Antioxidant and Antiproliferative Activities of Eclipta prostrata (L.) L. Extract and Isolated Compounds.

The primary objective of this study was to elucidate the chemical composition, antioxidant properties, and antiproliferative activities of Eclipta prostrata extracts. Two flavonoids, 3'-O-methylorobol and apigenin 7-sulfate, were isolated from the ethyl acetate (EtOAc) extract of E. prostrata. The total phenolic and flavonoid contents of the E. prostrata extracts, as well as their overall antioxidant activities as measured using the 2,2-diphenyl-1-picrylhydrazyl and reducing power assays, were investigated. The E. prostrata EtOAc extract exhibited significantly greater antioxidant activities in both assays and higher phenol and flavonoid contents than the other extracts. The potential antiproliferative properties of the E. prostrata extracts and isolated compounds were investigated in vitro against the AGS, A549, and HT-29 cancer cell lines and the normal human HEK-293 cell line using the MTT assay. Annexin V-FITC/PI staining analysis and quantitative real-time PCR were used to assess AGS cell apoptosis. At a concentration of 100 µg/mL, the EtOAc extract of E. prostrata reduced AGS cell viability and proliferation by inducing apoptosis through the alteration of gene expression in the apoptotic cascade. These results highlight E. prostrata as a promising source of anticancer compounds.

Impact of Molecular Drug Susceptibility Testing on the Time to Multidrug-resistant Tuberculosis Treatment Initiation.

BACKGROUND: The purpose of this study was to evaluate the current status and trends in the coverage of molecular drug susceptibility testing (mDST), and the impact of mDST on the time to multidrug-resistant tuberculosis (MDR-TB) treatment initiation in Korea. METHODS: We included confirmed rifampin-resistant (RR)/MDR-TB patients who submitted application forms for novel drug uses to the National TB Expert Review Committee from September 1, 2016 to November 30, 2019. We retrospectively reviewed their medical records. RESULTS: Of the 621 MDR/RR-TB patients, mDST was performed in 442 (71.2%); Xpert MTB/RIF (Xpert) alone in 109 (17.6%), MTBDRplus line probe assay (LPA) alone in 199 (32.0%), and both Xpert and LPA in 134 (21.6%) patients. The coverage rate of mDST has gradually increased to 70% in 2015, 50.7% in 2016, 67.9% in 2017, 75.2% in 2018, and 79.4% in 2019 (P for trend < 0.001). Median time to MDR-TB treatment initiation was 35 days (interquartile range25-75 0-72), which has gradually decreased during the study period (P < 0.001). Independent predictors of shorter time to MDR-TB treatment initiation were retreatment case (adjusted hazard ratio [aHR], 1.30; 95% confidence interval [CI], 1.10-1.54), Xpert testing (aHR, 2.42; 95% CI, 2.03-2.88), and LPA testing (aHR, 1.83; 95% CI, 1.55-2.16). Transfer to another healthcare facility was inversely related to shorter time to treatment initiation (aHR, 0.74; 95% CI, 0.63-0.88). CONCLUSION: mDST coverage is gradually increasing and contributes to reducing the time to MDR-TB treatment initiation. Further efforts are needed to achieve universal access to mDST and to properly integrate mDST into routine clinical practice.

In silico screening of GABA aminotransferase inhibitors from the constituents of Valeriana officinalis by molecular docking and molecular dynamics simulation study.

Modulation of γ-aminobutyric acid (GABA) levels has been required in various disorders. GABA itself cannot be directly introduced into central nervous system (CNS) because of the blood brain barrier; inhibition of GABA aminotransferase (GABA-AT), which degrades GABA in CNS, has been the target for the modulation of GABA levels in CNS. Given that root extract of valerian (Valeriana officinalis) has been used for millennia as anti-anxiolytic and sedative, in silico approach was carried out to investigate valerian compounds exhibiting GABA-AT inhibiting activity. The 3D structure of human GABA-AT was created from pig crystal structure via homology modeling. Inhibition of GABA-AT by 18 valerian compounds was analyzed using molecular docking and molecular dynamics simulations and compared with known GABA-AT inhibitors such as vigabatrin and valproic acid. Isovaleric acid and didrovaltrate exhibited GABA-AT inhibiting activity in computational analysis, albeit less potent compared with vigabatrin. However, multiple compounds with low activity may have additive effects when the total extract of valeriana root was used in traditional usage. In addition, isovaleric acid shares similar backbone structure to GABA, suggesting that isovaleric acid might be a valuable starting structure for the development of more efficient GABA-AT inhibitors for disorders related with low level of GABA in the CNS.

