Contamination of the traditional medicine Radix Dipsaci with aflatoxin B1 impairs hippocampal neurogenesis and cognitive function in a mouse model of osteoporosis.

BACKGROUND: Aflatoxin B1, which can penetrate the blood-brain barrier and kill neural cells, can contaminate traditional herbal medicines, posing a significant risk to human health. The present study examined cellular, cognitive and behavioral consequences of aflatoxin B1 contamination of the anti-osteoporotic medicine Radix Dipsaci. METHODS: A mouse model of osteoporosis was created by treating the animals with all-trans-retinoic acid. Then the animals were treated intragastically with water decoctions of Radix Dipsaci that contained detectable aflatoxin B1 or not. The animals were compared in terms of mineral density and mineral salt content of bone, production of pro-inflammatory factors, neurogenesis and microglial activation in hippocampus, as well as behavior and cognitive function. RESULTS: Contamination of Radix Dipsaci with aflatoxin B1 significantly reduced the medicine's content of bioactive saponins. It destroyed the ability of the herbal decoction to improve mineral density and mineral salt content in the bones of diseased mice, and it induced the production of the oxidative stress marker malondialdehyde as well as the pro-inflammatory cytokines interleukin-1ß and tumor necrosis factor-α. Aflatoxin B1 contamination inhibited formation of new neurons and increased the proportion of activated microglia in the hippocampus. These neurological changes were associated with anhedonia, behavioral despair, and deficits in short-term memory and social memory. CONCLUSION: Contamination of Radix Dipsaci with aflatoxin B1 not only eliminates the herbal decoction's anti-osteoporotic effects, but it also induces neurotoxicity that can lead to cognitive decline and behavioral abnormalities. Such contamination should be avoided through tightly regulated production and quality control of medicinal herbs.

An exploratory metabolomic study reveals the Dipsacus asper-Achyranthes bidentate herb pair against osteoarthritis by modulating imbalance in polyunsaturated fatty acids and energy metabolism.

Osteoarthritis (OA) is a degenerative joint disease primarily affecting the cartilage. The therapeutic potential of the Dipsacus asper-Achyranthes bidentate herb pair for OA has been acknowledged, yet its precise mechanism remains elusive. In this study, we conducted a comprehensive analysis of metabolomic changes and therapeutic outcomes in osteoarthritic rats, employing a gas chromatography-mass spectrometry-based metabolomics approach in conjunction with histopathological and biochemical assessments. The rats were divided into six groups: control, model, positive control, Dipsacus asper treated, Achyranthes bidentata treated, and herb pair treated groups. Compared to the model group, significant reductions in levels of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), interleukin-6 (IL-6), and iNOS were observed in the treated groups. Multivariate statistical analyses were employed to investigate metabolite profile changes in serum samples and identify potential biomarkers, revealing 45 differential biomarkers, with eighteen validated using standard substances. These analytes exhibited excellent linearity across a wide concentration range (R2>0.9990), with intra- and inter-day precision RSD values below 4.69% and 4.83%, respectively. Recoveries of the eighteen analytes ranged from 93.97% to 106.59%, with RSD values under 5.72%, underscoring the method's reliability. Treatment with the herbal pair effectively restored levels of unsaturated fatty acids such as linoleic acid and arachidonic acid, along with glucogenic amino acids. Additionally, levels of phosphoric acid and citric acid were reversed, indicating restoration of energy metabolism. Collectively, these findings highlight the utility of metabolomic analysis in evaluating therapeutic efficacy and elucidating the underlying molecular mechanisms of herb pairs in OA treatment.

Dipsacus Asperoides-Derived Exosomes-Like Nanoparticles Inhibit the Progression of Osteosarcoma via Activating P38/JNK Signaling Pathway.

