Daphnepedunins G and H, anti-HIV macrocyclic 3,4-seco-daphnane orthoesters from Daphne pedunculata.

Daphnepedunins G (1) and H (2) with unusual macrocyclic 3,4-seco-daphnane orthoester structure were isolated from Daphne pedunculata. Their structures were determined by physicochemical and spectroscopic analyses combined with synthetic methods, including methyl esterification, derivatization reaction using a chiral anisotropic agent, and biomimetic conversion. Compounds 1 and 2 along with their methyl esters 1a and 2a were evaluated for anti-HIV activity, among which 1a and 2a exhibited potent activity with IC50 values of 1.08 and 1.17 µM, respectively.

Identification of Daphne genkwa and Its Vinegar-Processed Products by Ultraperformance Liquid Chromatography-Quadrupole Time-of-Flight Mass Spectrometry and Chemometrics.

Crude herbs of Daphne genkwa (CHDG) are often used in traditional Chinese medicine to treat scabies baldness, carbuncles, and chilblain owing to their significant purgation and curative effects. The most common technique for processing DG involves the use of vinegar to reduce the toxicity of CHDG and enhance its clinical efficacy. Vinegar-processed DG (VPDG) is used as an internal medicine to treat chest and abdominal water accumulation, phlegm accumulation, asthma, and constipation, among other diseases. In this study, the changes in the chemical composition of CHDG after vinegar processing and the inner components of the changed curative effects were elucidated using optimized ultrahigh-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS). Untargeted metabolomics, based on multivariate statistical analyses, was also used to profile differences between CHDG and VPDG. Eight marker compounds were identified using orthogonal partial least-squares discrimination analysis, which indicated significant differences between CHDG and VPDG. The concentrations of apigenin-7-O-ß-d-methylglucuronate and hydroxygenkwanin were considerably higher in VPDG than those in CHDG, whereas the amounts of caffeic acid, quercetin, tiliroside, naringenin, genkwanines O, and orthobenzoate 2 were significantly lower. The obtained results can indicate the transformation mechanisms of certain changed compounds. To the best of our knowledge, this study is the first to employ mass spectrometry to detect the marker components of CHDG and VPDG.

Daphne genkwa flower extract promotes the neuroprotective effects of microglia.

BACKGROUND: Microglia are innate immune cells in the central nervous system that play a crucial role in neuroprotection by releasing neurotrophic factors, removing pathogens through phagocytosis, and regulating brain homeostasis. The constituents extracted from the roots and stems of the Daphne genkwa plant have shown neuroprotective effects in an animal model of Parkinson's disease. However, the effect of Daphne genkwa plant extract on microglia has yet to be demonstrated. PURPOSE: To study the anti-inflammatory and neuroprotective effects of Daphne genkwa flower extract (GFE) in microglia and explore the underlying mechanisms. METHODS: In-vitro mRNA expression levels of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), inducible nitric oxide synthase, Arginase1, and brain derived neurotropic factor (BDNF) were analyzed by reverse transcription polymerase chain reaction in microglia cells. Nitric oxide (NO) and TNF-α protein were respectively analyzed by Griess reagent and Enzyme Linked Immunosorbent Assay. Immunoreactivity of Iba-1, Neu-N, and BDNF in mouse brain were analyzed by immunofluorescence staining. Phagocytosis capacity of microglia was examined using fluorescent zymosan-red particles. RESULTS: GFE significantly inhibited lipopolysaccharide (LPS)-induced neuroinflammation and promoted neuroprotection both in vitro and in vivo. First, GFE inhibited the LPS-induced inflammatory factors NO, iNOS, and TNF-α in microglial cell lines and primary glial cells, thus demonstrating anti-inflammatory effects. Arginase1 and BDNF mRNA levels were increased in primary glial cells treated with GFE. Phagocytosis was also increased in microglia treated with GFE, suggesting a neuroprotective effect of GFE. In vivo, neuroprotective and anti-neuroinflammatory effects of GFE were also found in the mouse brain, as oral administration of GFE significantly inhibited LPS-induced neuronal loss and inflammatory activation of microglia. CONCLUSION: GFE has anti-inflammatory effects and promotes microglial neuroprotective effects. GFE inhibited the pro-inflammatory mediators and enhanced neuroprotective microglia activity by increasing BDNF expression and phagocytosis. These novel findings of the GFE effect on microglia show an innovative approach that can potentially promote neuroprotection for the prevention of neurodegenerative diseases.