Flavonoids from Scutellaria baicalensis inhibit senescence-associated secretory phenotype production by interrupting IκBζ/C/EBPß pathway: Inhibition of age-related inflammation.

BACKGROUND: Prolonged exposure to the senescence-associated secretory phenotype (SASP) with age leads to chronic low-grade inflammation in neighboring cells and tissues, causing many chronic degenerative diseases. PURPOSE: The effects on SASP production of the ethanol extract from Scutellaria radix and 17 isolated flavonoid constituents were examined in vitro and in vivo. METHODS: Cellular senescence was induced by bleomycin. Expression of the SASP and cell signaling molecules was detected using ELISA, RT-qPCR, Western blotting, and immunofluorescence staining. To investigate the in vivo effects, 21-month-old aged rats were used. RESULTS: The ethanol extract and 5 compounds including 1 (Oroxylin A; 5,7-dihydroxy-6-methoxyflavone), 5 (2',6',5,7-tetrahydroxy-8-methoxyflavone), 8 (2',5,7-trihydroxyflavone), 10 (2',5,7-trihydroxy-8-methoxyflavone) and 11 (2',5,7-trihydroxy-6-methoxyflavone) potently reduced IL-6 and IL-8 production and gene expression of the SASP, including IL-1α, IL-1ß, IL-6, IL-8, GM-CSF, CXCL1, MCP-2, and MMP-3. This finding indicates the important role of the B-ring 2'­hydroxyl group in flavonoid molecules. Furthermore, compounds 8 and 11, the strongest SASP inhibitors, decreased the expression of IκBζ and C/EBPß protein without affecting either BrdU uptake or the expression of senescence markers, such as pRb and p21. Finally, the oral administration of compound 8 to aged rats at 2 and 4 mg/kg/day for 10 days significantly inhibited the gene expression of SASP and IκBζ in kidneys. This is the first report of the strong SASP inhibitory action of flavonoids from Scutellaria radix on in vitro and in vivo senescence models. The inhibitory action was shown to be mediated mainly by interfering with the IκBζ/C/EBPß signaling pathway. CONCLUSION: Targeting production of the SASP using flavonoids from Scutellaria radix or its extract might help reduce low-grade sterile inflammation and control age-related diseases.

Inhibitory Mechanisms of Water Extract of Oplopanax elatus on Lipopolysaccharide-Induced Inflammatory Responses in RAW 264.7 Murine Macrophage Cells.

OBJECTIVE: To study the anti-inflammatory action and cellular mechanism of Oplopanax elatus. METHODS: A hot water extract of OE (WOE) was prepared and a major constituent, syringin, was successfully isolated. Its content in WOE was found to be 214.0 µg/g dried plant (w/w). Their anti-inflammatory activities were examined using RAW 264.7 macrophages and a mouse model of croton oil-induced ear edema. RESULTS: In lipopolysaccharide (LPS)-treated RAW 264.7 cells, a mouse macrophage cell line, WOE was found to significantly and strongly inhibit cyclooxygenase-2 (COX-2)-induced prostaglandin E2 (PGE2) production [half maximal inhibitory concentration (IC50)=135.2 µg/mL] and inducible nitric oxide synthase (iNOS)-induced NO production (IC50=242.9 µg/mL). In the same condition, WOE was revealed to inhibit NO production by down-regulating iNOS expression, mainly by interrupting mitogen activated protein kinases (MAPKs)/activator protein-1 (AP-1) pathway. The activation of all three major MAPKs, p38 MAPK, extracellular signal-regulated kinase (ERK), and c-Jun N-terminal kinase, was inhibited by WOE (50-300 µg/mL). On the other hand, WOE reduced PGE2 production by inhibiting COX-2 enzyme activity, but did not affect COX-2 expression levels. In addition, WOE inhibited the production of proinflammatory cytokines such as interleukin-6 and tumor necrosis factor-α. In croton oil-induced ear edema in mice, oral administration of WOE (50-300 mg/kg) dose-dependently inhibited edematic inflammation. CONCLUSION: Water extract of OE exhibited multiple anti-inflammatory action mechanisms and may have potential for treating inflammatory disorders.