Introduction: Osteosarcoma is a prevalent and highly malignant primary bone tumor. However, current clinical therapeutic drugs for osteosarcoma are not suitable for long-term use due to significant side effects. Therefore, there is an urgent need to develop new drugs with fewer side effects. Dipsacus asperoides C. Y. Cheng et T. M. Ai, a traditional Chinese medicine, is commonly used for its anti-inflammatory, anti-pain, bone fracture healing, and anti-tumor effects. In this study, we investigated the effects of exosome-like nanoparticles derived from Dipsacus asperoides (DAELNs) on osteosarcoma cells in vitro and in vivo. Methods: DAELNs were isolated and purified from Dipsacus asperoides and their physical and chemical properties were characterized using transmission electron microscopy (TEM) and nanoparticle tracking analysis (NTA). The cellular uptake of DAELNs in osteosarcoma cells was analyzed by PKH26 staining. The proliferation, invasion, migration, and apoptosis of osteosarcoma cells were assessed using CCK8 assay, EdU assay, colony-formation assay, transwell assay, wound healing assay, and mitochondrial membrane potential measurement, respectively. The regulatory mechanism of DAELNs inhibiting the progression of osteosarcoma via activating P38/JNK signaling pathway was investigated using Western blotting and immunohistochemistry. Moreover, the therapeutic effects of DAELNs were evaluated using in vivo small animal imaging assay, HE staining, and immunohistochemistry. Results: Our results showed that DAELNs inhibited the proliferation, invasion, migration, and fostered the apoptosis of osteosarcoma cells in vitro and suppressed the tumor growth of osteosarcoma cells in a xenograft nude mouse model. Furthermore, the bio-distribution of DiD-labeled DAELNs showed preferential targeting of osteosarcoma tumors and excellent biosafety in histological analysis of the liver and kidney. Mechanistically, DAELNs activated the P38/JNK signaling pathway-induced apoptosis. Conclusion: Taken together, DAELNs are novel, natural, and osteosarcoma-targeted agents that can serve as safe and effective therapeutic approaches for the treatment of osteosarcoma.

Four new triterpene saponins from Cephalaria speciosa and their potent cytotoxic and immunomodulatory activities.

Four new triterpene saponins, namely speciosides A-D (1-4) along with six known saponins were isolated from the n-butanol extract of Cephalaria speciosa. In addition to these, three new prosapogenins (2a-4a) were obtained after alkaline hydrolysis. Elucidation of the structures of the isolated compounds was carried out by 1D, 2D NMR, HR-ESI/MS and GC-MS analyses. Cytotoxic activity was investigated on A549, CCD34-Lu, MDA-MB-231, PC-3, U-87MG, HeLa, HepG-2 cells by MTT method. Additionally, the immunomodulatory effect of compounds was evaluated for macrophage polarization with/without inactivated IBV D274 antigen treatment on THP-1 cells originated macrophage cells in terms of M1 or M2. According to the cytotoxicity results, compound 1 and prosapogenin 2a exhibit significant cytotoxicity than doxorubicin by comparison. The results demonstrated that saponin molecules treated THP-1 originated macrophages were induced M1 and/or M2 polarization. Additionally, macrophage cells treated with/without IBV D274 antigen contained saponin compounds were triggered significantly M2 polarization relative to M1. Notably, monodesmosidic saponins (1 and 2a-4a) in comparison with bisdesmosidic ones (2-4) demonstrated the most effect on M2 polarization. In conclusion, the results showed that all the isolated new saponins and their prosapogenins have immunomodulatory potential on macrophage cells increasing immune response without significant cytotoxic effect on THP-1 originated macrophages.

Oleanolic Acid Glycosides from Scabiosa caucasica and Scabiosa ochroleuca: Structural Analysis and Cytotoxicity.