Chemical constituents from Daphne giraldii and their cytotoxicities and inhibitory activities against acetylcholinesterase.

Seven triterpenoids (1-7), two prenylated coumarins (8 and 9), and one diphenylpropane (10), including five previously undescribed compounds (1-3, 8, and 10), were obtained from the stem and root barks of Daphne giraldii. The structures and absolute configurations of the new triterpenoids were established by NMR, HRESIMS, ECD calculations, and single-crystal X-ray diffraction analysis. All identified compounds were tested for cytotoxicities (human tumour cell line Hep3B) and inhibitory effects on AChE in vitro. Notably, prenylated coumarins (8 and 9) exhibited moderate cytotoxic activities and 3-hydroxy-substituted triterpenoids (2 and 4) showed mild inhibitory effects on AChE. Furthermore, compounds 2 and 4 have also been subjected to molecular docking studies to investigate the inhibitory mechanism.

Daphnes Cortex and its licorice-processed products suppress inflammation via the TLR4/NF-κB/NLRP3 signaling pathway and regulation of the metabolic profile in the treatment of rheumatoid arthritis.

ETHNOPHARMACOLOGICAL RELEVANCE: Daphnes Cortex (Daphne Giraldii Nitsche, DGN) is a popular traditional Chinese herbal medicine for traumatic injuries and rheumatoid arthritis (RA) in the Shaanxi and Gansu provinces of China. Due to skin irritation caused by raw DGN (RDGN), licorice-processed DGN products are usually used in clinical practice. However, the efficacy and mechanisms of action between DGN and its licorice-processed DGN products in treating RA have not been compared. AIMS: This study compared the efficacy and elucidated the mechanisms in vitro and in vivo between RDGN and its licorice-processed DGN products in treating RA. MATERIALS AND METHODS: A collagen-induced RA rat model was established, and treated with different doses of RDGN and its licorice-processed DGN products for 4 weeks to explore the therapeutic effects. The anti-inflammatory effects were assessed in RAW 264.7 macrophages stimulated by lipopolysaccharide (LPS). Analyses of the differential quality markers (DQMs) between DGN and its licorice-processed DGN products using ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry, and non-targeted metabolomics analyses of rat synovial tissues were used to systematically explore correlations between DGN processing and its efficacy. RESULTS: Licorice-processed DGN products significantly ameliorated RA symptoms in CIA rats. Licorice-processed DGN products also regulated inflammatory cytokines, matrix metalloproteinases, and vascular endothelial growth factor in the serum and cell supernatants. Licorice-processed DGN products significantly inhibited Toll-like receptor 4/nuclear factor kappa B/NOD-like receptor family, pyrin domain containing 3 (TLR4/NF-κB/NLRP3) signaling in CIA rats and LPS-induced RAW264.7 cells. The DQMs between RDGN and its licorice-processed DGN products were identified, most of which were amino acids or energy-related metabolites present in licorice-processed DGN products. Correlations between DQMs with differential metabolites and differential metabolic pathways were established. CONCLUSIONS: Licorice-processed DGN products displayed better anti-inflammatory effects via the TLR4/NF-κB/NLRP3 signaling pathway on CIA rats and LPS-induced RAW264.7 cells, and regulation of the metabolic profile in treating RA.

Ethnomedicinal use, phytochemistry, pharmacology, and toxicology of Daphne gnidium: A review.