Antimicrobial activity and active compounds of a Rhus verniciflua Stokes extract.

The Rhus verniciflua Stokes (RVS) extract is used as a traditional herbal medicine in Southeast Asian countries such as Korea and China. In the present study, one phenolic acid and six flavonoids were isolated from an 80% ethanol RVS extract to examine their antimicrobial activities. These compounds were identified as 3',4',7-trihydroxyflavone (1), methyl gallate (2), gallic acid (3), fusti (4), fisetin (5), butin (6), and sulfuretin (7) by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and nuclear magnetic resonance spectroscopy. The antimicrobial activities of compounds 5 and 6 (at a dose of 16 µg/mL each) were superior to that of the control, cycloheximide (at a dose of 25 µg/mL), against Hypocrea nigricans; additionally, the activities of compounds 1 and 2 (at a dose of 8 µg/mL each) were superior to the control against Penicillium oxalicum. Also, chemical compounds 1 and 5 (at a dose of 16 µg/mL each) had higher activities than the control (25 µg/mL) against Trichoderma virens. Chemical compound 1 (at a dose of 8 µg/mL) had a similar activity to that of the control against Bacillus subtilis. The obtained results suggest that the RVS extract could be a promising food and nutraceutical source because of the antimicrobial properties of its phenolic compounds.

Therapeutic Potential of the Rhizomes of Anemarrhena asphodeloides and Timosaponin A-III in an Animal Model of Lipopolysaccharide-Induced Lung Inflammation.

Investigations into the development of new therapeutic agents for lung inflammatory disorders have led to the discovery of plant-based alternatives. The rhizomes of Anemarrhena asphodeloides have a long history of use against lung inflammatory disorders in traditional herbal medicine. However, the therapeutic potential of this plant material in animal models of lung inflammation has yet to be evaluated. In the present study, we prepared the alcoholic extract and derived the saponin-enriched fraction from the rhizomes of A. asphodeloides and isolated timosaponin A-III, a major constituent. Lung inflammation was induced by intranasal administration of lipopolysaccharide (LPS) to mice, representing an animal model of acute lung injury (ALI). The alcoholic extract (50-200 mg/kg) inhibited the development of ALI. Especially, the oral administration of the saponin-enriched fraction (10-50 mg/kg) potently inhibited the lung inflammatory index. It reduced the total number of inflammatory cells in the bronchoalveolar lavage fluid (BALF). Histological changes in alveolar wall thickness and the number of infiltrated cells of the lung tissue also indicated that the saponin-enriched fraction strongly inhibited lung inflammation. Most importantly, the oral administration of timosaponin A-III at 25-50 mg/kg significantly inhibited the inflammatory markers observed in LPS-induced ALI mice. All these findings, for the first time, provide evidence supporting the effectiveness of A. asphodeloides and its major constituent, timosaponin A-III, in alleviating lung inflammation.

Determination of Five Chemical Markers in DF Formula, the Herbal Composition of Ephedra intermedia, Rheum palmatum, and Lithospermum erythrorhizon, Using High-performance Liquid Chromatography-ultraviolet Detection.

