ABSTRACT
RATIONALE: Isopsoralen (ISO), a quality control marker (Q-marker) in Psoraleae Fructus, is proven to present an obvious anti-osteoporosis effect. Until now, the metabolism and anti-osteoporosis mechanisms of ISO have not been fully elucidated, greatly restricting its drug development. METHODS: The metabolites of ISO in rats were profiled by using ultrahigh-performance liquid chromatography coupled with time-of-flight mass spectrometry. The potential anti-osteoporosis mechanism of ISO in vivo was predicted by using network pharmacology. RESULTS: A total of 15 metabolites were characterized in rats after ingestion of ISO (20 mg/kg/day, by gavage), including 2 in plasma, 12 in urine, 6 in feces, 1 in heart, 3 in liver, 1 in spleen, 1 in lung, 3 in kidney, and 2 in brain. The pharmacology network results showed that ISO and its metabolites could regulate AKT1, SRC, NFKB1, EGFR, MAPK3, etc., involved in the prolactin signaling pathway, ErbB signaling pathway, thyroid hormone pathway, and PI3K-Akt signaling pathway. CONCLUSIONS: This is the first time for revealing the in vivo metabolism features and potential anti-osteoporosis mechanism of ISO by metabolite profiling and network pharmacology, providing data for further verification of pharmacological mechanism.
Subject(s)
Furocoumarins , Network Pharmacology , Psoralea , Rats, Sprague-Dawley , Animals , Furocoumarins/pharmacology , Furocoumarins/chemistry , Psoralea/chemistry , Rats , Chromatography, High Pressure Liquid/methods , Male , Osteoporosis/drug therapy , Osteoporosis/metabolism , Quality Control , Biomarkers/analysis , Biomarkers/metabolism , Biomarkers/urine , Fruit/chemistry , Drugs, Chinese Herbal/pharmacology , Drugs, Chinese Herbal/administration & dosage , Mass Spectrometry/methods , Bone Density Conservation Agents/pharmacology , Metabolome/drug effects , Metabolomics/methodsABSTRACT
BACKGROUND: Cirrhosis leads to portal hypertension (PHT), affecting survival with limited treatment options. This study investigated Imperatorin (IMP), a furanocoumarin with anti-inflammatory and hypotensive properties, for its therapeutic role and mechanisms in cirrhotic PHT. METHODS: Hepatic stellate cells (HSCs) inhibition by IMP was evaluated using LX-2 cell line. Rat cirrhosis was induced via CCl4 for 16 weeks. Experimental group were orally administered IMP (15/25 mg/kg/day) for 4 weeks. We subsequently examined portal pressure (PP), cirrhosis, inflammation, angiogenesis, and vascular remodeling. Network pharmacology was employed for mechanistic insights. RESULTS: IMP significantly inhibited the fibrogenesis in HSCs and suppressed cell viability. CCl4 exposure induced cirrhosis, inflammation, angiogenesis, vascular remodeling and PHT. IMP significantly reduced PP from 22.85 ± 3.88 mmHg to 6.67 ± 0.6 mmHg, diminished collagen deposition and pro-fibrotic factor expression, alleviated inflammation, and improved liver function. Vessel wall thickness in superior mesenteric arteries was restored, and intra-/extrahepatic angiogenesis was inhibited via VEGF and vWF. Furthermore, IMP induced sinusoidal vasodilation by upregulating eNOS and GCH1. Enrichment analysis indicated that IMP was involved in various biological processes associated with cirrhosis, such as the regulation of blood pressure, tissue remodeling, response to inflammation, and regulation of angiogenesis, etc. Additionally, IMP suppressed hepatic expression of TGF-ß both in vitro and in vivo, which was further supported by KEGG analysis. CONCLUSION: Our research demonstrated that IMP significantly mitigated cirrhosis PHT by reducing hepatic fibrosis and inflammation, curbing angiogenesis and vascular remodeling, and promoting vasodilation. This protective mechanism appears to be facilitated through the downregulation of TGF-ß.
Subject(s)
Carbon Tetrachloride , Furocoumarins , Hepatic Stellate Cells , Hypertension, Portal , Liver Cirrhosis , Vascular Remodeling , Furocoumarins/pharmacology , Furocoumarins/therapeutic use , Animals , Hypertension, Portal/drug therapy , Hypertension, Portal/physiopathology , Vascular Remodeling/drug effects , Hepatic Stellate Cells/drug effects , Hepatic Stellate Cells/metabolism , Hepatic Stellate Cells/pathology , Male , Rats , Liver Cirrhosis/drug therapy , Liver Cirrhosis/pathology , Liver Cirrhosis/metabolism , Liver Cirrhosis/chemically induced , Humans , Rats, Sprague-Dawley , Cell Line , Signal Transduction/drug effects , Network Pharmacology , Neovascularization, Pathologic/drug therapy , Liver/drug effects , Liver/pathology , Liver/metabolism , Liver/blood supplyABSTRACT
Dabrafenib is a BRAF inhibitor that has been demonstrated to be efficacious in the treatment of melanoma and non-small-cell lung cancer patients with BRAF V600E mutations. The objective of this study was to investigate the effects of 51 traditional Chinese medicines on the metabolism of dabrafenib and to further investigate the inhibitory effect of imperatorin. The quantification of dabrafenib and its metabolite hydroxy-dabrafenib was carried out using a sensitive, rapid, and accurate assay method based on ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). The results of in vitro experiments showed that 20 drugs inhibited the metabolism of dabrafenib by more than 80 %. In a further study of imperatorin on dabrafenib, the half-maximal inhibitory concentration (IC50) values of imperatorin on dabrafenib were 0.22 µM and 3.68 µM in rat liver microsomes (RLM) and human liver microsomes (HLM), respectively, while the inhibition mechanisms were non-competitive and mixed type inhibition, respectively. The results of in vivo experiments demonstrated that in the presence of imperatorin, the AUC(0-t), AUC(0-∞), Cmax, and Tmax of dabrafenib were increased by 2.38-, 2.26-, 1.05-, and 6.10-fold, respectively, while CLz/F was decreased by 67.9 %. In addition, Tmax of hydroxy-dabrafenib was increased by 1.4-fold. The results of the research showed that imperatorin had a consistent inhibitory effect on dabrafenib in vitro and in vivo. When the concurrent use of dabrafenib and imperatorin is unavoidable, clinicians should closely monitor for potential adverse events and make timely adjustments to the administered dosage.
