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Sample records for active metabolite retinoic

  1. Structure-dependent activities of polybrominated diphenyl ethers and hydroxylated metabolites on zebrafish retinoic acid receptor.

    PubMed

    Zhao, Jing; Zhu, Xiangwei; Xu, Ting; Yin, Daqiang

    2015-02-01

    Polybrominated diphenyl ethers (PBDEs), a group of potential endocrine-disrupting chemicals (EDCs) have been shown to disrupt retinoid homeostasis in different species in both laboratory and field studies. However, the molecular mechanisms of interactions with the retinoic acid receptor (RAR) are not fully understood. Zebrafish have proven useful for investigating mechanisms of chemical toxicity. In the present study, a reporter gene assay was used to investigate the activities of 11 PBDEs and six OH-PBDEs with different degrees of bromination on zebrafish RAR. All tested OH-PBDEs induced RAR transcriptional activity; however, of the 11 PBDEs examined, only BDE28 and BDE154 affected the RAR transcriptional activity. Homology modeling and molecular docking were employed to simulate the interactions of PBDEs/OH-PBDEs with zebrafish RARs and to identify binding affinities to analyze the specialization of the interaction between RARs and PBDEs/OH-PBDEs. The results showed that although these compounds could bind with RARs, the effects of PBDEs/OH-PBDEs on RAR transcriptional activity did not depend on their RAR-binding abilities. The present study is the first attempt to demonstrate that OH-PBDEs could induce RAR transcriptional activity by binding directly with RAR; these effects are possibly related to the structure of the compounds, especially their hydroxylation and bromination. Most of the PBDEs could not directly interact with the RAR. PMID:25077655

  2. Metabolic characteristics of 13-cis-retinoic acid (isotretinoin) and anti-tumour activity of the 13-cis-retinoic acid metabolite 4-oxo-13-cis-retinoic acid in neuroblastoma

    PubMed Central

    Sonawane, Poonam; Cho, Hwang Eui; Tagde, Ashujit; Verlekar, Dattesh; Yu, Alice L; Reynolds, C Patrick; Kang, Min H

    2014-01-01

    Background and Purpose Isotretinoin (13-cis-retinoic acid; 13-cRA) is a differentiation inducer used to treat minimal residual disease after myeloablative therapy for high-risk neuroblastoma. However, more than 40% of children develop recurrent disease during or after 13-cRA treatment. The plasma concentrations of 13-cRA in earlier studies were considered subtherapeutic while 4-oxo-13-cis-RA (4-oxo-13-cRA), a metabolite of 13-cRA considered by some investigators as inactive, were greater than threefold higher than 13-cRA. We sought to define the metabolic pathways of 13-cRA and investigated the anti-tumour activity of its major metabolite, 4-oxo-13-cRA. Experimental Approach Effects of 13-cRA and 4-oxo-13-cRA on human neuroblastoma cell lines were assessed by DIMSCAN and flow cytometry for cell proliferation, MYCN down-regulation by reverse transcription PCR and immunoblotting, and neurite outgrowth by confocal microscopy. 13-cRA metabolism was determined using tandem MS in human liver microsomes and in patient samples. Key Results Six major metabolites of 13-cRA were identified in patient samples. Of these, 4-oxo-13-cRA was the most abundant, and 4-oxo-13-cRA glucuronide was also detected at a higher level in patients. CYP3A4 was shown to play a major role in catalysing 13-cRA to 4-oxo-13-cRA. In human neuroblastoma cell lines, 4-oxo-13-cRA and 13-cRA were equi-effective at inducing neurite outgrowth, inhibiting proliferation, decreasing MYCN mRNA and protein, and increasing the expression of retinoic acid receptor-β mRNA and protein levels. Conclusions and Implications We showed that 4-oxo-13-cRA is as active as 13-cRA against neuroblastoma cell lines. Plasma levels of both 13-cRA and 4-oxo-13-cRA should be evaluated in pharmacokinetic studies of isotretinoin in neuroblastoma. PMID:25039756

  3. Induction of CYP26A1 by Metabolites of Retinoic Acid: Evidence That CYP26A1 Is an Important Enzyme in the Elimination of Active Retinoids

    PubMed Central

    Topletz, Ariel R.; Tripathy, Sasmita; Foti, Robert S.; Shimshoni, Jakob A.; Nelson, Wendel L.

    2015-01-01

    All-trans-retinoic acid (atRA), the active metabolite of vitamin A, induces gene transcription via binding to nuclear retinoic acid receptors (RARs). The primary hydroxylated metabolites formed from atRA by CYP26A1, and the subsequent metabolite 4-oxo-atRA, bind to RARs and potentially have biologic activity. Hence, CYP26A1, the main atRA hydroxylase, may function either to deplete bioactive retinoids or to form active metabolites. This study aimed to determine the role of CYP26A1 in modulating RAR activation via formation and elimination of active retinoids. After treatment of HepG2 cells with atRA, (4S)-OH-atRA, (4R)-OH-atRA, 4-oxo-atRA, and 18-OH-atRA, mRNAs of CYP26A1 and RARβ were increased 300- to 3000-fold, with 4-oxo-atRA and atRA being the most potent inducers. However, >60% of the 4-OH-atRA enantiomers were converted to 4-oxo-atRA in the first 12 hours of treatment, suggesting that the activity of the 4-OH-atRA was due to 4-oxo-atRA. In human hepatocytes, atRA, 4-OH-atRA, and 4-oxo-atRA induced CYP26A1 and 4-oxo-atRA formation was observed from 4-OH-atRA. In HepG2 cells, 4-oxo-atRA formation was observed even in the absence of CYP26A1 activity and this formation was not inhibited by ketoconazole. In human liver microsomes, 4-oxo-atRA formation was supported by NAD+, suggesting that 4-oxo-atRA formation is mediated by a microsomal alcohol dehydrogenase. Although 4-oxo-atRA was not formed by CYP26A1, it was depleted by CYP26A1 (Km = 63 nM and intrinsic clearance = 90 μl/min per pmol). Similarly, CYP26A1 depleted 18-OH-atRA and the 4-OH-atRA enantiomers. These data support the role of CYP26A1 to clear bioactive retinoids, and suggest that the enzyme forming active 4-oxo-atRA may be important in modulating retinoid action. PMID:25492813

  4. A paradoxical teratogenic mechanism for retinoic acid.

    PubMed

    Lee, Leo M Y; Leung, Chun-Yin; Tang, Walfred W C; Choi, Heung-Ling; Leung, Yun-Chung; McCaffery, Peter J; Wang, Chi-Chiu; Woolf, Adrian S; Shum, Alisa S W

    2012-08-21

    Retinoic acid, an active metabolite of vitamin A, plays essential signaling roles in mammalian embryogenesis. Nevertheless, it has long been recognized that overexposure to vitamin A or retinoic acid causes widespread teratogenesis in rodents as well as humans. Although it has a short half-life, exposure to high levels of retinoic acid can disrupt development of yet-to-be formed organs, including the metanephros, the embryonic organ which normally differentiates into the mature kidney. Paradoxically, it is known that either an excess or a deficiency of retinoic acid results in similar malformations in some organs, including the mammalian kidney. Accordingly, we hypothesized that excess retinoic acid is teratogenic by inducing a longer lasting, local retinoic acid deficiency. This idea was tested in an established in vivo mouse model in which exposure to excess retinoic acid well before metanephric rudiments exist leads to failure of kidney formation several days later. Results showed that teratogen exposure was followed by decreased levels of Raldh transcripts encoding retinoic acid-synthesizing enzymes and increased levels of Cyp26a1 and Cyp26b1 mRNAs encoding enzymes that catabolize retinoic acid. Concomitantly, there was significant reduction in retinoic acid levels in whole embryos and kidney rudiments. Restoration of retinoic acid levels by maternal supplementation with low doses of retinoic acid following the teratogenic insult rescued metanephric kidney development and abrogated several extrarenal developmental defects. This previously undescribed and unsuspected mechanism provides insight into the molecular pathway of retinoic acid-induced teratogenesis. PMID:22869719

  5. Biological activity of all-trans retinol requires metabolic conversion to all-trans retinoic acid and is mediated through activation of nuclear retinoid receptors in human keratinocytes.

    PubMed

    Kurlandsky, S B; Xiao, J H; Duell, E A; Voorhees, J J; Fisher, G J

    1994-12-30

    The biological activity of all-trans retinol, in human keratinocytes, was investigated through metabolic and functional analyses that assessed the capacity for retinol uptake and metabolism and the mechanism of retinol-induced activation of gene transcription. Human keratinocytes converted all-trans retinol predominantly to retinyl esters, which accounted for 60 and 90% of cell-associated radiolabel after a 90-min pulse and a 48-h chase, respectively. Human keratinocytes also metabolized all-trans retinol to low levels of all-trans retinoic acid (11.47-131.3 ng/mg of protein) in a dose-dependent manner, between 0.3 and 10 microM added retinol. Small amounts of 13-cis retinoic acid (5.47-8.62 ng/mg of protein) were detected, but 9-cis retinoic acid was detected only when keratinocytes were incubated with radiolabeled retinol. There was no accumulation of the oxidized catabolic metabolites 4-hydroxy- or 4-oxoretinoic acid; however, 5,6-epoxy retinoic acid was detected at pharmacological levels (10 and 30 microM) of added retinol. Biological activity of retinol was assessed through analysis of two known retinoic acid-mediated responses: 1) reduction of type I epidermal transglutaminase and 2) activation of a retinoic acid receptor-dependent reporter gene, beta RARE3-tk-CAT. Both all-trans retinol and all-trans retinoic acid reduced type I epidermal transglutaminase in a dose-dependent manner; however, the ED50 for all-trans retinol (10 nM) was 10 times greater than for all-trans retinoic acid (1 nM). All-trans retinol also stimulated beta RARE3-tk-CAT reporter gene activity in a dose-dependent manner. Half-maximal induction was observed at 30 nM retinol, which was again 10-fold greater than observed with all-trans retinoic acid. Cotransfection of human keratinocytes with expression vectors for dominant negative mutant retinoic acid and retinoid X receptors reduced retinol-induced beta RARE3-tk-CAT reporter gene activation by 80%. Inhibition of conversion of all

  6. Retinoic acid modulation of ultraviolet light-induced epidermal ornithine decarboxylase activity

    SciTech Connect

    Lowe, N.J.; Breeding, J.

    1982-02-01

    Irradiation of skin with ultraviolet light of sunburn range (UVB) leads to a large and rapid induction of the polyamine biosynthetic enzyme ornithine decarboxylase in the epidermis. Induction of epidermal ornithine decarboxylase also occurs following application of the tumor promoting agent 12-0-tetradecanoylphorbol-13 acetate and topical retinoic acid is able to block both this ornithine decarboxylase induction and skin tumor promotion. In the studies described below, topical application of retinoic acid to hairless mouse skin leads to a significant inhibition of UVB-induced epidermal ornithine decarboxylase activity. The degree of this inhibition was dependent on the dose, timing, and frequency of the application of retinoic acid. To show significant inhibition of UVB-induced ornithine decarboxylase the retinoic acid had to be applied within 5 hr of UVB irradiation. If retinoic acid treatment was delayed beyond 7 hr following UVB, then no inhibition of UVB-induced ornithine decarboxylase was observed. The quantities of retinoic acid used (1.7 nmol and 3.4 nmol) have been shown effective at inhibiting 12-0-tetradecanoyl phorbol-13 acetate induced ornithine decarboxylase. The results show that these concentrations of topical retinoic acid applied either before or immediately following UVB irradiation reduces the UVB induction of epidermal ornithine decarboxylase. The effect of retinoic acid in these regimens on UVB-induced skin carcinogenesis is currently under study.

  7. Activation of RAS/ERK alone is insufficient to inhibit RXRα function and deplete retinoic acid in hepatocytes

    SciTech Connect

    Wang, Ai-Guo Song, Ya-Nan; Chen, Jun; Li, Hui-Ling; Dong, Jian-Yi; Cui, Hai-Peng; Yao, Liang; Li, Xue-Feng; Gao, Wen-Ting; Qiu, Ze-Wen; Wang, Fu-Jin; Wang, Jing-Yu

    2014-09-26

    Highlights: • The activation of RAS/ERK is insufficient to inhibit RXRα function and deplete RA. • The retinoid metabolism-related genes are down-regulated by ras oncogene. • The atRA has no effect on preventing hepatic tumorigenesis or curing the developed hepatic nodules. - Abstract: Activation of RAS/ERK signaling pathway, depletion of retinoid, and phosphorylation of retinoid X receptor alpha (RXRα) are frequent events found in liver tumors and thought to play important roles in hepatic tumorigenesis. However, the relationships among them still remained to be elucidated. By exploring the transgenic mouse model of hepatic tumorigenesis induced by liver-specific expression of H-ras12V oncogene, the activation of RAS/ERK, the mRNA expression levels of retinoid metabolism-related genes, the contents of retinoid metabolites, and phosphorylation of RXRα were determined. RAS/ERK signaling pathway was gradually and significantly activated in hepatic tumor adjacent normal liver tissues (P) and hepatic tumor tissues (T) of H-ras12V transgenic mice compared with normal liver tissues (Wt) of wild type mice. On the contrary, the mRNA expression levels of retinoid metabolism-related genes were significantly reduced in P and T compared with Wt. Interestingly, the retinoid metabolites 9-cis-retinoic acid (9cRA) and all-trans-retinoic acid (atRA), the well known ligands for nuclear transcription factor RXR and retinoic acid receptor (RAR), were significantly decreased only in T compared with Wt and P, although the oxidized polar metabolite of atRA, 4-keto-all-trans-retinoic-acid (4-keto-RA) was significantly decreased in both P and T compared with Wt. To our surprise, the functions of RXRα were significantly blocked only in T compared with Wt and P. Namely, the total protein levels of RXRα were significantly reduced and the phosphorylation levels of RXRα were significantly increased only in T compared with Wt and P. Treatment of H-ras12V transgenic mice at 5-week

  8. Retinoic Acid-mediated Nuclear Receptor Activation and Hepatocyte Proliferation

    PubMed Central

    Bushue, Nathan; Wan, Yu-Jui Yvonne

    2016-01-01

    Due to their well-known differentiation and apoptosis-inducing abilities, retinoic acid (RA) and its analogs have strong anti-cancer efficacy in human cancers. However, in vivo RA is a liver mitogen. While speculation has persisted that RA-mediated signaling is likely involved in hepatocyte proliferation during liver regeneration, direct evidence is still required. Findings in support of this proposition include observations that a release of retinyl palmitate (the precursor of RA) occurs in liver stellate cells following liver injury. Nevertheless, the biological action of this released vitamin A is virtually unknown. More likely is that the released vitamin A is converted to RA, the biological form, and then bound to a specific receptor (retinoid x receptor; RXRα), which is most abundantly expressed in the liver. Considering the mitogenic effects of RA, the RA-activated RXRα would likely then influence hepatocyte proliferation and liver tissue repair. At present, the mechanism by which RA stimulates hepatocyte proliferation is largely unknown. This review summarizes the activation of nuclear receptors (peroxisome proliferator activated receptor-α, pregnane x receptor, constitutive androstane receptor, and farnesoid x receptor) in an RXRα dependent manner to induce hepatocyte proliferation, providing a link between RA and its proliferative role.

  9. All-trans retinoic acid negatively regulates cytotoxic activities of nature killer cell line 92

    SciTech Connect

    Li Ang . E-mail: liang3829@sina.com.cn; He Meilan; Wang Hui; Qiao Bin; Chen Ping; Gu Hua; Zhang Mengjie; He Shengxiang

    2007-01-05

    NK cells are key components of innate immune systems and their activities are regulated by cytokines and hormones. All-trans retinoic acid (ATRA), as a metabolite of vitamin A and an immunomodulatory hormone, plays an important role in regulating immune responses. In the present study, we investigated the effect of ATRA on human NK cell line NK92. We found that ATRA dose-dependently suppressed cytotoxic activities of NK92 cells without affecting their proliferation. To explore the mechanisms underlying the ATRA influence on NK92 cells, we examined the production of cytokines (TNF-{alpha}, IFN-{gamma}), gene expression of cytotoxic-associated molecules (perforin, granzyme B, nature killer receptors (NCRs), and NKG2D), and the activation of NF-{kappa}B pathways related with immune response. Our results demonstrated that ATRA suppressed NF-{kappa}B activity and prevented I{kappa}B{alpha} degradation in a dose-dependent way, inhibited IFN-{gamma} production and gene expression of granzyme B and NKp46. Our findings suggest that ATRA is a negative regulator of NK92 cell activation and may act as a potential regulator of anti-inflammatory functions in vivo.

  10. Redox control of retinoic acid receptor activity: a novel mechanism for retinoic acid resistance in melanoma cells.

    PubMed

    Demary, K; Wong, L; Liou, J S; Faller, D V; Spanjaard, R A

    2001-06-01

    Retinoic acid (RA) slows growth and induces differentiation of tumor cells through activation of RA receptors (RARs). However, melanoma cell lines display highly variable responsiveness to RA, which is a poorly understood phenomenon. By using Northern and Western blot analyses, we show that RA-resistant A375 and RA-responsive S91 melanoma cells express comparable levels of major components of RAR-signaling pathways. However, A375 cells have substantially higher intracellular reactive oxygen species (ROS) levels than S91 cells. Lowering ROS levels in A375 cells through hypoxic culture conditions restores RAR-dependent trans-activity, which could be further enhanced by addition of the antioxidant N-acetyl-cysteine. Hypoxia also enhances RAR activity in the moderately RA-responsive C32 cells, which have intermediate ROS levels. Conversely, increasing oxidative stress in highly RA-responsive S91 and B16 cells, which have low ROS levels, by treatment with H(2)O(2) impairs RAR activity. Consistent with these observations, RA more potently inhibited the proliferation of hypoxic A375 cells than that of normoxic cells. Oxidative states diminish, whereas reducing conditions enhance, DNA binding of retinoid X receptor/RAR heterodimers in vitro, providing a molecular basis for the observed inverse correlation between RAR activity and ROS levels. The redox state of melanoma cells provides a novel, epigenetic control mechanism of RAR activity and RA resistance. PMID:11356710

  11. The Retinoic Acid Receptor-a Mediates Human T-Cell Activation and Th2 Cytokine Production

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We have recently demonstrated that all-trans-retinoic acid (ATRA) and 9-cis-retinoic acid (9-cis RA) promote IL-4, IL-5 and IL-13 synthesis, while decreasing IFN-g and TNF-a expression by activated human T cells and reducing the synthesis of IL-12p70 from accessory cells. Here, we have demonstrated ...

  12. IL-1 signaling modulates STAT activation to antagonize retinoic acid signaling and control Th17–iTreg balance

    PubMed Central

    Basu, Rajatava; Whitley, Sarah K.; Bhaumik, Suniti; Zindl, Carlene L.; Schoeb, Trenton R.; Benveniste, Etty N.; Pear, Warren S.; Hatton, Robin D.; Weaver, Casey T.

    2016-01-01

    Interleukin 17 (IL-17)-producing helper (TH17) and inducible regulatory CD4+ T (iTreg) cells emerge from an overlapping developmental program. In the intestines, the vitamin A metabolite retinoic acid (RA) is produced at steady state and acts as an important cofactor to induce iTreg cell development while potently inhibiting TH17 development. Here, we found that IL-1 was required to fully override RA-mediated Foxp3 expression and induce protective TH17 responses. Through induction of an NF-κB-dependent repression of SOCS3 expression, IL-1 increased the amplitude and duration of STAT3 phosphorylation induced by TH17-polarizing cytokines, leading to an altered balance of STAT3–STAT5 binding to shared consensus sequences in developing T cells. Thus, IL-1 signaling differentially modulated STAT activation downstream of cytokine receptors to control TH17–iTreg developmental fate. PMID:25642823

  13. All-trans retinoic acid modulates mitogen-activated protein kinase pathway activation in human scleral fibroblasts through retinoic acid receptor beta

    PubMed Central

    Huo, Lijun; Cui, Dongmei; Yang, Xiao; Gao, Zhenya; Trier, Klaus

    2013-01-01

    Purpose All-trans retinoic acid (ATRA) is known to inhibit the proliferation of human scleral fibroblasts (HSFs) and to modulate the scleral intercellular matrix composition, and may therefore serve as a mediator for controlling eye growth. Cell proliferation is regulated by the mitogen-activated protein kinase (MAPK) pathway. The aim of the current study was to investigate whether changed activation of the MAPK pathway could be involved in the response of HSFs exposed to ATRA. Methods HSFs were cultured in Dulbecco Modified Eagle's Medium/F12 (DMEM/F12) and exposed to 1 μmol/l ATRA for 10 min, 30 min, 1 h, 8 h, or 24 h. The activation of extracellular signal-regulated kinase (ERK 1/2), p38, and c-Jun N-terminal kinase (JNK) in HSFs was assessed with western blot analysis and immunocytofluorescence. Results After exposure to ATRA for 24 h, the HSFs appeared shrunken and thinner than the control cells. The intercellular spaces were wider, and the HSFs appeared less numerous than in the control culture. Western blot showed decreased activation of ERK 1/2 in the HSFs from 30 min (p=0.01) to 24 h (p<0.01) after the start of exposure to ATRA, and increased activation of the JNK protein from 10 to 30 min (p<0.01) after the start of exposure to ATRA. Indirect immunofluorescence confirmed changes in activation of ERK 1/2 and JNK in HSFs exposed to ATRA. No change in activation of p38 in HSFs was observed after exposure to ATRA. Pretreatment of the HSFs with LE135, an antagonist of retinoic acid receptor beta (RARβ), abolished the ATRA-induced changes inactivation of ERK 1/2 and JNK. Conclusions ATRA inhibits HSF proliferation by a mechanism associated with modulation of ERK 1/2 and JNK activation and depends on stimulation of retinoic acid receptor beta. PMID:23946634

  14. Effect of retinoic acid and vitamin D3 on osteoblast differentiation and activity in aging.

    PubMed

    Bosetti, Michela; Sabbatini, Maurizio; Calarco, Anna; Borrone, Alessia; Peluso, Gianfranco; Cannas, Mario

    2016-01-01

    Several studies have evidenced that in aging, osteoblast functional activity is impaired: osteoblast proliferation is slower and matrix deposition is less efficient. Because peroxisome-proliferator-activated receptor γ2 (PPARγ2) and fatty acids are important inhibitory signals in osteoblast development, we have investigated in human primary osteoblasts obtained from patients of different ages, the influence of retinoic acid and calcitriol on enzymes involved in differentiative (PPARγ2, β-catenin, and insulin-like growth factor 1) and metabolic (carnitine palmitoyltransferase 1) intracellular pathways, and on transglutaminase 2, as enzyme fundamental for stabilizing the newly deposited extracellular matrix in bone. Retinoic acid and calcitriol influenced, respectively, proliferation and differentiation of osteoblasts, and an increase in PPARγ2 expression was observed following retinoic acid administration, whereas a decrease was observed following calcitriol administration. Aging widely influenced all parameters analyzed (the proliferation, differentiation, and new matrix deposition are significantly reduced in aged osteoblasts), with the exception of PPARγ2, which we found to be constitutively overexpressed and not modulated by retinoic acid or calcitriol administration. Our findings show the impaired ability of aged osteoblasts to perform adequate functional response and draw attention to the therapeutic approaches for bone healing in elderly patients. PMID:25691285

  15. Retinol metabolism in LLC-PK1 Cells. Characterization of retinoic acid synthesis by an established mammalian cell line.

    PubMed

    Napoli, J L

    1986-10-15

    Specific assays, based on gas chromatography-mass spectrometry and high-performance liquid chromatography, were used to quantify the conversion of retinol and retinal into retinoic acid by the pig kidney cell line LLC-PK1. Retinoic acid synthesis was linear for 2-4 h as well as with graded amounts of either substrate to at least 50 microM. Retinoic acid concentrations increased through 6-8 h, but decreased thereafter because of substrate depletion (t1/2 of retinol = 13 h) and product metabolism (1/2 = 2.3 h). Retinoic acid metabolism was accelerated by treating cells with 100 nM retinoic acid for 10 h (t1/2 = 1.7 h) and was inhibited by the antimycotic imidazole ketoconazole. Feedback inhibition was not indicated since retinoic acid up to 100 nM did not inhibit its own synthesis. Retinol dehydrogenation was rate-limiting. The reduction and dehydrogenation of retinal were 4-8-fold and 30-60-fold faster, respectively. Greater than 95% of retinol was converted into metabolites other than retinoic acid, whereas the major metabolite of retinal was retinoic acid. The synthetic retinoid 13-cis-N-ethylretinamide inhibited retinoic acid synthesis, but 4-hydroxylphenylretinamide did not. 4'-(9-Acridinylamino)methanesulfon-m-anisidide, an inhibitor of aldehyde oxidase, and ethanol did not inhibit retinoic acid synthesis. 4-Methylpyrazole was a weak inhibitor: disulfiram was a potent inhibitor. These data indicate that retinol dehydrogenase is a sulfhydryl group-dependent enzyme, distinct from ethanol dehydrogenase. Homogenates of LLC-PK1 cells converted retinol into retinoic acid and retinyl palmitate and hydrolyzed retinyl palmitate. This report suggests that substrate availability, relative to enzyme activity/amount, is a primary determinant of the rate of retinoic acid synthesis, identifies inhibitors of retinoic acid synthesis, and places retinoic acid synthesis into perspective with several other known pathways of retinoid metabolism. PMID:3759984

  16. Molecular recognition of CYP26A1 binding pockets and structure-activity relationship studies for design of potent and selective retinoic acid metabolism blocking agents.

    PubMed

    Sun, Bin; Song, Shuai; Hao, Chen-Zhou; Huang, Wan-Xu; Liu, Chun-Chi; Xie, Hong-Lei; Lin, Bin; Cheng, Mao-Sheng; Zhao, Dong-Mei

    2015-03-01

    All-trans-retinoic acid (ATRA), the biologically most active metabolite of vitamin A, plays a major role in the regulation of cellular differentiation and proliferation, and it is also an important pharmacological agent particularly used in the treatment of cancer, skin, neurodegenerative and autoimmune diseases. However, ATRA is very easy to be metabolized into 4-hydroxyl-RA in vivo by CYP26A1, an inducible cytochrome P450 enzyme, eventually into more polar metabolites. Therefore, it is vital to develop specific retinoic acid metabolism blocking agents (RAMBAs) to inhibit the metabolic enzyme CYP26A1 in the treatment of relevant diseases aforementioned. In this study, CYP26A1 and its interactions with retinoic acid-competitive metabolism blocking agents were investigated by a combined ligand- and structure-based approach. First, since the crystal structure of CYP26A1 protein has not been determined, we constructed the 3D structure of CYP26A1 using homology modeling. In order to achieve a deeper insight into the mode of action of RAMBAs in the active site, the molecular superimposition model and the common feature pharmacophore model were constructed, and molecular docking was performed. The molecular superimposition model is composed of three features: the main chain groups, side chain groups, and azole groups. The common feature pharmacophore model consists of five chemical features: four hydrophobic groups and one hydrogen acceptor (HHHHA). The results of molecular docking show that the characteristic groups of RAMBAs were mapped into three different active pockets, respectively. A structure-activity relationship (SAR) was obtained by a combination of the molecular superimposition and docking results with the pharmacophore model. This study gives more insight into the interaction model inside the CYP26A1 active site and provides guidance for the design of more potent and possibly more selective RAMBAs. PMID:25541526

  17. Thyroid hormone activation of retinoic acid synthesis in hypothalamic tanycytes

    PubMed Central

    Stoney, Patrick N.; Helfer, Gisela; Rodrigues, Diana; Morgan, Peter J.

    2015-01-01

    Thyroid hormone (TH) is essential for adult brain function and its actions include several key roles in the hypothalamus. Although TH controls gene expression via specific TH receptors of the nuclear receptor class, surprisingly few genes have been demonstrated to be directly regulated by TH in the hypothalamus, or the adult brain as a whole. This study explored the rapid induction by TH of retinaldehyde dehydrogenase 1 (Raldh1), encoding a retinoic acid (RA)‐synthesizing enzyme, as a gene specifically expressed in hypothalamic tanycytes, cells that mediate a number of actions of TH in the hypothalamus. The resulting increase in RA may then regulate gene expression via the RA receptors, also of the nuclear receptor class. In vivo exposure of the rat to TH led to a significant and rapid increase in hypothalamic Raldh1 within 4 hours. That this may lead to an in vivo increase in RA is suggested by the later induction by TH of the RA‐responsive gene Cyp26b1. To explore the actions of RA in the hypothalamus as a potential mediator of TH control of gene regulation, an ex vivo hypothalamic rat slice culture method was developed in which the Raldh1‐expressing tanycytes were maintained. These slice cultures confirmed that TH did not act on genes regulating energy balance but could induce Raldh1. RA has the potential to upregulate expression of genes involved in growth and appetite, Ghrh and Agrp. This regulation is acutely sensitive to epigenetic changes, as has been shown for TH action in vivo. These results indicate that sequential triggering of two nuclear receptor signalling systems has the capability to mediate some of the functions of TH in the hypothalamus. GLIA 2016;64:425–439 PMID:26527258

  18. Retinoic acid activates human inducible nitric oxide synthase gene through binding of RAR{alpha}/RXR{alpha} heterodimer to a novel retinoic acid response element in the promoter

    SciTech Connect

    Zou Fang; Liu Yan; Liu Li; Wu Kailang; Wei Wei; Zhu Ying . E-mail: yingzhu@whu.edu.cn; Wu Jianguo . E-mail: wu9988@vip.sina.com

    2007-04-06

    Human inducible nitric oxide synthase (hiNOS) catalyzes nitric oxide (NO) which has a significant effect on tumor suppression and cancer therapy. Here we revealed the detailed molecular mechanism involved in the regulation of hiNOS expression induced by retinoic acid (RA). We showed that RAR{alpha}/RXR{alpha} heterodimer was important in hiNOS promoter activation, hiNOS protein expression, and NO production. Serial deletion and site-directed mutation analysis revealed two half-sites of retinoic acid response element (RARE) spaced by 5 bp located at -172 to -156 in the hiNOS promoter. EMSA and ChIP assays demonstrated that RAR{alpha}/RXR{alpha} directly bound to this RARE of hiNOS promoter. Our results suggested the identification of a novel RARE in the hiNOS promoter and the roles of the nuclear receptors (RAR{alpha}/RXR{alpha}) in the induction of hiNOS by RA.