BACKGROUND: DF formula is a herbal preparation comprised three medicinal herbs, namely, Ephedra intermedia, Rheum palmatum, and Lithospermum erythrorhizon, which is being used for the treatment of obesity and liver fibrosis in Korean local clinics. OBJECTIVE: Since the abovementioned three herbs exist with different proportions in DF formula and their chemical markers have different physiochemical properties; it is quite challenging to develop an analytical methodology for the determination of these chemical markers. MATERIALS AND METHODS: For the analysis of the three herbs, five chemicals, (+)-pseudoephedrine (1) and (-)-ephedrine (2) for E. intermedia, aloe-emodin (3), and chrysophanol (4) for R. palmatum, and shikonin (5) for L. erythrorhizon, were selected for method validation of DF formula, and the analytical conditions were optimized and validated using high-performance liquid chromatography coupled with an ultraviolet detector (HPLC-UV). RESULTS: The specificities for the five compounds 1-5 were determined by their UV absorption spectra (1-4: 215 nm and 5: 520 nm). Their calibration curves showed good linear regressions with high correlation coefficient values (R2 > 0.9997). The limits of detection of these five markers were in the range 0.4-2.1 ng/mL, with the exception of 5 (12.7 ng/mL). The intraday variability for all the chemical markers was less than a Relative standard deviation (RSD) of 3%, except for 5 (RSD = 12.6%). In the case of interday analysis, 1 (1.0%), 2 (3.1%), and 4 (3.7%) showed much lower variabilities (RSD < 5%) than 3 (7.6%) and 5 (8.2%). Moreover, the five chemical markers showed good recoveries with good accuracies in the range of 90%-110%. CONCLUSIONS: The developed HPLC-UV method for the determination of the five chemical markers of the components of DF formula was validated. SUMMARY: DF formula, the herbal composition of Ephedra intermedia, Rheum palmatum and Lithospermum erythrorhizonFive chemical markers in DF formula were (+)-pseudoephedrine (1) and (-)-ephedrine (2) for E. intermedia, aloe-emodin (3) and chrysopanol (4) for R. palmatum, and shikonin (5) for L. erythrorhizon, with quite different physico-chemical propertiesFive chemical markers in DF formula were determined by HPLC-UV Abbreviations used: EP: (-)-ephedrine; PSEP: (+)-pseudoephedrine; HPLC: High-performance liquid chromatography; UV: Ultraviolet; LOD: Limit of detection; LOQ: Limit of quantification; RSD: Relative standard deviation.

Anticancer Activity of an Oplopanax elatus Stem Extract and Biologically Active Isolated Compounds.

BACKGROUND AND METHODS: Oplopanax elatus (Nakai) Nakai is used in folk medicine in China. In this study, the antiproliferative activity of an O. elatus fraction extracted by ethyl acetate (EF) was tested on human breast cancer MCF-7 cells, human colon cancer HCT-116 cells, and human stomach cancer AGS cells. The potential mechanism of antiproliferation was also investigated using an apoptosis assay. RESULTS: The results showed that the EF remarkably suppressed proliferation of human breast, stomach, and colon cancer cells. Further apoptosis tests by flow cytometry and immunoblot analyses showed the EF inhibited HCT-116 cell proliferation by inducing apoptosis. The bioassay-monitored fractionation of the EF resulted in the isolation of two polyacetylenes, falcarindiol (compound 1) and oplopandiol (compound 2), with falcarindiol possessing the strongest antiproliferative activity in colon cancer cells. CONCLUSION: Together, this study evaluated the anticancer activity of an O. elatus extract against human cancer cells, and provided the basis for further development of this herbal extract for the treatment of cancer.

Antioxidant Activity, α-glucosidase Inhibitory Activity and Chemoprotective Properties of Rhododendron brachycarpum Leaves Extracts.