In the field of research on medicinal plants from the Armenian flora, the phytochemical study of two Scabiosa L. species, S. caucasica M. Bieb. and S. ochroleuca L. (Caprifoliaceae), has led to the isolation of five previously undescribed oleanolic acid glycosides from an aqueous-ethanolic extract of the roots: 3-O-α-L-rhamnopyranosyl-(1→3)-ß-D-glucopyranosyl-(1→4)-ß-D-glucopyranosyl-(1→4)-ß-D-xylopyranosyl-(1→3)-α-L-rhamnopyranosyl-(1→2)-α-L-arabinopyranosyloleanolic acid 28-O-ß-D-glucopyranosyl-(1→6)-ß-D-glucopyranosyl ester, 3-O-ß-D-xylopyranosyl-(1→2)-[α-L-rhamnopyranosyl-(1→4)]-ß-D-glucopyranosyl-(1→4)-ß-D-glucopyranosyl-(1→4)-ß-D-xylopyranosyl-(1→3)-α-L-rhamnopyranosyl-(1→2)-α-L-arabinopyranosyloleanolic acid 28-O-ß-D-glucopyranosyl-(1→6)-ß-D-glucopyranosyl ester, 3-O-ß-D-xylopyranosyl-(1→2)-[α-L-rhamnopyranosyl-(1→4)]-ß-D-glucopyranosyl-(1→4)-ß-D-glucopyranosyl-(1→4)-ß-D-xylopyranosyl-(1→3)-α-L-rhamnopyranosyl-(1→2)-α-L-arabinopyranosyloleanolic acid, 3-O-ß-D-xylopyranosyl-(1→2)-[α-L-rhamnopyranosyl-(1→4)]-ß-D-xylopyranosyl-(1→4)-ß-D-glucopyranosyl-(1→4)-ß-D-xylopyranosyl-(1→3)-α-L-rhamnopyranosyl-(1→2)-α-L-arabinopyranosyloleanolic acid 28-O-ß-D-glucopyranosyl-(1→6)-ß-D-glucopyranosyl ester, 3-O-α-L-rhamnopyranosyl-(1→4)-ß-D-glucopyranosyl-(1→4)-ß-D-glucopyranosyl-(1→4)-ß-D-xylopyranosyl-(1→3)-α-L-rhamnopyranosyl-(1→2)-α-L-arabinopyranosyloleanolic acid 28-O-ß-D-glucopyranosyl-(1→6)-ß-D-glucopyranosyl ester. Their full structural elucidation required extensive 1D and 2D NMR experiments, as well as mass spectrometry analysis. For the biological activity of the bidesmosidic saponins and the monodesmosidic saponin, their cytotoxicity on a mouse colon cancer cell line (MC-38) was evaluated.

Dipsacus and Scabiosa Species-The Source of Specialized Metabolites with High Biological Relevance: A Review.

The genera Dipsacus L. and Scabiosa L. of the Caprifoliaceae family are widely distributed in Europe, Asia, and Africa. This work reviews the available literature on the phytochemical profiles, ethnomedicinal uses, and biological activities of the most popular species. These plants are rich sources of many valuable specialized metabolites with beneficial medicinal properties, such as triterpenoid derivatives, iridoids, phenolic acids, and flavonoids. They are also sources of essential oils. The genus Dipsacus has been used for centuries in Chinese and Korean folk medicines to treat bone (osteoporosis) and joint problems (rheumatic arthritis). The Korean Herbal Pharmacopoeia and Chinese Pharmacopoeia include Dipsaci radix, the dried roots of D. asperoides C.Y.Cheng & T.M.Ai. In addition, S. comosa Fisch. ex Roem & Schult. and S. tschiliiensis Grunning are used in traditional Mongolian medicine to treat liver diseases. The current scientific literature data indicate that these plants and their constituents have various biological properties, including inter alia antiarthritic, anti-neurodegenerative, anti-inflammatory, antioxidant, anticancer, and antimicrobial activities; they have also been found to strengthen tendon and bone tissue and protect the liver, heart, and kidney. The essential oils possess antibacterial, antifungal, and insecticidal properties. This paper reviews the key biological values of Dipsacus and Scabiosa species, as identified by in vitro and in vivo studies, and presents their potential pharmacological applications.

Exploring active ingredients of anti-osteoarthritis in raw and wine-processed Dipsaci Radix based on spectrum-effect relationship combined with chemometrics.

ETHNOPHARMACOLOGICAL RELEVANCE: Dipsaci Radix (DR) is the dry root of the Dipsacus asper Wall. ex DC., which has the function of tonifying the liver and kidney, continuing tendons and bones, and regulating blood vessels. However, there are few reports on the main active ingredients. AIM OF THE STUDY: This study aimed to find the main active components of DR in the treatment of osteoarthritis (OA) by spectrum-effect relationship and compare the differences between RDR and WDR. MATERIALS AND METHODS: Firstly, the high-performance liquid chromatography (HPLC) method was used to establish the fingerprint of DR, and 10 peaks of them were determined by UPLC-Q-TOF/MS. Then, the OA rat model was established by injecting sodium iodoacetate to study the effect of DR on OA. The spectrum-effect relationship was analyzed by grey relational analysis (GRA) and Pearson correlation analysis. RESULTS: According to the pharmacological results, compared with the model group, the cartilage score, tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and interleukin-6 (IL-6), and Mankin score of rats in low, medium and high dose groups were decreased, and the therapeutic effect of wine-processed DR tended to be better than raw DR at the same dose. Finally, the active components of DR were preliminarily determined as 4 (loganic acid), 6 (chlorogenic acid), 8 (caffeic acid), 14 (dipsanoside B), 16, and 17 (asperosaponin VI) which had a large correlation in GRA and Pearson correlation analysis. CONCLUSION: This study established the spectrum-effect relationship between the raw and wine-processed DR for the first time, which provided a theoretical basis for the study of the pharmacodynamic substance basis of DR before and after processing. This research provided a reference for the subsequent study of DR.

