polysaccharide PRM3 from Rhynchosia minima root enhances immune function through TLR4-NF-κB pathway.

BACKGROUND: Polysaccharides, one of the active ingredients in herbal medicine, are proved to enhance innate immunity against infections. The aim of this study is to explore the immunoregulatory ability of polysaccharides from Rhynchosia minima root in vitro and in vivo. METHODS: Polysaccharide fractions of R. minima root were obtained by chromatographic column. The content of NO was measured by spectrophotometry. The levels of cytokines (tumor necrosis factor-α, TNF-α; interleukin-6, IL-6; and monocyte chemoattractant protein-1, MCP-1) were determined by enzyme-linked immuno-sorbent assay (ELISA) kits. The translocation of p65 into the nucleus was imaged by confocal microscopy. The mRNA expression of TNF-α, IL-6, and MCP-1 was determined by quantitative real-time PCR. T-lymphocyte subgroups of spleen from immunosuppressive mouse were evaluated by flow cytometry. RESULTS: PRM3 remarkably enhanced the phagocytic ability of macrophages and promoted the release of NO and the secretion of cytokines (TNF-α, IL-6, and MCP-1) from macrophages. Simultaneously, PRM3 potently activated NF-κB signaling pathway via Toll-like receptor 4 (TLR4). In addition, PRM3 obviously increased the levels of serum cytokines, markedly up-regulated the percentages of CD3+ and CD4+ T lymphocytes and the CD4+/CD8+ ratio of splenocytes, and effectively attenuated cyclophosphamide induced immunosuppression in mice. CONCLUSIONS: PRM3 profoundly enhanced the immune function in vitro and in vivo through TLR4-NF-κB pathway and is a promising candidate of immunopotentiator which could be applied in functional foods or drugs. GENERAL SIGNIFICANCE: This study reported a polysaccharide PRM3 from R. minima root exhibited potent immunoenhancing activity and significantly alleviated cyclophosphamide-induced immunosuppression through TLR4-NF-κB pathway.

A new sesquiterpene glucoside from Lysionotus pauciflorus.

A new acorane sesquiterpene glucoside, 1R,3S,4R,5R,10R-3,10-dihydroxyacoronene-3-O-beta-D-gluc-oside (1), was isolated from the EtOAc-soluble partition of the ethanol extract of Lysionotus pauciflorus, together with six known compounds, namely p-hydroxybenzoic acid (2), vanillic acid (3). caffeic acid (4). beta-hydroxypropiosyringone (5), alpha,beta-dihydroxypropiosyringone (6), and lyoniresinol (7). The structure of the new compound was elucidated on the basis of spectroscopic methods, including 1D and 2D NMR, and high-resolution MS analysis. The absolute configuration of 1 was determined from CD spectra. When evaluated against several bacterial and fungal strains, and human cancer cell lines, compound 1 and its aglycone were inactive.