BACKGROUND: Rhododendron brachycarpum (RB) is a genus of flowering plants generally used for traditional medicine in Korea to treat hypertension, neuralgia, and sterility. Previous studies have shown that RB extract alleviate inflammation and antimicrobial activity. OBJECTIVE: In this study, the effects of RB and its different fractions (n-hexane, ethyl-acetate and n-butanol) on antioxidant activity, DNA damage prevention and the activity of α-glucosidase were studied. RESULTS: The antioxidant ability of RB was investigated in vitro, including that of DPPH-radical and reducing power. As expected, scavenging effect against DPPH-radical of ethyl acetate fraction (IC50 = 17.7 ± 0.5 µg/ml) of RB had the highest DPPH radical scavenging activity, and it was superior to the positive control, butylated hydroxytoluene (BHT) (IC50 = 80.8 ± 1.5 µg/ml). And the reducing power of RB was 3.18 at 1.0 mg/ml. Meanwhile, the α-glucosidase inhibitory activity and prevention of oxidation stress-induced DNA damage were also highest in the ethyl acetate fraction. Pretreatment of pancreatic ß-cells from Syrian golden hamster (HIT-TI5) with the ethyl acetate fraction at concentrations of 300 µg/ml significantly protected the cells from high glucose-induced cell death. CONCLUSION: Our results indicate that ethyl acetate fraction of RB leaves extract has strong antioxidant, α-glucosidase, and prevention of DNA damage activities, and furthermore, ethyl acetate fraction significantly protected the cells from high glucose-induced cell death.

Anti-inflammatory Effects of the Natural Compounds Cortex Phellodendri and Humulus japonicus on Pelvic Inflammatory Disease in Mice.

Pelvic inflammatory disease (PID) is an inflammatory and/or infectious disorder of the upper female genital tract, including the uterus, fallopian tubes, and adjacent pelvic structures, that may spread upward to the peritoneum. Currently available treatment options have presented to produce adverse effects of various degrees, such as increased antimicrobial resistance and a limited effective duration of hormones. In the study, the Cortex Phellodendri (CP) and Humulus japonicus (HJ) among natural compounds that are believed to present biological activities with fewer side effects were tested in a PID animal model. The results suggested that the administration CP and HJ reduced clinical signs, inflammatory cytokine expression as well as secretion in uterine tissue, and neutrophil infiltration into the tissue.

Therapeutic Potential of Medicinal Plants and Their Constituents on Lung Inflammatory Disorders.

Acute bronchitis and chronic obstructive pulmonary diseases (COPD) are essentially lung inflammatory disorders. Various plant extracts and their constituents showed therapeutic effects on several animal models of lung inflammation. These include coumarins, flavonoids, phenolics, iridoids, monoterpenes, diterpenes and triterpenoids. Some of them exerted inhibitory action mainly by inhibiting the mitogen-activated protein kinase pathway and nuclear transcription factor-κB activation. Especially, many flavonoid derivatives distinctly showed effectiveness on lung inflammation. In this review, the experimental data for plant extracts and their constituents showing therapeutic effectiveness on animal models of lung inflammation are summarized.

Choice between Levofloxacin and Moxifloxacin and Multidrug-Resistant Tuberculosis Treatment Outcomes.

RATIONALE: We previously showed that the choice of levofloxacin or moxifloxacin for the treatment of patients with fluoroquinolone-sensitive multidrug-resistant tuberculosis (MDR-TB) did not affect sputum culture conversion at 3 months of treatment. OBJECTIVES: To compare final treatment outcomes between patients with MDR-TB randomized to levofloxacin or moxifloxacin. METHODS: A total of 151 participants with MDR-TB who were included for the final analysis in our previous trial were followed through the end of treatment. Treatment outcomes were compared between 77 patients in the levofloxacin group and 74 in the moxifloxacin group, based on the 2008 World Health Organization definitions as well as 2013 revised definitions of treatment outcomes. In addition, the time to culture conversion was compared between the two groups. MEASUREMENTS AND MAIN RESULTS: Treatment outcomes were not different between the two groups, based on 2008 World Health Organization definitions as well as 2013 definitions. With 2008 definitions, cure was achieved in 54 patients (70.1%) in the levofloxacin group and 54 (73.0%) in the moxifloxacin group (P = 0.72). Treatment success rates, including cure and treatment completed, were not different between the two groups (87.0 vs. 81.1%, P = 0.38). With 2013 definitions, cure rates (83.1 vs. 78.4%, P = 0.54) and treatment success rates (84.4 vs. 79.7%, P = 0.53) were also similar between the levofloxacin and moxifloxacin groups. Time to culture conversion was also not different between the two groups (27.0 vs. 45.0 d, P = 0.11 on liquid media; 17.0 vs. 42.0 d, P = 0.14 on solid media). Patients in the levofloxacin group had more adverse events than those in the moxifloxacin group (79.2 vs. 63.5%, P = 0.03), especially musculoskeletal ones (37.7 vs. 14.9%, P = 0.001). CONCLUSIONS: The choice of levofloxacin or moxifloxacin made no difference to the final treatment outcome among patients with fluoroquinolone-sensitive MDR-TB. Clinical trial registered with www.clinicalrials.gov (NCT01055145).