In vitro anticancer, antioxidant and enzyme inhibitory potentials of endemic Cephalaria elazigensis var. purpurea with in silico studies.

In this study, the therapeutic potential and phytochemical composition of ethanolic extract of Cephalaria elazigensis var. purpurea (CE), an endemic species, were investigated. For this purpose, the antiproliferative effect of CE on the MCF-7 human breast cancer cell line was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, and its effectiveness on colony formation and cell migration was analyzed with clonogenic assay and wound healing assay, respectively. In addition, the cell death detection ELISA (CDDE) assay was conducted to determine the pro-apoptotic capacity of CE. The IC50 value of the CE was determined as 324.2 ± 14.7 µg/mL. Furthermore, upon 1000 µg/mL CE treatment, there was 4.96-fold increase in the population of cells undergoing apoptosis compared to the untreated control cells. The antioxidant activity tests were performed by DPPH free radical, ABTS cation radical, ferric-ion reducing power (FRAP) and ferrous-ion chelating power (FCAP) assays. Antioxidant activity values for the DPPH, ABTS and FRAP assays were found to be 125.6 ± 6.3, 34.09 ± 0.1 and 123.4 ± 4.2 µmol TE/mg DE, respectively. We further determined the effect of CE ethanolic extract against acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) enzymes. CE plays an effective inhibitory role in AChE and BuChE (AChE: IC50: 10.54 µg/mL, BuChE: IC50: 6.84 µg/mL) respectively. Further, molecular docking stuy was conducted to understand the nature of the all compound against AChE an BChE. It is revealed that α-Linolenic acid shows lowest binding energy (-7.90 kcal/mol) towards AChE, on the other side, Linoleic acid shows good binding affinity (-7.40 kcal/mol) for BChE.Communicated by Ramaswamy H. Sarma.

Extraction of bioactive compounds from Dipsacus fullonum leaves using deep eutectic solvents.

In this study, deep eutectic solvent (DES) based systems were evaluated for selective extraction and optimized for increased recovery of chlorogenic acid derivatives, flavone glycosides and iridoid glycosides from Dipsacus fullonum L. leaves. Bioactives from Dipsacus plants has shown great antioxidant and antimicrobial activities as well as effectiveness against several cancer strains and a source for anti-Borrelia compounds. Twelve different hydrophilic and hydrophobic DESs were tested to find the best solvent composition. Choline chloride and betaine were used as hydrogen bond acceptors (HBA) for the preparation of hydrophilic DESs and menthol for hydrophobic DESs. The tested hydrogen bond donors (HBD) were various organic acids and glycerol. The composition of most effective DES was optimized using the Box-Behnken design for each of the three main group of analytes from D. fullonum L. to evaluate possible selectivity and highest recovery. HPLC-DAD-MS was used to identify and quantify the main bioactive compounds extracted from plant material. The optimal extraction for highest overall recovery was achieved using a molar ratio of choline chloride and lactic acid of 1:2.4 with 35% water and 27 mL of the solvent per one gram of dry material. The optimized DES extract gave concentrations 1.8 to 2.2 times higher than traditional organic solvent extracts depending on the group of analytes.

[To explore mechanism of Scabiosa comosa against liver fibrosis based on pharmacodynamics and network pharmacology].