  19. Improved In Vitro Antileukemic Activity of All-Trans Retinoic Acid Loaded in Cholesteryl Butyrate Solid Lipid Nanoparticles.

    PubMed

    Silva, Elton Luiz; Lima, Flávia Alves; Carneiro, Guilherme; Ramos Jonas Periera; Gomes, Dawidson Assis; de Souza-Fagundes, Elaine Maria; Ferreira, Lucas Antônio Miranda

    2016-02-01

    All-trans retinoic acid, a hydrophobic drug, has become one of the most successful examples of differentiation agents used for treatment of acute promyelocytic leukemia. On the other hand, histone deacetylase inhibitors, such as cholesteryl butyrate, present differentiating activity and.can potentiate action of drugs such as all-trans retinoic acid. Solid lipid nanoparticles represent a promising alternative for administration of hydrophobic drugs such as ATRA. This study aimed to develop, characterize, and evaluate the cytotoxicity of all-trans retinoic acid-loaded solid lipid nanoparticles for leukemia treatment. The influence of in situ formation of an ion pairing between all-trans retinoic acid and lipophilic amines on the characteristics of the particles (size, zeta potential, encapsulation efficiency) was evaluated. Cholesteryl butyrate, a butyric acid donor, was used as a component of the lipid matrix. In vitro activity on cell viability and distribution of cell cycle phases were evaluated for HL-60, Jurkat, and THP-1 cell lines. The encapsulation efficiency of all-trans retinoic acid in cholesteryl butyrate-solid lipid nanoparticles was significantly increased by the presence of the amine. Inhibition of cell viability by all-trans retinoic acid-loaded solid lipid nanoparticles was more pronounced than the free drug. Analysis of the distribution of cell cycle phases also showed increased activity for all-trans retinoic acid-loaded cholesteryl butyrate-solid lipid nanoparticles, with a clear increase in subdiploid DNA content. The ion pair formation in SLN containing cholesteryl butyrate can be explored as a simple and inexpensive strategy to improve the efficacy and bioavail-ability of ATRA in the treatment of the cancer and metabolic diseases in which this retinoid plays an important role. PMID:27433579

  20. Phytol metabolites are circulating dietary factors that activate the nuclear receptor RXR.

    PubMed Central

    Kitareewan, S; Burka, L T; Tomer, K B; Parker, C E; Deterding, L J; Stevens, R D; Forman, B M; Mais, D E; Heyman, R A; McMorris, T; Weinberger, C

    1996-01-01

    RXR is a nuclear receptor that plays a central role in cell signaling by pairing with a host of other receptors. Previously, 9-cis-retinoic acid (9cRA) was defined as a potent RXR activator. Here we describe a unique RXR effector identified from organic extracts of bovine serum by following RXR-dependent transcriptional activity. Structural analyses of material in active fractions pointed to the saturated diterpenoid phytanic acid, which induced RXR-dependent transcription at concentrations between 4 and 64 microM. Although 200 times more potent than phytanic acid, 9cRA was undetectable in equivalent amounts of extract and cannot be present at a concentration that could account for the activity. Phytanic acid, another phytol metabolite, was synthesized and stimulated RXR with a potency and efficacy similar to phytanic acid. These metabolites specifically displaced [3H]-9cRA from RXR with Ki values of 4 microM, indicating that their transcriptional effects are mediated by direct receptor interactions. Phytol metabolites are compelling candidates for physiological effectors, because their RXR binding affinities and activation potencies match their micromolar circulating concentrations. Given their exclusive dietary origin, these chlorophyll metabolites may represent essential nutrients that coordinate cellular metabolism through RXR-dependent signaling pathways. PMID:8856661

  1. Retinoic Acid Receptor α Mediates All-trans-retinoic Acid-induced Klf4 Gene Expression by Regulating Klf4 Promoter Activity in Vascular Smooth Muscle Cells*

    PubMed Central

    Shi, Jian-hong; Zheng, Bin; Chen, Si; Ma, Guo-yan; Wen, Jin-kun

    2012-01-01

    The transcription factor Krüppel-like factor 4 (KLF4) plays a critical role in vascular smooth muscle cell (VSMC) differentiation induced by all-trans-retinoic acid (ATRA). Although it has been demonstrated that ATRA stimulation augments both KLF4 protein and mRNA levels in VSMCs, the molecular mechanisms by which ATRA regulates Klf4 transcription are unknown. In this study, we examined the roles of ATRA-selective nuclear retinoic acid receptors (RARs) in the transcriptional regulation of Klf4. The introduction of small interfering RNA and an RAR antagonist demonstrated that RARα, but not RARβ or RARγ, mediated ATRA-induced Klf4 expression. A luciferase assay for the Klf4 promoter showed that three GC boxes in the proximal Klf4 promoter were indispensible for ATRA-induced Klf4 transcription and that RARα enhanced Klf4 promoter activity in a GC box-dependent manner. Furthermore, chromatin immunoprecipitation and oligonucleotide pulldown assays demonstrated that the transcription factors KLF4, Sp1, and YB1 directly bound to the GC boxes of the proximal Klf4 promoter. Upon RARα agonist stimulation, RARα was recruited to the Klf4 promoter through its interaction with KLF4, Sp1, and YB1 to form a transcriptional activation complex on the three GC boxes of the Klf4 promoter. These results suggest that RARα serves as an essential co-activator for ATRA signaling and that the recruitment of RARα to the KLF4-Sp1-YB1 complex, which leads to Klf4 expression in VSMCs, is independent of a retinoic acid response element. PMID:22337869

  2. Retinoic acid affects calcium signaling in adult molluscan neurons.

    PubMed

    Vesprini, Nicholas D; Dawson, Taylor F; Yuan, Ye; Bruce, Doug; Spencer, Gaynor E

    2015-01-01

    Retinoic acid, the active metabolite of vitamin A, is important for nervous system development, regeneration, as well as cognitive functions of the adult central nervous system. These central nervous system functions are all highly dependent on neuronal activity. Retinoic acid has previously been shown to induce changes in the firing properties and action potential waveforms of adult molluscan neurons in a dose- and isomer-dependent manner. In this study, we aimed to determine the cellular pathways by which retinoic acid might exert such effects, by testing the involvement of pathways previously shown to be affected by retinoic acid. We demonstrated that the ability of all-trans retinoic acid (atRA) to induce electrophysiological changes in cultured molluscan neurons was not prevented by inhibitors of protein synthesis, protein kinase A or phospholipase C. However, we showed that atRA was capable of rapidly reducing intracellular calcium levels in the same dose- and isomer-dependent manner as shown previously for changes in neuronal firing. Moreover, we also demonstrated that the transmembrane ion flux through voltage-gated calcium channels was rapidly modulated by retinoic acid. In particular, the peak current density was reduced and the inactivation rate was increased in the presence of atRA, over a similar time course as the changes in cell firing and reductions in intracellular calcium. These studies provide further evidence for the ability of atRA to induce rapid effects in mature neurons. PMID:25343782

  3. Biologically Active Metabolites Synthesized by Microalgae

    PubMed Central

    de Morais, Michele Greque; Vaz, Bruna da Silva; de Morais, Etiele Greque; Costa, Jorge Alberto Vieira

    2015-01-01

    Microalgae are microorganisms that have different morphological, physiological, and genetic traits that confer the ability to produce different biologically active metabolites. Microalgal biotechnology has become a subject of study for various fields, due to the varied bioproducts that can be obtained from these microorganisms. When microalgal cultivation processes are better understood, microalgae can become an environmentally friendly and economically viable source of compounds of interest, because production can be optimized in a controlled culture. The bioactive compounds derived from microalgae have anti-inflammatory, antimicrobial, and antioxidant activities, among others. Furthermore, these microorganisms have the ability to promote health and reduce the risk of the development of degenerative diseases. In this context, the aim of this review is to discuss bioactive metabolites produced by microalgae for possible applications in the life sciences. PMID:26339647

  4. Prospective teratology of retinoic acid metabolic blocking agents (RAMBAs) and loss of CYP26 activity.

    PubMed

    McCaffery, P; Simons, C

    2007-01-01

    All-trans retinoic acid (atRA) is the transcriptionally active product of vitamin A and induces gene expression via specific receptors at nM concentrations. Essential enzymes that regulate the local levels of atRA are the CYP26 members of the cytochrome P450 family, which catabolize atRA. Compounds that have been designed to inhibit these enzymes are known as Retinoic Acid Metabolic Blocking Agents (RAMBAs). Treatment with these compounds will raise endogenous atRA levels and may be therapeutic for the treatment of diseases that respond to high atRA concentrations, including several types of cancer as well as skin conditions such as psoriasis and acne. This review describes the mechanism of action of the RAMBAs and discusses the potential side effects of these compounds. atRA is highly teratogenic and the potential teratogenicity of the RAMBAs is described by comparison with the abnormalities resulting from null mutation of individual CYP26 genes. The possible effects of RAMBAs on the adult brain are also described that have the potential for harm but, in the right circumstances, may also be beneficial. PMID:17979744

  5. Antimycobacterial activity of lichen metabolites in vitro.

    PubMed

    Ingólfsdóttir, K; Chung, G A; Skúlason, V G; Gissurarson, S R; Vilhelmsdóttir, M

    1998-04-01

    Several compounds, whose structures represent the most common chemical classes of lichen metabolites, were screened for in vitro activity against Mycobacterium aurum, a non-pathogenic organism with a similar sensitivity profile to M. tuberculosis. Of the compounds tested, usnic acid from Cladonia arbuscula exhibited the highest activity with an MIC value of 32 microg/ml. Atranorin and lobaric acid, both isolated from Stereocaulon alpinum, salazinic acid from Parmelia saxatilis and protolichesterinic acid from Cetraria islandica all showed MIC values >/=125 microg/ml. PMID:9795033

  6. The Retinoic Acid Receptor-α mediates human T-cell activation and Th2 cytokine and chemokine production

    PubMed Central

    Dawson, Harry D; Collins, Gary; Pyle, Robert; Key, Michael; Taub, Dennis D

    2008-01-01

    Background We have recently demonstrated that all-trans-retinoic acid (ATRA) and 9-cis-retinoic acid (9-cis RA) promote IL-4, IL-5 and IL-13 synthesis, while decreasing IFN-γ and TNF-α expression by activated human T cells and reduces the synthesis of IL-12p70 from accessory cells. Here, we have demonstrated that the observed effects using ATRA and 9-cis RA are shared with the clinically useful RAR ligand, 13-cis retinoic acid (13-cis RA), and the retinoic acid receptor-α (RAR-α)-selective agonist, AM580 but not with the RAR-β/γ ligand, 4-hydroxyphenylretinamide (4-HPR). Results The increase in type 2 cytokine production by these retinoids correlated with the expression of the T cell activation markers, CD69 and CD38. The RAR-α-selective agonist, AM580 recapitulated all of the T cell activation and type 2 cytokine-inducing effects of ATRA and 9-cis-RA, while the RAR-α-selective antagonist, RO 41–5253, inhibited these effects. Conclusion These results strongly support a role for RAR-α engagement in the regulation of genes and proteins involved with human T cell activation and type 2 cytokine production. PMID:18416830

  7. Evaluation of immunomodulatory and antitumor activity of all trans retinoic acid (ATRA) in solid tumor bearing mice.

    PubMed

    Siddikuzzaman; Berlin, Grace V M

    2013-02-01

    Natural or synthetic agents can modify the immune system and, in some cases, impart a therapeutic benefit. Cancer, a disease of uncontrolled growth and spread of abnormal cells, is a major cause of death. The Vitamin A metabolite all-trans retinoic acid (ATRA) and its other active derivatives are potent modulators of cell growth and differentiation, and because it has an influence on cancer, it can be used as a chemotherapeutic and -preventive agent. To evaluate the immunomodulatory activity of ATRA, the impact of treatment on various parameters, e.g. delayed-type hypersensitivity (DTH), bone marrow cellularity, hematology, and levels of esterase-positive cells, was assessed in Balb/c mice. To evaluate antitumor effects of ATRA, tumor volume and host survival rate were monitored in B16F10 melanoma cell-injected mice. The results showed that administration of ATRA (0.60 mg/kg/dose, IP) caused a decrease in DTH (footpad thickness) in response to challenge with sheep red blood cells (SRBC) in SRBC-sensitized hosts. ATRA also caused increases in WBC counts and bone marrow cell numbers. In tumor-inoculated mice, ATRA caused tumor growth suppression and gave rise to a heightened survival rate. It was also found that ATRA had differential effects on serum levels of reduced glutathione (GSH) and nitric oxide (NO) was reduced in serum. Based on these results, we conclude that ATRA has a potent immunomodulatory potential but also could significantly impact upon solid tumor growth and prolong host survival. PMID:22900644

  8. Nutrigenomic regulation of adipose tissue development - role of retinoic acid: A review.

    PubMed

    Wang, Bo; Yang, Qiyuan; Harris, Corrine L; Nelson, Mark L; Busboom, Jan R; Zhu, Mei-Jun; Du, Min

    2016-10-01

    To improve the efficiency of animal production, livestock have been extensively selected or managed to reduce fat accumulation and increase lean growth, which reduces intramuscular or marbling fat content. To enhance marbling, a better understanding of the mechanisms regulating adipogenesis is needed. Vitamin A has recently been shown to have a profound impact on all stages of adipogenesis. Retinoic acid, an active metabolite of vitamin A, activates both retinoic acid receptors (RAR) and retinoid X receptors (RXR), inducing epigenetic changes in key regulatory genes governing adipogenesis. Additionally, Vitamin D and folates interact with the retinoic acid receptors to regulate adipogenesis. In this review, we discuss nutritional regulation of adipogenesis, focusing on retinoic acid and its impact on epigenetic modifications of key adipogenic genes. PMID:27086067

  9. Identification of COUP-TFII Orphan Nuclear Receptor as a Retinoic Acid-Activated Receptor

    SciTech Connect

    Kruse, Schoen W; Suino-Powell, Kelly; Zhou, X Edward; Kretschman, Jennifer E; Reynolds, Ross; Vonrhein, Clemens; Xu, Yong; Wang, Liliang; Tsai, Sophia Y; Tsai, Ming-Jer; Xu, H Eric

    2010-01-12

    The chicken ovalbumin upstream promoter-transcription factors (COUP-TFI and II) make up the most conserved subfamily of nuclear receptors that play key roles in angiogenesis, neuronal development, organogenesis, cell fate determination, and metabolic homeostasis. Although the biological functions of COUP-TFs have been studied extensively, little is known of their structural features or aspects of ligand regulation. Here we report the ligand-free 1.48 {angstrom} crystal structure of the human COUP-TFII ligand-binding domain. The structure reveals an autorepressed conformation of the receptor, where helix {alpha}10 is bent into the ligand-binding pocket and the activation function-2 helix is folded into the cofactor binding site, thus preventing the recruitment of coactivators. In contrast, in multiple cell lines, COUP-TFII exhibits constitutive transcriptional activity, which can be further potentiated by nuclear receptor coactivators. Mutations designed to disrupt cofactor binding, dimerization, and ligand binding, substantially reduce the COUP-TFII transcriptional activity. Importantly, retinoid acids are able to promote COUP-TFII to recruit coactivators and activate a COUP-TF reporter construct. Although the concentration needed is higher than the physiological levels of retinoic acids, these findings demonstrate that COUP-TFII is a ligand-regulated nuclear receptor, in which ligands activate the receptor by releasing it from the autorepressed conformation.

  10. Fungistatic activity of all-trans retinoic acid against Aspergillus fumigatus and Candida albicans

    PubMed Central

    Campione, Elena; Gaziano, Roberta; Marino, Daniele; Orlandi, Augusto

    2016-01-01

    Purpose Fungal infections are a major complication in hematologic and neoplastic patients causing severe morbidity and mortality. Aspergillus fumigatus and Candida albicans are among the most invasive opportunistic pathogens in immunocompromised patients, and classic antifungal drugs are frequently unsuccessful in these patients. Recent reports hypothesize that the antifungal efficacy of all-trans retinoic acid (ATRA) is mainly related to its strong capacity to stimulate monocyte-mediated immunity, but no consideration was given to its potential direct fungistatic activity. Moreover, ATRA offers the opportunity for systemic therapy. Methods and results We investigated the efficacy of ATRA at different concentrations for its antifungal activity against opportunistic A. fumigatus and C. albicans obtained from clinical samples according to standard protocols. A fungistatic activity of ATRA on A. fumigatus and C. albicans at 0.5–1 mM concentration was documented up to 7 days. Conclusion This is the first evidence of a direct and strong fungistatic activity of ATRA against A. fumigatus and C. albicans. The potential adjuvant therapeutic application of ATRA might be useful in the treatment and/or prevention of systemic mycoses in immunocompromised patients. The discovery of a direct fungistatic activity, in association with its reported immunomodulatory properties, makes ATRA an excellent candidate for new combined antifungal strategies for systemic mycoses in immunocompromised and cancer patients. PMID:27199548

  11. Metabolite

    MedlinePlus

    A metabolite is any substance produced during metabolism (digestion or other bodily chemical processes). The term metabolite may also refer to the product that remains after a drug is broken down (metabolized) by the body.

  12. Effect of retinoic acid on transglutaminase and ornithine decarboxylase activities during liver regeneration.

    PubMed

    Ohtake, Yosuke; Maruko, Akiko; Ohishi, Nao; Kawaguchi, Masasumi; Satoh, Tetsuharu; Ohkubo, Yasuhito

    2008-04-01

    Liver regeneration is regulated by several factors, including growth factors, cytokines, and post-translational modifications of several proteins. It is suggested that transglutaminase 2 (TG2) and ornithine decarboxylase (ODC) are involved in liver regeneration. To investigate the role of TG2 and ODC activities in regenerating liver, we used retinoic acid (RA), an inducer of TG2 and a suppressor of ODC. Regenerating rat liver was prepared by 70% partial hepatectomy (PH). Rats were sacrificed at 1, 2, 3, 4, and 6 days after surgery. RA was intraperitoneally injected immediately after PH. TG2 and ODC activities and products (epsilon-(gamma-glutamyl) lysine isopeptide (Gln-Lys) and polyamines, respectively) were examined at the indicated times. In RA-treated rat, DNA synthesis and ODC activity declined and the peak shifted to 2 days after PH, whereas TG2 activity increased at 1 day after PH. At that time, protein-polyamine, especially the protein-spermidine (SPD) bond, transiently decreased, whereas the formation of the Gln-Lys bond increased after PH. These results suggested that in regenerating liver, enhanced the formation of Gln-Lys bonds catalyzed by TG2 led to reduced DNA synthesis, whereas when ODC produced newly synthesized SPD, the inhibition of Gln-Lys bond production by the preferential formation of protein-SPD bonds led to an increase in DNA synthesis. PMID:18008394

  13. All-trans retinoic acid triggered antimicrobial activity against Mycobacterium tuberculosis is dependent on NPC2

    PubMed Central

    Inkeles, Megan S.; De Leon, Avelino; Pellegrini, Matteo; Krutzik, Stephan R.; Liu, Philip T.

    2014-01-01

    A role for vitamin A in host defense against Mycobacterium tuberculosis has been suggested through epidemiological and in vitro studies; however, the mechanism is unclear. Here, we demonstrate that vitamin A-triggered antimicrobial activity against M. tuberculosis requires expression of Niemann-Pick disease type C2 (NPC2). Comparison of monocytes stimulated with all-trans retinoic acid (ATRA) or 1,25-dihydroxyvitamin D3 (1,25D3), the biologically active forms of vitamin A and vitamin D, respectively, indicates that ATRA and 1,25D3 induce mechanistically distinct antimicrobial activities. Stimulation of primary human monocytes with ATRA did not result in expression of the antimicrobial peptide cathelicidin, which is required for 1,25D3 antimicrobial activity. In contrast, ATRA triggers a reduction in the total cellular cholesterol concentration, whereas 1,25D3 did not. Blocking ATRA-induced cellular cholesterol reduction inhibits antimicrobial activity as well. Bioinformatic analysis of ATRA and 1,25D3 induced gene profiles suggests Niemann-Pick disease type C2 (NPC2) is a key gene in ATRA-induced cholesterol regulation. Knockdown experiments demonstrate that ATRA-mediated decrease of total cellular cholesterol content and increase in lysosomal acidification are both dependent upon expression of NPC2. Expression of NPC2 was lower in caseous tuberculosis granulomas and M. tuberculosis-infected monocytes compared to normal lung and uninfected cells, respectively. Loss of NPC2 expression ablated ATRA-induced antimicrobial activity. Taken together, these results suggest that the vitamin A-mediated antimicrobial mechanism against M. tuberculosis requires NPC2-dependent expression and function, indicating a key role for cellular cholesterol regulation in the innate immune response. PMID:24501203

  14. Immunohistochemical analysis of retinoic acid receptor-alpha in human breast tumors: retinoic acid receptor-alpha expression correlates with proliferative activity.

    PubMed Central

    van der Leede, B. M.; Geertzema, J.; Vroom, T. M.; Décimo, D.; Lutz, Y.; van der Saag, P. T.; van der Burg, B.

    1996-01-01

    Retinoids are known to prevent mammary carcinogenesis in rodents and inhibit the growth of human breast cancer cells in vitro. Previously we demonstrated that retinoid inhibition of proliferation of human breast cancer cell lines is largely mediated by retinoic acid receptor (RAR)-alpha. In this study we describe for the first time the histological distribution of RAR-alpha in 33 breast lesion specimens as determined by immunostaining with RAR-alpha antibody. Nuclear staining was observed in tumor tissue and normal portions of the breast samples. Connective tissue exhibited relative uniform staining, whereas a wide range of RAR-alpha expression was found in the epithelial tumor cells. RAR-alpha protein was expressed at significantly higher levels in tumors with greater proliferative activity as determined by immunostaining with Ki-67 antibody. This suggests that RAR-alpha expression may be altered with tumor progression. Although a positive correlation between RAR-alpha mRNA levels and estrogen receptor status of breast tumors has previously been documented, we did not find such a relationship at the protein level. As RAR-alpha plays a major role in retinoid-mediated growth inhibition of human breast cancer cell in vitro, our findings suggest that patients with highly proliferating tumors could be responsive to retinoid independently of their responsiveness to (anti)-estrogens. Images Figure 1 Figure 2 PMID:8669476

  15. Spatiotemporal manipulation of retinoic acid activity in zebrafish hindbrain development via photo-isomerization.

    PubMed

    Xu, Lijun; Feng, Zhiping; Sinha, Deepak; Ducos, Bertrand; Ebenstein, Yuval; Tadmor, Arbel D; Gauron, Carole; Le Saux, Thomas; Lin, Shuo; Weiss, Shimon; Vriz, Sophie; Jullien, Ludovic; Bensimon, David

    2012-09-01

    All-trans retinoic acid (RA) is a key player in many developmental pathways. Most methods used to study its effects in development involve continuous all-trans RA activation by incubation in a solution of all-trans RA or by implanting all-trans RA-soaked beads at desired locations in the embryo. Here we show that the UV-driven photo-isomerization of 13-cis RA to the trans-isomer (and vice versa) can be used to non-invasively and quantitatively control the concentration of all-trans RA in a developing embryo in time and space. This facilitates the global or local perturbation of developmental pathways with a pulse of all-trans RA of known concentration or its inactivation by UV illumination. In zebrafish embryos in which endogenous synthesis of all-trans RA is impaired, incubation for as little as 5 minutes in 1 nM all-trans RA (a pulse) or 5 nM 13-cis RA followed by 1-minute UV illumination is sufficient to rescue the development of the hindbrain if performed no later than bud stage. However, if subsequent to this all-trans RA pulse the embryo is illuminated (no later than bud stage) for 1 minute with UV light (to isomerize, i.e. deactivate, all-trans RA), the rescue of hindbrain development is impaired. This suggests that all-trans RA is sequestered in embryos that have been transiently exposed to it. Using 13-cis RA isomerization with UV light, we further show that local illumination at bud stage of the head region (but not the tail) is sufficient to rescue hindbrain formation in embryos whose all-trans RA synthetic pathway has been impaired. PMID:22874920

  16. ANALYSIS OF ARACHIDONIC ACID METABOLITE AND PLATELET ACTIVATING FACTOR PRODUCTION

    EPA Science Inventory

    Metabolites of arachidonic acid ("eicosanoids") and platelet activating factor are important bioactive lipids that may be involved in the pathobiological alterations in animals induced by pollutant exposure. nalysis of these substances in biological tissue and fluids is important...

  17. EMBO Retinoids 2011: mechanisms, biology and pathology of signaling by retinoic acid and retinoic acid receptors

    PubMed Central

    McKenna, Neil J.

    2012-01-01

    Retinoic acid (RA) is one of the principal active metabolites of vitamin A (retinol) which mediates a spectrum of critical physiological and developmental processes. Transcriptional regulation by RA is mediated primarily by members of the retinoic acid receptor (RAR) subfamily of the nuclear receptor (NR) superfamily of transcription factors. NRs bind specific genomic DNA sequence motifs and engage coregulators and components of the basal transcription machinery to effect transcriptional regulation at target gene promoters. Disruption of signaling by retinoic acid is thought to underlie the etiology of a number of inflammatory and neoplastic diseases including breast cancer and haematological malignancies. A meeting of international researchers in retinoid signaling was convened in Strasbourg in September 2011 under the auspices of the European Molecular Biology Organization (EMBO). Retinoids 2011 encompassed myriad mechanistic, biological and pathological aspects of these hormones and their cognate receptors, as well as setting these advances in the context of wider current questions on signaling by members of the NR superfamily. PMID:22438793

  18. DHEA metabolites activate estrogen receptors alpha and beta

    PubMed Central

    Michael Miller, Kristy K.; Al-Rayyan, Numan; Ivanova, Margarita M.; Mattingly, Kathleen A.; Ripp, Sharon L.; Klinge, Carolyn M.; Prough, Russell A.

    2012-01-01

    Dehydroepiandrosterone (DHEA) levels were reported to associate with increased breast cancer risk in postmenopausal women, but some carcinogen-induced rat mammary tumor studies question this claim. The purpose of this study was to determine how DHEA and its metabolites affect estrogen receptors α or β (ERα or ERβ) -regulated gene transcription and cell proliferation. In transiently transfected HEK-293 cells, androstenediol, DHEA, and DHEA-S activated ERα. In ERβ transfected HepG2 cells, androstenedione, DHEA, androstenediol, and 7-oxo DHEA stimulated reporter activity. ER antagonists ICI 182,780 (fulvestrant) and 4-hydroxytamoxifen, general P450 inhibitor miconazole, and aromatase inhibitor exemestane inhibited activation by DHEA or metabolites in transfected cells. ERβ-selective antagonist R,R-THC (R,R-cis-diethyl tetrahydrochrysene) inhibited DHEA and DHEA metabolite transcriptional activity in ERβ-transfected cells. Expression of endogenous estrogen-regulated genes: pS2, progesterone receptor, cathepsin D1, and nuclear respiratory factor-1 was increased by DHEA and its metabolites in an ER-subtype, gene, and cell-specific manner. DHEA metabolites, but not DHEA, competed with 17β-estradiol for ERα and ERβ binding and stimulated MCF-7 cell proliferation, demonstrating that DHEA metabolites interact directly with ERα and ERβ in vitro, modulating estrogen target genes in vivo. PMID:23123738

  19. Retinoic acid impairs estrogen signaling in breast cancer cells by interfering with activation of LSD1 via PKA.

    PubMed

    Ombra, Maria Neve; Di Santi, Annalisa; Abbondanza, Ciro; Migliaccio, Antimo; Avvedimento, Enrico Vittorio; Perillo, Bruno

    2013-05-01

    More than 70% of breast cancers in women require estrogens for cell proliferation and survival. 17β-estradiol (E2) effect on mammary target cells is almost exclusively mediated by its binding to the estrogen receptor-α (ERα) that joins chromatin where it assembles active transcription complexes. The proliferative and pro-survival action of estrogens is antagonized in most cases by retinoic acid (RA), even though the cognate retinoic acid receptor-α (RARα) cooperates with ERα on promoters of estrogen-responsive genes. We have examined at the molecular level the crosstalk between these nuclear receptors from the point of view of their control of cell growth and show here that RA reverts estrogen-stimulated transcription of the pivotal anti-apoptotic bcl-2 gene by preventing demethylation of dimethyl lysine 9 in histone H3 (HeK9me2). As we previously reported, this is obtained by means of E2-triggered activation of the lysine-specific demethylase 1 (LSD1), an enzyme that manages chromatin plasticity in order to allow specific movements of chromosomal regions within the nucleus. We find that E2 fuels LSD1 by inducing migration of the catalytic subunit of protein kinase A (PKA) into the nucleus, where it targets estrogen-responsive loci. RA rescues LSD1-dependent disappearance of H3K9me2 at bcl-2 regulatory regions upon the prevention of PKA assembly to the same sites. PMID:23507259

  20. Regulation of Retinoic Acid Inducible Gene-I (RIG-I) Activation by the Histone Deacetylase 6.

    PubMed

    Liu, Helene Minyi; Jiang, Fuguo; Loo, Yueh Ming; Hsu, ShuZhen; Hsiang, Tien-Ying; Marcotrigiano, Joseph; Gale, Michael

    2016-07-01

    Retinoic acid inducible gene-I (RIG-I) is a cytosolic pathogen recognition receptor that initiates the immune response against many RNA viruses. Upon RNA ligand binding, RIG-I undergoes a conformational change facilitating its homo-oligomerization and activation that results in its translocation from the cytosol to intracellular membranes to bind its signaling adaptor protein, mitochondrial antiviral-signaling protein (MAVS). Here we show that RIG-I activation is regulated by reversible acetylation. Acetyl-mimetic mutants of RIG-I do not form virus-induced homo-oligomers, revealing that acetyl-lysine residues of the RIG-I repressor domain prevent assembly to active homo-oligomers. During acute infection, deacetylation of RIG-I promotes its oligomerization upon ligand binding. We identify histone deacetylase 6 (HDAC6) as the deacetylase that promotes RIG-I activation and innate antiviral immunity to recognize and restrict RNA virus infection. PMID:27372014

  1. Metabolites and DNA adduct formation from flavoenzyme-activated porfiromycin.

    PubMed

    Pan, S S; Iracki, T

    1988-08-01

    Porfiromycin was reductively metabolized by NADPH cytochrome P-450 reductase and xanthine oxidase under anaerobic conditions. The production of metabolites varied with the pH and the contents of the reaction buffer. In Tris buffer, two major metabolites were produced at pH 7.5 and above, whereas one major metabolite was produced at pH 6.5. The three major metabolites were separated and isolated by HPLC. Identification by californium-252 plasma desorption mass spectrometry showed that the two major metabolites from pH 7.5 were (trans) and (cis)-forms of 7-amino-1-hydroxyl-2-methylaminomitosene and the major metabolite from pH 6.5 was 7-amino-2-methylaminomitosene. All three major metabolites showed substitutions at the C-1 position. DNA was alkylated readily by enzyme-activated porfiromycin. Digestion of porfiromycin-alkylated DNA by DNase, snake venom phosphodiesterase, and alkaline phosphatase resulted in an insoluble nuclease-resistant fraction and a soluble fraction. The nuclease-resistant fraction reflected a high content of cross-linked adducts. Upon HPLC analysis, the solubilized fraction contained two monofunctionally linked porfiromycin adducts and a possibly cross-linked dinucleotide. The major adduct was isolated by HPLC and identified by NMR, as N2-(2'-deoxyguanosyl)-7-amino-2-methylaminomitosene. The N2 position of deoxyguanosine appeared as the major monofunctional alkylating site for DNA alkylation by porfiromycin. Thus, mitomycin C and porfiromycin (which differs from mitomycin C only by the addition of a methyl group to the aziridine nitrogen) share the same enzymatic activating mechanism that leads to the formation of the same types of metabolites and the same specificity of DNA alkylation. PMID:3412325

  2. Activation of AMP-activated protein kinase by retinoic acid sensitizes hepatocellular carcinoma cells to apoptosis induced by sorafenib

    PubMed Central

    Ishijima, Naoki; Kanki, Keita; Shimizu, Hiroki; Shiota, Goshi

    2015-01-01

    To improve the outcome of cancer chemotherapy, strategies to enhance the efficacy of anticancer drugs are required. Sorafenib is the only drug to prolong overall survival of the patients with hepatocellular carcinoma (HCC), however, the outcome is still not satisfactory. Retinoids, vitamin A derivatives, have been known to exhibit inhibitory effects on various cancers including HCC. In this study, we investigated the effects of combined treatment using sorafenib and retinoids including all-trans retinoic acid (ATRA), NIK-333, and Am80 on HCC cells. Cell viability assays in six HCC cell lines, HepG2, PLC/PRF/5, HuH6, HLE, HLF, and Hep3B, revealed that 5 and 10 μM ATRA, concentrations that do not exert cytotoxic effects, enhanced the cytotoxicity of sorafenib, being much more effective than NIK-333 and Am80. We found that ATRA induced AMP-activated protein kinase activation, which was followed by reduced intracellular ATP level. Gene expression analysis revealed that ATRA decreased the expression of glycolytic genes such as GLUT-1 and LDHA. In the combination treatment using ATRA and sorafenib, increased apoptosis, followed by the activation of p38 MAPK and JNK, the upregulation and translocation of Bax to mitochondria, and the activation of caspase-3, was observed. Suppression of AMP-activated protein kinase by siRNA restored the viability of the cells treated with ATRA and sorafenib. Our results thus indicate that ATRA is useful for enhancing the cytotoxicity of sorafenib against HCC cells by regulating the energy metabolism of HCC cells. PMID:25683251

  3. Activation of AMP-activated protein kinase by retinoic acid sensitizes hepatocellular carcinoma cells to apoptosis induced by sorafenib.