Antiviral effects of Phyllanthus urinaria containing corilagin against human enterovirus 71 and Coxsackievirus A16 in vitro.

Human enterovirus 71 (EV71) and Coxsackievirus A16 (CA16) are major causative agents of hand, foot, and mouth disease (HFMD) especially in infants and children under 5 years of age. Despite recent outbreaks of HFMD, there are no approved therapeutics against EV71 and CA16 infection. Moreover, in a small percentage of cases, the disease progression can lead to serious complications of the central nervous system. In this study, we investigated the antiviral effect of corilagin and Phyllanthus urinaria extract, which contains corilagin as a major component, on EV71 and CA16 infection in vitro. Our results indicate that corilagin reduces the cytotoxicity induced by EV71 or CA16 on Vero cells with and IC50 value of 5.6 and 32.33 µg/mL, respectively. We confirmed the presence of corilagin in EtOAc and BuOH fractions from P. urinaria extract and this correlated with antiviral activity of the fractions against EV71 or CA16. Future studies will be required to confirm the antiviral activity of corilagin and P. urinaria extract in vivo. Challenging a model with a lethal dose of viral infection will be required to test this. Collectively, our work provides potential candidates for the development of novel drugs to treat HFMD.

Comparison of levofloxacin versus moxifloxacin for multidrug-resistant tuberculosis.

RATIONALE: Levofloxacin (LFX) and moxifloxacin (MXF) are the two most frequently recommended fluoroquinolones for treatment of patients with multidrug-resistant tuberculosis (MDR-TB). However, studies comparing the effectiveness of LFX and MXF among patients with MDR-TB are lacking. OBJECTIVES: To compare the effectiveness of LFX and MXF in terms of culture conversion after 3 months of treatment for MDR-TB. METHODS: In this prospective multicenter randomized open label trial, we randomly assigned 182 patients with MDR-TB (sensitive to LFX and MXF) to receive either LFX (750 mg/day; 90 patients) or MXF (400 mg/day; 92 patients) with a background drug regimen. The primary outcome was the proportion of patients who achieved sputum culture conversion at 3 months of treatment. Secondary outcomes were time to culture conversion and time to smear conversion, with data censored at 3 months, and the proportions of adverse drug reactions. MEASUREMENTS AND MAIN RESULTS: At 3 months of treatment, 68 (88.3%) of the 77 patients in the LFX group and 67 (90.5%) of the 74 in the MXF group showed conversion to negative sputum cultures (odds ratio for LFX compared with MXF, 0.78; 95% confidence interval, 0.27-2.20). Adverse drug reactions were reported in six patients (7.7%) in the LFX group and four (5.2%) in the MXF group (P = 0.75). CONCLUSIONS: The choice of LFX or MXF for treatment of patients with MDR-TB may not affect sputum culture conversion at 3 months of treatment. Clinical trial registered with www.clinicaltrials.gov (NCT 01055145).

The Cyt P450 enzyme CYP716A47 catalyzes the formation of protopanaxadiol from dammarenediol-II during ginsenoside biosynthesis in Panax ginseng.