ETHNOPHARMACOLOGICAL RELEVANCE: Daphne gnidium L., (Lazaz or Metnan) is a perennial plant that grows around the Mediterranean basin, in Southern Europe, North Africa and the Middle East. It is used in different countries for hair care and to treat several diseases including skin cancer, diabetes, nervous breakdowns, sinusitis, poisoning, rheumatic disorders, odontalgia, muscular pain, and gastrointestinal infections. It is also used as anti-inflammatory, insecticide, and anti-parasitic remedy. AIM OF THE REVIEW: In this review, previous studies on D. gnidium including its botanical description, taxonomy, geographical distribution, medicinal use, phytochemistry, and pharmacological properties were critically highlighted and discussed for suggesting the exploration of this specie and its bioactive compounds in medical applications. MATERIALS AND METHODS: Data on D. gnidium were gathered from Scientific search engines including PubMed, ScienceDirect, SpringerLink, Web of Science, Scopus, Wiley Online, SciFinder, and Google Scholar. Reports on D. gnidium written in English published before September 2020 were summarized. RESULTS: In traditional medicine, D. gnidium is used to treat diabetes, gastrointestinal infections, skin cancer, nervous breakdowns, and sinusitis. The extracts and essential oil of D. gnidium exhibited several biological properties such as antibacterial, antifungal, antiviral, antigenotoxic, hemolytic, anti-inflammatory, immunomodulatory, neuroprotective, allelopathic, and insecticidal effects. Phytochemical investigations identified several chemical classes of secondary metabolites in D. gnidium essential oil and extracts including terpenoids, coumarins, flavonoids, fatty acids, and alkanes. CONCLUSIONS: The findings presented in this study showed a link between the traditional medicinal use and scientific biological results about D. gnidium. However, further investigations should be carried out to support medical and cosmetic applications of this species. Indeed, D. gnidium and its main compounds should be confirmed concerning their safety and their bioavailability. Moreover, pharmacodynamic studies should be conducted to support their efficacy in medical applications.

Phytochemical Analysis and Biological Activities of the Ethanolic Extract of Daphne sericea Vahl Flowering Aerial Parts Collected in Central Italy.

In this paper, the first phytochemical analysis of the ethanolic extract of Daphne sericea Vahl flowering aerial parts collected in Italy and its biological activities were reported. Eleven compounds were identified i.e., α-linolenic acid (1), tri-linoleoyl-sn-glycerol (2), pheophorbide a ethyl ester (3), pilloin (4), sinensetin (5), yuanhuanin (6), rutamontine (7), syringin (8), p-coumaric acid (9), p-anisic acid (10) and caffeic acid (11). To the best of our knowledge, compounds (1-4, 7-8 and 10) were isolated from D. sericea for the first time during this work, whereas sinensetin (5) represents a newly identified component of the entire Thymelaeaceae family. The extract was found to possess radical scavenging against both DPPH• and 2,2'-azino-bis(3-thylbenzothiazoline-6-sulfonic acid (ABTS•+) radicals, with at least a 40-fold higher potency against the latter. Moreover, chelating abilities against both ferrous and ferric ions have been highlighted, thus suggesting a possible indirect antioxidant power of the extract. Although the precise bioactive compounds remain to be discovered, the polyphenolic constituents, including phenolic acids, tannins and flavonoids, seem to contribute to the antioxidant power of the phytocomplex. In addition, the extract produced cytotoxic effects in MDA-MB-231 and U87-MG cancer cell lines, especially at the concentration of 625 µg/mL and after 48-72 h. Further studies are required to clarify the contribution of the identified compounds in the bioactivities of the extract and to support possible future applications.

Acutissimalignan B from traditional herbal medicine Daphne kiusiana var. atrocaulis (Rehd.) F. Maekawa inhibits neuroinflammation via NF-κB Signaling pathway.