Subject(s)
Furocoumarins , Imidazoles , Microsomes, Liver , Oximes , Rats, Sprague-Dawley , Oximes/pharmacology , Imidazoles/pharmacology , Imidazoles/metabolism , Animals , Furocoumarins/pharmacology , Furocoumarins/metabolism , Microsomes, Liver/metabolism , Humans , Rats , Male , Tandem Mass Spectrometry , Chromatography, High Pressure LiquidABSTRACT
The Order Lepidoptera contains nearly 160,000 described species and most of them are specialist herbivores that use restricted plant species as hosts. Speciation that originated from host shift is one of the important factors for the diversification of Lepidoptera. Because plants prepare secondary metabolites for defense against herbivores, with varying profiles of the components among different plant taxa, the specialist herbivores need to be adapted to the toxic substances unique to their host plants. Swallowtail butterflies of the genus Papilio consist of over 200 species. Approximately 80% of them utilize Rutaceae plants, and among the remaining species, a specific subgroup uses phylogenetically distant Apiaceae plants as larval hosts. Rutaceae and Apiaceae commonly contain toxic secondary metabolites, furanocoumarins, and molecular phylogenetic studies support the concept that Apiaceae feeders were derived from Rutaceae feeders. Molecular mechanisms underlying furanocoumarin tolerance in Papilio butterflies have been investigated almost exclusively in an Apiaceae feeder by an in vitro assay. In contrast, there is little information regarding the Rutaceae feeders. Here, we focused on a Rutaceae feeder, Papilio xuthus, and identified two furanocoumarin-responsive cytochrome P450-6B (CYP6B) genes, of which one was an ortholog of a furanocoumarin-metabolizing enzyme identified in the Apiaceae-feeding Papilio while the other was previously unreported. We further conducted in vivo functional analysis using the CRISPR/Cas9 system, revealing a contribution of these CYP6Bs to furanocoumarin tolerance of P. xuthus larvae. Our findings suggest that co-option of furanocoumarin-metabolizing CYP6B enzymes at least partially contributed to the host shift from Rutaceae to Apiaceae in Papilio butterflies.
Subject(s)
Butterflies , Cytochrome P-450 Enzyme System , Furocoumarins , Rutaceae , Animals , Butterflies/enzymology , Butterflies/genetics , Butterflies/metabolism , Cytochrome P-450 Enzyme System/metabolism , Cytochrome P-450 Enzyme System/genetics , Furocoumarins/metabolism , Furocoumarins/chemistry , Rutaceae/metabolism , Rutaceae/genetics , Rutaceae/chemistry , Larva/metabolism , Insect Proteins/metabolism , Insect Proteins/genetics , Insect Proteins/chemistry , Phylogeny , HerbivoryABSTRACT
The establishment of near infrared (NIR) spectroscopy model mostly relies on chemometrics, and spectral analysis combined with artificial intelligence (AI) provides a new way of thinking for pharmaceutical quality inspection, new algorithms such as back propagation artificial neural networks (BP-ANN) and swarm intelligence optimization algorithms such as sparrow search algorithm (SSA) provide core technical support. In order to explore the application of AI in the pharmaceutical field, in this study, Angelica dahurica formula granules with a relatively complex system were selected as the research object. Quantitative analysis models were established by using partial least squares regression (PLSR) with a micro-NIR spectrometer, and BP-ANN modeling results were compared. For the best PLSR models of six characteristic components in the continuous counter-current extract of Angelica dahurica, R2v of imperatorin was lower than 0.90, and the RPD values of imperatorin, phellopterin, and isoimperatorin were even lower than 1. When the prediction model established by SSA-BP-ANN was used for quantitative analysis, R2v of six components were all higher than 0.92, and the RPD values all higher than 1.5, which proved that the BP-ANN method was better than PLSR. This study confirmed that in the continuous counter-current extraction progress of Angelica dahurica formula granules, the use of micro-NIR spectrometer combined with AI could realize the rapid prediction of the contents of six characteristic components. The comparison results provided a scientific reference for the process analysis and on-line monitoring in the production process of traditional Chinese medicine by micro-NIR spectrometer combined with AI.