    PubMed

    Ishijima, Naoki; Kanki, Keita; Shimizu, Hiroki; Shiota, Goshi

    2015-05-01

    To improve the outcome of cancer chemotherapy, strategies to enhance the efficacy of anticancer drugs are required. Sorafenib is the only drug to prolong overall survival of the patients with hepatocellular carcinoma (HCC), however, the outcome is still not satisfactory. Retinoids, vitamin A derivatives, have been known to exhibit inhibitory effects on various cancers including HCC. In this study, we investigated the effects of combined treatment using sorafenib and retinoids including all-trans retinoic acid (ATRA), NIK-333, and Am80 on HCC cells. Cell viability assays in six HCC cell lines, HepG2, PLC/PRF/5, HuH6, HLE, HLF, and Hep3B, revealed that 5 and 10 μM ATRA, concentrations that do not exert cytotoxic effects, enhanced the cytotoxicity of sorafenib, being much more effective than NIK-333 and Am80. We found that ATRA induced AMP-activated protein kinase activation, which was followed by reduced intracellular ATP level. Gene expression analysis revealed that ATRA decreased the expression of glycolytic genes such as GLUT-1 and LDHA. In the combination treatment using ATRA and sorafenib, increased apoptosis, followed by the activation of p38 MAPK and JNK, the upregulation and translocation of Bax to mitochondria, and the activation of caspase-3, was observed. Suppression of AMP-activated protein kinase by siRNA restored the viability of the cells treated with ATRA and sorafenib. Our results thus indicate that ATRA is useful for enhancing the cytotoxicity of sorafenib against HCC cells by regulating the energy metabolism of HCC cells. PMID:25683251

  4. Neutrophils are immune cells preferentially targeted by retinoic acid in elderly subjects

    PubMed Central

    2010-01-01

    Background The immune system gradually deteriorates with age and nutritional status is a major factor in immunosenescence. Of the many nutritional factors implicated in age-related immune dysfunction, vitamin A may be a good candidate, since vitamin A concentrations classically decrease during aging whereas it may possess important immunomodulatory properties via its active metabolites, the retinoic acids. This prompted us to investigate the immune response induced by retinoids in adults and elderly healthy subjects. Before and after oral supplementation with 13cis retinoic acid (0.5 mg/kg/day during 28 days), whole blood cells were phenotyped, and functions of peripheral blood mononuclear cells (PBMC) and polymorphonuclear cells (PMN) were investigated by flow cytometry and ELISA tests. Results In both young adults (n = 20, 25 ± 4 years) and older subjects (n = 20, 65 ± 4 years), retinoic acid supplementation had no effect on the distribution of leukocyte subpopulations or on the functions of PBMC (Il-2 and sIl-2R production, membrane expression of CD25). Concerning PMN, retinoic acid induced an increase in both spontaneous migration and cell surface expression of CD11b in the two different age populations, whereas bactericidal activity and phagocytosis remained unchanged. Conclusions We demonstrated that retinoic acid induces the same intensity of immune response between adult and older subjects, and more specifically affects PMN functions, i.e. adhesion and migration, than PBMC functions. PMID:20727130

  5. Secondary Metabolites from Three Florida Sponges with Antidepressant Activity

    PubMed Central

    Kochanowska, Anna J.; Rao, Karumanchi V.; Childress, Suzanne; El-Alfy, Abir; Matsumoto, Rae R.; Kelly, Michelle; Stewart, Gina S.; Sufka, Kenneth J.; Hamann, Mark T.

    2016-01-01

    Brominated indole alkaloids are a common class of metabolites reported from sponges of the order Verongida. Herein we report the isolation, structure determination, and activity of metabolites from three Florida sponges, namely, Verongula rigida (order Verongida, family Aplysinidae), Smenospongia aurea, and S. cerebriformis (order Dictyoceratida, family Thorectidae). All three species were investigated chemically, revealing similarities in secondary metabolites. Brominated compounds, as well as sesquiterpene quinones and hydroquinones, were identified from both V. rigida and S. aurea despite their apparent taxonomic differences at the ordinal level. Similar metabolites found in these distinct sponge species of two different genera provide evidence for a microbial origin of the metabolites. Isolated compounds were evaluated in the Porsolt forced swim test (FST) and the chick anxiety–depression continuum model. Among the isolated compounds, 5,6-dibromo-N,N-dimethyltryptamine (1) exhibited significant antidepressant-like action in the rodent FST model, while 5-bromo-N,N-dimethyltryptamine (2) caused significant reduction of locomotor activity indicative of a potential sedative action. The current study provides ample evidence that marine natural products with the diversity of brominated marine alkaloids will provide potential leads for antidepressant and anxiolytic drugs. PMID:18217716

  6. Synthesis of the alkylated active metabolite of tipidogrel.

    PubMed

    Zhi, Shuang; Xia, Guangping; Liu, Ying; Tao, Zunwei; Chen, Ligong; Liu, Dengke

    2015-04-15

    Tipidogrel (3), an effective anti-platelet drug candidate working by irreversibly inhibiting P2Y12 receptor, holds great promise in overcoming clopidogrel resistance and increasing bioavailability. As a prodrug like other thienopyridines, it metabolizes through thiophene ring opening to form active metabolites 3a and 3b, nevertheless they are easily to form disulfide bond. Derivatization of 3a and 3b via alkylation with MPBr can prevent disulfide conjugation and ensure reliable pharmacokinetic results. Thus, in order to support its pre-clinical studies on efficiencies in the formation of tipidogrel active metabolites, 13a and 13b were synthesized via seven steps of chemosynthesis and incubation with MPBr in rat plasma in vitro. The resulting crude productions were purified by semi-preparative HPLC to give Z configuration 13a and E configuration 13b. In LC-MS/MS spectra, they showed identical fragmentation pattern and retention time with M-13a and M-13b, the MPBr-derivatives of active metabolites of tipidogrel in rats. Thus, 13a and 13b were the anticipated alkylated active metabolite of tipidogrel. In addition, in the nucleophilic substitution of thioacetate with compound 11, besides the anticipated compounds 12a and 12b, their isomers compounds 12c and 12d were detected, whose structures were confirmed and the corresponding mechanism was presented. PMID:25801935

  7. A transition in transcriptional activation by the glucocorticoid and retinoic acid receptors at the tumor stage of dermal fibrosarcoma development.

    PubMed Central

    Vivanco, M D; Johnson, R; Galante, P E; Hanahan, D; Yamamoto, K R

    1995-01-01

    In transgenic mice harboring the bovine papillomavirus genome, fibrosarcomas arise along an experimentally accessible pathway in which normal dermal fibroblasts progress through two pre-neoplastic stages, mild and aggressive fibromatosis, followed by a final transition to the tumor stage. We found that the glucocorticoid receptor (GR) displays only modest transcriptional regulatory activity in cells derived from the three non-tumor stages, whereas it is highly active in fibrosarcoma cells. Upon inoculation into mice, the aggressive fibromatosis cells progress to tumor cells that have high GR activity; thus, the increased transcriptional regulatory activity of GR correlates with the cellular transition to the tumor stage. The intracellular levels of GR, as well as its hormone-dependent nuclear translocation and specific DNA binding activities, are unaltered throughout the progression. Strikingly, the low GR activity observed in the pre-neoplastic stages cannot be overcome by exogenous GR introduced by co-transfection. Moreover, comparisons of primary embryo fibroblasts and their transformed derivatives revealed a similar pattern--modest GR activity, unresponsive to overexpressed GR protein, in the normal cells was strongly increased in the transformed cells. Likewise, the retinoic acid receptor (RAR) displayed similar differential activity in the fibrosarcoma pathway. Thus, the oncogenic transformation of fibroblasts, and likely other cell types, is accompanied by a striking increase in the activities of transcriptional regulators such as GR and RAR. We suggest that normal primary cells have a heretofore unrecognized capability to limit the magnitude of induction of gene expression. Images PMID:7774580

  8. Activation of protein phosphatase 2A is responsible for increased content and inactivation of respiratory chain complex i induced by all-trans retinoic acid in human keratinocytes.

    PubMed

    Papa, F; Sardaro, N; Lippolis, R; Panelli, D; Scacco, S

    2016-01-01

    This study presents the effect of all-trans retinoic acid (ATRA) on cell growth and respiratory chain complex I in human keratinocyte cultures. Keratinocyte treatment results in increased level of GRIM-19 and other subunits of complex I, in particular of their carbonylated forms, associated with inhibition of its enzymatic activity. The results show that in keratinocytes ATRA-promoted phosphatase activity controls the proteostasis and activity of complex I. PMID:27358125

  9. Activation of Akt pathway by transcription-independent mechanisms of retinoic acid promotes survival and invasion in lung cancer cells

    PubMed Central

    2013-01-01

    Background All-trans retinoic acid (ATRA) is currently being used in clinical trials for cancer treatment. The use of ATRA is limited because some cancers, such as lung cancer, show resistance to treatment. However, little is known about the molecular mechanisms that regulate resistance to ATRA treatment. Akt is a kinase that plays a key role in cell survival and cell invasion. Akt is often activated in lung cancer, suggesting its participation in resistance to chemotherapy. In this study, we explored the hypothesis that activation of the Akt pathway promotes resistance to ATRA treatment at the inhibition of cell survival and invasion in lung cancer. We aimed to provide guidelines for the proper use of ATRA in clinical trials and to elucidate basic biological mechanisms of resistance. Results We performed experiments using the A549 human lung adenocarcinoma cell line. We found that ATRA treatment promotes PI3k-Akt pathway activation through transcription-independent mechanisms. Interestingly, ATRA treatment induces the translocation of RARα to the plasma membrane, where it colocalizes with Akt. Immunoprecipitation assays showed that ATRA promotes Akt activation mediated by RARα-Akt interaction. Activation of the PI3k-Akt pathway by ATRA promotes invasion through Rac-GTPase, whereas pretreatment with 15e (PI3k inhibitor) or over-expression of the inactive form of Akt blocks ATRA-induced invasion. We also found that treatment with ATRA induces cell survival, which is inhibited by 15e or over-expression of an inactive form of Akt, through a subsequent increase in the levels of the active form of caspase-3. Finally, we showed that over-expression of the active form of Akt significantly decreases expression levels of the tumor suppressors RARβ2 and p53. In contrast, over-expression of the inactive form of Akt restores RARβ2 expression in cells treated with ATRA, indicating that activation of the PI3k-Akt pathway inhibits the expression of ATRA target genes

  10. Oxidative and reductive metabolism of 9-cis-retinoic acid in the rat. Identification of 13,14-dihydro-9-cis-retinoic acid and its taurine conjugate.

    PubMed

    Shirley, M A; Bennani, Y L; Boehm, M F; Breau, A P; Pathirana, C; Ulm, E H

    1996-03-01

    9-cis-Retinoic acid (9-cis-RA), a hormone that binds and activates all known retinoid receptor subtypes, is structurally similar to all-trans-retinoic acid and may share common metabolic fates. Both oral and intravenous doses of 9-cis-RA to rats led to hydroxylation and ketone formation at carbon-4. 9-Cis-RA also isomerized in vivo to 13-cis-retinoic acid, 9-cis, 13-cis-retinoic acid, and all-trans-retinoic acid. After administration of [11-3H]9-cis-RA, the proportion of plasma radioactivity that was volatile increased over time, which suggested that beta-oxidative chain-shortening of 9-cis-RA might occur. An equimolar mixture of [1-13C2H3]9-cis-RA and 9-cis-RA was administered to rats for stable-isotope-labeled metabolite production. A chromatographic peak that had a lambdamax = 290 nm vs. 348 nm for the parent compound, had a retention time similar to the parent, and yielded a 1:1 positive-ion isotope cluster at m/z 303/307 in its mass spectrum. NMR analysis revealed 9-cis and 13,14-dihydro configurations, indicating that 9-cis-RA can be metabolized in rat by reduction to 13,14-dihydro-9-cis-RA. An earlier-eluting HPLC peak that exhibited a lambdamax at 290 nm, and a negative-ion-MS isotope cluster at m/z 408/412 was observed during separations of rat liver extracts. LC/MS/MS analysis revealed product ions for this peak diagnostic for carboxylic acid taurine conjugates. In rats, reduction of 9-cis-RA to 13,14-dihydro-9-cis-RA may represent an initial step leading to beta-oxidation, although available data demonstrate it is conjugated with taurine to form a novel metabolite. PMID:8820419

  11. Metabolism of a highly selective gelatinase inhibitor generates active metabolite.

    PubMed

    Lee, Mijoon; Villegas-Estrada, Adriel; Celenza, Giuseppe; Boggess, Bill; Toth, Marta; Kreitinger, Gloria; Forbes, Christopher; Fridman, Rafael; Mobashery, Shahriar; Chang, Mayland

    2007-11-01

    (4-Phenoxyphenylsulfonyl)methylthiirane (inhibitor 1) is a highly selective inhibitor of gelatinases (matrix metalloproteinases 2 and 9), which is showing considerable promise in animal models for cancer and stroke. Despite demonstrated potent, selective, and effective inhibition of gelatinases both in vitro and in vivo, the compound is rapidly metabolized, implying that the likely activity in vivo is due to a metabolite rather than the compound itself. To this end, metabolism of inhibitor 1 was investigated in in vitro systems. Four metabolites were identified by LC/MS-MS and the structures of three of them were further validated by comparison with authentic synthetic samples. One metabolite, 4-(4-thiiranylmethanesulfonylphenoxy)phenol (compound 21), was generated by hydroxylation of the terminal phenyl group of 1. This compound was investigated in kinetics of inhibition of several matrix metalloproteinases. This metabolite was a more potent slow-binding inhibitor of gelatinases (matrix metalloproteinase-2 and matrix metalloproteinase-9) than the parent compound 1, but it also served as a slow-binding inhibitor of matrix metalloproteinase-14, the upstream activator of matrix metalloproteinase-2. PMID:17927722

  12. Detection of retinoic acid receptor agonistic activity and identification of causative compounds in municipal wastewater treatment plants in Japan.

    PubMed

    Sawada, Kazuko; Inoue, Daisuke; Wada, Yuichiro; Sei, Kazunari; Nakanishi, Tsuyoshi; Ike, Michihiko

    2012-02-01

    Retinoic acid (RA) receptor (RAR) agonists are potential toxicants that can cause teratogenesis in vertebrates. To determine the occurrence of RAR agonists in municipal wastewater treatment plants (WWTPs), we examined the RARα agonistic activities of influent and effluent samples from several municipal WWTPs in Osaka, Japan, using a yeast two-hybrid assay. Significant RARα agonistic activity was detected in all the influent samples investigated, suggesting that municipal wastewater consistently contains RAR agonists. Fractionations using high-performance liquid chromatography, directed by the bioassay, found several bioactive peaks from influent samples. The RAR agonists, all-trans RA (atRA), 13-cis RA (13cRA), 4-oxo-atRA, and 4-oxo-13cRA, possibly arising from human urine, were identified by liquid chromatography ion trap time-of-flight mass spectrometry. Quantification of the identified compounds in municipal WWTPs confirmed that they were responsible for the majority of RARα agonistic activity in WWTP influents, and also revealed they were readily removed from wastewater by activated sludge treatment. Simultaneous measurement of the RARα agonistic activity revealed that although total activity typically declined concomitant with the reduction of the four identified compounds, it remained high after the decline of RAs and 4-oxo-RAs in one WWTP, suggesting the occurrence of unidentified RAR agonists during the activated sludge treatment. PMID:22095885

  13. Involvement of protein kinase C α and δ activities on the induction of the retinoic acid system in mammary cancer cells.

    PubMed

    Berardi, Damián E; Bessone, María I Díaz; Motter, Andrea; Bal de Kier Joffé, Elisa D; Urtreger, Alejandro J; Todaro, Laura B

    2015-10-01

    It has been established that retinoids exert some of their effects on cell differentiation and malignant phenotype reversion through the interaction with different members of the protein kinase C (PKC) family. Till nowadays the nature and extension of this interaction is not well understood. Due to the cytostatic and differentiating effects of retinoids, in the present study we propose to evaluate whether the crosstalk between the retinoid system and the PKC pathway could become a possible target for breast cancer treatment. We could determine that ATRA (all-trans retinoic) treatment showed a significant growth inhibition due to (G1 or G2) cell cycle arrest both in LM3 and SKBR3, a murine and human mammary cell line respectively. ATRA also induced a remarkable increase in PKCα and PKCδ expression and activity. Interestingly, the pharmacological inhibition of these two PKC isoforms prevented the activation of retinoic acid receptors (RARs) by ATRA, indicating that both PKC isoforms are required for RARs activation. Moreover, PKCδ inhibition also impaired ATRA-induced RARα translocation to the nucleus. In vivo assays revealed that a combined treatment using ATRA and PKCα inhibitors prevented lung metastatic dissemination in an additive way. Our results clearly indicate that ATRA modulates the expression and activity of different PKCs. Besides inducing cell arrest, the activity of both PKC is necessary for the induction of the retinoic acid system. The combined ATRA and PKCα inhibitors could be an option for the hormone-independent breast cancer treatment. PMID:24838400

  14. Nanostructured lipid carriers loaded with tributyrin as an alternative to improve anticancer activity of all-trans retinoic acid

    PubMed Central

    Silva, Elton Luiz; Carneiro, Guilherme; Caetano, Priscila Albuquerque; Costa, Daniel Ferreira; de Souza-Fagundes, Elaine Maria; Gomes, Dawidson Assis; Ferreira, Lucas Antônio Miranda

    2015-01-01

    Objectives All-trans retinoic acid (ATRA) is one of the most successful examples of differentiation agents and histone deacetylase inhibitors, such as tributyrin (TB), are known for their antitumor activity and potentiating action of drugs such as ATRA. Nanostructured lipid carriers (NLC) represent a promising alternative to the encapsulation of lipophilic drugs such as ATRA. This study aimed to develop, characterize, and evaluate the cytotoxicity of ATRA-TB-loaded nanostructured lipid carriers (NLC) for cancer treatment. Methods The influence of in situ formation of an ion pairing between ATRA and a lipophilic amine (benethamine; BNT) on the characteristics of NLC (size, zeta potential, encapsulation efficiency) was evaluated. Tributyrin (TB), a butyric acid donor, was used as a component of the lipid matrix. In vitro activity on cell viability and distribution of cell cycle phases were evaluated for MCF-7, MDA-MB-231, HL-60, and Jurkat cell lines. Results The presence of the amine significantly increased the encapsulation efficiency of ATRA in NLC. Inhibition of cell viability by TB-ATRA-loaded NLC was more pronounced than the free drug. Analysis of the distribution of cell cycle phases also showed increased activity for TB-ATRA-loaded NLC, with the clear effect of cell cycle arrest in G0/G1 phase transition. The presence of TB played an important role in the activity of the formulation. Conclusion Taken together, these findings suggest that TB-ATRA-loaded NLC represent a promising alternative to intravenous administration of ATRA in cancer treatment. PMID:25611812

  15. All-trans retinoic acid decreases susceptibility of a gastric cancer cell line to lymphokine-activated killer cytotoxicity.

    PubMed Central

    Chao, T. Y.; Jiang, S. Y.; Shyu, R. Y.; Yeh, M. Y.; Chu, T. M.

    1997-01-01

    All-trans retinoic acid (RA) was previously shown to regulate the growth of gastric cancer cells derived from the cell line SC-M1. This study was designed to investigate the effect of RA on the sensitivity of SC-M1 cells to lymphokine-activated killer (LAK) activity. RA at the concentration range of 0.001-10 microM was shown to induce SC-M1 cells to exhibit resistance to LAK activity in a dose-dependent manner. A kinetics study indicated that a significantly increased resistance was detected after 2 days of co-culturing SC-M1 cells with RA and reached a maximum after 6 days of culture. Similar results were obtained from two other cancer cell lines: promyelocytic leukaemia HL-60 and hepatic cancer Hep 3B. A binding assay demonstrated that the binding efficacy between target SC-M1 cells and effector LAK cells was not altered by RA. Flow cytometric analyses revealed that RA exhibited no effect on the expression of cell surface molecules, including HLA class I and class II antigens, intercellular adhesion molecule-1 and -2, and lymphocyte function antigen-3. Cell cycle analysis revealed that culture of SC-M1 cells with RA resulted in an increase in G0/G1 phase and a decrease in S phase, accompanied by a decrease in cyclin A and cyclin B1 mRNA as determined by Northern blot analysis. Additionally, RA was shown to enhance the expression of retinoic acid receptor alpha (RAR alpha) in SC-M1 cells, and to have no effect on the expression of RARbeta or RARgamma. Taken together, these results indicate that RA can significantly increase gastric cancer cells SC-M1 to resist LAK cytotoxicity by means of a cytostatic effect through a mechanism relating to cell cycle regulation. The prevailing ideas, such as a decrease in effector to target cell binding, a reduced MHC class I antigen expression or an altered RARbeta expression, are not involved. Images Figure 4 Figure 5 PMID:9155047

  16. Retinoid X receptors stimulate and 9-cis retinoic acid inhibits 1,25-dihydroxyvitamin D3-activated expression of the rat osteocalcin gene.

    PubMed Central

    MacDonald, P N; Dowd, D R; Nakajima, S; Galligan, M A; Reeder, M C; Haussler, C A; Ozato, K; Haussler, M R

    1993-01-01

    The vitamin D receptor (VDR) binds the vitamin D-responsive element (VDRE) as a heterodimer with an unidentified receptor auxiliary factor (RAF) present in mammalian cell nuclear extracts. VDR also interacts with the retinoid X receptors (RXRs), implying that RAF may be related to the RXRs. Here we demonstrate that highly purified HeLa cell RAF contained RXR beta immunoreactivity and that both activities copurified and precisely coeluted in high-resolution hydroxylapatite chromatography. Furthermore, an RXR beta-specific antibody disrupted VDR-RAF-VDRE complexes in mobility shift assays. These data strongly indicate that HeLa RAF is highly related to or is identical to RXR beta. Consequently, the effect of the 9-cis retinoic acid ligand for RXRs was examined in 1,25-dihydroxyvitamin D3 [1,25(OH)2D3]-activated gene expression systems. Increasing concentrations of 9-cis retinoic acid (1 nM to 1 microM) markedly reduced 1,25(OH)2D3-dependent accumulation of osteocalcin mRNA in osteoblast-like ROS 17/2.8 cells. All-trans retinoic acid also interfered with vitamin D responsiveness, but it was consistently less potent than the 9-cis isomer. Transient transfection studies revealed that attenuation by 9-cis retinoic acid was at the transcriptional level and was mediated through interactions at the osteocalcin VDRE. Furthermore, overexpression of both RXR beta and RXR alpha augmented 1,25(OH)2D3 responsiveness in transient expression studies. Direct analysis of VDRE binding in mobility shift assays demonstrated that heteromeric interactions between VDR and RXR were enhanced by 1,25(OH)2D3 and were not affected appreciably by 9-cis retinoic acid, except that inhibition was observed at high retinoid concentrations. These data suggest a regulatory mechanism for osteocalcin gene expression that involves 1,25(OH)2D3-induced heterodimerization of VDR and unliganded RXR. 9-cis retinoic acid may attenuate 1,25(OH)2D3 responsiveness by diverting RXRs away from VDR

  17. Investigations of fungal secondary metabolites with potential anticancer activity.

    PubMed

    Balde, ElHadj Saidou; Andolfi, Anna; Bruyère, Céline; Cimmino, Alessio; Lamoral-Theys, Delphine; Vurro, Maurizio; Damme, Marc Van; Altomare, Claudio; Mathieu, Véronique; Kiss, Robert; Evidente, Antonio

    2010-05-28

    Fourteen metabolites, isolated from phytopathogenic and toxigenic fungi, were evaluated for their in vitro antigrowth activity for six distinct cancer cell lines, using the MTT colorimetric assay. Bislongiquinolide (1) and dihydrotrichodimerol (5), which belong to the bisorbicillinoid structural class, displayed significant growth inhibitory activity against the six cancer cell lines studied, while the remaining compounds displayed weak or no activity. The data show that 1 and 5 have similar growth inhibitory activities with respect to those cancer cell lines that display certain levels of resistance to pro-apoptotic stimuli or those that are sensitive to apoptosis. Quantitative videomicroscopy analysis revealed that 1 and 5 exert their antiproliferative effect through cytostatic and not cytotoxic activity. The preliminary results from the current study have stimulated further structure-activity investigations with respect to the growth inhibitory activity of compounds belonging to the bisorbicillinoid group. PMID:20415482

  18. All-Trans Retinoic Acid Activity in Acute Myeloid Leukemia: Role of Cytochrome P450 Enzyme Expression by the Microenvironment

    PubMed Central

    Su, Meng; Alonso, Salvador; Jones, Jace W.; Yu, Jianshi; Kane, Maureen A.; Jones, Richard J.; Ghiaur, Gabriel

    2015-01-01

    Differentiation therapy with all-trans retinoic acid (atRA) has markedly improved outcome in acute promyelocytic leukemia (APL) but has had little clinical impact in other AML sub-types. Cell intrinsic mechanisms of resistance have been previously reported, yet the majority of AML blasts are sensitive to atRA in vitro. Even in APL, single agent atRA induces remission without cure. The microenvironment expression of cytochrome P450 (CYP)26, a retinoid-metabolizing enzyme was shown to determine normal hematopoietic stem cell fate. Accordingly, we hypothesized that the bone marrow (BM) microenvironment is responsible for difference between in vitro sensitivity and in vivo resistance of AML to atRA-induced differentiation. We observed that the pro-differentiation effects of atRA on APL and non-APL AML cells as well as on leukemia stem cells from clinical specimens were blocked by BM stroma. In addition, BM stroma produced a precipitous drop in atRA levels. Inhibition of CYP26 rescued atRA levels and AML cell sensitivity in the presence of stroma. Our data suggest that stromal CYP26 activity creates retinoid low sanctuaries in the BM that protect AML cells from systemic atRA therapy. Inhibition of CYP26 provides new opportunities to expand the clinical activity of atRA in both APL and non-APL AML. PMID:26047326

  19. Active Pin1 is a key target of all-trans retinoic acid in acute promyelocytic leukemia and breast cancer

    PubMed Central

    Wei, Shuo; Kozono, Shingo; Kats, Lev; Nechama, Morris; Li, Wenzong; Guarnerio, Jlenia; Luo, Manli; You, Mi-Hyeon; Yao, Yandan; Kondo, Asami; Hu, Hai; Bozkurt, Gunes; Moerke, Nathan J.; Cao, Shugeng; Reschke, Markus; Chen, Chun-Hau; Rego, Eduardo M.; LoCoco, Francesco; Cantley, Lewis; Lee, Tae Ho; Wu, Hao; Zhang, Yan; Pandolfi, Pier Paolo; Zhou, Xiao Zhen; Lu, Kun Ping

    2015-01-01

    A common key regulator of oncogenic signaling pathways in multiple tumor types is the unique isomerase Pin1. However, available Pin1 inhibitors lack the required specificity and potency. Using mechanism-based screening, here we find that all-trans retinoic acid (ATRA)--a therapy for acute promyelocytic leukemia (APL) that is considered the first example of targeted therapy in cancer, but its drug target remains elusive--inhibits and degrades active Pin1 selectively in cancer cells by directly binding to the substrate phosphate- and proline-binding pockets in the Pin1 active site. ATRA-induced Pin1 ablation degrades the fusion oncogene PML-RARα and treats APL in cell and animal models and human patients. ATRA-induced Pin1 ablation also inhibits triple negative breast cancer cell growth in human cells and in animal models by acting on many Pin1 substrate oncogenes and tumor suppressors. Thus, ATRA simultaneously blocks multiple Pin1-regulated cancer-driving pathways, an attractive property for treating aggressive and drug-resistant tumors. PMID:25849135

  20. Inhibition of LN-308 glioma cell proliferation and migration by retinoic acid amide through activation of Akt pathway

    PubMed Central

    Zhu, Jun; Lu, Xiang-Dong; Si, Feng; Song, Chun-Yu; Meng, Qing-Hai

    2015-01-01

    The present study was performed to investigate the effect of retinoic acid amide (RAA) on the expression of integrin α3β1, rate of cell proliferation and migration in p53-deficient glioma cell line, LN-308. The results revealed promotion of integrin α3 expression, reduction in proliferation and migration in RAA treated cells compared to the control LN-308 glioma cells. Promotion of RAA induced integrin α3β1 expression led to the enhancement in cyclin-dependent kinase nuclear localization and activation of Akt pathway. In addition, RAA treatment inhibited the expression of nuclear factor-κB, Bcl-2 and epidermal growth factor receptor (EGFR). These factors are responsible for promoting the rate of cell proliferation and survival in the carcinoma cells. Thus RAA treatment inhibits rate of LN-308 glioma cell proliferation and migration through increase in integrin α3β1 expression and activation of Akt pathway. Therefore, RAA can be of therapeutic importance for the treatment of glioma. PMID:26823704

  1. All-Trans Retinoic Acid Activity in Acute Myeloid Leukemia: Role of Cytochrome P450 Enzyme Expression by the Microenvironment.