BACKGROUND: Emerging evidence indicates the important role of herbal medicine for neuroinflammation, which is closely associated with neurodegenerative diseases. OBJECTIVE: To clarify the characteristics and primary mechanisms of action of the traditional herbal medicine Daphne kiusiana var. atrocaulis (Rehd.) F. Maekawa in neuroinflammation by phytochemistry and bioassays using both in vitro and in vivo assays. METHODS: The chemical composition of D. kiusiana var. atrocaulis was clarified using multiple chromatography technologies and spectroscopic analysis. The anti-neuroinflammatory effects of the identified components were evaluated in LPS-induced BV-2 cells by monitoring the production of nitric oxide. C57BL/6 mice were used to construct a neuroinflammatory model by injecting LPS into the lateral ventricle of the brain. The most promising component was evaluated in vivo by measuring the number of Iba-1 cells and expression of inflammatory factors. Furthermore, the anti-neuroinflammatory mechanism involved in the activation of the NF-κB pathway was investigated using western blot and immunofluorescence. RESULTS: Thirty-two constituents (1-32), including five new compounds, were successfully identified from D. kiusiana var. atrocaulis. Compounds 3, 5, 12-15, and 20 (IC50 values from 5.41 to 57.27 µM) could considerably inhibit the LPS-induced production of NO in BV-2 cells, displaying stronger anti-neuroinflammatory activities than that of minocycline (IC50 = 67.08 µM). The concentration of the most potential compound 13 (IC50 5.41 µM) was 5.4% of the ethyl acetate fraction. Acutissimalignan B (13) could reduce the mRNA expression of iNOs, TNF-α, IL-1ß, and IL-6, inhibit the phosphorylation of IκBα, and inhibit the nuclear translocation of NK-κB p65 in BV-2 cells induced by LPS. Moreover, in the LPS-induced mouse model, compound 13 was found to exert anti-neuroinflammatory activity by attenuating the activation of microglia in the cortex and hippocampus, repressing the phosphorylation of IκBα, inhibiting the nuclear translocation of NK-κB p65, and decreasing the mRNA expression of iNOs, TNF-α, IL-1ß, and IL-6 in the cortex. CONCLUSION: We found that D. kiusiana var. atrocaulis had an inhibitory activity on neuroinflammation. In addition, the main active component (-)-acutissimalignan B (13) showed anti-neuroinflammatory effects in both in vivo and in vitro assays. Its mechanism of action may be associated with the inhibition of the NF-κB signaling pathway. Our current findings provide new information on D. kiusiana var. atrocaulis in the treatment of neuroinflammation.

Biflavonoid-rich fraction from Daphne pseudomezereum var. koreana Hamaya exerts anti-inflammatory effect in an experimental animal model of allergic asthma.

ETHNOPHARMACOLOGICAL RELEVANCE: Daphne pseudomezereum var. koreana Hamaya is distributed in the Gangwon-do of South Korea and is traditionally used to treat chronic inflammatory diseases, including rheumatoid arthritis. AIM OF THE STUDY: We investigated the anti-inflammatory effect of biflavonoid-rich fraction (BF) obtained from an extract of D. pseudomezereum leaves on lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages and mouse model of ovalbumin (OVA)-induced allergic asthma. MATERIALS AND METHODS: Neochamaejasmin B (NB) and chamaejasmin D (CD) were spectroscopically characterized as major components of BF obtained from the leaves of D. pseudomezereum. RAW264.7 cells pretreated with NB, CD and BF and activated by LPS (500 ng/ml) were used to assess the anti-inflammatory effects of these materials in vitro. To evaluate the protective effect of BF on allergic asthma, female BALB/c mice were sensitized to OVA by intraperitoneal (i.p.) injection and treated with BF by oral administration (15 or 30 mg/kg). RESULTS: Pretreatment with BF inhibited LPS-stimulated nitric oxide (NO), TNF-α and IL-6, and led to upregulation of heme oxygenase-1 (HO-1) in RAW264.7 macrophages. Orally administered BF significantly inhibited the recruitment of eosinophils and the production of IL-5, IL-6, IL-13 and MCP-1 as judged by the analysis of BALF from OVA-induced asthma animal model. BF also decreased the levels of IgE in the serum of asthmatic mice. BF suppressed the influx of inflammatory cells into nearby airways and the hypersecretion of mucus by the airway epithelium of asthmatic mice. In addition, the increase in Penh in asthmatic mice was reduced by BF administration. Furthermore, BF led to Nrf2 activation and HO-1 induction in the lungs of mice. CONCLUSIONS: These data have shown the anti-asthmatic effects of BF, and therefore we expect that BF may be a potential candidate as a natural drug/nutraceutical for the prevention and treatment of allergic asthma.