This study aims to systematically elucidate the pharmacodynamics and network pharmacological mechanism of Mongolian medicinal plants Scabiosa comosa, explore their key targets and related pathways, and further clarify the mechanism of the plants in treating liver fibrosis. Wistar rats were assigned into the blank group, carbon tetrachloride-induced liver fibrosis model group, and low-, medium-, and high-dose S. comosa groups. HE staining and Masson staining were performed for the observation of liver tissue under a microscope. Further, Wistar rats were assigned into a control group and a S. comosa group for administration. Seven days later, blood was collected from the abdominal aorta, and different doses of drug-containing serum samples were used to treat hepatic stellate cell-T6(HSC-T6). Flow cytometry was adopted to detect the apoptosis of HSC-T6 cells. Ultra-high performance liquid chromatography-time of flight-mass spectrometry(UHPLC-TOF-MS) was employed to determine the components in Scabiosa comosa. The target of S. comosa and liver fibrosis were obtained from SwissTargetPrediction and GeneCards, respectively, and the common targets were selected as the anti-liver fibrosis targets. Protein-protein interaction was analyzed via STRING. Gene Ontology(GO) annotation and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment were carried out via Metascape. Phosphatidylinosital 3-kinase(PI3 K), protein kinase B(AKT), p-AKT, p38, and p-p38 targets which are involved in the top-ranked PI3 K/AKT and mitogen activated kinase-like protein(MAPK) signaling pathways were selected for validation via Western blot. The HE and Masson staining results showed that Scabiosa alleviated the hyperplasia of connective tissue and the fibrosis. The serum containing Scabiosa significantly promoted the apoptosis of HSC-T6 in a concentration-dependent manner. A total of 76 chemical components were identified by UHPLC-TOF-MS, among which flavonoids, alkaloids, terpenoids, phenols, and fatty acids were the main components. According to the prediction, there were 63 anti-liver fibrosis targets in Scabiosa comosa, the annotated GO terms of which involved biological processes, cell components, and molecular functions. The KEGG pathway enrichment showed that the targets were mainly involved in PI3 K/AKT, epidermal growth factor receptor(EGFR), RAS-associated protein 1(Rap1), hypoxia-inducible factor 1(HIF-1), resistance to audiogenic seizures(Ras), and MAPK signaling pathways. Western blot results showed that compared with the model group, S. comosa down-regulated the protein levels of α-smooth muscle actin(α-SMA), collagen Ⅰ, PI3 K, AKT, p-AKT, p38, and p-p38 in liver tissue. Compared with the control group, the low-, medium-, and high-dose S. comosa significantly down-regulated the protein levels of α-SMA, collagen Ⅰ, PI3 K, AKT, p-AKT, p38, and p-p38 in HSC-T6. The evidence of pharmacodynamics, network pharmacology, and molecular biology indicated that the plants of S. comosa had significant activity against liver fibrosis, the mechanism of which may involve the regulation of the key targets PI3 K, AKT, and MAPK14 p38 in the PI3 K/AKT and MAPK signaling pathways.

A hederagenin-type triterpene saponin, sumbulianoside a from Cephalaria sumbuliana and its potent immunomodulatory activity against seasonal flu virus H3N2.

A new hederagenin-type triterpene saponin; hederagenin 28-O-ß-D-galactopyranosyl-(1→6)-ß-D-glucopyranosyl ester named sumbulianoside A (1), together with twelve known saponins were isolated from the n-butanol extract of Cephalaria sumbuliana (Caprifoliaceae) from which, one known saponin, dipsacus saponin A (2) was isolated, for the first time from Cephalaria species. The structures of the isolated compounds were elucidated by 1 D and 2 D NMR and HRESIMS analyses. Cytotoxic activities were investigated on A549, Hela, PANC1, SHSY5Y cells and non-cancerous cell HEK293 by MTT method and immunomodulatory activities were evaluated against activated H3N2 seasonal virus in whole blood by measuring IL-4, IFN-γ, IL-1ß cytokine level with ELISA. According to the cytotoxicity results, compounds 1 and 2 did not possess significant cytotoxicity, while only compound 2 induced significant IL-4 production (** p<.001) against H3N2 showing a possible Th2 response and antibody production. All compounds had no effect on cytokine release (p > 0.5).

A Methanol Extract of Scabiosa atropurpurea Enhances Doxorubicin Cytotoxicity against Resistant Colorectal Cancer Cells In Vitro.