    PubMed

    Su, Meng; Alonso, Salvador; Jones, Jace W; Yu, Jianshi; Kane, Maureen A; Jones, Richard J; Ghiaur, Gabriel

    2015-01-01

    Differentiation therapy with all-trans retinoic acid (atRA) has markedly improved outcome in acute promyelocytic leukemia (APL) but has had little clinical impact in other AML sub-types. Cell intrinsic mechanisms of resistance have been previously reported, yet the majority of AML blasts are sensitive to atRA in vitro. Even in APL, single agent atRA induces remission without cure. The microenvironment expression of cytochrome P450 (CYP)26, a retinoid-metabolizing enzyme was shown to determine normal hematopoietic stem cell fate. Accordingly, we hypothesized that the bone marrow (BM) microenvironment is responsible for difference between in vitro sensitivity and in vivo resistance of AML to atRA-induced differentiation. We observed that the pro-differentiation effects of atRA on APL and non-APL AML cells as well as on leukemia stem cells from clinical specimens were blocked by BM stroma. In addition, BM stroma produced a precipitous drop in atRA levels. Inhibition of CYP26 rescued atRA levels and AML cell sensitivity in the presence of stroma. Our data suggest that stromal CYP26 activity creates retinoid low sanctuaries in the BM that protect AML cells from systemic atRA therapy. Inhibition of CYP26 provides new opportunities to expand the clinical activity of atRA in both APL and non-APL AML. PMID:26047326

  2. All-trans retinoic acid-triggered antimicrobial activity against Mycobacterium tuberculosis is dependent on NPC2.

    PubMed

    Wheelwright, Matthew; Kim, Elliot W; Inkeles, Megan S; De Leon, Avelino; Pellegrini, Matteo; Krutzik, Stephan R; Liu, Philip T

    2014-03-01

    A role for vitamin A in host defense against Mycobacterium tuberculosis has been suggested through epidemiological and in vitro studies; however, the mechanism is unclear. In this study, we demonstrate that vitamin A-triggered antimicrobial activity against M. tuberculosis requires expression of NPC2. Comparison of monocytes stimulated with all-trans retinoic acid (ATRA) or 1,25-dihydroxyvitamin D3 (1,25D3), the biologically active forms of vitamin A and vitamin D, respectively, indicates that ATRA and 1,25D3 induce mechanistically distinct antimicrobial activities. Stimulation of primary human monocytes with ATRA did not result in expression of the antimicrobial peptide cathelicidin, which is required for 1,25D3 antimicrobial activity. In contrast, ATRA triggered a reduction in the total cellular cholesterol concentration, whereas 1,25D3 did not. Blocking ATRA-induced cellular cholesterol reduction inhibits antimicrobial activity as well. Bioinformatic analysis of ATRA- and 1,25D3-induced gene profiles suggests that NPC2 is a key gene in ATRA-induced cholesterol regulation. Knockdown experiments demonstrate that ATRA-mediated decrease in total cellular cholesterol content and increase in lysosomal acidification are both dependent upon expression of NPC2. Expression of NPC2 was lower in caseous tuberculosis granulomas and M. tuberculosis-infected monocytes compared with normal lung and uninfected cells, respectively. Loss of NPC2 expression ablated ATRA-induced antimicrobial activity. Taken together, these results suggest that the vitamin A-mediated antimicrobial mechanism against M. tuberculosis requires NPC2-dependent expression and function, indicating a key role for cellular cholesterol regulation in the innate immune response. PMID:24501203

  3. Activity of Praziquantel Enantiomers and Main Metabolites against Schistosoma mansoni

    PubMed Central

    Meister, Isabel; Ingram-Sieber, Katrin; Cowan, Noemi; Todd, Matthew; Robertson, Murray N.; Meli, Claudia; Patra, Malay; Gasser, Gilles

    2014-01-01

    A racemic mixture of R and S enantiomers of praziquantel (PZQ) is currently the treatment of choice for schistosomiasis. Though the S enantiomer and the metabolites are presumed to contribute only a little to the activity of the drug, in-depth side-by-side studies are lacking. The aim of this study was to investigate the in vitro activities of PZQ and its main metabolites, namely, R- and S-cis- and R- and S-trans-4′-hydroxypraziquantel, against adult worms and newly transformed schistosomula (NTS). Additionally, we explored the in vivo activity and hepatic shift (i.e., the migration of the worms to the liver) produced by each PZQ enantiomer in mice. Fifty percent inhibitory concentrations of R-PZQ, S-PZQ, and R-trans- and R-cis-4′-hydroxypraziquantel of 0.02, 5.85, 4.08, and 2.42 μg/ml, respectively, for adult S. mansoni were determined in vitro. S-trans- and S-cis-4′-hydroxypraziquantel were not active at 100 μg/ml. These results are consistent with microcalorimetry data and studies with NTS. In vivo, single 400-mg/kg oral doses of R-PZQ and S-PZQ achieved worm burden reductions of 100 and 19%, respectively. Moreover, worms treated in vivo with S-PZQ displayed an only transient hepatic shift and returned to the mesenteric veins within 24 h. Our data confirm that R-PZQ is the main effector molecule, while S-PZQ and the metabolites do not play a significant role in the antischistosomal properties of PZQ. PMID:24982093

  4. Cholinergic activation enhances retinoic acid-induced differentiation in the human NB-4 acute promyelocytic leukemia cell line.

    PubMed

    Chotirat, Sadudee; Suriyo, Tawit; Hokland, Marianne; Hokland, Peter; Satayavivad, Jutamaad; Auewarakul, Chirayu U

    2016-07-01

    The non-neuronal cholinergic system (NNCS) has been shown to play a role in regulating hematopoietic differentiation. We determined the expression of cholinergic components in leukemic cell lines by Western blotting and in normal leukocyte subsets by flow cytometry and found a heterogeneous expression of choline acetyltransferase (ChAT), acetylcholinesterase (AChE), choline transporter (CHT), M3 muscarinic acetylcholine receptor (M3-mAChR) and α7 nicotinic acetylcholine receptor (α7-nAChR). We then evaluated NNCS role in differentiation of human NB-4 acute promyelocytic leukemia cell line and discovered a dramatic induction of M3-mAChR after all-trans retinoic acid (ATRA) treatment (p<0.0001). Adding carbachol which is a cholinergic agonist to the ATRA treatment resulted in an increase of a granulocytic differentiation marker (CD11b) as compared with ATRA treatment alone (p<0.05), indicating that cholinergic activation enhanced ATRA in inducing NB-4 maturation. The combination of carbachol and ATRA treatment for 72h also resulted in decreased viability and increased cleaved caspase-3 expression when compared with ATRA treatment alone (p<0.05). However, this combination did not cause poly (ADP-ribose) polymerase (PARP) cleavage. Overall, we have shown that NB-4 cells expressed M3-mAChR in a differentiation-dependent manner and cholinergic stimulation induced maturation and death of ATRA-induced differentiated NB-4 cells. PMID:27282572

  5. Hsp90 Activity Modulation by Plant Secondary Metabolites.

    PubMed

    Dal Piaz, Fabrizio; Terracciano, Stefania; De Tommasi, Nunziatina; Braca, Alessandra

    2015-09-01

    Hsp90 is an evolutionarily conserved adenosine triphosphate-dependent molecular chaperone and is one of the most abundant proteins in the cells (1-3 %). Hsp90 is induced when a cell undergoes various types of environmental stresses such as heat, cold, or oxygen deprivation. It is involved in the turnover, trafficking, and activity of client proteins, including apoptotic factors, protein kinases, transcription factors, signaling proteins, and a number of oncoproteins. Most of the Hsp90 client proteins are involved in cell growth, differentiation, and survival, and include kinases, nuclear hormone receptors, transcription factors, and other proteins associated with almost all the hallmarks of cancer. Consistent with these diverse activities, genetic and biochemical studies have demonstrated the implication of Hsp90 in a range of diseases, including cancer, making this chaperone an interesting target for drug research.During the last few decades, plant secondary metabolites have been studied as a major source for lead compounds in drug discovery. Recently, several plant-derived small molecules have been discovered exhibiting inhibitory activity towards Hsp90, such as epigallocatechin gallate, gedunin, lentiginosine, celastrol, and deguelin. In this work, an overview of plant secondary metabolites interfering with Hsp90 activities is provided. PMID:26227505

  6. Retinoic Acid Activity in Undifferentiated Neural Progenitors Is Sufficient to Fulfill Its Role in Restricting Fgf8 Expression for Somitogenesis

    PubMed Central

    Cunningham, Thomas J.; Brade, Thomas; Sandell, Lisa L.; Lewandoski, Mark; Trainor, Paul A.; Colas, Alexandre; Mercola, Mark; Duester, Gregg

    2015-01-01

    Bipotent axial stem cells residing in the caudal epiblast during late gastrulation generate neuroectodermal and presomitic mesodermal progeny that coordinate somitogenesis with neural tube formation, but the mechanism that controls these two fates is not fully understood. Retinoic acid (RA) restricts the anterior extent of caudal fibroblast growth factor 8 (Fgf8) expression in both mesoderm and neural plate to control somitogenesis and neurogenesis, however it remains unclear where RA acts to control the spatial expression of caudal Fgf8. Here, we found that mouse Raldh2-/- embryos, lacking RA synthesis and displaying a consistent small somite defect, exhibited abnormal expression of key markers of axial stem cell progeny, with decreased Sox2+ and Sox1+ neuroectodermal progeny and increased Tbx6+ presomitic mesodermal progeny. The Raldh2-/- small somite defect was rescued by treatment with an FGF receptor antagonist. Rdh10 mutants, with a less severe RA synthesis defect, were found to exhibit a small somite defect and anterior expansion of caudal Fgf8 expression only for somites 1–6, with normal somite size and Fgf8 expression thereafter. Rdh10 mutants were found to lack RA activity during the early phase when somites are small, but at the 6-somite stage RA activity was detected in neural plate although not in presomitic mesoderm. Expression of a dominant-negative RA receptor in mesoderm eliminated RA activity in presomitic mesoderm but did not affect somitogenesis. Thus, RA activity in the neural plate is sufficient to prevent anterior expansion of caudal Fgf8 expression associated with a small somite defect. Our studies provide evidence that RA restriction of Fgf8 expression in undifferentiated neural progenitors stimulates neurogenesis while also restricting the anterior extent of the mesodermal Fgf8 mRNA gradient that controls somite size, providing new insight into the mechanism that coordinates somitogenesis with neurogenesis. PMID:26368825

  7. Retinoic acid-induced AP-1 transcriptional activity regulates B16 mouse melanoma growth inhibition and differentiation.

    PubMed

    Huang, Ying; Boskovic, Goran; Niles, Richard M

    2003-02-01

    Retinoic acid (RA) inhibits growth and induces differentiation of B16 mouse melanoma cells. These effects are accompanied by a large increase in PKCalpha mRNA and protein levels and surprisingly an increase in activating protein-1 (AP-1) transcriptional activity. To further investigate the RA-induced AP-1 activity we established clones of B16 cells stably expressing an AP-1-luciferase reporter gene. Treatment of these clones with phorbol dibutyrate increased AP-1 activity which peaked at 2-4 h and returned to baseline level by 24 h. In contrast, RA treatment resulted in a slow increase in AP-1 activity that reached a maximum level at 48 h and was maintained for the duration of the treatment. We tested the importance of the RA-induced AP-1 activity by establishing clones which stably express a dominant negative fos gene (A-fos) and have greatly diminished AP-1 activity. Growth rates of untreated A-fos expressing cells were similar to wt B16 and clones not expressing A-fos. However, clones expressing the dominant-negative fos had a markedly decreased sensitivity to RA-induced inhibition of anchorage-dependent and -independent growth. Treatment of wt B16 cells for 48 h with RA increased melanin production by two to fourfold, but this effect was completely lost in the A-fos clones. The ability of RA to induce RARbeta and PKCalpha expression was retained in A-fos clones, suggesting that A-fos was not interfering with RAR transcription activation functions. We tested whether the RA-induced AP-1 activity might be mediated by the ERK1/2 MAPK pathway. Inhibition of ERK1/2 phosphorylation stimulated AP-1 activity, which was not additive to that induced by RA. This finding raises the possibility that this MAPK pathway may be a target of retinoid action. Our observations suggest that AP-1 transcriptional activity induced by RA likely plays an important role in the biological changes mediated by this retinoid in B16 melanoma cells. PMID:12494454

  8. Antioxidant activity of nimesulide and its main metabolites.

    PubMed

    Facino, R M; Carini, M; Aldini, G

    1993-01-01

    The antioxidant activity of nimesulide and its main metabolites, 4'-hydroxynimesulide (M1) and 2-(4'-hydroxyphenoxy)-4-N-acetylamino-methansulfonanilide (M2), was investigated using 2 in vitro models: NADPH-supported lipid peroxidation in rat liver microsomes (marker MDA formation) and xanthine/xanthine oxidase, iron-promoted depolymerisation of hyaluronic acid, determined by gel permeation chromatographic analysis (marker molecular weight distribution). In the lipid peroxidation model, all the compounds inhibited MDA formation in a concentration-dependent manner, although with different potencies; the maximum scavenging effect was observed for M1 [50% inhibitory concentration (IC50) = 30 mumol/L; M2 IC50 = 0.5 mmol/L; nimesulide = 0.8 mmol/L]. Nimesulide was more active than its metabolites in preventing OH-induced depolymerisation of hyaluronic acid, with a 50% effective concentration of approximately 230 mumol/L, which was fairly comparable to that of tenoxicam. This protective effect was due to the OH.-entrapping capacity of the drug, which, in the Fenton-driven model, is easily converted, via OH. attack, to M1 and putatively to 2-hydroxy-4-nitro-methansulfonanilide. PMID:7506157

  9. Retinoic acid inhibits the proliferative response induced by CD40 activation and interleukin-4 in mantle cell lymphoma.

    PubMed

    Guidoboni, Massimo; Zancai, Paola; Cariati, Roberta; Rizzo, Silvana; Dal Col, Jessica; Pavan, Alessandro; Gloghini, Annunziata; Spina, Michele; Cuneo, Antonio; Pomponi, Fabrizio; Bononi, Antonio; Doglioni, Claudio; Maestro, Roberta; Carbone, Antonino; Boiocchi, Mauro; Dolcetti, Riccardo

    2005-01-15

    Mantle cell lymphoma (MCL) is an aggressive B-cell non-Hodgkin's lymphoma with poor response to therapy and unfavorable prognosis. Here, we show that retinoic acid (RA) isomers significantly inhibit the proliferation of both primary MCL cultures (n = 7) and established cell lines (Granta 519 and SP-53) as shown by [(3)H]thymidine uptake and carboxyfluorescein diacetate succinimidyl ester labeling coupled with cyclin D1 staining. RA induces cell accumulation in G(0)-G(1) together with a marked up-regulation of p27(Kip1) by inhibiting ubiquitination and proteasome-dependent degradation of the protein. The p21(Cip1) inhibitor was also up-regulated by RA in Granta 519 cells, whereas the expression of cyclin D1 is unaffected. Most of RA-induced p27(Kip1) was bound to cyclin D1/cyclin-dependent kinase 4 complexes, probably contributing to the decreased cyclin-dependent kinase 4 kinase activity and pRb hypophosphorylation observed in RA-treated cells. Experiments with receptor-selective ligands indicate that RA receptor alpha cooperates with retinoid X receptors in mediating RA-dependent MCL cell growth inhibition. Notably, RA isomers, and particularly 9-cis-RA, also inhibited the growth-promoting effect induced in primary MCL cells by CD40 activation alone or in combination with interleukin-4. Immunohistochemical analysis showed that significant numbers of CD40L-expressing lymphoid cells are present in lymph node biopsies of MCL patients. These results therefore further strengthen the possibility that triggering of CD40 by infiltrating CD40L+ cells may continuously promote the growth of MCL cells in vivo. On these grounds, our findings that RA inhibits basal MCL proliferation as well as MCL growth-promoting effects exerted by microenvironmental factors make these compounds highly attractive in terms of potential clinical efficacy in this setting. PMID:15695403

  10. Liposome encapsulated all trans retinoic acid (ATRA) has enhanced immunomodulatory and inflammation reducing activities in mice model.

    PubMed

    Grace, V M Berlin; Siddikuzzaman; Rimashree, B

    2015-01-01

    The all trans retinoic acid (ATRA) is found to have a promising regulatory effect on immune system and inflammatory responses in experimental research. The purpose of this study was to investigate whether this therapeutic efficiency of ATRA could be enhanced by encapsulating into a liposome formulation composed of Distearoyl-L-phosphatidylcholine (DSPC) and cholesterol utilizing a well-established mice model. The humoral antibody titer (HA), delayed-type hypersensitivity (DTH), bone marrow cellularity, hematology, and levels of α- esterase-positive cells, were taken as parameters to assess the level of immunomodulation in the sheep red blood cells (SRBC) immunized and challenged BALB/c mice. The anti-inflammatory effect of encapsulated ATRA was evaluated by the size changes in the induced inflammation edema in the mice paw as well as its histopathology. The results showed a significant immunostimulatory effect for both the free and encapsulated ATRA as indicated by the increase in the levels of total leukocyte, bone marrow and α-esterase positive cells and decreased Hb level respectively. We have also observed an enhanced specific antibody hemagglutinin titre value and the DTH response developed in response to SRBC challenge in these treatments. Both the immunostimulatory as well as inflammation reducing property were significantly higher in encapsulated ATRA treated group of mice over that of in free ATRA treated group of mice. Based on these results, we conclude that the encapsulated ATRA has a higher potency over free ATRA in its immunomodulatory activity and also has a significant impact on reducing inflammation in BALB/c mice model. PMID:25594892

  11. Mutagenic activity of austocystins - secondary metabolites of Aspergillus ustus

    SciTech Connect

    Kfir, R.; Johannsen, E.; Vleggaar, R.

    1986-11-01

    Mycotoxins constitute a group of toxic secondary fungal metabolites. Fungi that produce these toxins frequently contaminate food and feed, creating a potential threat to human and animal health. Biological activities of mycotoxins include, amongst others: toxicity, mutagenicity and carcinogenicity, which can be expressed with or without metabolic activation. Austocystins are similar in structure to aflatoxin B/sup 1/ and are probably synthesized in a similar manner. The Ames Salmonella test, a widely accepted method employed for the detection of mutagenic activity of various chemical compounds was used for testing the mutagenic activity of different mycotoxins. As aflatoxin B/sup 1/ was found by the Ames test to be highly mutagenic, the same test was applied for the study of possible mutagenicity of the austocystins. The mutagenic activity of these compounds was studied with and without metabolic activation using two tester strains of S. typhimurium, one capable of detecting frame shift mutation (strain TA98) and the other capable of detecting base pair substitution (strain TA100).

  12. Depsides: Lichen Metabolites Active against Hepatitis C Virus

    PubMed Central

    Vu, Thi Huyen; Le Lamer, Anne-Cécile; Lalli, Claudia; Boustie, Joël; Samson, Michel

    2015-01-01

    A thorough phytochemical study of Stereocaulon evolutum was conducted, for the isolation of structurally related atranorin derivatives. Indeed, pilot experiments suggested that atranorin (1), the main metabolite of this lichen, would interfere with the lifecycle of hepatitis C virus (HCV). Eight compounds, including one reported for the first time (2), were isolated and characterized. Two analogs (5, 6) were also synthesized, to enlarge the panel of atranorin-related structures. Most of these compounds were active against HCV, with a half-maximal inhibitory concentration of about 10 to 70 µM, with depsides more potent than monoaromatic phenols. The most effective inhibitors (1, 5 and 6) were then added at different steps of the HCV lifecycle. Interestingly, atranorin (1), bearing an aldehyde function at C-3, inhibited only viral entry, whereas the synthetic compounds 5 and 6, bearing a hydroxymethyl and a methyl function, respectively, at C-3 interfered with viral replication. PMID:25793970

  13. Retinoic Acid-Mediated Regulation of GLI3 Enables Efficient Motoneuron Derivation from Human ESCs in the Absence of Extrinsic SHH Activation

    PubMed Central

    Calder, Elizabeth L.; Steinbeck, Julius A.; Tu, Edmund; Keros, Sotirios; Ying, Shui-Wang; Jaiswal, Manoj K.; Cornacchia, Daniela; Goldstein, Peter A.; Tabar, Viviane

    2015-01-01

    The derivation of somatic motoneurons (MNs) from ES cells (ESCs) after exposure to sonic hedgehog (SHH) and retinoic acid (RA) is one of the best defined, directed differentiation strategies to specify fate in pluripotent lineages. In mouse ESCs, MN yield is particularly high after RA + SHH treatment, whereas human ESC (hESC) protocols have been generally less efficient. In an effort to optimize yield, we observe that functional MNs can be derived from hESCs at high efficiencies if treated with patterning molecules at very early differentiation steps before neural induction. Remarkably, under these conditions, equal numbers of human MNs were obtained in the presence or absence of SHH exposure. Using pharmacological and genetic strategies, we demonstrate that early RA treatment directs MN differentiation independently of extrinsic SHH activation by suppressing the induction of GLI3. We further demonstrate that neural induction triggers a switch from a poised to an active chromatin state at GLI3. Early RA treatment prevents this switch by direct binding of the RA receptor at the GLI3 promoter. Furthermore, GLI3 knock-out hESCs can bypass the requirement for early RA patterning to yield MNs efficiently. Our data demonstrate that RA-mediated suppression of GLI3 is sufficient to generate MNs in an SHH-independent manner and that temporal changes in exposure to patterning factors such as RA affect chromatin state and competency of hESC-derived lineages to adopt specific neuronal fates. Finally, our work presents a streamlined platform for the highly efficient derivation of human MNs from ESCs and induced pluripotent stem cells. SIGNIFICANCE STATEMENT Our study presents a rapid and efficient protocol to generate human motoneurons from embryonic and induced pluripotent stem cells. Surprisingly, and in contrast to previous work, motoneurons are generated in the presence of retinoic acid but in the absence of factors that activate sonic hedgehog signaling. We show that early

  14. Biologically Active Metabolites Produced by the Basidiomycete Quambalaria cyanescens

    PubMed Central

    Stodůlková, Eva; Císařová, Ivana; Kolařík, Miroslav; Chudíčková, Milada; Novák, Petr; Man, Petr; Kuzma, Marek; Pavlů, Barbora; Černý, Jan; Flieger, Miroslav

    2015-01-01

    Four strains of the fungus Quambalaria cyanescens (Basidiomycota: Microstromatales), were used for the determination of secondary metabolites production and their antimicrobial and biological activities. A new naphthoquinone named quambalarine A, (S)-(+)-3-(5-ethyl-tetrahydrofuran-2-yliden)-5,7,8-trihydroxy-2-oxo-1,4-naphthoquinone (1), together with two known naphthoquinones, 3-hexanoyl-2,5,7,8-tetrahydroxy-1,4-naphthoquinone (named here as quambalarine B, 2) and mompain, 2,5,7,8-tetrahydroxy-1,4-naphthoquinone (3) were isolated. Their structures were determined by single-crystal X-ray diffraction crystallography, NMR and MS spectrometry. Quambalarine A (1) had a broad antifungal and antibacterial activity and is able inhibit growth of human pathogenic fungus Aspergillus fumigatus and fungi co-occurring with Q. cyanescens in bark beetle galleries including insect pathogenic species Beauveria bassiana. Quambalarine B (2) was active against several fungi and mompain mainly against bacteria. The biological activity against human-derived cell lines was selective towards mitochondria (2 and 3); after long-term incubation with 2, mitochondria were undetectable using a mitochondrial probe. A similar effect on mitochondria was observed also for environmental competitors of Q. cyanescens from the genus Geosmithia. PMID:25723150

  15. Retinoids increase human apolipoprotein A-11 expression through activation of the retinoid X receptor but not the retinoic acid receptor.

    PubMed Central

    Vu-Dac, N; Schoonjans, K; Kosykh, V; Dallongeville, J; Heyman, R A; Staels, B; Auwerx, J

    1996-01-01

    Considering the link between plasma high-density lipoprotein (HDL) cholesterol levels and a protective effect against coronary artery disease as well as the suggested beneficial effects of retinoids on the production of the major HDL apolipoprotein (apo), apo A-I, the goal of this study was to analyze the influence of retinoids on the expression of apo A-II, the other major HDL protein. Retinoic acid (RA) derivatives have a direct effect on hepatic apo A-II production, since all-trans (at) RA induces apo A-II mRNA levels and apo A-II secretion in primary cultures of human hepatocytes. In the HepG2 human hepatoblastoma cell line, both at-RA and 9-cis RA as well as the retinoid X receptor (RXR)-specific agonist LGD 1069, but not the RA receptor (RAR) agonist ethyl-p-[(E)-2-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthyl)-l-pro penyl]-benzoic acid (TTNPB), induce apo A-II mRNA levels. Transient-transfection experiments with a reporter construct driven by the human apo A-II gene promoter indicated that 9-cis RA and at-RA, as well as the RXR agonists LGD 1069 and LG 100268, induced apo A-II gene expression at the transcriptional level. Only minimal effects of the RAR agonist TTNPB were observed on the apo A-II promoter reporter construct. Unilateral deletions and site-directed mutagenesis identified the J site of the apo A-II promoter mediating the responsiveness to RA. This element contains two imperfect half-sites spaced by 1 oligonucleotide. Cotransfection assays in combination with the use of RXR or RAR agonists showed that RXR but not RAR transactivates the apo A-II promoter through this element. By contrast, RAR inhibits the inductive effects of RXR on the apo A-II J site in a dose-dependent fashion. Gel retardation assays demonstrated that RXR homodimers bind, although with a lower affinity than RAR-RXR heterodimers, to the AH-RXR response element. In conclusion, retinoids induce hepatic apo A-II production at the transcriptional level via the interaction of

  16. Prediction and Validation of Gene Regulatory Elements Activated During Retinoic Acid Induced Embryonic Stem Cell Differentiation.

    PubMed

    Simandi, Zoltan; Horvath, Attila; Nagy, Peter; Nagy, Laszlo

    2016-01-01

    Embryonic development is a multistep process involving activation and repression of many genes. Enhancer elements in the genome are known to contribute to tissue and cell-type specific regulation of gene expression during the cellular differentiation. Thus, their identification and further investigation is important in order to understand how cell fate is determined. Integration of gene expression data (e.g., microarray or RNA-seq) and results of chromatin immunoprecipitation (ChIP)-based genome-wide studies (ChIP-seq) allows large-scale identification of these regulatory regions. However, functional validation of cell-type specific enhancers requires further in vitro and in vivo experimental procedures. Here we describe how active enhancers can be identified and validated experimentally. This protocol provides a step-by-step workflow that includes: 1) identification of regulatory regions by ChIP-seq data analysis, 2) cloning and experimental validation of putative regulatory potential of the identified genomic sequences in a reporter assay, and 3) determination of enhancer activity in vivo by measuring enhancer RNA transcript level. The presented protocol is detailed enough to help anyone to set up this workflow in the lab. Importantly, the protocol can be easily adapted to and used in any cellular model system. PMID:27403939

  17. Pharmacologically active drug metabolites: therapeutic and toxic activities, plasma and urine data in man, accumulation in renal failure.