Daphne mucronata enhances cell proliferation and protects human adipose stem cells against monosodium iodoacetate induced oxidative stress in vitro.

Mesenchymal stem cells (MSCs) are being used to treat many diseases as they exhibit great regenerative potential. However, MSC's transplantation sometimes does not yield the maximum regenerative outcome as they are unable to survive in inflammatory conditions. Several approaches including preconditioning are used to improve the survival rate of mesenchymal stem cells. One such recently reported approach is preconditioning MSCs with plant extracts. The present study was designed to evaluate the effect of Daphne mucronata extract on stressed human adipose-derived mesenchymal stem cells (hADMSCs). Isolated hADMSCs were preconditioned with different concentrations of Daphne muconata extract and the protective, proliferative, antioxidant and anti-inflammatory effect was assessed through various assays and expression analysis of inflammatory markers regulated through NF-κB pathway. Results suggest that preconditioning hADMSCs with Daphne mucronata increased the cell viability, proliferative and protective potential of hADMSCs with a concomitant reduction in LDH, ROS and elevation in SOD activity. Moreover, both the ELISA and gene expression analysis demonstrated down regulations of inflammatory markers (IL1-ß, TNF-α, p65, p50, MMP13) in Daphne mucronata preconditioned hADMSCs as compared to stress. This is the first study to report the use of MIA induced oxidative stress against hADMSC's and effect of Daphne mucronata on stressed hADMSCs. Results of these studies provided evidence that Daphne mucronata protects the hADMSCs during stress conditions by down regulating the inflammatory markers and hence increase the viability and proliferative potential of hADMSCs that is crucial for transplantation purposes.

Effects of an ethyl acetate extract of Daphne altaica stem bark on the cell cycle, apoptosis and expression of PPARγ in Eca­109 human esophageal carcinoma cells.

Daphne altaica Pall. (D. altaica; Thymelaeaceae) has long been used in traditional Kazakh medicine for the treatment of cancer and respiratory diseases. Previous studies have demonstrated the in vitro anticancer effects of D. altaica extract and its constituents in certain cancer cell lines; however, the underlying molecular mechanisms are not completely understooD. The present study aimed to investigate the molecular mechanisms underlying the activity of an ethyl acetate extract of D. altaica (Da­Ea) by assessing its effects on cell morphology, cell apoptosis, cell cycle progression and the expression levels of peroxisome proliferator­activated receptor γ (PPARγ) in Eca­109 cells. Cell morphology was observed under a phase contrast microscope. Cell apoptosis and cell cycle progression were assessed by flow cytometry following Annexin V/propidium iodide (PI) double staining and PI single staining, respectively. The mRNA and protein expression levels of PPARγ were determined by reverse transcription­quantitative PCR and western blotting, respectively. Compared with the control group, the percentage of apoptotic cells, cell cycle arrest at S phase and apoptotic morphological cell characteristics were increased in Da­Ea­treated Eca­109 cells. Furthermore, Da­Ea treatment upregulated the mRNA and protein expression levels of PPARγ compared with the control cells. High­performance liquid chromatography with diode­array detection indicated that daphnetin­7­O­ß­D­glucoside, daphnetin, demethyldaphnoretin­7­O­ß­D­glucopyranoside and genkwanol A were the main constituents of Da­Ea. Collectively, the results suggested that Da­Ea displayed antiproliferative activities in Eca­109 cells by inducing apoptosis and S phase cell cycle arrest, as well as upregulating PPARγ expression levels.