Subject(s)
Angelica , Spectroscopy, Near-Infrared , Spectroscopy, Near-Infrared/methods , Angelica/chemistry , Least-Squares Analysis , Drugs, Chinese Herbal/chemistry , Drugs, Chinese Herbal/isolation & purification , Drugs, Chinese Herbal/analysis , Neural Networks, Computer , Artificial Intelligence , Algorithms , Furocoumarins/isolation & purification , Furocoumarins/chemistry , Furocoumarins/analysisABSTRACT
Ammi majus L. (Apiaceae) is a medicinal plant with a well-documented history in phytotherapy. The aim of the present work was to isolate isopimpinellin (5,8-methoxypsoralen; IsoP) from the fruit of this plant and evaluate its biological activity against selected tumor cell lines. The methanol extract obtained with the use of an accelerated solvent extraction (ASE) method was the most suitable for the quantitative analysis of coumarins in the A. majus fruit matrix. The coumarin content was estimated by RP-HPLC/DAD, and the amount of IsoP was found to be 404.14 mg/100 g dry wt., constituting 24.56% of the total coumarin fraction (1.65 g/100 g). This, along with the presence of xanthotoxin (368.04 mg/100 g, 22.36%) and bergapten (253.05 mg/100 g, 15.38%), confirmed A. majus fruits as an excellent source of these compounds. IsoP was isolated (99.8% purity) by combined liquid chromatography/centrifugal partition chromatography (LC/CPC) and tested for the first time on its antiproliferative activity against human colorectal adenocarcinoma (HT29, SW620), osteosarcoma (Saos-2, HOS), and multiple myeloma (RPMI8226, U266) cell lines. MTT assay results (96 h incubation) demonstrated a dose- and cell line-dependent decrease in cell proliferation/viability, with the strongest effect of IsoP against the Saos-2 cell line (IC50; 42.59 µM), medium effect against U266, HT-29, and RPMI8226 (IC50 = 84.14, 95.53, and 105.0 µM, respectively), and very weak activity against invasive HOS (IC50; 321.6 µM) and SW620 (IC50; 711.30 µM) cells, as well as normal human skin fibroblasts (HSFs), with IC50; 410.7 µM. The mechanistic study on the Saos-2 cell line showed that IsoP was able to reduce DNA synthesis and trigger apoptosis via caspase-3 activation. In general, IsoP was found to have more potency towards cancerous cells (except for HOS and SW620) than against healthy cells. The Selective Index (SI) was determined, underlining the higher selectivity of IsoP towards cancer cells compared to healthy cells (SI = 9.62 against Saos-2). All these results suggest that IsoP might be a promising molecule in the chemo-prevention and treatment of primary osteosarcoma.
Subject(s)
Ammi , Fruit , Furocoumarins , Plant Extracts , Humans , Fruit/chemistry , Cell Line, Tumor , Furocoumarins/pharmacology , Furocoumarins/chemistry , Plant Extracts/pharmacology , Plant Extracts/chemistry , Ammi/chemistry , Cell Proliferation/drug effects , Antineoplastic Agents, Phytogenic/pharmacology , Antineoplastic Agents, Phytogenic/chemistry , Apoptosis/drug effects , Chromatography, High Pressure Liquid , Cell Survival/drug effectsABSTRACT
BACKGROUND: The Xin-yi-san herbal decoction (XYS) is commonly used to treat patients with allergic rhinitis in Taiwan. Theophylline is primarily oxidized with high affinity by human cytochrome P450 (CYP)1A2, and has a narrow therapeutic index. PURPOSE: This study aimed to investigate the inhibition of human CYP1A2-catalyzed theophylline oxidation (THO) by XYS and its adverse effects in patients. METHODS: Human CYPs were studied in recombinant enzyme systems. The influence of concurrent XYS usage in theophylline-treated patients was retrospectively analyzed. RESULTS: Among the major human hepatic and respiratory CYPs, XYS inhibitors preferentially inhibited CYP1A2 activity, which determined the elimination and side effects of theophylline. Among the herbal components of XYS decoction, Angelicae Dahuricae Radix contained potent THO inhibitors. Furanocoumarin imperatorin was abundant in XYS and Angelicae Dahuricae Radix decoctions, and non-competitively inhibited THO activity with Ki values of 77â84 nM, higher than those (20â52 nM) of fluvoxamine, which clinically interacted with theophylline. Compared with imperatorin, the intestinal bacterial metabolite xanthotoxol caused weaker THO inhibition. Consistent with the potency of the inhibitory effects, the docking analysis generated Gold fitness values in the order-fluvoxamine > imperatorin > xanthotoxol. During 2017â2018, 2.6 % of 201,093 theophylline users consumed XYS. After inverse probability weighting, XYS users had a higher occurrence of undesired effects than non-XYS users; in particular, there was an approximately two-fold higher occurrence of headaches (odds ratio (OR), 2.14; 95 % confidence interval (CI), 1.99â2.30; p < 0.001) and tachycardia (OR, 1.83; 95 % CI, 1.21â2.77; p < 0.05). The incidence of irregular heartbeats increased (OR, 1.36; 95 % CI, 1.07â1.72; p < 0.05) only in the theophylline users who took a high cumulative dose (≥ 24 g) of XYS. However, the mortality in theophylline users concurrently taking XYS was lower than that in non-XYS users (OR, 0.24; 95 % CI, 0.14â0.40; p < 0.001). CONCLUSION: XYS contains human CYP1A2 inhibitors, and undesirable effects were observed in patients receiving both theophylline and XYS. Further human studies are essential to reduce mortality and to adjust the dosage of theophylline in XYS users.