Ginseng (Panax ginseng C.A. Meyer) is one of the most popular medicinal herbs and contains pharmacologically active components, ginsenosides, in its roots. Ginsenosides, a class of tetracyclic triterpene saponins, are thought to be synthesized from dammarenediol-II after hydroxylation by the Cyt P450 (CYP) enzyme and then glycosylation by glycosyltransferase (GT). However, no genes encoding the hydroxylation and glycosylation in ginsenoside biosynthesis have been identified. Here, we identify protopanaxadiol synthase, which is a CYP enzyme (CYP716A47), to be involved in the hydroxylation of dammarenediol-II at the C-12 position to yield protopanaxadiol. Nine putative full CYP sequences were isolated from the expressed sequence tags (ESTs) of methyl jasmonate (MeJA)-treated adventitious ginseng roots. The CYP716A47 gene product was selected as the putative protopanaxadiol synthase because this gene was transcriptionally activated not only by MeJA treatment but also in transgenic ginseng that overexpresses squalene synthase and overproduces ginsenosides. In vitro enzymatic activity assays revealed that CYP716A47 catalyzed the oxidation of dammarenediol-II to produce protopanaxadiol. Ectopic expression of CYP716A47 in recombinant WAT21 yeasts that were fed dammarenediol-II yielded protopanaxadiol. Furthermore, co-expression of the dammarenediol synthase gene (PgDDS) and CYP716A47 in yeast yielded protopanaxadiol without adding dammarenediol-II. The chemical structures of the protopanaxadiol products from dammarenediol-II were confirmed using liquid chromatography-atmospheric pressure chemical ionization mass spectrometry (LC/APCIMS). Thus, CYP716A47 is a dammarenediol 12-hydroxylase that produces protopanaxadiol from dammarenediol-II.

Effects of oxypeucedanin on global gene expression and MAPK signaling pathway in mouse neuroblastoma Neuro-2A cells.

Oxypeucedanin is a major coumarin aglycone that can be extracted from Ostericum koreanum. Coumarin aglycones have demonstrated various pharmacological effects, including anti-proliferation, anti-inflammation, and anti-pain. In this study, in order to understand the pharmacological properties of oxypeucedanin, we investigated global gene expression alteration in mouse neuroblastoma Neuro-2A cells. Results from the MTT assay indicated no decrease of cell viability up to 100 µM for 24 h. We measured gene expression profiles in Neuro-2A cells treated with either 10 µM or no oxypeucedanin for 24 h. We selected 128 differentially expressed genes (DEGs) for comparison of gene expression profiles by Bonferroni-adjusted p values (p < 0.1). Analysis of Gene Ontology (GO) biological process terms using the DEGs demonstrated the importance of protein metabolism, particularly ribosomal protein synthesis and protein degradation, intramembrane protein trafficking, and electron transport. Treatment with oxypeucedanin resulted in the downregulation of most DEGs for ribosomal protein synthesis and the electron transport chain (ETC). In contrast, most DEGs for protein degradation and cellular trafficking systems were upregulated. In addition, we found five upregulated DEGs for core and regulatory proteins involved in the mitogen-activated protein kinase (MAPK) signaling pathway. Independent translational validation of DEGs for MAPK signaling by immunoblot analysis showed consistent agreement with microarray data. Overall protein levels of Erk2 and p38MAPK were elevated, and their phosphorylated forms were also increased. These functional categories, based on transcriptional alteration and complicated modulation of MAPK signaling, might be underlying mechanisms responsible for the various pharmacological effects of oxypeucedanin.

Downregulation of matrix metalloproteinase-13 by the root extract of Cyathula officinalis Kuan and its constituents in IL-1ß-treated chondrocytes.