[Correlation between chemical composition,ecological factors and soil factors of Chinese herbal medicine Daphnes Cortex].

In this paper, the correlation between the chemical constituents of Chinese herbal medicines Daphnes Cortex and the ecological factors and soil factors was studied, which provided a reference for the selection of suitable areas for artificial cultivation of Daphne giraldii and wild tending. The geographic information system(GIS) was applied to obtain the ecological factor information of 23 collection sites of Daphnes Cortex, and the soil factor information was determined by the standard procedure in the soil test standard manual. Combining the information of 93 chemical constituents of Daphnes Cortex in 23 collection sites the correlation between components and ecological factors and soil factors was analyzed by statistical methods. The correlation analysis showed that the longitude, annual average rainfall, annual sunshine intensity, annual average temperature in the ecological factors, soil type, effective copper and pH value were the dominant factors affecting the chemical composition of Daphnes Cortex.

The genus Daphne: A review of its traditional uses, phytochemistry and pharmacology.

The genus Daphne belongs to the Thymeleaceae family and contains over 90 species that are distributed in Asia, Europe and parts of North Africa. The species of the genus Daphne are used in the traditional medicine of China, Tibet, Korea, and the Middle East for the treatment of various conditions. A broad range of studies has shown the significant biological potential of these species as sources of biologically and pharmacologically active compounds. Daphne species are a source of several classes of valuable phytochemicals such as coumarins, flavonoids, lignans, steroids and different classes of terpenes. The phytochemical diversity of this genus is demonstrated by over 350 secondary metabolites isolated from various species. The genus possesses a broad spectrum of biological activities including antibacterial, antifungal, antioxidant, analgesic, anti-inflammatory, cytotoxic, antiviral, abortive and haemostatic effects. A variety of bioactive secondary metabolites found in this genus may have potential use in pharmaceutical, cosmetic and food industries. Thus, species belonging to the genus Daphne can be considered an important source both for the treatment of various disorders, due to the presence of a plethora of bioactive constituents with potent bioactivities, and as possible leads in the discovery and synthesis of new medications.

ANATOMICAL CHARACTERISTICS AND BIOLOGICAL ACTIVITY OF THE STEMS AND LEAVES OF DAPHNE ALBOWIANA WORONOW EX POBED.

The aim of the study was to determine the stable diagnostic traits and the biological activity of the stems and leaves of Daphne albowiana Woronow ex Pobed., a plant native to Georgia. Biological assays of the methanol, chloroform and hexane extracts show the plant to possess cytotoxic and antioxidant activities, but no noteworthy antibacterial or antifungal activities. All extracts show cytotoxic activity on A549 (lung carcinoma) cells. The following stable diagnostic characteristic were identified during the microstructural analysis: leaf surface glabrous, hypostomatic, dorsoventral; epidermal cells chaotic; curved with curved walls on both the upper and lower epidermis; stomata paracytic; well visible spherical crystals of inulin in leaf epidermis; leaf vascular bundles reverse-collateral; vascular system monocyclic, bilateral; wood diffuse-porous; xylem parenchyma is apotracheal, slightly diffuse; vessel walls are predominantly spirally thickened; collenchyma lamellar; radial rays in single rows, heterogeneous. The identified cytotoxic and antioxidant activity showcase this species to be of significant interest to the medicinal field. The identified anatomical peculiarities provide valuable information for the correct identification and standardization of the Daphne albowiana plant material.

Identification of the metabolites in normal and AA rat plasma, urine and feces after oral administration of Daphne genkwa flavonoids by LC-Q-TOF-MS spectrometry.