Subject(s)
Angelica , Cytochrome P-450 CYP1A2 Inhibitors , Cytochrome P-450 CYP1A2 , Drugs, Chinese Herbal , Furocoumarins , Theophylline , Theophylline/pharmacology , Humans , Drugs, Chinese Herbal/pharmacology , Cytochrome P-450 CYP1A2/metabolism , Cytochrome P-450 CYP1A2 Inhibitors/pharmacology , Angelica/chemistry , Furocoumarins/pharmacology , Male , Herb-Drug Interactions , Retrospective Studies , Female , Taiwan , Middle Aged , Adult , Oxidation-Reduction , Rhinitis, Allergic/drug therapy , Rhinitis, Allergic/chemically inducedABSTRACT
Natural products are important sources of seed compounds for drug discovery. However, it has become difficult in recent years to discover new compounds with valuable pharmacological activities. On the other hand, among the vast number of natural products that have been isolated so far, a considerable number of compounds with specific biological activities are thought to be overlooked in screening that uses biological activity as an index. Therefore, it is conceivable that such overlooked useful compounds may be found by screening compound libraries that have been amassed previously through specific assays. Previously, NPD723, a member of the Natural Products Depository library comprised of a mixture of natural and non-natural products developed at RIKEN, and its metabolite H-006 were found to inhibit growth of various cancer cells at low nanomolar half-maximal inhibitory concentration. Subsequent analysis revealed that H-006 strongly inhibited human dihydroorotate dehydrogenase (DHODH), the rate-limiting enzyme in the de novo pyrimidine biosynthetic pathway. Here, we elucidated the crystal structure of the DHODH-flavin mononucleotide-orotic acid-H-006 complex at 1.7 Å resolution to determine that furocoumavirin, the S-enantiomer of H-006, was the actual inhibitor. The overall mode of interaction of furocoumavirin with the inhibitor binding pocket was similar to that described for previously reported tight-binding inhibitors. However, the structural information together with kinetic characterizations of site-specific mutants identified key unique features that are considered to contribute to the sub-nanomolar inhibition of DHODH by furocoumavirin. Our finding identified new chemical features that could improve the design of human DHODH inhibitors.
Subject(s)
Antiviral Agents , Dihydroorotate Dehydrogenase , Humans , Antiviral Agents/pharmacology , Antiviral Agents/chemistry , Crystallography, X-Ray , Dihydroorotate Dehydrogenase/antagonists & inhibitors , Dihydroorotate Dehydrogenase/chemistry , Enzyme Inhibitors/pharmacology , Enzyme Inhibitors/chemistry , Furocoumarins/pharmacology , Furocoumarins/chemistry , Models, MolecularABSTRACT
This study aimed to explore the antioxidant and prooxidative activity of two natural furanocoumarin derivatives, Bergaptol (4-Hydroxy-7H-furo [3,2-g] [1]benzopyran-7-one, BER) and Xanthotoxol (9-Hydroxy-7H-furo [3,2-g] [1]benzopyran-7-one, XAN). The collected thermodynamic and kinetic data demonstrate that both compounds possess substantial antiradical activity against HO⢠and CCl3OO⢠radicals in physiological conditions. BER exhibited better antiradical activity in comparison to XAN, which can be attributed to the enhanced deprotonation caused by the positioning of the -OH group on the psoralen ring. In contrast to highly reactive radical species, newly formed radical species BER⢠and XAN⢠exhibited negligible reactivity towards the chosen constitutive elements of macromolecules (fatty acids, amino acids, nucleobases). Furthermore, in the presence of O2â¢â, the ability to regenerate newly formed radicals BER⢠and XAN⢠was observed. Conversely, in physiological conditions in the presence of Cu(II) ions, both compounds exhibit prooxidative activity. Nevertheless, the prooxidative activity of both compounds is less prominent than their antioxidant activity. Furthermore, it has been demonstrated that anionic species can engage in the creation of a chelate complex, which restricts the reduction of metal ions when reducing agents are present (O2â¢â and Ascâ). Moreover, studies have demonstrated that these chelating complexes can be coupled with other radical species, hence enhancing their ability to inactivate radicals. Both compounds exhibited substantial inhibitory effects against enzymes involved in the direct or indirect generation of ROS: Xanthine Oxidase (XOD), Lipoxygenase (LOX), Myeloperoxidase (MPO), NADPH oxidase (NOX).