    PubMed

    Drayer, D E

    1976-01-01

    Drugs that are administered to man may be biotransformed to yield metabolites that are pharmacologically active. The therapeutic and toxic activities of drug metabolites and the species in which this activity was demonstrated are compiled for the metabolites of 58 drugs. The metabolite to parent drug ratio in the plasma of non-uraemic man and the percentage urinary excretion of the metabolite in non-uraemic man are also tabulated. Those active metabolites with significant pharmacological activity and high plasma levels, both relative to that of the parent drug, will probably contribute substantially to the pharmacological effect ascribed to the parent drug. Active metabolites may accumulate in patients with end stage renal disease if renal excretion is a major elimination pathway for the metabolite. This is true even if the active metabolite is a minor metabolite of the parent drug, as long as the minor metabolite is not further biotransformed and is mainly excreted in the urine. Minor metabolite accumulation may also occur if it is further biotransformed by a pathway inhibited in uraemia. Some clinical examples of the accumulation of active drug metabolites in patients with renal failure are: (a) The abolition of premature ventricular contractions and prevention of paroxysmal atrial tachycardia in some cardiac patients with poor renal function treated with procainamide are associated with high levels of N-acetylprocainamide. (b) The severe irritability and twitching seen in a uraemic patient treated with pethidine (meperidine) are associated with high levels of norpethidine. (c) The severe muscle weakness and tenderness seen in patients with renal failure receiving clofibrate are associated with excessive accumulation of the free acid metabolite of clofibrate. (d) Patients with severe renal insufficiency taking allopurinol appear to experience a higher incidence of side reactions, possibly due to the accumulation of oxipurinol. (e) Accumulation of free and

  18. All-Trans Retinoic Acid Modulates ORMDL3 Expression via Transcriptional Regulation

    PubMed Central

    Zhuang, Li-Li; Huang, Bo-Xian; Feng, Jie; Zhu, Liang-Hua; Jin, Rui; Qiu, Ling-Zhi; Zhou, Guo-Ping

    2013-01-01

    All-trans retinoic acid (ATRA) is an active metabolite of Vitamin A, it shows protective effects on asthma, including maintains airway epithelial integrity, inhibits asthma effector cells differentiation, modulates immune response, et al. However, the promoting effect of ATRA on Th2 response has restricted the clinical application of ATRA in asthma treatment. ORMDL3 is a candidate gene of childhood onset asthma, and high-transcript of ORMDL3 is associated with the development of asthma. Here we show that ATRA increases ORMDL3 production in vitro via inducing PKA-dependent CREB phosphorylation which in turn binds to the CRE element in promoter region of ORMDL3 and initiates ORMDL3 transcription. This finding is in consistent with the previous reports that ATRA could regulate target genes without the presence of retinoic acid response element (RARE) in promoter region but through other signals such as PKA/CREB. Nevertheless, in the present study, the traditional signal pathway of ATRA, retinoic acid receptor (RAR) signal transduction pathway, indirectly modulated ORMDL3 expression. RAR-α agonist (Am-80) increased ORMDL3 production even though there was no RARE in ORMDL3 promoter, introns or 3′-downstream region. Besides, the signal of RAR might differ from that of ATRA since Am-80 failed to induce CREB activation. In conclusion, our data indicate that ATRA facilitates ORMDL3 production probable through PKA/CREB, and this may be a starting point for more detailed mechanism researches on ATRA and asthma. PMID:24204796

  19. All-trans retinoic acid modulates ORMDL3 expression via transcriptional regulation.

    PubMed

    Zhuang, Li-Li; Huang, Bo-Xian; Feng, Jie; Zhu, Liang-Hua; Jin, Rui; Qiu, Ling-Zhi; Zhou, Guo-Ping

    2013-01-01

    All-trans retinoic acid (ATRA) is an active metabolite of Vitamin A, it shows protective effects on asthma, including maintains airway epithelial integrity, inhibits asthma effector cells differentiation, modulates immune response, et al. However, the promoting effect of ATRA on Th2 response has restricted the clinical application of ATRA in asthma treatment. ORMDL3 is a candidate gene of childhood onset asthma, and high-transcript of ORMDL3 is associated with the development of asthma. Here we show that ATRA increases ORMDL3 production in vitro via inducing PKA-dependent CREB phosphorylation which in turn binds to the CRE element in promoter region of ORMDL3 and initiates ORMDL3 transcription. This finding is in consistent with the previous reports that ATRA could regulate target genes without the presence of retinoic acid response element (RARE) in promoter region but through other signals such as PKA/CREB. Nevertheless, in the present study, the traditional signal pathway of ATRA, retinoic acid receptor (RAR) signal transduction pathway, indirectly modulated ORMDL3 expression. RAR-α agonist (Am-80) increased ORMDL3 production even though there was no RARE in ORMDL3 promoter, introns or 3'-downstream region. Besides, the signal of RAR might differ from that of ATRA since Am-80 failed to induce CREB activation. In conclusion, our data indicate that ATRA facilitates ORMDL3 production probable through PKA/CREB, and this may be a starting point for more detailed mechanism researches on ATRA and asthma. PMID:24204796

  20. Three Conazoles Increase Hepatic Microsomal Retinoic Acid Metabolism and Decrease Mouse Hepatic Retinoic Acid Levels In Vivo

    EPA Science Inventory

    Conazoles are fungicides used in agriculture and as pharmaceuticals. In a previous toxicogenomic study of triazole-containing conazoles we found gene expression changes consistent with the alteration of the metabolism of all trans-retinoic acid (atRA), a vitamin A metabolite with...

  1. Cilostazol suppresses β-amyloid production by activating a disintegrin and metalloproteinase 10 via the upregulation of SIRT1-coupled retinoic acid receptor-β.

    PubMed

    Lee, Hye Rin; Shin, Hwa Kyoung; Park, So Youn; Kim, Hye Young; Lee, Won Suk; Rhim, Byung Yong; Hong, Ki Whan; Kim, Chi Dae

    2014-11-01

    The accumulation of plaques of β-amyloid (Aβ) peptides, a hallmark of Alzheimer's disease, results from the sequential cleavage of amyloid precursor protein (APP) by activation of β- and γ-secretases. However, the production of Aβ can be avoided by alternate cleavage of APP by α-and γ-secretases. We hypothesized that cilostazol attenuates Aβ production by increasing a disintegrin and metalloproteinase 10 (ADAM10)/α-secretase activity via SIRT1-coupled retinoic acid receptor-β (RARβ) activation in N2a cells expressing human APP Swedish mutation (N2aSwe). To evoke endogenous Aβ overproduction, the culture medium was switched from medium containing 10% fetal bovine serum (FBS) to medium containing 1% FBS, and cells were cultured for 3∼24 hr. After depletion of FBS in media, N2aSwe cells showed increased accumulations of full-length APP (FL-APP) and Aβ in a time-dependent manner (3-24 hr) in association with decreased ADAM10 protein expression. When pretreated with cilostazol (10-30 μM), FL-APP and Aβ levels were significantly reduced, and ADAM10 and α-secretase activities were restored. Furthermore, the effect of cilostazol on ADAM10 expression was antagonized by pretreating Rp-cAMPS and sirtinol and by SIRT1-gene silencing. In the N2aSwe cells overexpressing the SIRT1 gene, ADAM10, and sAPPα levels were significantly elevated. In addition, like all-trans retinoic acid, cilostazol enhanced the protein expressions of RARβ and ADAM10, and the cilostazol-stimulated ADAM10 elevation was significantly attenuated by LE135 (a RARβ inhibitor), sirtinol, and RARβ-gene silencing. In conclusion, cilostazol suppresses the accumulations of FL-APP and Aβ by activating ADAM10 via the upregulation of SIRT1-coupled RARβ. PMID:24903973

  2. Evolution of retinoic acid receptors and retinoic acid signaling.

    PubMed

    Gutierrez-Mazariegos, Juliana; Schubert, Michael; Laudet, Vincent

    2014-01-01

    Retinoic acid (RA) is a vitamin A-derived morphogen controlling important developmental processes in vertebrates, and more generally in chordates, including axial patterning and tissue formation and differentiation. In the embryo, endogenous RA levels are controlled by RA synthesizing and degrading enzymes and the RA signal is transduced by two retinoid receptors: the retinoic acid receptor (RAR) and the retinoid X receptor (RXR). Both RAR and RXR are members of the nuclear receptor superfamily of ligand-activated transcription factors and mainly act as heterodimers to activate the transcription of target genes in the presence of their ligand, all-trans RA. This signaling pathway was long thought to be a chordate innovation, however, recent findings of gene homologs involved in RA signaling in the genomes of a wide variety of non-chordate animals, including ambulacrarians (sea urchins and acorn worms) and lophotrochozoans (annelids and mollusks), challenged this traditional view and suggested that the RA signaling pathway might have a more ancient evolutionary origin than previously thought. In this chapter, we discuss the evolutionary history of the RA signaling pathway, and more particularly of the RARs, which might have experienced independent gene losses and duplications in different animal lineages. In sum, the available data reveal novel insights into the origin of the RA signaling pathway as well as into the evolutionary history of the RARs. PMID:24962881

  3. Integrating Retinoic Acid Signaling with Brain Function

    ERIC Educational Resources Information Center

    Luo, Tuanlian; Wagner, Elisabeth; Drager, Ursula C.

    2009-01-01

    The vitamin A derivative retinoic acid (RA) regulates the transcription of about a 6th of the human genome. Compelling evidence indicates a role of RA in cognitive activities, but its integration with the molecular mechanisms of higher brain functions is not known. Here we describe the properties of RA signaling in the mouse, which point to…

  4. COMPARISON OF CHOLINESTERASE ACTIVITY, RESIDUE LEVELS, AND URINARY METABOLITE EXCRETION OF RATS EXPOSED TO ORGANOPHOSPHORUS PESTICIDES

    EPA Science Inventory

    Blood cholinesterase activity, urinary levels of phenolic and organophosphorus metabolites, and residues of intact compounds in blood and fat were determined following exposure of rats to organophosphorus pesticides. The eight pesticides studied included representative halogenate...

  5. Widespread occurrence of neuro-active pharmaceuticals and metabolites in 24 Minnesota rivers and wastewaters.

    PubMed

    Writer, Jeffrey H; Ferrer, Imma; Barber, Larry B; Thurman, E Michael

    2013-09-01

    Concentrations of 17 neuro-active pharmaceuticals and their major metabolites (bupropion, hydroxy-bupropion, erythro-hydrobupropion, threo-hydrobupropion, carbamazepine, 10,11,-dihydro-10,11,-dihydroxycarbamazepine, 10-hydroxy-carbamazepine, citalopram, N-desmethyl-citalopram, fluoxetine, norfluoxetine, gabapentin, lamotrigine, 2-N-glucuronide-lamotrigine, oxcarbazepine, venlafaxine and O-desmethyl-venlafaxine), were measured in treated wastewater and receiving surface waters from 24 locations across Minnesota, USA. The analysis of upstream and downstream sampling sites indicated that the wastewater treatment plants were the major source of the neuro-active pharmaceuticals and associated metabolites in surface waters of Minnesota. Concentrations of parent compound and the associated metabolite varied substantially between treatment plants (concentrations±standard deviation of the parent compound relative to its major metabolite) as illustrated by the following examples; bupropion and hydrobupropion 700±1000 ng L(-1), 2100±1700 ng L(-1), carbamazepine and 10-hydroxy-carbamazepine 480±380 ng L(-1), 360±400 ng L(-1), venlafaxine and O-desmethyl-venlafaxine 1400±1300 ng L(-1), 1800±2300 ng L(-1). Metabolites of the neuro-active compounds were commonly found at higher or comparable concentrations to the parent compounds in wastewater effluent and the receiving surface water. Neuro-active pharmaceuticals and associated metabolites were detected only sporadically in samples upstream from the effluent outfall. Metabolite to parent ratios were used to evaluate transformation, and we determined that ratios in wastewater were much lower than those reported in urine, indicating that the metabolites are relatively more labile than the parent compounds in the treatment plants and in receiving waters. The widespread occurrence of neuro-active pharmaceuticals and metabolites in Minnesota effluents and surface waters indicate that this is likely a global environmental issue

  6. All-trans retinoic acid inhibits vascular endothelial growth factor expression in a cell model of neutrophil activation.

    PubMed

    Tee, Meng Kian; Vigne, Jean-Louis; Taylor, Robert N

    2006-03-01

    Infiltrating neutrophil granulocytes are a particularly rich source of vascular endothelial growth factor (VEGF) in the endometrium and may contribute to the angiogenesis of endometriosis lesions. The objective of this study is to evaluate the expression and regulation of VEGF in endometrial neutrophils and in a model of neutrophil differentiation relevant to endometriosis. Immunohistochemistry was performed on endometriosis patient biopsies and cultured neutrophil-like HL-60 cells were assessed. The study was set in a reproductive biology division within an academic medical center. Endometrial biopsies were performed on women with endometriosis and HL-60 cells were treated with all-trans retinoic acid (atRA) and dimethyl sulfoxide in vitro. Immunofluorescence histochemistry, VEGF mRNA and protein quantification, and transfection studies of VEGF gene promoter-luciferase constructs were all main outcome measures. Immunofluorescence studies verified the presence of neutrophils in eutopic endometrium from women with endometriosis. Examination of the regulation of VEGF using differentiated HL-60 cells as a model, revealed that atRA induced a dose- and time-dependent suppression of VEGF mRNA and protein. Transient transfection, truncation, EMSA, and site-directed mutagenesis of human VEGF promoter-luciferase constructs in HL-60 cells indicated that atRA repressed VEGF gene transcription via a direct repeat 1 element located between -443 and -431 bp relative to the transcription initiation site. Because retinoic acid is synthesized de novo in endometrial cells under the influence of progesterone, our findings suggest that the up-regulated VEGF and angiogenesis in tissue from women with endometriosis may reflect failure of neutrophil differentiation in these cases, and provide a rationale for retinoid therapy in this condition. PMID:16322068

  7. Ardipusilloside-I Metabolites from Human Intestinal Bacteria and Their Antitumor Activity.

    PubMed

    Cao, Wei-Yu; Wang, Ya-Nan; Wang, Peng-Yuan; Lei, Wan; Feng, Bin; Wang, Xiao-Juan

    2015-01-01

    Ardipusilloside-I (ADS-I) is a triterpenoid saponin extracted from Ardisia pusilla DC, and has been demonstrated to have potent antitumor activity. However, ADS-I metabolism in humans has not been investigated. In this study, we studied the biotransformation of ADS-I in human intestinal bacteria, and examined the in vitro antitumor activity of the major metabolites. Ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) was used to detect ADS-I biotransformation products, and their chemical structures were identified by high performance liquid chromatography-nuclear magnetic resonance (HPLC-NMR). The antitumor activity of the major metabolites was determined by the MTT assay. Here, we show that main reaction seen in the metabolism of ADS-I in human intestinal bacteria was deglycosylation, which produced a total of four metabolites. The structures of the two major metabolites M1 and M2 were confirmed by using NMR. MTT assay showed that ADS-I metabolites M1 and M2 have the same levels of inhibitory activities as ADS-I in cultured SMMC-7721 cells and MCF-7 cells. In conclusion, this study demonstrates deglycosylation as a primary pathway of ADS-I metabolism in human intestinal bacteria, and suggests that the pharmacological activity of ADS-I may be mediated, at least in part, by its metabolites. PMID:26610438

  8. Antiproliferative and hepatoprotective activity of metabolites from Corynebacterium xerosis against Ehrlich Ascites Carcinoma cells

    PubMed Central

    Islam, Farhadul; Ghosh, Soby; Khanam, Jahan Ara

    2014-01-01

    Objective To find out the effective anticancer drugs from bacterial products, petroleum ether extract of Corynebacterium xerosis. Methods Antiproliferative activity of the metabolite has been measured by monitoring the parameters like tumor weight measurement, tumor cell growth inhibition in mice and survival time of tumor bearing mice, etc. Hepatoprotective effect of the metabolites was determined by observing biochemical, hematological parameters. Results It has been found that the petroleum ether extract bacterial metabolite significantly decrease cell growth (78.58%; P<0.01), tumor weight (36.04 %; P<0.01) and increase the life span of tumor bearing mice (69.23%; P<0.01) at dose 100 mg/kg (i.p.) in comparison to those of untreated Ehrlich ascites carcinoma (EAC) bearing mice. The metabolite also alters the depleted hematological parameters like red blood cell, white blood cell, hemoglobin (Hb%), etc. towards normal in tumor bearing mice. Metabolite show no adverse effect on liver functions regarding blood glucose, serum alkaline phosphatases, glutamic pyruvic transaminase, glutamic oxaloacetic transaminase activity and serum billirubin, etc. in normal mice. Histopathological observation of these mice organ does not show any toxic effect on cellular structure. But in the case of EAC bearing untreated mice these hematological and biochemical parameters deteriorate extremely with time whereas petroleum ether extract bacterial metabolite receiving EAC bearing mice nullified the toxicity induced by EAC cells. Conclusion Study results reveal that metabolite possesses significant antiproliferative and hepatoprotective effect against EAC cells. PMID:25183099

  9. Retinoic acid receptors: from molecular mechanisms to cancer therapy.

    PubMed

    di Masi, Alessandra; Leboffe, Loris; De Marinis, Elisabetta; Pagano, Francesca; Cicconi, Laura; Rochette-Egly, Cécile; Lo-Coco, Francesco; Ascenzi, Paolo; Nervi, Clara

    2015-02-01

    Retinoic acid (RA), the major bioactive metabolite of retinol or vitamin A, induces a spectrum of pleiotropic effects in cell growth and differentiation that are relevant for embryonic development and adult physiology. The RA activity is mediated primarily by members of the retinoic acid receptor (RAR) subfamily, namely RARα, RARβ and RARγ, which belong to the nuclear receptor (NR) superfamily of transcription factors. RARs form heterodimers with members of the retinoid X receptor (RXR) subfamily and act as ligand-regulated transcription factors through binding specific RA response elements (RAREs) located in target genes promoters. RARs also have non-genomic effects and activate kinase signaling pathways, which fine-tune the transcription of the RA target genes. The disruption of RA signaling pathways is thought to underlie the etiology of a number of hematological and non-hematological malignancies, including leukemias, skin cancer, head/neck cancer, lung cancer, breast cancer, ovarian cancer, prostate cancer, renal cell carcinoma, pancreatic cancer, liver cancer, glioblastoma and neuroblastoma. Of note, RA and its derivatives (retinoids) are employed as potential chemotherapeutic or chemopreventive agents because of their differentiation, anti-proliferative, pro-apoptotic, and anti-oxidant effects. In humans, retinoids reverse premalignant epithelial lesions, induce the differentiation of myeloid normal and leukemic cells, and prevent lung, liver, and breast cancer. Here, we provide an overview of the biochemical and molecular mechanisms that regulate the RA and retinoid signaling pathways. Moreover, mechanisms through which deregulation of RA signaling pathways ultimately impact on cancer are examined. Finally, the therapeutic effects of retinoids are reported. PMID:25543955

  10. Evidence for impaired retinoic acid receptor-thyroid hormone receptor AF-2 cofactor activity in human lung cancer.

    PubMed Central

    Moghal, N; Neel, B G

    1995-01-01

    Retinoic acid (RA) is required for normal airway epithelial cell growth and differentiation both in vivo and in vitro. One of the earliest events following the exposure of bronchial epithelial cells to RA is the strong induction of RA receptor beta (RAR beta) mRNA. Previous work established that many lung cancer cell lines and primary tumors display abnormal RAR beta mRNA expression, most often absence or weak expression of the RAR beta 2 isoform, even after RA treatment. Restoration of RAR beta 2 into RAR beta-negative lung cancer cell lines has been reported to inhibit tumorigenicity. Since RAR beta 2 inactivation may contribute to lung cancer, we have investigated the molecular mechanism of defective RAR beta 2 expression. Nuclear run-on assays and transient transfections with RAR beta 2 promoter constructs indicate the presence of trans-acting transcriptional defects in most lung cancer cell lines, which map to the RA response element (RARE). These defects cannot be complemented by RAR-retinoid X receptor cotransfection and can be separated into two types: (i) one affecting transcription from direct repeat RAREs, but not palindromic RAREs, and (ii) another affecting transcription from both types of RARE. Studies using chimeras between RAR alpha, TR alpha, and other transcription factors suggest the existence of novel RAR-thyroid hormone receptor AF-2-specific cofactors, which are necessary for high levels of transcription. Furthermore, these factors may be frequently inactivated in human lung cancer. PMID:7791800

  11. NIR and visible investigation of some potential SERS-active substrates for studying antitumour agent all- trans retinoic acid

    NASA Astrophysics Data System (ADS)

    Beljebbar, A.; Sockalingum, G. D.; Morjani, H.; Angiboust, J. F.; Manfait, M.

    1997-01-01

    Red and near-infrared excited Fourier transform surface-enhanced Raman spectra of an anticancer agent, all- trans retinoic acid (ATRA), adsorbed on gold island films are reported. Best results have been obtained with plates 80 Å and 40 Å thick respectively in the red and near-infrared and at concentrations of 10 -5 and 5 × 10 -6 M with a spinning system. The use of near-infrared laser excitation with low photon energy, allows us to overcome the problems of isomerisation when the sample is exposed for a long time to the laser radiation. Comparison between the Raman and SERS spectra in the visible shows that the adsorption on the surface does not perturb the structure of ATRA and confirms the long range enhancement of the island films with this type of molecule. Spectral data show that while gold island films and colloids are appropriate substrates for use with red excitation, silver and gold colloids as well as gold island films exhibit satisfactory enhancement levels in the near-infrared. This study will in the future allow us to choose the appropriate system that will serve to investigate the interaction of ATRA with its target in vitro and the effect of this differentiating agent in human leukaemia cell lines such as K562 and HL60.

  12. Antiviral activity of human oligoadenylate synthetases-like (OASL) is mediated by enhancing retinoic acid-inducible gene I (RIG-I) signaling

    PubMed Central

    Zhu, Jianzhong; Zhang, Yugen; Ghosh, Arundhati; Cuevas, Rolando A.; Forero, Adriana; Dhar, Jayeeta; Ibsen, Mikkel Søes; Schmid-Burgk, Jonathan Leo; Schmidt, Tobias; Ganapathiraju, Madhavi K.; Fujita, Takashi; Hartmann, Rune; Barik, Sailen; Hornung, Veit; Coyne, Carolyn B.; Sarkar, Saumendra N.

    2014-01-01

    SUMMARY Virus infection is sensed in the cytoplasm by retinoic acid-inducible gene I (RIG-I, also known as DDX58), which requires RNA and polyubiquitin binding to induce type I interferon (IFN), and activate cellular innate immunity. We show that the human IFN-inducible oligoadenylate synthetases-like (OASL) protein had antiviral activity and mediated RIG-I activation by mimicking polyubiquitin. Loss of OASL expression reduced RIG-I signaling and enhanced virus replication in human cells. Conversely, OASL expression suppressed replication of a number of viruses in a RIG-I-dependent manner and enhanced RIG-I-mediated IFN induction. OASL interacted and colocalized with RIG-I, and through its C-terminal ubiquitin-like domain specifically enhanced RIG-I signaling. Bone marrow derived macrophages from mice deficient for Oasl2 showed that among the two mouse orthologs of human OASL; Oasl2 is functionally similar to human OASL. Our findings show a mechanism by which human OASL contributes to host antiviral responses by enhancing RIG-I activation. PMID:24931123

  13. The dual nature of retinoic acid in pemphigus and its therapeutic potential: Special focus on all-trans Retinoic Acid.

    PubMed

    Tavakolpour, Soheil; Daneshpazhooh, Maryam; Mahmoudi, Hamid Reza; Balighi, Kamran

    2016-07-01

    The efficient treatment of pemphigus with no certain side effect remained a controversial issue. Although there are various options for controlling disease severity, the majority of them may cause serious side effects. Retinoic acid (RA), an active metabolite converted from vitamin A, plays an active role in immune functions. Effects of RA, especially all-trans-Retinoic Acid (ATRA) on different types of cells involved in immune responses were analyzed in vitro and in vivo. RAs could affect the differentiation of T helper (Th) cells, B cells responses, stabilization of both natural regulatory T cells (nTregs) and regulatory B cells (Bregs) populations, and regulating the expression of critical genes in immune responses. The role of RA, based on major immune cells involved in pemphigus has not been addressed so far. In this study, we sought to determine the possible effects of RA, with a special focus on ATRA in pemphigus. All the evidences of ATRA effects on the immune system were collected and their association with the pemphigus was analyzed. According to the previous results, ATRA causes a decline in Th17 populations; increase in CD4+ induced regulatory T cells (iTregs), stabilization of nTregs, and promotion of suppressive B cells, which are critical in the improvement of pemphigus. Nevertheless, it also causes shifting of the Th1:Th2 balance toward Th2 cells, which is not favorable for pemphigus patients. In conclusion, ATRA acts via different ways in pemphigus. Due to increase in the suppressive function via iTregs, nTregs, and Bregs, it is suggested that patients with pemphigus may benefit from systemic ATRA therapy. To clarify this issue, further studies, such as clinical trials are needed. PMID:27156125

  14. Association between cytoplasmic CRABP2, altered retinoic acid signaling, and poor prognosis in glioblastoma.

    PubMed

    Liu, Rong-Zong; Li, Shuai; Garcia, Elizabeth; Glubrecht, Darryl D; Yin Poon, Ho; Easaw, Jacob C; Godbout, Roseline

    2016-06-01

    Retinoic acid (RA), a metabolite of vitamin A, is required for the regulation of growth and development. Aberrant expression of molecules involved in RA signaling has been reported in various cancer types including glioblastoma multiforme (GBM). Cellular retinoic acid-binding protein 2 (CRABP2) has previously been shown to play a key role in the transport of RA to retinoic acid receptors (RARs) to activate their transcription regulatory activity. Here, we demonstrate that CRABP2 is predominantly located in the cytoplasm of GBM tumors. Cytoplasmic, but not nuclear, CRABP2 levels in GBM tumors are associated with poor patient survival. Treatment of malignant glioma cell lines with RA results in a dose-dependent increase in accumulation of CRABP2 in the cytoplasm. CRABP2 knockdown reduces proliferation rates of malignant glioma cells, and enhances RA-induced RAR activation. Levels of CRYAB, a small heat shock protein with anti-apoptotic activity, and GFAP, an astrocyte-specific intermediate filament protein, are greatly reduced in CRABP2-depleted cells. Restoration of CRYAB expression partially but significantly reversed the effect of CRABP2 depletion on RAR activation. Our combined in vivo and in vitro data indicate that: (i) CRABP2 is an important determinant of clinical outcome in GBM patients, and (ii) the mechanism of action of CRABP2 in GBM involves sequestration of RA in the cytoplasm and activation of an anti-apoptotic pathway, thereby enhancing proliferation and preventing RA-mediated cell death and differentiation. We propose that reducing CRABP2 levels may enhance the therapeutic index of RA in GBM patients. GLIA 2016;64:963-976. PMID:26893190

  15. Retinoic Acid Protects Cardiomyocytes from High Glucose-Induced Apoptosis via Inhibition of Sustained Activation of NF-κB Signaling

    PubMed Central

    Nizamutdinova, Irina T.; Guleria, Rakeshwar S.; Singh, Amar B.; Kendall, Jonathan A.; Baker, Kenneth M.; Pan, Jing

    2012-01-01

    We have previously shown that retinoic acid (RA) has protective effects on high glucose (HG)-induced cardiomyocyte apoptosis. To further elucidate the molecular mechanisms of RA effects, we determined the interaction between nuclear factor (NF)-κB and RA signaling. HG induced a sustained phosphorylation of IKK/IκBα and transcriptional activation of NF-κB in cardiomyocytes. Activated NF-κB signaling has an important role in HG-induced cardiomyocyte apoptosis and gene expression of interleukin-6 (IL-6), tumor necrosis factor (TNF)-α and monocyte chemoattractant protein-1 (MCP-1). All-trans RA (ATRA) and LGD1069, through activation of RAR/RXR-mediated signaling, inhibited the HG-mediated effects in cardiomyocytes. The inhibitory effect of RA on NF-κB activation was mediated through inhibition of IKK/IκBα phosphorylation. ATRA and LGD1069 treatment promoted protein phosphatase 2A (PP2A) activity, which was significantly suppressed by HG stimulation. The RA effects on IKK and IκBα were blocked by okadaic acid or silencing the expression of PP2Ac-subunit, indicating that the inhibitory effect of RA on NF-κB is regulated through activation of PP2A and subsequent dephosphorylation of IKK/IκBα. Moreover, ATRA and LGD1069 reversed the decreased PP2A activity and inhibited the activation of IKK/IκBα and gene expression of MCP-1, IL-6 and TNF-α in the hearts of Zucker diabetic fatty rats. In summary, our findings suggest that the suppressed activation of PP2A contributed to sustained activation of NF-κB in HG-stimulated cardiomyocytes; and that the protective effect of RA on hyperglycemia-induced cardiomyocyte apoptosis and inflammatory responses is partially regulated through activation of PP2A and suppression of NF-κB-mediated signaling and downstream targets. PMID:22718360

  16. Diversity of Secondary Metabolites from Marine Bacillus Species: Chemistry and Biological Activity

    PubMed Central

    Mondol, Muhammad Abdul Mojid; Shin, Hee Jae; Islam, Mohammad Tofazzal

    2013-01-01

    Marine Bacillus species produce versatile secondary metabolites including lipopeptides, polypeptides, macrolactones, fatty acids, polyketides, and isocoumarins. These structurally diverse compounds exhibit a wide range of biological activities, such as antimicrobial, anticancer, and antialgal activities. Some marine Bacillus strains can detoxify heavy metals through reduction processes and have the ability to produce carotenoids. The present article reviews the chemistry and biological activities of secondary metabolites from marine isolates. Side by side, the potential for application of these novel natural products from marine Bacillus strains as drugs, pesticides, carotenoids, and tools for the bioremediation of heavy metal toxicity are also discussed. PMID:23941823

  17. Diversity of secondary metabolites from marine Bacillus species: chemistry and biological activity.

    PubMed

    Mondol, Muhammad Abdul Mojid; Shin, Hee Jae; Islam, Mohammad Tofazzal

    2013-08-01

    Marine Bacillus species produce versatile secondary metabolites including lipopeptides, polypeptides, macrolactones, fatty acids, polyketides, and isocoumarins. These structurally diverse compounds exhibit a wide range of biological activities, such as antimicrobial, anticancer, and antialgal activities. Some marine Bacillus strains can detoxify heavy metals through reduction processes and have the ability to produce carotenoids. The present article reviews the chemistry and biological activities of secondary metabolites from marine isolates. Side by side, the potential for application of these novel natural products from marine Bacillus strains as drugs, pesticides, carotenoids, and tools for the bioremediation of heavy metal toxicity are also discussed. PMID:23941823

  18. Transcriptional Factors Mediating Retinoic Acid Signals in the Control of Energy Metabolism.

    PubMed

    Zhang, Rui; Wang, Yueqiao; Li, Rui; Chen, Guoxun

    2015-01-01

    Retinoic acid (RA), an active metabolite of vitamin A (VA), is important for many physiological processes including energy metabolism. This is mainly achieved through RA-regulated gene expression in metabolically active cells. RA regulates gene expression mainly through the activation of two subfamilies in the nuclear receptor superfamily, retinoic acid receptors (RARs) and retinoid X receptors (RXRs). RAR/RXR heterodimers or RXR/RXR homodimers bind to RA response element in the promoters of RA target genes and regulate their expressions upon ligand binding. The development of metabolic diseases such as obesity and type 2 diabetes is often associated with profound changes in the expressions of genes involved in glucose and lipid metabolism in metabolically active cells. RA regulates some of these gene expressions. Recently, in vivo and in vitro studies have demonstrated that status and metabolism of VA regulate macronutrient metabolism. Some studies have shown that, in addition to RARs and RXRs, hepatocyte nuclear factor 4α, chicken ovalbumin upstream promoter-transcription factor II, and peroxisome proliferator activated receptor β/δ may function as transcriptional factors mediating RA response. Herein, we summarize current progresses regarding the VA metabolism and the role of nuclear receptors in mediating RA signals, with an emphasis on their implication in energy metabolism. PMID:26110391

  19. Transcriptional Factors Mediating Retinoic Acid Signals in the Control of Energy Metabolism

    PubMed Central

    Zhang, Rui; Wang, Yueqiao; Li, Rui; Chen, Guoxun

    2015-01-01

    Retinoic acid (RA), an active metabolite of vitamin A (VA), is important for many physiological processes including energy metabolism. This is mainly achieved through RA-regulated gene expression in metabolically active cells. RA regulates gene expression mainly through the activation of two subfamilies in the nuclear receptor superfamily, retinoic acid receptors (RARs) and retinoid X receptors (RXRs). RAR/RXR heterodimers or RXR/RXR homodimers bind to RA response element in the promoters of RA target genes and regulate their expressions upon ligand binding. The development of metabolic diseases such as obesity and type 2 diabetes is often associated with profound changes in the expressions of genes involved in glucose and lipid metabolism in metabolically active cells. RA regulates some of these gene expressions. Recently, in vivo and in vitro studies have demonstrated that status and metabolism of VA regulate macronutrient metabolism. Some studies have shown that, in addition to RARs and RXRs, hepatocyte nuclear factor 4α, chicken ovalbumin upstream promoter-transcription factor II, and peroxisome proliferator activated receptor β/δ may function as transcriptional factors mediating RA response. Herein, we summarize current progresses regarding the VA metabolism and the role of nuclear receptors in mediating RA signals, with an emphasis on their implication in energy metabolism. PMID:26110391

  20. Evaluation of Bacillus cereus and Bacillus pumilus metabolites for anthelmintic activity

    PubMed Central

    Kumar, M. L. Vijaya; Thippeswamy, B.; Kuppust, I. L.; Naveenkumar, K. J.; Shivakumar, C. K.