Colorectal cancer is a malignancy with a high incidence. Currently, the drugs used in chemotherapy are often accompanied by strong side effects. Natural secondary metabolites can interfere with chemotherapeutic drugs and intensify their cytotoxic effects. This study aimed to profile the secondary metabolites from the methanol extract of Scabiosa atropurpurea and investigate their in vitro activities, alone or in combination with the chemotherapeutic agent doxorubicin. MTT assay was used to determine the cytotoxic activities. Annexin-V/PI double-staining analysis was employed to evaluate the apoptotic concentration. Multicaspase assay, quantitative reverse transcription real-time PCR (RT-qPCR), and ABC transporter activities were also performed. LC-MS analysis revealed 31 compounds including phenolic acids, flavonoids, and saponins. S. atropurpurea extract intensified doxorubicin anti-proliferative effects against resistant tumor cells and enhanced the cytotoxic effects towards Caco-2 cells after 48 h. The mRNA expression levels of Bax, caspase-3, and p21 were increased significantly whereas Bcl-2 expression level was decreased. Furthermore, the methanol extract reversed P-glycoprotein or multidrug resistance-associated protein in Caco-2 cells. In conclusion, S. atropurpurea improved chemosensitivity and modulated multidrug resistance in Caco-2 cells which makes it a good candidate for further research in order to develop a new potential cancer treatment.

Cell Growth Inhibition of Saponin XII from Dipsacus japonicus Miq. on Acute Myeloid Leukemia Cells.

In previous studies, we isolated the known compound saponin XII from the roots of Dipsacus japonicus Miq. Here, we show that this compound reduced the number of acute myeloid leukemia OCI-AML3 cells as evaluated by a hemocytometer. Flow cytometry analyses demonstrated that the reported activity was associated with a significant increase of apoptosis and of cells in the G0/G1 phase of the cell cycle, with a decrease of cells in the S and G2/M phases. Thus, the inhibition of cell growth in OCI-AML3 cells was due to antiproliferative and pro-apoptotic effects. Interestingly, the bioactivity of saponin XII exerted its effect at a concentration as low as 1 µg/mL.

The effect of luteolin 7-glucoside, apigenin 7-glucoside and Succisa pratensis extracts on NF-κB activation and α-amylase activity in HepG2 cells.

The chemical composition of Succisa pratensis is not well known. The existing data indicate a substantial content of flavonoids, which include luteolin and apigenin 7-glucosides. The aim of this study was to elaborate the isolation protocol of these flavonoids from flowers and leaves of S. pratensis, to carry out their characterization, as well as evaluate the effect of S. pratensis extracts on activation of transcription factor NF-κB and α-amylase activity. The extraction protocol applied in this study allowed isolation and characterization of flavonoid fraction of S. pratensis. Their identity was confirmed by NMR spectra analysis, UV spectroscopy and electrospray ionization-tandem MS evaluation. Treatment of pancreatic α-amylase with S. pratensis extract inhibited this enzyme's activity to an extent comparable to that of isolated luteolin and apigenin 7-glucosides. Incubation of HepG2 cells for 24 h with S. pratensis extracts or isolated flavonoids resulted in moderate reduction in NF-κB transcription factor activation evaluated in terms of translocation of its active subunits from cytosol into nucleus and subsequently diminished expression of the COX-2 gene. Expression of NF-κB was also reduced. The most significantly diminished NF-κB activation and expression, as well as COX-2 expression, was found to result from treatment with isolated flavonoids and ethyl acetate extract of S. pratensis leaves. These results indicate that S. pratensis flavonoids may modulate the metabolic and signaling pathways whose deregulation is related to pathogenesis of liver cancer. Further studies are required to confirm these observations and assess the chemopreventive and/or therapeutic potential of the S. pratensis herb.

Systems pharmacology dissection of action mechanisms of Dipsaci Radix for osteoporosis.