Subject(s)
Antioxidants , Furocoumarins , Furocoumarins/chemistry , Furocoumarins/pharmacology , Kinetics , Antioxidants/chemistry , Antioxidants/pharmacology , Density Functional Theory , Oxidation-Reduction , Thermodynamics , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Lipoxygenase/metabolism , Xanthine Oxidase/metabolism , Xanthine Oxidase/antagonists & inhibitors , Biological Products/chemistry , Biological Products/pharmacology , Free Radical Scavengers/chemistry , Free Radical Scavengers/pharmacologyABSTRACT
Chronic hepatitis B virus (HBV) infection remains a significant global health challenge due to its link to severe conditions like HBV-related cirrhosis and hepatocellular carcinoma (HCC). Although current treatments effectively reduce viral levels, they have limited impact on certain HBV elements, namely hepatitis B surface antigen (HBsAg) and covalently closed circular DNA (cccDNA). This highlights the urgent need for innovative pharmaceutical and biological interventions that can disrupt HBsAg production originating from cccDNA. In this study, we identified a natural furanocoumarin compound, Imperatorin, which markedly inhibited the expression of HBsAg from cccDNA, by screening a library of natural compounds derived from Chinese herbal medicines using ELISA assay and qRT-PCR. The pharmacodynamics study of Imperatorin was explored on HBV infected HepG2-NTCP/PHHs and HBV-infected humanized mouse model. Proteome analysis was performed on HBV infected HepG2-NTCP cells following Imperatorin treatment. Molecular docking and bio-layer interferometry (BLI) were used for finding the target of Imperatorin. Our findings demonstrated Imperatorin remarkably reduced the level of HBsAg, HBV RNAs, HBV DNA and transcriptional activity of cccDNA both in vitro and in vivo. Additionally, Imperatorin effectively restrained the actions of HBV promoters responsible for cccDNA transcription. Mechanistic study revealed that Imperatorin directly binds to ERK and subsequently interfering with the activation of CAMP response element-binding protein (CREB), a crucial transcriptional factor for HBV and has been demonstrated to bind to the PreS2/S and X promoter regions of HBV. Importantly, the absence of ERK could nullify the antiviral impact triggered by Imperatorin. Collectively, the natural compound Imperatorin may be an effective candidate agent for inhibiting HBsAg production and cccDNA transcription by impeding the activities of HBV promoters through ERK-CREB axis.
Subject(s)
DNA, Circular , Furocoumarins , Hepatitis B Surface Antigens , Hepatitis B virus , Transcription, Genetic , Furocoumarins/pharmacology , Humans , Animals , Hepatitis B virus/drug effects , Hepatitis B virus/genetics , Hepatitis B Surface Antigens/metabolism , Hepatitis B Surface Antigens/genetics , Hep G2 Cells , Mice , DNA, Circular/genetics , DNA, Circular/metabolism , Transcription, Genetic/drug effects , Antiviral Agents/pharmacology , DNA, Viral , Molecular Docking Simulation , Virus Replication/drug effects , Cyclic AMP Response Element-Binding Protein/metabolism , Hepatitis B, Chronic/drug therapy , Hepatitis B, Chronic/virology , Disease Models, Animal , Promoter Regions, GeneticABSTRACT
The post-column reaction method enables the evaluation of the antiradical capacity of individual components in a mixture by separating the components using HPLC and measuring stable free radical (e.g., DPPHâ) scavenging that occurs after the chromatography column. The equipment typically consists of two detectors. The first records signals of the analytes leaving the column. The second records radical scavenging by the analytes, which appears as a negative band. The recorded signals are found on two separate chromatograms, which must be combined to interpret the results. In this study, a single DAD detector was used behind the post-column reactor, enabling the simultaneous recording of the analyte bands and negative signals, indicating radical scavenging. The objective of this study was to evaluate the antiradical capacity of key compounds found in two herbal raw materials used in traditional Chinese medicine. Saposhnikovia divaricata roots contain phenolic acids, chromones, and furanocoumarins. Chlorogenic acid, rosmarinic acid, and imperatorin demonstrated strong radical scavenging, while prim-O-glucoslocimifugin showed a weaker response, both in standards and in root extracts. However, scavenging was not observed for cimifugin and 4'-O-ß-D-glucosyl-5-O-methylvisamminol. Astragalus mongholicus roots contain astragalosides I-IV (triterpene saponins). None of these showed DPPHâ scavenging. Furthermore, additional signals were observed, indicating the presence of unidentified radical scavenging compounds.
Subject(s)
Free Radical Scavengers , Plant Extracts , Plants, Medicinal , Plants, Medicinal/chemistry , Plant Extracts/chemistry , Chromatography, High Pressure Liquid/methods , Free Radical Scavengers/chemistry , Apiaceae/chemistry , Plant Roots/chemistry , Chromones/analysis , Chromones/chemistry , Furocoumarins/chemistry , Furocoumarins/analysisABSTRACT
With the increasing use of oral contraceptives and estrogen replacement therapy, the incidence of estrogen-induced cholestasis (EC) has tended to rise. Psoralen (P) and isopsoralen (IP) are the major bioactive components in Psoraleae Fructus, and their estrogen-like activities have already been recognized. Recent studies have also reported that ERK1/2 plays a critical role in EC in mice. This study aimed to investigate whether P and IP induce EC and reveal specific mechanisms. It was found that P and IP increased the expression of esr1, cyp19a1b and the levels of E2 and VTG at 80 µM in zebrafish larvae. Exemestane (Exe), an aromatase antagonist, blocked estrogen-like activities of P and IP. At the same time, P and IP induced cholestatic hepatotoxicity in zebrafish larvae with increasing liver fluorescence areas and bile flow inhibition rates. Further mechanistic analysis revealed that P and IP significantly decreased the expression of bile acids (BAs) synthesis genes cyp7a1 and cyp8b1, BAs transport genes abcb11b and slc10a1, and BAs receptor genes nr1h4 and nr0b2a. In addition, P and IP caused EC by increasing the level of phosphorylation of ERK1/2. The ERK1/2 antagonists GDC0994 and Exe both showed significant rescue effects in terms of cholestatic liver injury. In conclusion, we comprehensively studied the specific mechanisms of P- and IP-induced EC and speculated that ERK1/2 may represent an important therapeutic target for EC induced by phytoestrogens.