    2015-01-01

    Objective: To assess the anthelmintic acivity of Bacillus cereus and Bacillus pumilus metabolites. Materials and Methods: The successive solvent extractions with petroleum ether, ethyl acetate and methanol. The solvent extracts were tested for anthelmintic activity against Pheretima posthuma at 20 mg/ml concentration. The time of paralysis and time of death of the worms was determined for all the extracts. Albendazole was taken as a standard reference and sterile water as a control. Results: All the sample extracts showed significant anthelmintic activity in paralyzing the worms comparable with that of the standard drug. The time of death exhibited by BP metabolites was close to the time exhibited by standard. Conclusion: The study indicates both bacteria Bacillus cereus and Bacillus pumilus have anthelmintic activity indicating potential metabolites in them. PMID:25598639

  1. Curcumin Pharmacokinetic and Pharmacodynamic Evidences in Streptozotocin-Diabetic Rats Support the Antidiabetic Activity to Be via Metabolite(s)

    PubMed Central

    Gutierres, Vânia Ortega; Campos, Michel Leandro; Arcaro, Carlos Alberto; Assis, Renata Pires; Baldan-Cimatti, Helen Mariana; Peccinini, Rosângela Gonçalves; Paula-Gomes, Silvia; Kettelhut, Isis Carmo; Baviera, Amanda Martins; Brunetti, Iguatemy Lourenço

    2015-01-01

    This study measures the curcumin concentration in rat plasma by liquid chromatography and investigates the changes in the glucose tolerance and insulin sensitivity of streptozotocin-diabetic rats treated with curcumin-enriched yoghurt. The analytical method for curcumin detection was linear from 10 to 500 ng/mL. The Cmax⁡ and the time to reach Cmax⁡ (tmax⁡) of curcumin in plasma were 3.14 ± 0.9 μg/mL and 5 minutes (10 mg/kg, i.v.) and 0.06 ± 0.01 μg/mL and 14 minutes (500 mg/kg, p.o.). The elimination half-time was 8.64 ± 2.31 (i.v.) and 32.70 ± 12.92 (p.o.) minutes. The oral bioavailability was about 0.47%. Changes in the glucose tolerance and insulin sensitivity were investigated in four groups: normal and diabetic rats treated with yoghurt (NYOG and DYOG, resp.) and treated with 90 mg/kg/day curcumin incorporated in yoghurt (NC90 and DC90, resp.). After 15 days of treatment, the glucose tolerance and the insulin sensitivity were significantly improved in DC90 rats in comparison with DYOG, which can be associated with an increase in the AKT phosphorylation levels and GLUT4 translocation in skeletal muscles. These findings can explain, at least in part, the benefits of curcumin-enriched yoghurt to diabetes and substantiate evidences for the curcumin metabolite(s) as being responsible for the antidiabetic activity. PMID:26064170

  2. Stress-induced NF-κB activation differentiates promyelocytic leukemia cells to macrophages in response to all-trans-retinoic acid.

    PubMed

    Imran, Muhammad; Park, Joon Seong; Lim, In Kyoung

    2015-03-01

    All-trans-retinoic acid (ATRA) has been known as a choice of treatment for inducing differentiation of promyelocytic leukemia cells to granulocytes. NF-κB plays a crucial role in inflammation and immunity and its activation is an important event for macrophage differentiation both in vivo and in vitro. We report here that NF-κB activation is critical for determining ATRA-induced lineage specific differentiation of myeloid leukemia cells. Our data revealed that ATRA treatment to HL-60 cells enhanced IκBα degradation and NF-κB nuclear translocation and the activated NF-κB potentiated the ability of ATRA for differentiation and switched differentiation to macrophages instead of granulocytes. Serum withdrawal and LPS treatment dampened IκBα expression via MAPK activation and reactive oxygen species generation leading to NF-κB nuclear translocation and ATRA treatment further corroborated these effects in myeloid leukemia cells. Activated NF-κB enhanced the degree of ATRA-induced differentiation of HL-60 cells to macrophages, rather than granulocytes, as assessed by morphologic examination and expressions of differentiation markers such as CD11b, CD38, CD68, MMP9 and Btg2. Employing LLnL or dominant negative IκBα attenuated NF-κB associated enhanced cell maturation and differentiation switch thus suggesting NF-κB as one of the factors that determines ATRA induced lineage specificity of myeloid leukemia cells. Furthermore, MAPK activation was observed to be central both for the differentiation of promyelocytic cells to macrophages or granulocytes regulating NF-κB or C/EBPα expressions, respectively; however, MAPK-mediated signals are modulated under various conditions affecting lineage specificity. In summary, our present data demonstrate that activation of NF-κB directly affects differentiation program of promyelocytes to macrophages, rather than granulocyte, in response to ATRA treatment. PMID:25435432

  3. Widespread occurrence of neuro-active pharmaceuticals and metabolites in 24 Minnesota rivers and wastewaters

    USGS Publications Warehouse

    Writer, Jeffrey; Ferrer, Imma; Barber, Larry B.; Thurman, E. Michael

    2013-01-01

    Concentrations of 17 neuro-active pharmaceuticals and their major metabolites (bupropion, hydroxy-bupropion, erythro-hydrobupropion, threo-hydrobupropion, carbamazepine, 10,11,-dihydro-10,11,-dihydroxycarbamazepine, 10-hydroxy-carbamazepine, citalopram, N-desmethyl-citalopram, fluoxetine, norfluoxetine, gabapentin, lamotrigine, 2-N-glucuronide-lamotrigine, oxcarbazepine, venlafaxine and O-desmethyl-venlafaxine), were measured in treated wastewater and receiving surface waters from 24 locations across Minnesota, USA. The analysis of upstream and downstream sampling sites indicated that the wastewater treatment plants were the major source of the neuro-active pharmaceuticals and associated metabolites in surface waters of Minnesota. Concentrations of parent compound and the associated metabolite varied substantially between treatment plants (concentrations ± standard deviation of the parent compound relative to its major metabolite) as illustrated by the following examples; bupropion and hydrobupropion 700 ± 1000 ng L−1, 2100 ± 1700 ng L−1, carbamazepine and 10-hydroxy-carbamazepine 480 ± 380 ng L−1, 360 ± 400 ng L−1, venlafaxine and O-desmethyl-venlafaxine 1400 ± 1300 ng L−1, 1800 ± 2300 ng L−1. Metabolites of the neuro-active compounds were commonly found at higher or comparable concentrations to the parent compounds in wastewater effluent and the receiving surface water. Neuro-active pharmaceuticals and associated metabolites were detected only sporadically in samples upstream from the effluent outfall. Metabolite to parent ratios were used to evaluate transformation, and we determined that ratios in wastewater were much lower than those reported in urine, indicating that the metabolites are relatively more labile than the parent compounds in the treatment plants and in receiving waters. The widespread occurrence of neuro-active pharmaceuticals and metabolites in Minnesota effluents and surface waters indicate that

  4. TRIM32 promotes neural differentiation through retinoic acid receptor-mediated transcription.

    PubMed

    Sato, Tomonobu; Okumura, Fumihiko; Kano, Satoshi; Kondo, Takeshi; Ariga, Tadashi; Hatakeyama, Shigetsugu

    2011-10-15

    Retinoic acid (RA), a metabolite of vitamin A, plays versatile roles in development, differentiation, cell cycles and regulation of apoptosis by regulating gene transcription through nuclear receptor activation. Ubiquitinylation, which is one of the post-translational modifications, appears to be involved in the transcriptional activity of intranuclear receptors including retinoic acid receptor α (RARα). Mutations in the tripartite motif-containing protein 32 gene (TRIM32; also known as E3 ubiquitin-protein ligase) have been reported to be responsible for limb-girdle muscular dystrophy type 2H in humans, and its encoded protein has been shown to interact with several other important proteins. In this study, we found that TRIM32 interacts with RARα and enhances its transcriptional activity in the presence of RA. We also found that overexpression of TRIM32 in mouse neuroblastoma cells and embryonal carcinoma cells promoted stability of RARα, resulting in enhancement of neural differentiation. These findings suggest that TRIM32 functions as one of the co-activators for RARα-mediated transcription, and thereby TRIM32 is a potential therapeutic target for developmental disorders and RA-dependent leukemias. PMID:21984809

  5. Anti-Oxidative Activity of Mytiloxanthin, a Metabolite of Fucoxanthin in Shellfish and Tunicates

    PubMed Central

    Maoka, Takashi; Nishino, Azusa; Yasui, Hiroyuki; Yamano, Yumiko; Wada, Akimori

    2016-01-01

    Anti-oxidative activities of mytiloxanthin, a metabolite of fucoxanthin in shellfish and tunicates, were investigated. Mytiloxanthin showed almost the same activities for quenching singlet oxygen and the inhibition of lipid peroxidation as those of astaxanthin, which is a well-known singlet oxygen quencher. Furthermore, mytiloxanthin showed excellent scavenging activity for hydroxyl radicals and this activity was markedly higher than that of astaxanthin. PMID:27187417

  6. Anti-Oxidative Activity of Mytiloxanthin, a Metabolite of Fucoxanthin in Shellfish and Tunicates.

    PubMed

    Maoka, Takashi; Nishino, Azusa; Yasui, Hiroyuki; Yamano, Yumiko; Wada, Akimori

    2016-01-01

    Anti-oxidative activities of mytiloxanthin, a metabolite of fucoxanthin in shellfish and tunicates, were investigated. Mytiloxanthin showed almost the same activities for quenching singlet oxygen and the inhibition of lipid peroxidation as those of astaxanthin, which is a well-known singlet oxygen quencher. Furthermore, mytiloxanthin showed excellent scavenging activity for hydroxyl radicals and this activity was markedly higher than that of astaxanthin. PMID:27187417

  7. All-Trans Retinoic Acid Induces Proliferation, Survival, and Migration in A549 Lung Cancer Cells by Activating the ERK Signaling Pathway through a Transcription-Independent Mechanism

    PubMed Central

    Quintero Barceinas, Reyna Sara; García-Regalado, Alejandro; Aréchaga-Ocampo, Elena; Villegas-Sepúlveda, Nicolás; González-De la Rosa, Claudia Haydée

    2015-01-01

    All-trans retinoic acid (ATRA) has been used as an antineoplastic because of its ability to promote proliferation, inhibition, and differentiation, primarily in leukemia; however, in other types of cancer, such as lung cancer, treatment with ATRA is restricted because not all the patients experience the same results. The ERK signaling pathway is dysregulated in cancer cells, including lung cancer, and this dysregulation promotes proliferation and cell invasion. In this study, we demonstrate that treatment with ATRA can activate the ERK signaling pathway by a transcription-independent mechanism through a signaling cascade that involves RARα and PI3K, promoting growth, survival, and migration in lung cancer cells. Until now, this mechanism was unknown in lung cancer cells. The inhibition of the ERK signaling pathway restores the beneficial effects of ATRA, reduces proliferation, increases apoptosis, and blocks the cell migration process in lung cancer cells. In conclusion, our results suggest that the combination of ATRA with ERK inhibitor in clinical trials for lung cancer is warranted. PMID:26557664

  8. Tissue-specific response of the human platelet-activating factor receptor gene to retinoic acid and thyroid hormone by alternative promoter usage.

    PubMed Central

    Mutoh, H; Fukuda, T; Kitamaoto, T; Masushige, S; Sasaki, H; Shimizu, T; Kato, S

    1996-01-01

    We have studied the effects of retinoic acid (RA) and thyroid hormone (3,3',5-triiodothyronine; T3) on platelet-activating factor receptor (PAFR) gene expression in intact rats and the ability of two human PAFR gene promoters (PAFR promoters 1 and 2) to generate two transcripts (PAFR transcripts 1 and 2). Northern blotting showed that RA and T3 regulated PAFR gene expression only in rat tissues that express PAFR transcript 2. Functional analysis of the human PAFR promoter 2 revealed that responsiveness to RA and T3 was conferred through a 24-bp element [PAFR-hormone response element (HRE) located from -67 to -44 bp of the transcription start site, whereas PAFR promoter 1 did not respond to these hormones. The PAFR-HRE is composed of three direct repeated TGACCT-like hexamer motifs with 2-and 4-bp spaces, and the two upstream and two downstream motifs were identified as response elements for RA and T3. Thus, the PAF-PAFR pathway is regulated by the PAFR level altered by a tissue-specific response to RA and T3 through the PAFR-HRE of the PAFR promoter 2. Images Fig. 1 Fig. 2 Fig. 4 Fig. 5 Fig. 6 PMID:8570633

  9. Antifeedant Activity of Ginkgo biloba Secondary Metabolites against Hyphantria cunea Larvae: Mechanisms and Applications.

    PubMed

    Pan, Long; Ren, Lili; Chen, Fang; Feng, Yuqian; Luo, Youqing

    2016-01-01

    Ginkgo biloba is a typical relic plant that rarely suffers from pest hazards. This study analyzed the pattern of G. biloba pest hazards in Beijing; tested the antifeedant activity of G. biloba extracts, including ginkgo flavonoids, ginkgolide, and bilobalide, against Hyphantria cunea larvae; determined the activities of glutathione transferase (GSTs), acetylcholinesterase (AChE), carboxylesterase (CarE) and mixed-functional oxidase (MFO), in larvae after feeding on these G. biloba secondary metabolites; and screened for effective botanical antifeedants in the field. In this study, no indicators of insect infestation were found for any of the examined leaves of G. biloba; all tested secondary metabolites showed significant antifeedant activity and affected the activity of the four larval detoxifying enzymes. Ginkgolide had the highest antifeedant activity and the most significant effect on the detoxifying enzymes (P<0.05). Spraying leaves with G. biloba extracts or ginkgolide both significantly repelled H. cunea larvae in the field (P<0.05), although the former is more economical and practical. This study investigated the antifeedant activity of G. biloba secondary metabolites against H. cunea larvae, and the results provide new insights into the mechanism of G. biloba pest resistance. This study also developed new applications of G. biloba secondary metabolites for effective pest control. PMID:27214257

  10. Antifeedant Activity of Ginkgo biloba Secondary Metabolites against Hyphantria cunea Larvae: Mechanisms and Applications

    PubMed Central

    Ren, Lili; Chen, Fang; Feng, Yuqian

    2016-01-01

    Ginkgo biloba is a typical relic plant that rarely suffers from pest hazards. This study analyzed the pattern of G. biloba pest hazards in Beijing; tested the antifeedant activity of G. biloba extracts, including ginkgo flavonoids, ginkgolide, and bilobalide, against Hyphantria cunea larvae; determined the activities of glutathione transferase (GSTs), acetylcholinesterase (AChE), carboxylesterase (CarE) and mixed-functional oxidase (MFO), in larvae after feeding on these G. biloba secondary metabolites; and screened for effective botanical antifeedants in the field. In this study, no indicators of insect infestation were found for any of the examined leaves of G. biloba; all tested secondary metabolites showed significant antifeedant activity and affected the activity of the four larval detoxifying enzymes. Ginkgolide had the highest antifeedant activity and the most significant effect on the detoxifying enzymes (P<0.05). Spraying leaves with G. biloba extracts or ginkgolide both significantly repelled H. cunea larvae in the field (P<0.05), although the former is more economical and practical. This study investigated the antifeedant activity of G. biloba secondary metabolites against H. cunea larvae, and the results provide new insights into the mechanism of G. biloba pest resistance. This study also developed new applications of G. biloba secondary metabolites for effective pest control. PMID:27214257

  11. Estrogenic activities of diuron metabolites in female Nile tilapia (Oreochromis niloticus).

    PubMed

    Pereira, Thiago Scremin Boscolo; Boscolo, Camila Nomura Pereira; Felício, Andreia Arantes; Batlouni, Sergio Ricardo; Schlenk, Daniel; de Almeida, Eduardo Alves

    2016-03-01

    Some endocrine disrupting chemicals (EDCs) can alter the estrogenic activities of the organism by directly interacting with estrogen receptors (ER) or indirectly through the hypothalamus-pituitary-gonadal axis. Recent studies in male Nile tilapia (Oreochromis niloticus) indicated that diuron may have anti-androgenic activity augmented by biotransformation. In this study, the effects of diuron and three of its metabolites were evaluated in female tilapia. Sexually mature female fish were exposed for 25 days to diuron, as well as to its metabolites 3,4-dichloroaniline (DCA), 3,4-dichlorophenylurea (DCPU) and 3,4-dichlorophenyl-N-methylurea (DCPMU), at concentrations of 100 ng/L. Diuron metabolites caused increases in E2 plasma levels, gonadosomatic indices and in the percentage of final vitellogenic oocytes. Moreover, diuron and its metabolites caused a decrease in germinative cells. Significant differences in plasma concentrations of the estrogen precursor and gonadal regulator17α-hydroxyprogesterone (17α-OHP) were not observed. These results show that diuron metabolites had estrogenic effects potentially mediated through enhanced estradiol biosynthesis and accelerated the ovarian development of O. niloticus females. PMID:26741556

  12. The antiproliferative activity of all-trans-retinoic acid catabolites and isomers is differentially modulated by liarozole-fumarate in MCF-7 human breast cancer cells.

    PubMed Central

    Van heusden, J.; Wouters, W.; Ramaekers, F. C.; Krekels, M. D.; Dillen, L.; Borgers, M.; Smets, G.

    1998-01-01

    The clinical use of all-trans-retinoic acid (ATRA) in the treatment of cancer is significantly hampered by the prompt emergence of resistance, believed to be caused by increased ATRA catabolism. Inhibitors of ATRA catabolism may therefore prove valuable for cancer therapy. Liarozole-fumarate is an anti-tumour drug that inhibits the cytochrome P450-dependent catabolism of ATRA. ATRA, but also its naturally occurring catabolites, 4-oxo-ATRA and 5,6-epoxy-ATRA, as well as its stereoisomers, 9-cis-RA and 13-cis-RA, show significant antiproliferative activity in MCF-7 human breast cancer cells. To further elucidate its mechanism of action, we investigated whether liarozole-fumarate was able to enhance the antiproliferative activity of ATRA catabolites and isomers. Liarozole-fumarate alone up to a concentration of 10(-6) M had no effect on MCF-7 cell proliferation. However, in combination with ATRA or the ATRA catabolites, liarozole-fumarate (10(-6) M) significantly enhanced their antiproliferative activity. On the contrary, liarozole-fumarate (10(-6) M) was not able to potentiate the antiproliferative activity of the ATRA stereoisomers, most probably because of the absence of cytochrome P450-dependent catabolism. Together, these findings show that liarozole-fumarate acts as a versatile inhibitor of retinoid catabolism in that it not only blocks the breakdown of ATRA, but also inhibits the catabolic pathway of 4-oxo-ATRA and 5,6-epoxy-ATRA, thereby enhancing their antiproliferative activity. PMID:9579827

  13. Retinoic Acid Ameliorates Pancreatic Fibrosis and Inhibits the Activation of Pancreatic Stellate Cells in Mice with Experimental Chronic Pancreatitis via Suppressing the Wnt/β-Catenin Signaling Pathway

    PubMed Central

    Yin, Guojian; Fan, Yuting; Wu, Deqing; Qiu, Lei; Yu, Ge; Xing, Miao; Hu, Guoyong; Wang, Xingpeng; Wan, Rong

    2015-01-01

    Pancreatic fibrosis, a prominent feature of chronic pancreatitis (CP), induces persistent and permanent damage in the pancreas. Pancreatic stellate cells (PSCs) provide a major source of extracellular matrix (ECM) deposition during pancreatic injury, and persistent activation of PSCs plays a vital role in the progression of pancreatic fibrosis. Retinoic acid (RA), a retinoid, has a broad range of biological functions, including regulation of cell differentiation and proliferation, attenuating progressive fibrosis of multiple organs. In the present study, we investigated the effects of RA on fibrosis in experimental CP and cultured PSCs. CP was induced in mice by repetitive cerulein injection in vivo, and mouse PSCs were isolated and activated in vitro. Suppression of pancreatic fibrosis upon administration of RA was confirmed based on reduction of histological damage, α-smooth muscle actin (α-SMA) expression and mRNA levels of β-catenin, platelet-derived growth factor (PDGF)-Rβ transforming growth factor (TGF)-βRII and collagen 1α1 in vivo. Wnt 2 and β-catenin protein levels were markedly down-regulated, while Axin 2 expression level was up-regulated in the presence of RA, both in vivo and in vitro. Nuclear translation of β-catenin was significantly decreased following RA treatment, compared with cerulein-induced CP in mice and activated PSCs. Furthermore, RA induced significant PSC apoptosis, inhibited proliferation, suppressed TCF/LEF-dependent transcriptional activity and ECM production of PSC via down-regulation of TGFβRII, PDGFRβ and collagen 1α1 in vitro. These results indicate a critical role of the Wnt/β-catenin signaling pathway in RA-induced effects on CP and PSC regulation and support the potential of RA as a suppressor of pancreatic fibrosis in mice. PMID:26556479

  14. Effect of all-trans retinoic acid on procoagulant and fibrinolytic activities of cultured blast cells from patients with acute promyelocytic leukemia.

    PubMed

    De Stefano, V; Teofili, L; Sica, S; Mastrangelo, S; Di Mario, A; Rutella, S; Salutari, P; Rumi, C; d'Onofrio, G; Leone, G

    1995-11-01

    The mechanisms underlying acute promyelocytic leukemia (APL) coagulopathy and its reversal by administration of all-trans retinoic acid (ATRA) have been investigated. Bone marrow promyelocytic blasts from nine patients with APL were cultured with or without ATRA 1 mumol/L. Cultured blasts (days 0, 3, 6, and 9) were washed, resuspended in phosphate buffer, lysed by freezing and thawing, and then assayed for procoagulant activity (PCA), elastase activity, tissue factor (TF) antigen, tissue-type plasminogen activator (t-PA) antigen and urokinase-type plasminogen activator (u-PA) antigen. PCA was determined by a recalcification assay. Elastase was measured by an amidolytic assay (S-2484). TF, t-PA, and u-PA antigens were measured by an enzyme-linked immunosorbent assay (ELISA). Malignant promyelocytes isolated from the patients had increased levels of PCA and TF as compared with the control polymorphonucleates, and low levels of elastase, t-PA, and u-PA; the patient blast PCA level was significantly related to the degree of hypofibrinogenemia. In this system, blast PCA depended on the tissue factor and was significantly correlated to the TF antigen values. In the cultures without ATRA, PCA, TF, and u-PA progressively increased, whereas elastase and t-PA levels remained essentially unchanged. In the presence of ATRA, all parameters (except u-PA) decreased during the culture time. Thus, a major role of the promyelocytic blast cell PCA in the pathogenesis of M3-related coagulopathy is suggested; the ATRA effect on coagulopathy seems mainly mediated by a downregulation of the PCA. PMID:7579461

  15. Substrate Specificity and Ligand Interactions of CYP26A1, the Human Liver Retinoic Acid Hydroxylase

    PubMed Central

    Thatcher, Jayne E.; Buttrick, Brian; Shaffer, Scott A.; Shimshoni, Jakob A.; Goodlett, David R.; Nelson, Wendel L.

    2011-01-01

    All-trans-retinoic acid (atRA) is the active metabolite of vitamin A. atRA is also used as a drug, and synthetic atRA analogs and inhibitors of retinoic acid (RA) metabolism have been developed. The hepatic clearance of atRA is mediated primarily by CYP26A1, but design of CYP26A1 inhibitors is hindered by lack of information on CYP26A1 structure and structure-activity relationships of its ligands. The aim of this study was to identify the primary metabolites of atRA formed by CYP26A1 and to characterize the ligand selectivity and ligand interactions of CYP26A1. On the basis of high-resolution tandem mass spectrometry data, four metabolites formed from atRA by CYP26A1 were identified as 4-OH-RA, 4-oxo-RA, 16-OH-RA and 18-OH-RA. 9-cis-RA and 13-cis-RA were also substrates of CYP26A1. Forty-two compounds with diverse structural properties were tested for CYP26A1 inhibition using 9-cis-RA as a probe, and IC50 values for 10 inhibitors were determined. The imidazole- and triazole-containing inhibitors [S-(R*,R*)]-N-[4-[2-(dimethylamino)-1-(1H-imidazole-1-yl)propyl]-phenyl]2-benzothiazolamine (R116010) and (R)-N-[4-[2-ethyl-1-(1H-1,2,4-triazol-1-yl)butyl]phenyl]-2-benzothiazolamine (R115866) were the most potent inhibitors of CYP26A1 with IC50 values of 4.3 and 5.1 nM, respectively. Liarozole and ketoconazole were significantly less potent with IC50 values of 2100 and 550 nM, respectively. The retinoic acid receptor (RAR) γ agonist CD1530 was as potent an inhibitor of CYP26A1 as ketoconazole with an IC50 of 530 nM, whereas the RARα and RARβ agonists tested did not significantly inhibit CYP26A1. The pan-RAR agonist 4-[(E)-2-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthalenyl)-1-propenyl]benzoic acid and the peroxisome proliferator-activated receptor ligands rosiglitazone and pioglitazone inhibited CYP26A1 with IC50 values of 3.7, 4.2, and 8.6 μM, respectively. These data demonstrate that CYP26A1 has high ligand selectivity but accepts structurally related nuclear

  16. Rapidly Probing Antibacterial Activity of Graphene Oxide by Mass Spectrometry-based Metabolite Fingerprinting

    PubMed Central

    Zhang, Ning; Hou, Jian; Chen, Suming; Xiong, Caiqiao; Liu, Huihui; Jin, Yulong; Wang, Jianing; He, Qing; Zhao, Rui; Nie, Zongxiu

    2016-01-01

    Application of nanomaterials as anti-bacteria agents has aroused great attention. To investigate the antibacterial activity and antibacterial mechanism of nanomaterials from a molecular perspective is important for efficient developing of nanomaterial antibiotics. In the current work, a new mass spectrometry-based method was established to investigate the bacterial cytotoxicity of graphene oxide (GO) by the metabolite fingerprinting of microbes. The mass spectra of extracted metabolites from two strains DH5α and ATCC25922 were obtained before and after the incubation with nanomaterials respectively. Then principal component analysis (PCA) of these spectra was performed to reveal the relationship between the metabolism disorder of microbes and bactericidal activity of GO. A parameter “D” obtained from PCA scores was proposed that is capable to quantitatively evaluate the antibacterial activity of GO in concentration and time-dependent experiments. Further annotation of the fingerprinting spectra shows the variabilities of important metabolites such as phosphatidylethanolamine, phosphatidylglycerol and glutathione. This metabolic perturbation of E. coli indicates cell membrane destruction and oxidative stress mechanisms for anti-bacteria activity of graphene oxide. It is anticipated that this mass spectrometry-based metabolite fingerprinting method will be applicable to other antibacterial nanomaterials and provide more clues as to their antibacterial mechanism at molecular level. PMID:27306507

  17. Influence of age and caloric restriction on liver glycolytic enzyme activities and metabolite concentrations in mice.