AIMS: Osteoporosis (OP) is a systemic metabolic bone disease characterized by bone mass decrease and microstructural degradation, which may increase the risk of bone fracture and leading to high morbidity. Dipsaci Radix (DR), one typical traditional Chinese medicine (TCM), which has been applied in the treatment of OP with good therapeutic effects and few side effects. However, the underlying molecular mechanisms of DR to treat OP have not been fully elucidated. In this study, we aim to dissect the molecular mechanism of DR in the treatment of OP. MATERIALS AND METHODS: A systems pharmacology approach was employed to comprehensively dissect the action mechanisms of DR for the treatment of OP. KEY FINDINGS: 10 compounds were screened out as the potential active ingredients with excellent biological activity based on in silico ADME (absorption, distribution, metabolism and excretion) prediction model. Then, 36 key protein targets of 6 compounds were identified by systems drug targeting model (SysDT) and they were involved in several biological processes, such as osteoclast differentiation, osteoblast differentiation and anti-inflammation. The target-pathway network indicated that targets are mainly mapped in multiple signaling pathways, i.e., MAPK, Tumor necrosis factor α (TNF-α), NF-κb and Toll-like receptor pathways. The in vitro results indicated that the compounds ursolic acid and beta-sitosterol effectively inhibited the osteoclast differentiation. SIGNIFICANCE: These results systematically dissected that DR exhibits the therapeutic effects of OP by the regulation of immune system-related pathways, which provide novel perspective to drug development of OP.

Protective effects and network analysis of natural compounds obtained from Radix dipsaci, Eucommiae cortex, and Rhizoma drynariae against RANKL-induced osteoclastogenesis in vitro.

ETHNOPHARMACOLOGICAL RELEVANCE: Osteoporosis is one of the most common bone diseases; it is characterized by bone loss and is a risk factor for hip fracture. Chinese herbal medicines (CHMs) and their related natural compounds have been used for treating many diseases, including bone diseases, since ancient times in China and are regarded as a cost-effective complementary therapy. AIM OF THE STUDY: The goal of this study was to investigate the osteoprotective mechanisms of these three Chinese herbs and their related natural compounds. The effects of CHMs and related natural compounds on RANKL-induced osteoclastogenesis in vitro were investigated. MATERIALS AND METHODS: A network pharmacology method was applied to study CHM-related natural compounds and their osteoporosis targets. In addition, their effect on RANKL-induced osteoclastogenesis in RAW264.7 cells was also investigated in vitro. RESULTS: Radix dipsaci, Eucommiae cortex, and Rhizoma drynariae exhibited protective effects against mortality in hip fracture patients. Furthermore, these three herbs inhibited RANKL-induced TRAP activities and reduced the expression of bone resorption-related genes in RAW264.7 cells. Network analysis of natural compound (ingredient)-target interactions identified 11 natural compounds. Signal pathway analyses suggested that these compounds may target cytokine-cytokine receptor interactions, including RANKL-induced osteoclastogenesis. Five novel natural compounds exhibited reduced RANKL-induced TRAP activities and bone resorption-related gene expression. CONCLUSION: The clinically used CHMs, Radix dipsaci, Eucommiae cortex, and Rhizoma drynariae, and natural compounds obtained from them may suppress RANKL-induced osteoclastogenesis in vitro.

Absolute structure assignment of an iridoid-monoterpenoid indole alkaloid hybrid from Dipsacus asper.

Iridoid-monoterpenoid indole alkaloid hybrids (IMIAHs) represent a rare class of natural products reported from only several plants of Rubiaceae and Dipsacaceae families, while their structural assignments remain a very challenging work due to complexity and flexibility. In the current study, a new IMIAH (1) was isolated from the roots of Dipsacus asper and its structure with absolute configuration was unambiguously established by a combination of spectroscopic analyses, chemical degradation and ECD calculation. A new oleanane-type triterpenoid saponin (2) and 15 known co-metabolites were also obtained and structurally characterized. Our biological evaluations showed that compound 2 exhibited moderate inhibition against acetylcholine esterase (AChE) with an IC50 value of 15.8 ±â€¯0.56 µM, and compound 15 displayed potent cytotoxicity selectively against human A549 and H157 lung cancer cells with IC50 values of 6.94 ±â€¯0.24 and 9.06 ±â€¯0.12 µM, respectively.

Protective effect of a polysaccharide from Dipsacus asper Wall on streptozotocin (STZ)-induced diabetic nephropathy in rat.