Subject(s)
Cholestasis , Ficusin , Furocoumarins , Psoralea , Animals , Biological Products/pharmacology , Biological Products/chemistry , Cholestasis/chemically induced , Cholestasis/metabolism , Estrogens/metabolism , Estrogens/pharmacology , Ficusin/pharmacology , Furocoumarins/pharmacology , Furocoumarins/chemistry , MAP Kinase Signaling System/drug effects , Psoralea/chemistry , ZebrafishABSTRACT
Furanocoumarins (FCs) are widely distributed secondary metabolites found in higher plants, including Apiaceae, Rutaceae, Moraceae, and Fabaceae. They play a crucial role in the physiological functions of plants and are well-known for their diverse pharmacological activities. As a representative plant of the Apiaceae family, Angelica sinensis is highly valued for its medicinal properties and FCs are one of the main ingredients of A. sinensis. However, the biosynthetic mechanism of FCs in A. sinensis remains poorly understood. In this study, we successfully cloned and verified three types of enzymes using genome analysis and in vitro functional verification, which complete the biosynthesis of the FCs core skeleton in A. sinensis. It includes a p-coumaroyl CoA 2'-hydroxylase (AsC2'H) responsible for umbelliferone formation, two UbiA prenyltransferases (AsPT1 and AsPT2) that convert umbelliferone to demethylsuberosin (DMS) and osthenol, respectively, and two CYP736 subfamily cyclases (AsDC and AsOD) that catalyze the formation of FCs core skeleton. Interestingly, AsOD was demonstrated to be a bifunctional cyclase and could catalyze both DMS and osthenol, but had a higher affinity to osthenol. The characterization of these enzymes elucidates the molecular mechanism of FCs biosynthesis, providing new insights and technologies for understanding the diverse origins of FCs biosynthesis.
Subject(s)
Angelica sinensis , Furocoumarins , Furocoumarins/chemistry , Furocoumarins/metabolism , Furocoumarins/biosynthesis , Angelica sinensis/chemistry , Angelica sinensis/metabolism , Molecular StructureABSTRACT
Nucleot(s)ide analogues, the current antiviral treatments against chronic hepatitis B (CHB) infection, are non-curative due to their inability to eliminate covalently closed circular DNA (cccDNA) from the infected hepatocytes. Preclinical studies have shown that coumarin derivatives can effectively reduce the HBV DNA replication. We evaluated the antiviral efficacy of thirty new coumarin derivatives in cell culture models for studying HBV. Furanocoumarins Fc-20 and Fc-31 suppressed the levels of pre-genomic RNA as well as cccDNA, and reduced the secretion of virions, HBsAg and HBeAg. The antiviral efficacies of Fc-20 and Fc31 improved further when used in combination with the hepatitis B antiviral drug Entecavir. There was a marked reduction in the intracellular HBx level in the presence of these furanocoumarins due to proteasomal degradation resulting in the down-regulation of HBx-dependent viral genes. Importantly, both Fc-20 and Fc-31 were non-cytotoxic to cells even at high concentrations. Further, our molecular docking studies confirmed a moderate to high affinity interaction between furanocoumarins and viral HBx via residues Ala3, Arg26 and Lys140. These data suggest that furanocoumarins could be developed as a new therapeutic for CHB infection.
Subject(s)
Antiviral Agents , DNA, Circular , Furocoumarins , Hepatitis B virus , Proteasome Endopeptidase Complex , Trans-Activators , Viral Regulatory and Accessory Proteins , Virus Replication , Hepatitis B virus/drug effects , Hepatitis B virus/genetics , Hepatitis B virus/physiology , Hepatitis B virus/metabolism , Virus Replication/drug effects , Humans , Trans-Activators/metabolism , Trans-Activators/genetics , DNA, Circular/metabolism , DNA, Circular/genetics , Viral Regulatory and Accessory Proteins/metabolism , Viral Regulatory and Accessory Proteins/genetics , Furocoumarins/pharmacology , Antiviral Agents/pharmacology , Proteasome Endopeptidase Complex/metabolism , DNA, Viral/metabolism , DNA, Viral/genetics , Down-Regulation/drug effects , Transcription, Genetic/drug effects , Proteolysis/drug effects , Gene Expression Regulation, Viral/drug effects , Hep G2 CellsABSTRACT
This study provides a comprehensive computational exploration of the inhibitory activity and metabolic pathways of 8-methoxypsoralen (8-MP), a furocoumarin derivative used for treating various skin disorders, on cytochrome P450 (P450). Employing quantum chemical DFT calculations, molecular docking, and molecular dynamics (MD) simulations analyses, the biotransformation mechanisms and the active site binding profile of 8-MP in CYP1B1 were investigated. Three plausible inactivation mechanisms were minutely scrutinized. Further analysis explored the formation of reactive metabolites in subsequent P450 metabolic processes, including covalent adduct formation through nucleophilic addition to the epoxide, 8-MP epoxide hydrolysis, and non-CYP-catalyzed epoxide ring opening. Special attention was paid to the catalytic effect of residue Phe268 on the mechanism-based inactivation (MBI) of P450 by 8-MP. Energetic profiles and facilitating conditions revealed a slight preference for the C4'=C5' epoxidation pathway, while recognizing a potential kinetic competition with the 8-OMe demethylation pathway due to comparable energy demands. The formation of covalent adducts via nucleophilic addition, particularly by phenylalanine, and the generation of potentially harmful reactive metabolites through autocatalyzed ring cleavage are likely to contribute significantly to P450 metabolism of 8-MP. Our findings highlight the key role of Phe268 in retaining 8-MP within the active site of CYP1B1, thereby facilitating initial oxygen addition transition states. This research offers crucial molecular-level insights that may guide the early stages of drug discovery and risk assessment related to the use of 8-MP.