The roots of Cyathula officinalis Kuan are widely used in Chinese medicine for the treatment of inflammatory disorders. Here, the ability of C. officinalis Kuan to downregulate matrix metalloproteinase (MMP)-13 was examined since MMP-13 is an important enzyme for the degradation of the cartilage collagen matrix, especially under arthritic conditions. The ethanol extract of C. officinalis Kuan as well as the N-hexane and chloroform soluble fractions were found to potently inhibit MMP-13 induction in IL-1 ß-treated SW1353 cells, a human chondrosarcoma cell line, at 50-200 µg/mL. Activity-guided separation led to the isolation of six compounds, palmitic acid (1), ß-sitosterol (2), α-spinasterol (3), atractylenolide I (4), 1,3-diacetoxy-tetradeca-6E,12E-dien-8,10-dyn (5), and N-trans-feruloyl-3-methyldopamine (6). Among these, 4 and 5 exhibited MMP-13 downregulating activity in IL-1 ß-treated SW1353 cells. And 4 also showed anti-oedematous activity against λ-carageenan-induced paw edema in mice at 20-200 mg/kg, p. o. The results of this study provide information that can help elucidate the action mechanism of C. officinalis Kuan. In addition, the results presented here suggest that C. officinalis Kuan and its constituents may have the potential for chondroprotection against cartilage degrading disorders.

Extraction and identification of three major aldose reductase inhibitors from Artemisia montana.

Aldose reductase inhibitors (ARIs) provide an important therapeutic and preventive opportunity against hyperglycemia associated diabetic complications. The methanolic extracts of 12 species from the genus Artemisia exhibited significant in vitro rat lens AR (RLAR) inhibitory activities with IC(50) values ranging from 0.51 to 13.45 µg/mL (quercetin, 0.64 µg/mL). Since the whole plant of Artemisia montana showed the highest RLAR inhibitory activity, bioassay-guided fractionation was performed to obtain ethyl acetate and n-butanol fractions. Repeated column chromatography of two active fractions, yielded fifteen compounds, including four chlorogenic acids (3,5-di-O-caffeoylquinic acid, chlorogenic acid, neochlorogenic acid, cryptochlorogenic acid), six flavonoids (apigenin, luteolin, quercetin, isoquercitrin, hyperoside, luteolin 7-rutinoside), and five coumarins (umbelliferone, scoparone, scopoletin, esculetin, and scopolin); their structures were confirmed by spectroscopic methods. 3,5-Di-O-caffeoylquinic acid and chlorogenic acid, as well as test flavonoids, displayed the most potent RLAR inhibitory activities with IC(50) values ranging from 0.19 to 5.37 µM. Furthermore, the HPLC profiles of the ethyl acetate and n-butanol fractions indicated that 3,5-di-O-caffeoylquinic acid, chlorogenic acid, and hyperoside, as major compounds, might play crucial roles in RLAR inhibition. The results suggest that A. montana and three key AR inhibitors therein would clearly be potential candidates as therapeutic or preventive agents for diabetic complications.

Isolation and identification of sea buckthorn (Hippophae rhamnoides) phenolics with antioxidant activity and α-glucosidase inhibitory effect.

This study was performed to evaluate the antioxidant and α-glucosidase inhibitory effects from the extract, fractions, and isolated compounds of sea buckthorn leaves. Six compounds, kaempferol-3-O-ß-D-(6''-O-coumaryl) glycoside, 1-feruloyl-ß-D-glucopyranoside, isorhamnetin-3-O-glucoside, quercetin 3-O-ß-D-glucopyranoside, quercetin 3-O-ß-D-glucopyranosyl-7-O-α-L-rhamnopyranoside, and isorhamnetin-3-O-rutinoside, were isolated from sea buckthorn leaf extracts. The butanol fraction (EC(50) = 1.81 µg/mL) along with quercetin 3-O-ß-D-glucopyranoside (EC(50) = 1.86 µg/mL) had a higher DPPH radical-scavenging activity and showed stronger reducing power (OD(700) = 1.83 and 1.78, respectively). The butanol fraction (477 mg GAE/g) contained the highest amount of phenolic compounds and also the most powerful α-glucosidase inhibitory effect (86%) at 5 µg/mL. The results indicate that sea buckthorn leaf extracts could potentially be used for food additives and the development of useful natural compounds.