    PubMed

    Hagopian, Kevork; Ramsey, Jon J; Weindruch, Richard

    2003-03-01

    The influence of caloric restriction (CR) from 2 months of age on the activities of liver glycolytic enzymes and metabolite levels was studied in young and old mice. Livers were sampled 48 h after the last scheduled feeding time. Old mice on CR showed significant decreases in the activities of all the enzymes studied, except for aldolase, triosephosphate isomerase and phosphoglycerate mutase, which were unchanged. The metabolites glucose, glucose-6-phosphate, fructose-6-phosphate, pyruvate and lactate were lower while fructose-1,6-bisphosphate, glyceraldehyde-3-phosphate, dihydroxyacetone phosphate, 3-phosphoglycerate and phosphoenolpyruvate were increased in old CR. Young mice on CR also showed reduced enzyme activities, except for aldolase, triosephosphate isomerase and enolase which were unchanged when compared with young controls. The metabolites glucose, glucose-6-phosphate, fructose-6-phosphate and pyruvate were decreased when compared with young controls, while phosphoenolpyruvate was increased. Ketone bodies increased (65%) in old, but not young, CR mice while fructose-2,6-bisphosphate decreased in both young (22%) and old CR (28%) mice. The results indicate that decreased hepatic glucose levels in CR mice are associated with decreased enzyme activities but not a uniform decrease in metabolite levels. Increased ketone body levels indicate increased utilization of non-carbohydrate fuels while decreased fructose-2,6-bisphosphate level suggests its importance in the control of glycolysis in CR. PMID:12581789

  18. Molecular complexes of cocaine, its active metabolites and some other stimulants with thiamine.

    PubMed

    Misra, A L; Vadlamani, N L

    1976-10-01

    Cocaine, its pharmacologically active metabolites, norcocaine benzoylnorecgonine, benzoylecgonine and other central nervous system stimulants e.g. dextrococaine, nicotine, caffeine and p-hydroxy norephedrine formed molecular complexes with thiamine. The possible implications of such an interaction are discussed. PMID:10608

  19. Oral administration of active vitamin D metabolites to low birthweight infants.

    PubMed Central

    Kovar, I Z; Mayne, P D; James, J J; Barnes, I C

    1986-01-01

    The active vitamin D metabolites 1 alpha, 25-dihydroxycholecalciferol (Rocaltrol) and the analogue 1 alpha-hydroxycholecalciferol (One-Alpha) are adequately absorbed after oral administration in the preterm infant. The absorption pattern is similar to that seen in adults. PMID:3755581

  20. Rapidly Probing Antibacterial Activity of Graphene Oxide by Mass Spectrometry-based Metabolite Fingerprinting.

    PubMed

    Zhang, Ning; Hou, Jian; Chen, Suming; Xiong, Caiqiao; Liu, Huihui; Jin, Yulong; Wang, Jianing; He, Qing; Zhao, Rui; Nie, Zongxiu

    2016-01-01

    Application of nanomaterials as anti-bacteria agents has aroused great attention. To investigate the antibacterial activity and antibacterial mechanism of nanomaterials from a molecular perspective is important for efficient developing of nanomaterial antibiotics. In the current work, a new mass spectrometry-based method was established to investigate the bacterial cytotoxicity of graphene oxide (GO) by the metabolite fingerprinting of microbes. The mass spectra of extracted metabolites from two strains DH5α and ATCC25922 were obtained before and after the incubation with nanomaterials respectively. Then principal component analysis (PCA) of these spectra was performed to reveal the relationship between the metabolism disorder of microbes and bactericidal activity of GO. A parameter "D" obtained from PCA scores was proposed that is capable to quantitatively evaluate the antibacterial activity of GO in concentration and time-dependent experiments. Further annotation of the fingerprinting spectra shows the variabilities of important metabolites such as phosphatidylethanolamine, phosphatidylglycerol and glutathione. This metabolic perturbation of E. coli indicates cell membrane destruction and oxidative stress mechanisms for anti-bacteria activity of graphene oxide. It is anticipated that this mass spectrometry-based metabolite fingerprinting method will be applicable to other antibacterial nanomaterials and provide more clues as to their antibacterial mechanism at molecular level. PMID:27306507

  1. CHARACTERIZATION ADN BIOLOGICAL ACTIVITY OF SECONDARY METABOLITES FROM ARMILLARIA TABESCENS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Ethyl acetate extracts from liquid cultures of Armillaria tabescens showed good antimicrobial activity against Candida albicans, Cryptococcus neoformans, Escherichia coli and Mycobacterium intracellulare. Chemical analyses of extract constituents led to the isolation and identification of two new co...

  2. Activity-dependent and graded BACE1 expression in the olfactory epithelium is mediated by the retinoic acid metabolizing enzyme CYP26B1.

    PubMed

    Login, Hande; Butowt, Rafal; Bohm, Staffan

    2015-07-01

    It is well established that environmental influences play a key role in sculpting neuronal connectivity in the brain. One example is the olfactory sensory map of topographic axonal connectivity. While intrinsic odorant receptor signaling in olfactory sensory neurons (OSN) determines anterior-posterior counter gradients of the axonal guidance receptors Neuropilin-1 and Plexin-A1, little is known about stimulus-dependent gradients of protein expression, which correlates with the functional organization of the olfactory sensory map along its dorsomedial (DM)-ventrolateral (VL) axis. Deficiency of the Alzheimer's β-secretase BACE1, which is expressed in a DM(low)-VL(high) gradient, results in OSN axon targeting errors in a DM > VL and gene dose-dependent manner. We show that expression of BACE1 and the all-trans retinoic acid (RA)-degrading enzyme Cyp26B1 form DM-VL counter gradients in the olfactory epithelium. Analyses of mRNA and protein levels in OSNs after naris occlusion, in mice deficient in the olfactory cyclic nucleotide-gated channel and in relation to onset of respiration, show that BACE1 and Cyp26B1 expression in OSNs inversely depend on neuronal activity. Overexpression of a Cyp26B1 or presence of a dominant negative RA receptor transgene selectively in OSNs, inhibit BACE1 expression while leaving the DM(low)-VL(high) gradient of the axonal guidance protein Neuropilin-2 intact. We conclude that stimulus-dependent neuronal activity can control the expression of the RA catabolic enzyme Cyp26B1 and downstream genes such as BACE1. This result is pertinent to an understanding of the mechanisms by which a topographic pattern of connectivity is achieved and modified as a consequence of graded gene expression and sensory experience. PMID:24797530

  3. Peroxisome proliferator-activated receptors and retinoic acid receptors differentially control the interactions of retinoid X receptor heterodimers with ligands, coactivators, and corepressors.

    PubMed Central

    DiRenzo, J; Söderstrom, M; Kurokawa, R; Ogliastro, M H; Ricote, M; Ingrey, S; Hörlein, A; Rosenfeld, M G; Glass, C K

    1997-01-01

    As the obligate member of most nuclear receptor heterodimers, retinoid X receptors (RXRs) can potentially perform two functions: cooperative binding to hormone response elements and coordinate regulation of target genes by RXR ligands. In this paper we describe allosteric interactions between RXR and two heterodimeric partners, retinoic acid receptors (RARs) and peroxisome proliferator-activated receptors (PPARs); RARs and PPARs prevent and permit activation by RXR-specific ligands, respectively. By competing for dimerization with RXR on response elements consisting of direct-repeat half-sites spaced by 1 bp (DR1 elements), the relative abundance of RAR and PPAR determines whether the RXR signaling pathway will be functional. In contrast to RAR, which prevents the binding of RXR ligands and recruits the nuclear receptor corepressor N-CoR, PPAR permits the binding of SRC-1 in response to both RXR and PPAR ligands. Overexpression of SRC-1 markedly potentiates ligand-dependent transcription by PPARgamma, suggesting that SRC-1 serves as a coactivator in vivo. Remarkably, the ability of RAR to both block the binding of ligands to RXR and interact with corepressors requires the CoR box, a structural motif residing in the N-terminal region of the RAR ligand binding domain. Mutations in the CoR box convert RAR from a nonpermissive to a permissive partner of RXR signaling on DR1 elements. We suggest that the differential recruitment of coactivators and corepressors by RAR-RXR and PPAR-RXR heterodimers provides the basis for a transcriptional switch that may be important in controlling complex programs of gene expression, such as adipocyte differentiation. PMID:9121466

  4. Secondary Metabolites Produced by an Endophytic Fungus Pestalotiopsis sydowiana and Their 20S Proteasome Inhibitory Activities.

    PubMed

    Xia, Xuekui; Kim, Soonok; Liu, Changheng; Shim, Sang Hee

    2016-01-01

    Fungal endophytes have attracted attention due to their functional diversity. Secondary metabolites produced by Pestalotiopsis sydowiana from a halophyte, Phragmites communis Trinus, were investigated. Eleven compounds, including four penicillide derivatives (1-4) and seven α-pyrone analogues (5-10) were isolated from cultures of P. sydowiana. The compounds were identified based on spectroscopic data. The inhibitory activities against the 20S proteasome were evaluated. Compounds 1-3, 5, and 9-10 showed modest proteasome inhibition activities, while compound 8 showed strong activity with an IC50 of 1.2 ± 0.3 μM. This is the first study on the secondary metabolites produced by P. sydowiana and their proteasome inhibitory activities. The endophytic fungus P. sydowiana might be a good resource for proteasome inhibitors. PMID:27447600

  5. Characterization of retinoic acid-induced neurobehavioral effects in developing zebrafish.

    PubMed

    Wang, Yujiang; Chen, Jiangfei; Du, Changchun; Li, Chunqi; Huang, Changjiang; Dong, Qiaoxiang

    2014-02-01

    Retinoic signaling plays an important role in cell proliferation and differentiation. Disruption of retinoic signaling via excessive or deficient retinoic acid can cause teratogenic effects on developing embryos. Similar to retinoic acid, many xenobiotic environmental pollutants have been found to disrupt retinoic signaling through binding and eliciting agonistic activity on retinoic acid receptors. Currently, studies of retinoic acid or retinoic acid-like compounds in aquatic organisms have mainly focused on teratogenicity and few studies have explored their neurobehavioral toxicity. In the present study, the authors used retinoic acid as an example to explore the neurobehavioral toxicity associated with developmental exposure of retinoic acid-like compounds in zebrafish. The findings confirmed retinoic acid's teratogenic effects such as bent spine, malformed tail, and pericardial edema in developing zebrafish with a median effective concentration of 2.47 nM. Retinoic acid-induced cell apoptosis at 24 h postfertilization was consistently found in the eye and tail regions of embryos. Spontaneous movement as characterized by tail bend frequency was significantly increased in zebrafish embryos following exposure to 2 nM and 8 nM retinoic acid. Relatively low-dose retinoic acid exposure of 2 nM led to fast locomotion behavior in the dark period and hyperactivity during light-dark photoperiod stimulation. The 2-nM retinoic acid exposure also led to alterations of neurobehavior- and optic nerve-related genes, with the transforming growth factor-β signal transduction inhibitor noggin (nog) and the spinal cord marker homeobox c3a (hox) being underexpressed and the retinal G protein-coupled receptor a (rgr), the photoreceptor cell marker rhodopsin (rho), and the short wave-sensitive cone pigment opsin 1 (opn1sw1) being overexpressed. Increased expression of opn1sw1 and rho was confirmed by whole-mount in situ hybridization. Whether the misexpression of these genes leads

  6. CONTROL OF ALKALINE PHOSPHATASE ACTIVITY IN C3H10T1/2 CELLS: ROLE OF RETINOIC ACID AND CELL DENSITY

    EPA Science Inventory

    The enzyme alkaline phosphatase (AP) has been shown to be lost or inappropriately expressed during carcinogenesis in some tissues. ecause retinoic acid (RA) appears to play a role in the normal regulation of the enzyme (RA up-regulates AP in a variety of cell types) we have sugge...

  7. Retinoic acid affects the expression of nuclear retinoic acid receptors in tissues of retinol-deficient rats.

    PubMed Central

    Haq, R; Pfahl, M; Chytil, F

    1991-01-01

    The multitude of biological effects of the vitamin A metabolite, retinoic acid, are mediated by nuclear retinoic acid receptors (RARs), which are members of the steroid/thyroid hormone receptor superfamily. RAR-alpha, -beta, and -gamma are encoded by three genes from which multiple isoforms can be generated. Recent studies suggest that the expression of at least some RAR isoforms can be regulated by retinoic acid in certain cell lines. Here we examined regulation of RAR expression in the adult animal. RARs were analyzed by Northern blots from lung, liver, and testes of retinol-deficient rats. Retinol deficiency caused a 65-70% decrease in the mRNA levels of lung and liver RAR-beta, whereas no change was observed in RAR-alpha and -gamma mRNA levels in these organs. In the testes of retinol-deficient animals, two transcripts, RAR-alpha 1 (3.7 kb) and RAR-alpha 2 (2.8 kb), were detected as compared with one RAR-alpha 1 (3.7 kb) transcript in retinol-sufficient testes. When retinol-deficient rats were orally administered 1 dose of retinoic acid (100 micrograms per rat), lung RAR-beta mRNA levels started to increase after 1 hr and reached a 16-fold higher level after 4 hr; after 4 hr these retinoic acid-fed rats also showed a 7-fold increase in liver RAR-beta mRNA levels as compared with levels in the retinol-deficient rats. In contrast, liver, lung, and testes RAR-alpha transcripts remained either unchanged or showed only a slight increase in response to retinoic acid. RAR-gamma was constitutively expressed in lung, and its mRNA levels were induced 2-fold by retinoic acid. These results show tissue diversity in the rapid induction of RAR-beta and RAR-gamma by retinoic acid in the adult animal and suggest distinct roles for the various receptor isoforms in the control of the retinoid response. Images PMID:1654565

  8. Antifouling Activity of Lipidic Metabolites Derived from Padina tetrastromatica.

    PubMed

    Suresh, Murugan; Iyapparaj, Palanisamy; Anantharaman, Perumal

    2016-07-01

    An attempt has been made to identify the potential seaweed for antifouling property due to the growing need for environmentally safe antifouling systems. The antibacterial, antimicroalgal, and antimussel foot adherence potentials of methanol, dichloromethane, and hexane extracts of the chosen seaweeds such as Padina tetrastromatica, Caulerpa taxifolia, and Amphiroa fragilissima have been compared against copper sulfate. Among the extracts, the maximum antibacterial activities were exhibited by the methanol extract of P. tetrastromatica. The minimum inhibitory concentration (MIC) of the methanolic extract of P. tetrastromatica was found to be 10 and 1 μg/ml against test biofilm bacteria and diatoms, respectively. The antimussel foot adherence assay indicated that the extract had inhibited the foot adherence of the green mussels Perna viridis with the effective concentration (EC50) of 25.51 ± 0.03 μg/ml, and lethal concentration for 50 % mortality (LC50) was recorded at 280.22 ± 0.12 μg/ml. Based on the prolific results, the crude methanolic extract of P. tetrastromatica was subjected to purification using silica gel column and thin-layer chromatography (TLC). Then, the active compounds of the bioassay-guided fraction (F13) were identified using gas chromatography coupled with mass spectroscopy (GC-MS), and it was observed that fatty acids were the major components, which may be responsible for the antifouling properties. PMID:26956575

  9. REACTIVITY PROFILE OF LIGANDS OF MAMMALIAN RETINOIC ACID RECEPTORS: A PRELIMINARY COREPA ANALYSIS

    EPA Science Inventory

    Retinoic acid and associated derivatives comprise a class of endogenous hormones that bind to and activate different families of retinoic acid receptors (RARs, RXRs), and control many aspects of vertebrate development. Identification of potential RAR and RXR ligands is of interes...

  10. Vitamin A and Retinoic Acid in the Regulation of B-Cell Development and Antibody Production

    PubMed Central

    Ross, A. Catharine; Chen, Qiuyan; Ma, Yifan

    2013-01-01

    Signaling by vitamin A through its active metabolite retinoic acid (RA) is critical for the normal development and functions of the hematopoietic and immune systems. B cells, as both factories for antibody production and part of the immune regulatory system, are critical to a successful vaccination response. RA is a factor in the development and competence of mature B cells, in B cell proliferation, and in the regulation of transcription factors associated with B cell differentiation, class switch recombination, and the generation of antibody-secreting plasma cells. Emerging evidence suggests that RA can function alone and in combination with other immune system stimuli to augment the formation of germinal centers, leading to increased primary and secondary antibody responses. Taken together, RA could be a useful component in vaccine strategies and/or for immunotherapy. PMID:21419269

  11. Global metabolite analysis of the land snail Theba pisana hemolymph during active and aestivated states.

    PubMed

    Bose, U; Centurion, E; Hodson, M P; Shaw, P N; Storey, K B; Cummins, S F

    2016-09-01

    The state of metabolic dormancy has fascinated people for hundreds of years, leading to research exploring the identity of natural molecular components that may induce and maintain this state. Many animals lower their metabolism in response to high temperatures and/or arid conditions, a phenomenon called aestivation. The biological significance for this is clear; by strongly suppressing metabolic rate to low levels, animals minimize their exposure to stressful conditions. Understanding blood or hemolymph metabolite changes that occur between active and aestivated animals can provide valuable insights relating to those molecular components that regulate hypometabolism in animals, and how they afford adaptation to their different environmental conditions. In this study, we have investigated the hemolymph metabolite composition from the land snail Theba pisana, a remarkably resilient mollusc that displays an annual aestivation period. Using LC-MS-based metabolomics analysis, we have identified those hemolymph metabolites that show significant changes in relative abundance between active and aestivated states. We show that certain metabolites, including some phospholipids [e.g. LysoPC(14:0)], and amino acids such as l-arginine and l-tyrosine, are present at high levels within aestivated snails. Further investigation of our T. pisana RNA-sequencing data elucidated the entire repertoire of phospholipid-synthesis genes in the snail digestive gland, as a precursor towards future comparative investigation between the genetic components of aestivating and non-aestivating species. In summary, we have identified a large number of metabolites that are elevated in the hemolymph of aestivating snails, supporting their role in protecting against heat or desiccation. PMID:27318654

  12. Culture condition-dependent metabolite profiling of Aspergillus fumigatus with antifungal activity.

    PubMed

    Kang, Daejung; Son, Gun Hee; Park, Hye Min; Kim, Jiyoung; Choi, Jung Nam; Kim, Hyang Yeon; Lee, Sarah; Hong, Seung-Beom; Lee, Choong Hwan

    2013-03-01

    Three sections of Aspergillus (five species, 21 strains) were classified according to culture medium-dependent and time-dependent secondary metabolite profile-based chemotaxonomy. Secondary metabolites were analysed by liquid chromatography-electrospray ionisation tandem mass spectrometry (LC-ESI-MS-MS) and multivariate statistical methods. From the Aspergillus sections that were cultured on malt extract agar (MEA) and Czapek yeast extract agar (CYA) for 7, 12, and 16 d, Aspergillus sections Fumigati (A. fumigatus), Nigri (A. niger), and Flavi (A. flavus, A. oryzae, and A. sojae) clustered separately on the basis of the results of the secondary metabolite analyses at 16 d regardless of culture medium. Based on orthogonal projection to latent structures discriminant analysis by partial least squares discriminant analysis (PLS-DA), we identified the secondary metabolites that helped differentiate sections between A. fumigatus and Aspergillus section Flavi to be gliotoxin G, fumigatin oxide, fumigatin, pseurotin A or D, fumiquinazoline D, fumagillin, helvolic acid, 1,2-dihydrohelvolic acid, and 5,8-dihydroxy-9,12-octadecadienoic acid (5,8-diHODE). Among these compounds, fumagillin, helvolic acid, and 1,2-dihydrohelvolic acid of A. fumigatus showed antifungal activities against Malassezia furfur, which is lipophilic yeast that causes epidermal skin disorders. PMID:23537878

  13. Structural analysis of DNA interaction with retinol and retinoic acid.

    PubMed

    Mandeville, J S; N'soukpoé-Kossi, C N; Neault, J F; Tajmir-Riahi, H A

    2010-06-01

    Dietary constituents of fresh fruits and vegetables may play a relevant role in DNA adduct formation by inhibiting enzymatic activities. Studies have shown the important role of antioxidant vitamins A, C, and E in the protection against cancer and cardiovascular diseases. The antioxidant activity of vitamin A and beta-carotene may consist of scavenging oxygen radicals and preventing DNA damage. This study was designed to examine the interaction of calf-thymus DNA with retinol and retinoic acid in aqueous solution at physiological conditions using a constant DNA concentration and various retinoid contents. Fourier transform infrared (FTIR), circular dichroism (CD), and fluorescence spectroscopic methods were used to determine retinoid binding mode, the binding constant, and the effects of retinol and retinoic acid complexation on DNA conformation and aggregation. Structural analysis showed that retinol and retinoic acid bind DNA via G-C and A-T base pairs and the backbone phosphate groups with overall binding constants of Kret = 3.0 (+/-0.50) x 10(3) (mol.L(-1))(-1) and Kretac = 1.0 (+/-0.20) x 10(4) (mol.L(-1))(-1). The number of bound retinoids per DNA were 0.84 for retinol and 1.3 for retinoic acid. Hydrophobic interactions were also observed at high retinol and retinoic acid contents. At a high retinoid concentration, major DNA aggregation occurred, while DNA remained in the B-family structure. PMID:20555389

  14. Interactive Roles of Ets-1, Sp1, and Acetylated Histones in the Retinoic Acid-dependent Activation of Guanylyl Cyclase/Atrial Natriuretic Peptide Receptor-A Gene Transcription*

    PubMed Central

    Kumar, Prerna; Garg, Renu; Bolden, Gevoni; Pandey, Kailash N.

    2010-01-01

    Cardiac hormones atrial and brain natriuretic peptides activate guanylyl cyclase/natriuretic peptide receptor-A (GC-A/NPRA), which plays a critical role in reduction of blood pressure and blood volume. Currently, the mechanisms responsible for regulating the Npr1 gene (coding for GC-A/NPRA) transcription are not well understood. The present study was conducted to examine the interactive roles of all-trans retinoic acid (ATRA), Ets-1, Sp1, and histone acetylation on the transcriptional regulation and function of the Npr1 gene. Deletion analysis of the Npr1 promoter and luciferase assays showed that ATRA enhanced a 16-fold Npr1 promoter activity and greatly stimulated guanylyl cyclase (GC) activity of the receptor protein in both atrial natriuretic peptide (ANP)-dependent and -independent manner. As confirmed by gel shift and chromatin immunoprecipitation assays, ATRA enhanced the binding of both Ets-1 and Sp1 to the Npr1 promoter. The retinoic acid receptor α (RARα) was recruited by Ets-1 and Sp1 to form a transcriptional activator complex with their binding sites in the Npr1 promoter. Interestingly, ATRA also increased the acetylation of histones H3 and H4 and enhanced their recruitment to Ets-1 and Sp1 binding sites within the Npr1 promoter. Collectively, the present results demonstrate that ATRA regulates Npr1 gene transcription and GC activity of the receptor by involving the interactive actions of Ets-1, Sp1, and histone acetylation. PMID:20864529

  15. Reproductive activity in the peninsular pronghorn determined from excreted gonadal steroid metabolites.

    PubMed

    Kersey, David C; Holland, Jeff; Eng, Curtis

    2015-01-01

    Fecal hormone monitoring was employed to better define annual patterns of reproductive steroid metabolites from a breeding pair of peninsular pronghorn (Antilocapra americana peninsularis) maintained at the Los Angeles Zoo. Notably in the female, increased excretion of estrogen metabolites occurred during the breeding season (Jun-Aug), and a biphasic pattern in progestagen activity was measured during gestation. Of additional interest, a preterm increase in estrogen that continued for an additional 64 days post partum. Male androgen activity correlated with the female estrogen patterns, with a single successful copulation occurring during the breeding season; interestingly however, the male exhibited no reproductive behaviors during the female's preterm/post partum estrogen increase. These data are the first reproductive steroid profiles for the peninsular pronghorn and provide valuable insight that will aid efforts that link the species' reproductive physiology with conservation management. PMID:25652944

  16. Secondary metabolites of plants from the genus chloranthus: chemistry and biological activities.

    PubMed

    Wang, An-Ran; Song, Hong-Chuan; An, Hong-Mei; Huang, Qian; Luo, Xie; Dong, Jin-Yan

    2015-04-01

    Chloranthus, a genus of the family Chloranthaceae, which is mainly distributed in eastern and southern Asia, has been used in Chinese folk medicine due to its antitumor, antifungal, and anti-inflammatory activities. This review compiles the research on isolation, structure elucidation, structural diversity, and bioactivities of Chloranthus secondary metabolites reported between 2007 and 2013. The metabolites listed encompass 82 sesquiterpenoids, 50 dimeric sesquiterpenoids, 15 diterpenoids, one coumarin, and five other compounds. Among them, dimeric sesquiterpenoids, the characteristic components of plants from the genus Chloranthus, have attracted considerable attention due to their complex structures and significant biological features, e.g., antitumor, antibacterial, antifungal, anti-inflammatory, and hepatoprotective activities, and potent and selective inhibition of the delayed rectifier (IK) K(+) current and tyrosinase. PMID:25879494

  17. RETINOIC ACID ALTERS EPITHELIAL DIFFERENTIATION DURING PALATOGENESIS

    EPA Science Inventory

    Retinoids are teratogenic in humans and animals, producing a syndrome of craniofacial malformations which includes cleft palate. his study investigates the mechanism through which retinoic acid induces cleft palate. urine palatogenesis after exposure to retinoic acid in utero is ...

  18. Synthesis of (11)C-labeled retinoic acid, [(11)C]ATRA, via an alkenylboron precursor by Pd(0)-mediated rapid C-[(11)C]methylation.

    PubMed

    Suzuki, Masaaki; Takashima-Hirano, Misato; Ishii, Hideki; Watanabe, Chika; Sumi, Kengo; Koyama, Hiroko; Doi, Hisashi

    2014-08-01

    Retinoids are a class of chemical compounds which include both natural dietary vitamin A (retinol) metabolites and active synthetic analogs. Both experimental and clinical studies have revealed that retinoids regulate a wide variety of essential biological processes. In this study, we synthesized (11)C-labeled all-trans-retinoic acid (ATRA), the most potent biologically active metabolite of retinol and used in the treatment of acute promyelocytic leukemia. The synthesis of (11)C-labeled ATRA was accomplished by a combination of rapid Pd(0)-mediated C-[(11)C]methylation of the corresponding pinacol borate precursor prepared by 8 steps and hydrolysis. [(11)C]ATRA will prove useful as a PET imaging agent, particularly for elucidating the improved therapeutic activity of ATRA (natural retinoid) for acute promyelocytic leukemia by comparing with the corresponding PET probe [(11)C]Tamibarotene (artificial retinoid). PMID:24930828

  19. Aryl Hydrocarbon Receptor Activity of Tryptophan Metabolites in Young Adult Mouse Colonocytes.

    PubMed

    Cheng, Yating; Jin, Un-Ho; Allred, Clint D; Jayaraman, Arul; Chapkin, Robert S; Safe, Stephen

    2015-10-01

    The tryptophan microbiota metabolites indole-3-acetate, indole-3-aldehyde, indole, and tryptamine are aryl hydrocarbon receptor (AhR) ligands, and in this study we investigated their AhR agonist and antagonist activities in nontransformed young adult mouse colonocyte (YAMC) cells. Using Cyp1a1 mRNA as an Ah-responsive end point, we observed that the tryptophan metabolites were weak AhR agonists and partial antagonists in YAMC cells, and the pattern of activity was different from that previously observed in CaCo2 colon cancer cells. However, expansion of the end points to other Ah-responsive genes including the Cyp1b1, the AhR repressor (Ahrr), and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-inducible poly(ADP-ribose) polymerase (TiParp) revealed a highly complex pattern of AhR agonist/antagonist activities that were both ligand- and gene-dependent. For example, the magnitude of induction of Cyp1b1 mRNA was similar for TCDD, tryptamine, and indole-3-acetate, whereas lower induction was observed for indole and indole-3-aldehyde was inactive. These results suggest that the tryptophan metabolites identified in microbiota are selective AhR modulators. PMID:25873348

  20. Oxidation of propylthiouracil to reactive metabolites by activated neutrophils. Implications for agranulocytosis.

    PubMed

    Waldhauser, L; Uetrecht, J

    1991-01-01

    Propylthiouracil (PTU) is associated with idiosyncratic agranulocytosis that may be due to reactive metabolites generated from oxidative metabolism by neutrophils. Therefore, the metabolism of PTU was investigated in activated neutrophils. Three oxidized metabolites were observed on HPLC: PTU-disulfide, propyluracil-2-sulfinate, and propyluracil-2-sulfonate (PTU-SO3-). No metabolism was detected in cells that had not been activated. Metabolism was inhibited by sodium azide and by catalase. The same products were produced by myeloperoxidase (MPO) in an MPO/H2O2/Cl- system. PTU inhibited its own metabolism; however, complete conversion to PTU-SO3- could be achieved with optimal PTU concentrations. MPO/H2O2 without Cl- produced only slight metabolism. The PTU-sulfenyl chloride is a postulated intermediate. In the absence of chloride, oxidation might proceed through propyluracil-2-sulfenic acid. The sulfenyl chloride and PTU-SO3- are both chemically reactive with sulfhydryl compounds such as N-acetylcysteine. Such reactive metabolites, generated by activated neutrophils, may be involved in hypersensitivity reactions associated with PTU, such as agranulocytosis. PMID:1676636

  1. Fungal transformation and T-cell proliferation inhibitory activity of melengestrol acetate and its metabolite.

    PubMed

    Baydoun, Elias; Bano, Saira; Atia-tul-Wahab; Jabeen, Almas; Yousuf, Sammer; Mesaik, Ahmed; Smith, Colin; Choudhary, M Iqbal

    2014-08-01

    Biotransformation of melengestrol acetate (MGA, 17α-acetoxy-6-methyl-16-methylenepregna-4,6-diene-3,20-dione) (1) was investigated for the first time by using fungal cultures. Incubation of compound 1 with Cunninghamella blakesleeana yielded a new major metabolite, 17α-acetoxy-11β-hydroxy-6-methyl-16-methylenepregna-4,6-diene-3,20-dione (2). The metabolite 2 was purified by using HPLC, followed by characterization through (1)H- and (13)C-NMR and other spectroscopic techniques. Single crystal X-ray diffraction analysis was used to deduce the three dimensional structures of melengestrol acetate (1) and metabolite 2 for the first time. T-cell proliferation assay was employed to evaluate the immunosuppressant effect of compounds 1 and 2 with IC50=0.5±0.07 and 0.6±0.08μg/mL, respectively. The results indicated that these compounds possess sixfold potent T-cell proliferation inhibitory activity as compared to the standard prednisolone (IC50<3.1μg/mL). Both compounds were found to be non-toxic in a 3T3 (mouse fibroblast) cell-based cytotoxicity assay. This discovery of potent anti-inflammatory activity of compounds 1 and 2 can lead the way to develop new immunosuppressant compounds for clinical application. PMID:24793568