Subject(s)
Furocoumarins , Methoxsalen , Methoxsalen/pharmacology , Molecular Docking Simulation , Secondary Metabolism , Furocoumarins/pharmacology , Epoxy CompoundsABSTRACT
Cnidium monnieri (L.) Cusson, a member of the Apiaceae family, is rich in coumarins, such as imperatorin and osthole. Cnidium monnieri fruit (CM) has a broad range of therapeutic potential that can be used in anti-bacterial, anti-cancer, and sexual dysfunction treatments. However, its efficacy in lowering blood pressure through vasodilation remains unknown. This study aimed to assess the potential therapeutic effect of CM 50% ethanol extract (CME) on hypertension and the mechanism of its vasorelaxant effect. CME (1-30 µg/mL) showed a concentration-dependent vasorelaxation on constricted aortic rings in Sprague Dawley rats induced by phenylephrine via an endothelium-independent mechanism. The vasorelaxant effect of CME was inhibited by blockers of voltage-dependent and Ca2+-activated K+ channels. Additionally, CME inhibited the vascular contraction induced by angiotensin II and CaCl2. The main active compounds of CM, i.e., imperatorin (3-300 µM) and osthole (1-100 µM), showed a concentration-dependent vasorelaxation effect, with half-maximal effective concentration values of 9.14 ± 0.06 and 5.98 ± 0.06 µM, respectively. Orally administered CME significantly reduced the blood pressure of spontaneously hypertensive rats. Our research shows that CME is a promising treatment option for hypertension. However, further studies are required to fully elucidate its therapeutic potential.
Subject(s)
Antihypertensive Agents , Blood Pressure , Cnidium , Ethanol , Fruit , Furocoumarins , Hypertension , Plant Extracts , Rats, Inbred SHR , Rats, Sprague-Dawley , Vasodilator Agents , Animals , Cnidium/chemistry , Plant Extracts/pharmacology , Plant Extracts/chemistry , Blood Pressure/drug effects , Rats , Fruit/chemistry , Vasodilator Agents/pharmacology , Male , Antihypertensive Agents/pharmacology , Ethanol/chemistry , Furocoumarins/pharmacology , Hypertension/drug therapy , Hypertension/physiopathology , Vasodilation/drug effects , Coumarins/pharmacology , Coumarins/chemistryABSTRACT
Though Isoimperatorin from Angelicae dahuricae is known to have antiviral, antidiabetic, anti-inflammatory and antitumor effects, its underlying antitumor mechanism remains elusive so far. Hence, the apoptotic mechanism of Isoimperatorin was explored in hepatocellular carcinomas (HCCs). In this study, Isoimperatorin inhibited the viability of Huh7 and Hep3B HCCs and increased the subG1 apoptotic portion and also abrogated the expression of pro-poly-ADP ribose polymerase (pro-PARP) and pro-caspase 3 in Huh7 and Hep3B cells. Also, Isoimperatorin abrogated the expression of cyclin D1, cyclin E1, CDK2, CDK4, CDK6 and increased p21 as G1 phase arrest-related proteins in Huh7 and Hep3B cells. Interestingly, Isoimperatorin reduced the expression and binding of c-Myc and Sirtuin 1 (SIRT1) by Immunoprecipitation (IP), with a binding score of 0.884 in Huh7 cells. Furthermore, Isoimperatorin suppressed the overexpression of c-Myc by the proteasome inhibitor MG132 and also disturbed cycloheximide-treated c-Myc stability in Huh7 cells. Overall, these findings support the novel evidence that the pivotal role of c-Myc and SIRT1 is critically involved in Isoimperatorin-induced apoptosis in HCCs as potent molecular targets in liver cancer therapy.