Daphne genkwa Sieb. et Zucc., as a traditional oriental herb, has been widely distributed and employed in China. The major bioactive components in D. genkwa are flavonoid compounds, which showed pharmacological activities such as anti-inflammatory, analgesic, anti-tumor and immunomodulatory activities. In this study, we analyzed total flavonoids in D. genkwa and their metabolites in normal and adjuvant arthritis (AA) rat plasma, urine and feces samples by liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (LC-Q-TOF-MS). A total of 4 metabolites in plasma, 9 metabolites in urine and 15 metabolites in feces were characterized respectively by LC-Q-TOF-MS technology in normal rat. And 9 of the metabolites were observed in the AA rat urine, while there was no prototype drug or its metabolites detected in plasma and fecal samples. The metabolic pathway mainly involves hydroxylation, methylation, glucuronide, sulfate conjugation, oxidation and reduction, during the phase I and phase II biotransformation pathway. All the information gained here will be greatly helpful in elucidating the potential biological and pharmacological mechanism of flavonoid in D. genkwa, thus providing new ideas for drug development.

Evaluating the inhibitory effect of eight compounds from Daphne papyracea against the NS3/4A protease of hepatitis C virus.

Hepatitis C virus (HCV) infection is a global threat to human health with an estimated 1.75 million new cases in 2015. Our previous studies showed that the ethyl acetate extraction of Daphne papyracea exhibited an inhibitory effect towards the HCV NS3/4A protease and eight compounds were identified from the extract. In this study, we investigated which of the eight compounds was responsible for the inhibitory effect of the extract against the HCV NS3/4A protease. From both molecular docking and enzyme inhibition studies, (+)-usnic acid was shown to be the most active compound and could be used as a lead compound in developing novel anti-HCV agents.

Suppression of Hepatocellular Carcinoma Progression through FOXM1 and EMT Inhibition via Hydroxygenkwanin-Induced miR-320a Expression.

Daphne genkwa, a Chinese medicinal herb, is used frequently in Southeast Asian countries to treat diseases; the flavonoid hydroxygenkwanin (HGK) is extracted from its flower buds. The bioactivity of HGK, particularly as an anti-liver cancer agent, has not been explored. In this study, human hepatocellular carcinoma (HCC) cell lines and an animal xenograft model were employed to investigate both the activity of HGK against liver cancer and its cellular signaling mechanisms. HCC cells treated with HGK were subjected to cell function assays. Whole transcriptome sequencing was used to identify genes whose expression was influenced by HGK, and the flavonoid's cancer suppression mechanisms were further investigated through gain- and loss-of-function assays. Finally, in vitro findings were tested in a mouse xenograft model. The data showed that HGK induced the expression of the microRNA miR-320a, which in turn inhibited the expression of the transcription factor 'forkhead box protein M1' (FOXM1) and downstream FOXM1-regulated proteins related to epithelial-mesenchymal transition, thereby leading to the suppression of liver cancer cell growth and invasion. Significant inhibition of tumor growth was also observed in HGK-treated mice. Hence, the present study demonstrated the activity of HGK against liver cancer and validated its potential use as a therapeutic agent.

Yuanhuatine from Daphne genkwa selectively induces mitochondrial apoptosis in estrogen receptor α-positive breast cancer cells in vitro.