The effects of a polysaccharide (DAP) from the roots of Dipsacus asper on renal complication in streptozotocin (STZ)-induced diabetic rats were investigated in the present study. Administration of DAP (100 and 300 mg/kg) for 4 weeks significantly decreased the fasting blood glucose (FBG) and glycosylated hemoglobin (HbA1c) levels, but increase the body weight of STZ-induced diabetic rats. Increased kidney weight index, serum creatinine (Scr), blood urea nitrogen (BUN), urine protein, and urinary albumin excretion (UAE) levels along with decreased creatinine clearance (Ccr) were nearly reversed to the normal levels by DAP. Moreover, DAP was able to normalize hyperlipidemia, improve oxidative stress, and down-regulate the formation of advanced glycation end products-receptor for advanced glycation end products (AGE-RAGE). In conclusion, our work demonstrates the preventive effect of DAP was achieved via antihyperglycemic, hypolipidemic, and antioxidant activity along with inhibition of AGE accumulation and RAGE expression in STZ-treated diabetic rats.

Protective effects of Dipsacus asper polysaccharide on osteoporosis in vivo by regulating RANKL/RANK/OPG/VEGF and PI3K/Akt/eNOS pathway.

A homogenous polysaccharide (DAP), with a molecular weight of 2.61 × 104 Da, was isolated from the roots of Dipsacus asper Wall. Gas chromatography (GC) indicated that DAP was composed of galactose and mannose with a molar ratio of 1:1. The purpose of this study was to evaluate the effect of DAP on the progress of bone loss in the ovariectomized (OVX) rat model of osteoporosis. Administration of DAP (50 and 200 mg/kg/body wt. day) for 12 weeks significantly prevented OVX-induced bone loss, biomechanical reduction, the body weight gain, the loss of the uterus weight, as well as increased U-Ca/Cr, U-P/Cr, ALP, TRAP, OC and DPD/Cr levels in rats, which was further supported by the histopathological examinations. Furthermore, we found that the mechanism by which DAP elicited anti-osteoporotic effects was mediated by up-regulation of VEGF and OPG, but down-regulation of RANK and RANKL in both protein and mRNA expression in OVX rats, as well as the activation of PI3K/Akt/eNOS signaling pathway, indicating that DAP can be clinically used as a potential alternative medicine for the prevention and treatment of postmenopausal osteoporosis.

Aqueous extract of Dipsacus asperoides suppresses lipopolysaccharide-stimulated inflammatory responses by inhibiting the ERK1/2 signaling pathway in RAW 264.7 macrophages.

ETHNOPHARMACOLOGICAL RELEVANCE: Dipsaci Radix, which is the dried root of Dipsacus asperoides C. Y. Cheng and T. M. Ai (Dipsacaceae), is used to treat back pain and blood stasis syndrome in Korean traditional medicine. AIM OF THE STUDY: To understand the mechanisms responsible for the pharmacological activities of D. asperoides, we investigated the inhibitory effect of D. asperoides on lipopolysaccharide (LPS)-induced inflammation in mouse macrophages RAW 264.7 cells. MATERIALS AND METHODS: Aqueous extract of D. asperoides (AEDA) was prepared by boiling D. asperoides in water and then administered to LPS treated RAW 264.7 cells. Cell viabilities were measured using an MTT assay, and protein levels were determined by western blotting. The ROS scavenging activity of AEDA was measured using a DCFH-DA assay and levels of nitric oxide (NO) were determined using a NO assay. The nuclear translocations of NF-κB and Nrf2 were investigated immunocytochemically, and pro-inflammatory cytokines in supernatant were evaluated by ELISA. RESULTS: Treatment with AEDA suppressed the expressions of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in LPS-stimulated RAW 264.7 macrophages. AEDA also reduced ROS, pro-inflammatory cytokine (IL-6 and IL-1ß) levels, and iNOS-derived NO and COX-2-derived prostaglandin E2 release to medium, and suppressed the phosphorylation and degradation of IκB and the activation of NF-κB in macrophages. Furthermore, treatment with AEDA inhibited the ERK1/2 pathway but not the JNK or p38 MAPK pathways. In addition, AEDA significantly promoted Nrf2 translocation from cytoplasm to nucleus and up-regulated the expression of HO-1. CONCLUSION: These results suggest that AEDA has anti-inflammatory and anti-oxidative effects through the inhibition of NF-κB and ERK1/2 and the activation of Nrf2/HO-1 in macrophages.