Subject(s)
Apoptosis , Carcinoma, Hepatocellular , Furocoumarins , Liver Neoplasms , Proto-Oncogene Proteins c-myc , Signal Transduction , Sirtuin 1 , Humans , Apoptosis/drug effects , Carcinoma, Hepatocellular/metabolism , Carcinoma, Hepatocellular/pathology , Carcinoma, Hepatocellular/drug therapy , Carcinoma, Hepatocellular/genetics , Cell Line, Tumor , Cell Proliferation/drug effects , Cell Survival/drug effects , Gene Expression Regulation, Neoplastic/drug effects , Liver Neoplasms/metabolism , Liver Neoplasms/pathology , Liver Neoplasms/drug therapy , Liver Neoplasms/genetics , Proto-Oncogene Proteins c-myc/drug effects , Proto-Oncogene Proteins c-myc/metabolism , Signal Transduction/drug effects , Sirtuin 1/drug effects , Sirtuin 1/metabolism , Furocoumarins/pharmacologyABSTRACT
A total of 11 active ingredients including psoralen, isopsoralen, bakuchiol, bavachalcone, bavachinin, corylin, coryfolin, isobavachalcone, neobavaisoflavone, bakuchalcone, and corylifol A from Psoraleae Fructus in the plasma samples of diabetic and normal rats were simultaneously determined by UHPLC-MS/MS. The pharmacokinetic parameters were calculated to elucidate the pharmacokinetic profiles of coumarins, flavonoids, and monoterpene phenols in normal and diabetic rats. The rat model of type 2 diabetes mellitus(T2DM) was induced by a high-sugar and high-fat diet combined with injection of 1% streptozotocin every two days. The plasma samples were collected at different time points after the rats were administrated with Psoraleae Fructus. The proteins in the plasma samples were precipitated by ethyl acetate, and the plasma concentrations of the 11 components of Psoraleae Fructus were determined by UHPLC-MS/MS. The pharmacokinetic parameters were calculated by DAS 3.0. The results showed that the pharmacokinetic beha-viors of 8 components including psoralen, isopsoralen, bakuchiol, and bavachinin from Psoraleae Fructus in both female and male mo-del rats were significantly different from those in normal rats. Among them, the coumarins including psoralen, isopsoralen, and corylin showed lowered levels in the blood of both female and male model rats. The flavonoids(bavachinin, corylifol A, and bakuchalcone) and the monoterpene phenol bakuchiol showed decreased levels in the female model rats but elevated levels in the male model rats. It is suggested that the dosage of Psoraleae Fructus should be reasonably adjusted for the patients of different genders at the time of clinical administration.
Subject(s)
Diabetes Mellitus, Experimental , Diabetes Mellitus, Type 2 , Drugs, Chinese Herbal , Furocoumarins , Phenols , Psoralea , Humans , Rats , Female , Male , Animals , Drugs, Chinese Herbal/pharmacokinetics , Tandem Mass Spectrometry/methods , Diabetes Mellitus, Experimental/drug therapy , Flavonoids/pharmacology , Ficusin , Coumarins , MonoterpenesABSTRACT
The use of plants such as giant hogweed as raw materials for the manufacture of dosage forms has been little explored. In this study, we utilized furanocoumarins from the Heracleum sosnowskyi plant to create an experimental emulsion dosage form (EmFHS). The EmFHS was finely dispersed (481.8 nm ± 71.1 nm), shelf-stable, and contained predominantly 8-methoxypsoralen at a concentration of 1 mg/ml. Phototoxicity analysis of EmFHS for THP-1 cells under UV (365 nm) irradiation showed an IC50 of 19.1 µg/ml (24 h) and 6.3 µg/ml (48 h). In relation to spheroids (L929), EmFHS exhibited a phototoxic effect in the concentration range of 31.25-125 µg/ml8-MOP. A full phototoxic effect was observed 48 h after UV irradiation. The phototoxic effect of EmFHS in vitro was dose-dependent and comparable to the effect of emulsion synthetic 8-methoxypsoralen and chlorin e6 solution. EmFHS cytotoxicity was caused solely by UV radiation, and toxicity in the dark was minimal. EmFHS, administered at a dose of 3 mg/kg8-MOP, was found to be safe after a single intravenous administration to rats. It had a photosensitizing effect in the form of local photodermatitis when exposed to UV irradiation at a dose of 44 J/cm2. The biokinetics of emulsion furanocoumarins showed that the phototoxic effect of EmFHS is due to the high penetration ability of the emulsion into cells of spheroids. At the same time, it has a low degree of cumulation when administered intravenously. The obtained data suggest that EmFHS may be a promising treatment for PUVA therapy of various dermatological diseases. Additionally, the plant Heracleum sosnowskyi shows potential as a basis for creating new dosage forms with phototherapeutic effects.
Subject(s)
Furocoumarins , Heracleum , Rats , Animals , Photosensitizing Agents , Methoxsalen , EmulsionsABSTRACT
ETHNOPHARMACOLOGICAL RELEVANCE: The adverse effects of Fructus Psoraleae (FP), especially liver injury, have attracted wide attention in recent years. AIM OF THE STUDY: To establish a system to explore potential hepatotoxic targets and the chief culprit of liver injury based on clinical experience, network pharmacological method, molecular docking, and in vitro and in vivo experiments. MATERIALS AND METHODS: Clinical applications and adverse reactions to FP were obtained from public literatures. Components absorbed in the blood were selected as candidates to search for potential active targets (PATs) of FP. Subsequently, potential pharmacological core targets (PPCTs) were screened through the "drug targets-disease targets" network. Non-drug active targets (NPATs) were obtained by subtracting the PPCTs from the PATs. The potential hepatotoxic targets (PHTs) of FP were the intersection targets obtained from Venn analysis using NPATs, hepatotoxic targets, and adverse drug reaction (ADR) targets provided by the databases. Then, potential hepatotoxic components and targets were obtained using the "NPATS-component" network relationship. Molecular docking and in vitro and in vivo hepatotoxicity experiments were performed to verify the targets and related components. RESULTS: Overall, 234 NPATs were acquired from our analysis, and 6 targets were identified as PHTs. Results from molecular docking and in vitro and in vivo experiments showed that angelicin is the leading cause of liver injury in FP, and VKORC1 plays an important role. CONCLUSION: The results indicate that six targets, especially VKORC1, are associated with the PHTs of FP, and angelicin is the leading culprit involved in FP liver injury via inhibition of VKORC1.