Breast cancer is one of the most common cancers diagnosed among women worldwide. Estrogen receptor alpha (ERα) is a transcriptional factor that plays an important role in the development and progression of breast cancer. Yuanhuatine, a natural daphnane-type diterpenoid extracted from Daphne genkwa, was reported to exhibit significant cytotoxicity against breast cancer cells. However, the underlying mechanism is still unclear. In this study, we evaluated the cytotoxicity of yuanhuatine on two breast cancer cell lines that are ERα-positive and -negative. The results show that yuanhuatine inhibits the growth of ERα-positive cells (MCF-7) with much stronger inhibitory activity (IC50 = 0.62 µM) compared with positive control tamoxifen (IC50 = 14.43 µM). However, no obvious cytotoxicity was observed in ERα-negative cells (MDA-MB-231). Subsequent experiment also indicated that yuanhuatine markedly induced mitochondrial dysfunction, leading to apoptosis in MCF-7 cells. Molecular docking studies suggest the potential interactions between yuanhuatine and ERα. Immunofluorescence staining and Western blot analysis indicated that yuanhuatine down-regulated the expression of ERα in MCF-7 cells. MPP, a specific ERα inhibitor, significantly enhanced yuanhuatine-induced mitochondrial dysfunction and apoptosis in MCF-7 cells. On the contrary, the treatment with yuanhuatine causes no apoptosis in MM231 cells. Altogether, in vitro and in silico results suggested that ERα down-regulation was involved in yuanhuatine-induced mitochondrial dysfunction and apoptosis in ERα-positive breast cancer cells. Thus, yuanhuatine could be a potential candidate for treating ERα-positive breast cancer.

Assignment of the stereostructures of sesquiterpenoids from the roots of Daphne genkwa via quantum chemical calculations.

Five new guaiane-type sesquiterpenoids were obtained from the roots of Daphne genkwa. Their gross structures were established by extensive spectroscopic analyses. Attempts on the assignment of the relative configurations were unsuccessful when based on the NOESY correlations. Therefore, NMR chemical shift calculations based on the gauge independent atomic orbital (GIAO) method in combination with the statistical method DP4+ were employed to establish their relative configurations. Furthermore, the absolute configurations were determined by comparing the experimental and calculated electronic circular dichroism (ECD) using time-dependent density functional theory (TDDFT). The isolated compounds were screened for their cytotoxicity in vitro against two human hepatocellular carcinoma, HepG2 and Hep3B cell lines.

Health risk of Licorice-Yuanhua combination through induction of colonic H2S metabolism.

ETHNOPHARMACOLOGICAL RELEVANCE: Licorice and Yuanhua are both famous herbs in Traditional Chinese Medicine (TCM), and their combination is used by some TCM doctors to treat renal and gastrointestinal diseases as well as tumors. On the other hand, the compatibility theory of TCM warns that toxic effects might be triggered by Licorice-Yuanhua combination. The usability of Licorice-Yuanhua combination has long been controversial due to lack of evidence and mechanism illustration. Colonic hydrogen sulfide (H2S) metabolism imbalance is closely related with colonic inflammation, tumor promotion and many other diseases. AIM OF THE STUDY: This study was carried out to investigate if licorice-Yuanhua combination could induce potential toxic effects in the aspect of colonic H2S metabolism. MATERIALS AND METHODS: Normal mice were treated with high or low doses of Licorice, Yuanhua and Licorice-Yuanhua combination. Fecal H2S concentration was measured by colorimetric method, colon sulfomucin production was compared through tissue staining, fecal microbiota and microbial metagenomes were analyzed by 16S rDNA sequencing and data mining. RESULTS: Data shows that although licorice cannot change colonic H2S concentration, it can exacerbate Yuanhua induced H2S rising. Licorice or Yuanhua increases colon sulfomucin production, and their combination further enhances this effect. 16S rDNA sequencing analysis revealed that licorice or Yuanhua has little influence on gut microbiota, however, licorice-Yuanhua combination can impact gut microbiota structural balance and increase the abundance of Desulfovibrio genus and other related genera. Moreover, the combination extensively changes microbial metagenomes, influencing 1172 genes that cannot be changed by individual licorice or Yuanhua. By searching in KEGG database, ten genes are annotated with H2S producing gene, and these genes are remarkably increased by licorice-Yuanhua combination, more significantly than licorice or Yuanhua. CONCLUSIONS: This study provides evidences and mechanisms for licorice induced risks, which is related with colonic H2S metabolism disturbance, gut microbiota and microbial metagenomes. More risk assessment should be evaluated when licorice was used in combination with foods, herbs or drugs. The study provides an example where healthy risks can be induced by combination of food additive, herbs or drugs, through regulating gut microbiota and its metagenomes.