  2. Marine Invertebrate Metabolites with Anticancer Activities: Solutions to the "Supply Problem".

    PubMed

    Gomes, Nelson G M; Dasari, Ramesh; Chandra, Sunena; Kiss, Robert; Kornienko, Alexander

    2016-05-01

    Marine invertebrates provide a rich source of metabolites with anticancer activities and several marine-derived agents have been approved for the treatment of cancer. However, the limited supply of promising anticancer metabolites from their natural sources is a major hurdle to their preclinical and clinical development. Thus, the lack of a sustainable large-scale supply has been an important challenge facing chemists and biologists involved in marine-based drug discovery. In the current review we describe the main strategies aimed to overcome the supply problem. These include: marine invertebrate aquaculture, invertebrate and symbiont cell culture, culture-independent strategies, total chemical synthesis, semi-synthesis, and a number of hybrid strategies. We provide examples illustrating the application of these strategies for the supply of marine invertebrate-derived anticancer agents. Finally, we encourage the scientific community to develop scalable methods to obtain selected metabolites, which in the authors' opinion should be pursued due to their most promising anticancer activities. PMID:27213412

  3. Marine Invertebrate Metabolites with Anticancer Activities: Solutions to the “Supply Problem”

    PubMed Central

    Gomes, Nelson G. M.; Dasari, Ramesh; Chandra, Sunena; Kiss, Robert; Kornienko, Alexander

    2016-01-01

    Marine invertebrates provide a rich source of metabolites with anticancer activities and several marine-derived agents have been approved for the treatment of cancer. However, the limited supply of promising anticancer metabolites from their natural sources is a major hurdle to their preclinical and clinical development. Thus, the lack of a sustainable large-scale supply has been an important challenge facing chemists and biologists involved in marine-based drug discovery. In the current review we describe the main strategies aimed to overcome the supply problem. These include: marine invertebrate aquaculture, invertebrate and symbiont cell culture, culture-independent strategies, total chemical synthesis, semi-synthesis, and a number of hybrid strategies. We provide examples illustrating the application of these strategies for the supply of marine invertebrate-derived anticancer agents. Finally, we encourage the scientific community to develop scalable methods to obtain selected metabolites, which in the authors’ opinion should be pursued due to their most promising anticancer activities. PMID:27213412

  4. Biologically active polyketide metabolites from an undetermined fungicolous hyphomycete resembling Cladosporium.

    PubMed

    Höller, Ulrich; Gloer, James B; Wicklow, Donald T

    2002-06-01

    Eight new polyketide-derived metabolites [cladoacetals A and B (1 and 2), 3-(2-formyl-3-hydroxyphenyl)propionic acid (3), 3-deoxyisoochracinic acid (4), isoochracinol (5), 7-hydroxy-3-(2,3-dihydroxybutyl)-1(3H)-isobenzofuranone (6), (+)-cyclosordariolone (10), and altersolanol J (11)] and six known metabolites [two isomeric 1-(1,3-dihydro-4-hydroxy-1-isobenzofuranyl)butan-2,3-diols (7a/b), 7-hydroxy-1(3H)-isobenzofuranone (8), isoochracinic acid (9), altersolanol A (12), and macrosporin (13)] have been isolated from solid-substrate fermentation cultures of an undetermined fungicolous isolate (NRRL 29097) that resembles Cladosporium sp. All structures were assigned primarily by analysis of 1D and/or 2D NMR data. Five of the compounds showed antibacterial activity. PMID:12088431

  5. Microbial transformation of (+)-nootkatone and the antiproliferative activity of its metabolites.

    PubMed

    Gliszczyńska, Anna; Łysek, Agnieszka; Janeczko, Tomasz; Świtalska, Marta; Wietrzyk, Joanna; Wawrzeńczyk, Czesław

    2011-04-01

    Six metabolites were obtained as a result of microbial transformation of (+)-nootkatone (1) by the fungal strains: Botrytis, Didymosphaeria, Aspergillus, Chaetomium and Fusarium. Their structure were established as (+)-(4R,5S,7R,9R)-9α-hydroxynootkatone (2), (+)-(4R,5S,7R)-13-hydroxynootkatone (3) and (+)-(4R,5S,7R,9R,11S)-11,12-epoxy-9α-hydroxynootkatone (4), (+)-(4R,5S,7R,11S)-11,12-epoksynootkatone (5), (+)-(4R,5S,7R)-11,12-dihydroxynootkatone (6) and (+)-(4R,5S,7R)-7,11,12-trihydroxynootkatone (7) on the basis of their spectral data. Two products: (4) and (7) were not previously reported in the literature. The antiproliferative activity of (+)-nootkatone (1) and isolated metabolites (2-7) of its biotransformation has been evaluated. PMID:21377882

  6. Evaluation of the pharmacological activity of the major mexiletine metabolites on skeletal muscle sodium currents

    PubMed Central

    De Bellis, M; De Luca, A; Rana, F; Cavalluzzi, M M; Catalano, A; Lentini, G; Franchini, C; Tortorella, V; Conte Camerino, D

    2006-01-01

    Background and purpose: Mexiletine (Mex), an orally effective antiarrhythmic agent used to treat ventricular arrhythmias, has also been found to be effective for myotonia and neuropathic pain. It is extensively metabolized in humans but little information exists about the pharmacodynamic properties of its metabolites. Experimental approach: To determine their contribution to the clinical activity of Mex, p-hydroxy-mexiletine (PHM), hydroxy-methyl-mexiletine (HMM), N-hydroxy-mexiletine (NHM) (phase I reaction products) and N-carbonyloxy β-D-glucuronide (NMG) (phase II reaction product) were tested on sodium currents (INa) of frog skeletal muscle fibres. Sodium currents were elicited with depolarizing pulses from different holding potentials (HP=−140, −100, −70 mV) and stimulation frequencies (0.25, 0.5, 1, 2, 5, 10 Hz) using the vaseline-gap voltage-clamp method. Key results: All the hydroxylated derivatives blocked the sodium channel in a voltage- and use-dependent manner. The PHM, HMM and NHM metabolites were up to 10-fold less effective than the parent compound. However, HMM showed a greater use-dependent behaviour (10 Hz), compared to Mex and the other metabolites. Similar to Mex, these products behaved as inactivating channel blockers. Conjugation with glucuronic acid (NMG) resulted in almost complete abolition of the pharmacological activity of the parent compound. Conclusions and Implications: Thus, although less potent, the phase I metabolites tested demonstrated similar pharmacological behaviour to Mex and might contribute to its clinical profile. PMID:16921388

  7. Antimicrobial activity of secondary metabolites from Streptomyces sp. K15, an endophyte in Houttuynia cordata Thunb.

    PubMed

    Chen, Huabao; Yang, Chunping; Ke, Tao; Zhou, Miaomiao; Li, Zhaojun; Zhang, Min; Gong, Guoshu; Hou, Taiping

    2015-01-01

    We isolated Streptomyces sp. K15 from the root tissue of Houttuynia cordata Thunb and found that some of its secondary metabolites exhibited significant antimicrobial activity against Botrytis cinerea. Moreover, we separated, purified and identified the major active ingredient to be 2-pyrrol formic acid by using silica gel column chromatography, high-performance liquid chromatography and NMR analysis of the spectral data. 2-Pyrrol formic acid critically inhibited the growth of some phytopathogenic bacteria. Therefore, it has potential value in agricultural applications. PMID:25675117

  8. Triterpenoid resinous metabolites from the genus Boswellia: pharmacological activities and potential species-identifying properties

    PubMed Central

    2013-01-01

    The resinous metabolites commonly known as frankincense or olibanum are produced by trees of the genus Boswellia and have attracted increasing popularity in Western countries in the last decade for their various pharmacological activities. This review described the pharmacological specific details mainly on anti-inflammatory, anti-carcinogenic, anti-bacterial and apoptosis-regulating activities of individual triterpenoid together with the relevant mechanism. In addition, species-characterizing triterpenic markers with the methods for their detection, bioavailability, safety and other significant properties were reviewed for further research. PMID:24028654

  9. Activation of the aryl hydrocarbon receptor by carcinogenic aromatic amines and modulatory effects of their N-acetylated metabolites.

    PubMed

    Juricek, Ludmila; Bui, Linh-Chi; Busi, Florent; Pierre, Stéphane; Guyot, Erwan; Lamouri, Aazdine; Dupret, Jean-Marie; Barouki, Robert; Coumoul, Xavier; Rodrigues-Lima, Fernando

    2015-12-01

    Aromatic amines (AAs) are an important class of chemicals which account for 12 % of known carcinogens. The biological effects of AAs depend mainly on their biotransformation into reactive metabolites or into N-acetylated metabolites which are generally considered as less toxic. Although the activation of the aryl hydrocarbon receptor (AhR) pathway by certain carcinogenic AAs has been reported, the effects of their N-acetylated metabolites on the AhR have not been addressed. Here, we investigated whether carcinogenic AAs and their N-acetylated metabolites may activate/modulate the AhR pathway in the absence and/or the presence of a bona fide AhR ligand (benzo[a]pyrene/B(a)P]. In agreement with previous studies, we found that certain AAs activated the AhR in human liver and lung cells as assessed by an increase in cytochrome P450 1A1 (CYP1A1) expression and activity. Altogether, we report for the first time that these properties can be modulated by the N-acetylation status of the AA. Whereas 2-naphthylamine significantly activated the AhR and induced CYP1A1 expression, its N-acetylated metabolite was less efficient. In contrast, the N-acetylated metabolite of 2-aminofluorene was able to significantly activate AhR, whereas the parent AA, 2-aminofluorene, did not. In the presence of B(a)P, activation of AhR or antagonist effects were observed depending on the AA or its N-acetylated metabolite. Activation and/or modulation of the AhR pathway by AAs and their N-acetylated metabolites may represent a novel mechanism contributing to the toxicological effects of AAs. More broadly, our data suggest biological interactions between AAs and other classes of xenobiotics through the AhR pathway. PMID:25224404

  10. Identification of metabolites from an active fraction of Cajanus cajan seeds by high resolution mass spectrometry.

    PubMed

    Tekale, Satishkumar S; Jaiwal, Bhimrao V; Padul, Manohar V

    2016-11-15

    Antioxidants are important food additives which prolong food storage due to their protective effects against oxidative degradation of foods by free radicals. However, the synthetic antioxidants show toxic properties. Alternative economical and eco-friendly approach is screening of plant extract for natural antioxidants. Plant phenolics are potent antioxidants. Hence, in present study Cajanus cajan seeds were analyzed for antioxidant activity, Iron chelating activity and total phenolic content. The antioxidant activity using DPPH (2,2-diphenyl-1-picrylhydrazyl) radical scavenging assay showed 71.3% inhibition and 65.8% Iron chelating activity. Total 37 compounds including some short peptides and five major abundant compounds were identified in active fraction of C. cajan seeds. This study concludes that C. cajan seeds are good source of antioxidants and Iron chelating activity. Metabolites found in C. cajan seeds which remove reactive oxygen species (ROS), may help to alleviate oxidative stress associated dreaded health problem like cancer and cardiovascular diseases. PMID:27283694

  11. Select steroid hormone glucuronide metabolites can cause toll-like receptor 4 activation and enhanced pain.

    PubMed

    Lewis, Susannah S; Hutchinson, Mark R; Frick, Morin M; Zhang, Yingning; Maier, Steven F; Sammakia, Tarek; Rice, Kenner C; Watkins, Linda R

    2015-02-01

    We have recently shown that several classes of glucuronide metabolites, including the morphine metabolite morphine-3-glucuronide and the ethanol metabolite ethyl glucuronide, cause toll like receptor 4 (TLR4)-dependent signaling in vitro and enhanced pain in vivo. Steroid hormones, including estrogens and corticosterone, are also metabolized through glucuronidation. Here we demonstrate that in silico docking predicts that corticosterone, corticosterone-21-glucuronide, estradiol, estradiol-3-glucuronide and estradiol-17-glucuronide all dock with the MD-2 component of the TLR4 receptor complex. In addition to each docking with MD-2, the docking of each was altered by pre-docking with (+)-naloxone, a TLR4 signaling inhibitor. As agonist versus antagonist activity cannot be determined from these in silico interactions, an in vitro study was undertaken to clarify which of these compounds can act in an agonist fashion. Studies using a cell line transfected with TLR4, necessary co-signaling molecules, and a reporter gene revealed that only estradiol-3-glucuronide and estradiol-17-glucuronide increased reporter gene product, indicative of TLR4 agonism. Finally, in in vivo studies, each of the 5 drugs was injected intrathecally at equimolar doses. In keeping with the in vitro results, only estradiol-3-glucuronide and estradiol-17-glucuronide caused enhanced pain. For both compounds, pain enhancement was blocked by the TLR4 antagonist lipopolysaccharide from Rhodobacter sphaeroides, evidence for the involvement in TLR4 in the resultant pain enhancement. These findings have implications for several chronic pain conditions, including migraine and temporomandibular joint disorder, in which pain episodes are more likely in cycling females when estradiol is decreasing and estradiol metabolites are at their highest. PMID:25218902

  12. Select steroid hormone glucuronide metabolites can cause Toll-like receptor 4 activation and enhanced pain

    PubMed Central

    Lewis, Susannah S.; Hutchinson, Mark R.; Frick, Morin M.; Zhang, Yingning; Maier, Steven F.; Sammakia, Tarek; Rice, Kenner C.; Watkins, Linda R.

    2014-01-01

    We have recently shown that several classes of glucuronide metabolites, including the morphine metabolite morphine-3-glucuronide and the ethanol metabolite ethyl glucuronide, cause toll like receptor 4 (TLR4)-dependent signalling in vitro and enhanced pain in vivo. Steroid hormones, including estrogens and corticosterone, are also metabolized through glucuronidation. Here we demonstrate that in silico docking predicts that corticosterone, corticosterone-21-glucuronide, estradiol, estradiol-3-glucuronide and estradiol-17-glucuronide all dock with the MD-2 component of the TLR4 receptor complex. In addition to each docking with MD-2, the docking of each was altered by pre-docking with (+)-naloxone, a TLR4 signaling inhibitor. As agonist versus antagonist activity cannot be determined from these in silico interactions, an in vitro study was undertaken to clarify which of these compounds can act in an agonist fashion. Studies using a cell line transfected with TLR4, necessary co-signaling molecules, and a reporter gene revealed that only estradiol-3-glucuronide and estradiol-17-glucuronide increased reporter gene product, indicative of TLR4 agonism. Finally, in in vivo studies, each of the 5 drugs was injected intrathecally at equimolar doses. In keeping with the in vitro results, only estradiol-3-glucuronide and estradiol-17-glucuronide caused enhanced pain. For both compounds, pain enhancement was blocked by the TLR4 antagonist lipopolysaccharide from Rhodobacter sphaeroides, evidence for the involvement in TLR4 in the resultant pain enhancement. These findings have implications for several chronic pain conditions, including migraine and tempromandibular joint disorder, in which pain episodes are more likely in cycling females when estradiol is decreasing and estradiol metabolites are at their highest. PMID:25218902

  13. Biotransformation of dianabol with the filamentous fungi and β-glucuronidase inhibitory activity of resulting metabolites.

    PubMed

    Khan, Naik T; Zafar, Salman; Noreen, Shagufta; Al Majid, Abdullah M; Al Othman, Zeid A; Al-Resayes, Saud Ibrahim; Atta-ur-Rahman; Choudhary, M Iqbal

    2014-07-01

    Biotransformation of the anabolic steroid dianabol (1) by suspended-cell cultures of the filamentous fungi Cunninghamella elegans and Macrophomina phaseolina was studied. Incubation of 1 with C. elegans yielded five hydroxylated metabolites 2-6, while M. phaseolina transformed compound 1 into polar metabolites 7-11. These metabolites were identified as 6β,17β-dihydroxy-17α-methylandrost-1,4-dien-3-one (2), 15α,17β-dihydroxy-17α-methylandrost-1,4-dien-3-one (3), 11α,17β-dihydroxy-17α-methylandrost-1,4-dien-3-one (4), 6β,12β,17β-trihydroxy-17α-methylandrost-1,4-dien-3-one (5), 6β,15α,17β-trihydroxy-17α-methylandrost-1,4-dien-3-one (6), 17β-hydroxy-17α-methylandrost-1,4-dien-3,6-dione (7), 7β,17β,-dihydroxy-17α-methylandrost-1,4-dien-3-one (8), 15β,17β-dihydroxy-17α-methylandrost-1,4-dien-3-one (9), 17β-hydroxy-17α-methylandrost-1,4-dien-3,11-dione (10), and 11β,17β-dihydroxy-17α-methylandrost-1,4-dien-3-one (11). Metabolite 3 was also transformed chemically into diketone 12 and oximes 13, and 14. Compounds 6 and 12-14 were identified as new derivatives of dianabol (1). The structures of all transformed products were deduced on the basis of spectral analyses. Compounds 1-14 were evaluated for β-glucuronidase enzyme inhibitory activity. Compounds 7, 13, and 14 showed a strong inhibition of β-glucuronidase enzyme, with IC50 values between 49.0 and 84.9 μM. PMID:24755238

  14. Low water activity induces the production of bioactive metabolites in halophilic and halotolerant fungi.

    PubMed

    Sepcic, Kristina; Zalar, Polona; Gunde-Cimerman, Nina

    2011-01-01

    The aim of the present study was to investigate indigenous fungal communities isolated from extreme environments (hypersaline waters of solar salterns and subglacial ice), for the production of metabolic compounds with selected biological activities: hemolysis, antibacterial, and acetylcholinesterase inhibition. In their natural habitats, the selected fungi are exposed to environmental extremes, and therefore the production of bioactive metabolites was tested under both standard growth conditions for mesophilic microorganisms, and at high NaCl and sugar concentrations and low growth temperatures. The results indicate that selected halotolerant and halophilic species synthesize specific bioactive metabolites under conditions that represent stress for non-adapted species. Furthermore, adaptation at the level of the chemical nature of the solute lowering the water activity of the medium was observed. Increased salt concentrations resulted in higher hemolytic activity, particularly within species dominating the salterns. The appearance of antibacterial potential under stress conditions was seen in the similar pattern of fungal species as for hemolysis. The active extracts exclusively affected the growth of the Gram-positive bacterium tested, Bacillus subtilis. None of the extracts tested showed inhibition of acetylcholinesterase activity. PMID:21339946

  15. Low Water Activity Induces the Production of Bioactive Metabolites in Halophilic and Halotolerant Fungi

    PubMed Central

    Sepcic, Kristina; Zalar, Polona; Gunde-Cimerman, Nina

    2011-01-01

    The aim of the present study was to investigate indigenous fungal communities isolated from extreme environments (hypersaline waters of solar salterns and subglacial ice), for the production of metabolic compounds with selected biological activities: hemolysis, antibacterial, and acetylcholinesterase inhibition. In their natural habitats, the selected fungi are exposed to environmental extremes, and therefore the production of bioactive metabolites was tested under both standard growth conditions for mesophilic microorganisms, and at high NaCl and sugar concentrations and low growth temperatures. The results indicate that selected halotolerant and halophilic species synthesize specific bioactive metabolites under conditions that represent stress for non-adapted species. Furthermore, adaptation at the level of the chemical nature of the solute lowering the water activity of the medium was observed. Increased salt concentrations resulted in higher hemolytic activity, particularly within species dominating the salterns. The appearance of antibacterial potential under stress conditions was seen in the similar pattern of fungal species as for hemolysis. The active extracts exclusively affected the growth of the Gram-positive bacterium tested, Bacillus subtilis. None of the extracts tested showed inhibition of acetylcholinesterase activity. PMID:21339946

  16. Activation and Products of the Cryptic Secondary Metabolite Biosynthetic Gene Clusters by Rifampin Resistance (rpoB) Mutations in Actinomycetes

    PubMed Central

    Tanaka, Yukinori; Kasahara, Ken; Hirose, Yutaka; Murakami, Kiriko; Kugimiya, Rie

    2013-01-01

    A subset of rifampin resistance (rpoB) mutations result in the overproduction of antibiotics in various actinomycetes, including Streptomyces, Saccharopolyspora, and Amycolatopsis, with H437Y and H437R rpoB mutations effective most frequently. Moreover, the rpoB mutations markedly activate (up to 70-fold at the transcriptional level) the cryptic/silent secondary metabolite biosynthetic gene clusters of these actinomycetes, which are not activated under general stressful conditions, with the exception of treatment with rare earth elements. Analysis of the metabolite profile demonstrated that the rpoB mutants produced many metabolites, which were not detected in the wild-type strains. This approach utilizing rifampin resistance mutations is characterized by its feasibility and potential scalability to high-throughput studies and would be useful to activate and to enhance the yields of metabolites for discovery and biochemical characterization. PMID:23603745

  17. All-Trans-Retinoic Acid Enhances Mitochondrial Function in Models of Human Liver.

    PubMed

    Tripathy, Sasmita; Chapman, John D; Han, Chang Y; Hogarth, Cathryn A; Arnold, Samuel L M; Onken, Jennifer; Kent, Travis; Goodlett, David R; Isoherranen, Nina

    2016-05-01

    All-trans-retinoic acid (atRA) is the active metabolite of vitamin A. The liver is the main storage organ of vitamin A, but activation of the retinoic acid receptors (RARs) in mouse liver and in human liver cell lines has also been shown. AlthoughatRA treatment improves mitochondrial function in skeletal muscle in rodents, its role in modulating mitochondrial function in the liver is controversial, and little data are available regarding the human liver. The aim of this study was to determine whetheratRA regulates hepatic mitochondrial activity.atRA treatment increased the mRNA and protein expression of multiple components of mitochondrialβ-oxidation, tricarboxylic acid (TCA) cycle, and respiratory chain. Additionally,atRA increased mitochondrial biogenesis in human hepatocytes and in HepG2 cells with and without lipid loading based on peroxisome proliferator activated receptor gamma coactivator 1αand 1βand nuclear respiratory factor 1 mRNA and mitochondrial DNA quantification.atRA also increasedβ-oxidation and ATP production in HepG2 cells and in human hepatocytes. Knockdown studies of RARα, RARβ, and PPARδrevealed that the enhancement of mitochondrial biogenesis andβ-oxidation byatRA requires peroxisome proliferator activated receptor delta. In vivo in mice,atRA treatment increased mitochondrial biogenesis markers after an overnight fast. Inhibition ofatRA metabolism by talarozole, a cytochrome P450 (CYP) 26 specific inhibitor, increased the effects ofatRA on mitochondrial biogenesis markers in HepG2 cells and in vivo in mice. These studies show thatatRA regulates mitochondrial function and lipid metabolism and that increasingatRA concentrations in human liver via CYP26 inhibition may increase mitochondrial biogenesis and fatty acidβ-oxidation and provide therapeutic benefit in diseases associated with mitochondrial dysfunction. PMID:26921399

  18. Endophytic Streptomyces in the traditional medicinal plant Arnica montana L.: secondary metabolites and biological activity.

    PubMed

    Wardecki, Tina; Brötz, Elke; De Ford, Christian; von Loewenich, Friederike D; Rebets, Yuriy; Tokovenko, Bogdan; Luzhetskyy, Andriy; Merfort, Irmgard

    2015-08-01

    Arnica montana L. is a medical plant of the Asteraceae family and grows preferably on nutrient poor soils in mountainous environments. Such surroundings are known to make plants dependent on symbiosis with other organisms. Up to now only arbuscular mycorrhizal fungi were found to act as endophytic symbiosis partners for A. montana. Here we identified five Streptomyces strains, microorganisms also known to occur as endophytes in plants and to produce a huge variety of active secondary metabolites, as inhabitants of A. montana. The secondary metabolite spectrum of these strains does not contain sesquiterpene lactones, but consists of the glutarimide antibiotics cycloheximide and actiphenol as well as the diketopiperazines cyclo-prolyl-valyl, cyclo-prolyl-isoleucyl, cyclo-prolyl-leucyl and cyclo-prolyl-phenylalanyl. Notably, genome analysis of one strain was performed and indicated a huge genome size with a high number of natural products gene clusters among which genes for cycloheximide production were detected. Only weak activity against the Gram-positive bacterium Staphylococcus aureus was revealed, but the extracts showed a marked cytotoxic activity as well as an antifungal activity against Candida parapsilosis and Fusarium verticillioides. Altogether, our results provide evidence that A. montana and its endophytic Streptomyces benefit from each other by completing their protection against competitors and pathogens and by exchanging plant growth promoting signals with nutrients. PMID:26036671

  19. In vitro formation of retinoic acid from retinal in rat liver.

    PubMed

    Hupert, J; Mobarhan, S; Layden, T J; Papa, V M; Lucchesi, D J

    1991-08-01

    Enzymatic conversion of retinal to retinoic acid in rat liver cytosol was detected using a rapid and sensitive assay based on high pressure liquid chromatography (HPLC). This retinal oxidase assay system did not require extraction steps or any other manipulation of the sample mixture once the sample vial was sealed for incubation. The product (retinoic acid) and the reactant (retinal) were separated by HPLC in 14.0 min with a sensitivity of 15 and 40 pmol per injection for retinoic acid and retinal, respectively. Enzymatic activity was observed to be linear with protein concentration (0-2.4 mg/mL) and time (0-30 min) and displayed a broad pH maximum of 7.7-9.7. The enzyme exhibited Michaelis-Menten single-substrate kinetics with an apparent Km of 0.25 mM. The average specific activity in nine normal rats was 35.6 +/- 3.3 nmol retinoic acid formed/h per mg protein. Incubation of the enzyme with zinc did not affect the rate of retinoic acid synthesis. Dithiothreitol inhibited the reaction. Both NAD and NADH stimulated retinoic acid formation. Formation of retinol was also observed when these pyridine nucleotides were added to the reaction mixture, indicating the presence of retinal reductase activity. The results of kinetic studies suggest that NADH may act indirectly to stimulate retinoic acid formation. PMID:1760155

  20. [The pharmacokinetics of the dipeptide analog of piracetam with nootropic activity GVS-111 and of its basic metabolites].

    PubMed

    Boĭko, S S; Zherdev, V P; Dvorianinov, A A; Gudasheva, T A; Ostrovskaia, R U; Voronina, T A; Rozantsev, G G; Seredenin, S B

    1997-01-01

    The pharmacokinetics of a new nootropic dipeptide analog of piracetam-N-phenylacetyl-L-prolylglycine (GWS-111) and its main metabolites were studied in rats by means of high performance liquid chromatography and gas-liquid chromatography. The compound under study showed a greater resistance to an enzymatic effect than natural neuropeptides. In addition to an unchanged compound three of its metabolites were found in the blood plasma of the rats. One of them, cyclo-Pro-Gly was an active metabolite of GWS-111. PMID:9206571

  1. Green Tea Catechin Metabolites Exert Immunoregulatory Effects on CD4(+) T Cell and Natural Killer Cell Activities.

    PubMed

    Kim, Yoon Hee; Won, Yeong-Seon; Yang, Xue; Kumazoe, Motofumi; Yamashita, Shuya; Hara, Aya; Takagaki, Akiko; Goto, Keiichi; Nanjo, Fumio; Tachibana, Hirofumi

    2016-05-11

    Tea catechins, such as (-)-epigallocatechin-3-O-gallate (EGCG), have been shown to effectively enhance immune activity and prevent cancer, although the underlying mechanism is unclear. Green tea catechins are instead converted to catechin metabolites in the intestine. Here, we show that these green tea catechin metabolites enhance CD4(+) T cell activity as well as natural killer (NK) cell activity. Our data suggest that the absence of a 4'-hydroxyl on this phenyl group (B ring) is important for the effect on immune activity. In particular, 5-(3',5'-dihydroxyphenyl)-γ-valerolactone (EGC-M5), a major metabolite of EGCG, not only increased the activity of CD4(+) T cells but also enhanced the cytotoxic activity of NK cells in vivo. These data suggest that EGC-M5 might show immunostimulatory activity. PMID:27112424

  2. Potent Antidiabetic Activity and Metabolite Profiling of Melicope Lunu-ankenda Leaves.

    PubMed

    Al-Zuaidy, Mizher Hezam; Hamid, Azizah Abdul; Ismail, Amin; Mohamed, Suhaila; Abdul Razis, Ahmad Faizal; Mumtaz, Muhammad Waseem; Salleh, Syafiq Zikri

    2016-05-01

    Diabetes mellitus is normally characterized by chronic hyperglycemia associated with disturbances in the fat, carbohydrate, and protein metabolism. There is an increasing trend of using natural products instead of synthetic agents as alternative therapy for disorders due to their fewer side effects. In this study, antidiabetic and antioxidant activities of different Melicope lunu-ankenda (ML) ethanolic extracts were evaluated using inhibition of α-glucosidase and 2,2-diphenyl-l-picrylhydrazyl (DPPH) radicals scavenging activity, respectively; whereas, proton nuclear magnetic resonance ((1) H NMR) and ultra-high performance liquid chromatography-tandem mass spectrometric (UHPLC-MS/MS) techniques were used for metabolite profiling of ML leaf extracts at different concentrations of ethanol and water. Sixty percent of ethanolic ML extract showed highest inhibitory effect against α-glucosidase enzyme (IC50 of 37 μg/mL) and DPPH scavenging activity (IC50 of 48 μg/mL). Antidiabetic effect of ML extracts was also evaluated in vivo and it was found that the high doses (400 mg/Kg BW) of ML extract exhibited high suppression in fasting blood glucose level by 62.75%. The metabolites responsible for variation among ML samples with variable ethanolic levels have been evaluated successfully using (1) H-NMR-based metabolomics. The principal component analysis (PCA) and partial least squares(PLS) analysis scores depicted clear and distinct separations into 4 clusters representing the 4 ethanolic concentrations by PC1 and PC2, with an eigenvalue of 69.9%. Various (1) H-NMR chemical shifts related to the metabolites responsible for sample difference were also ascribed. The main bioactive compounds identified attributing toward the separation included: isorhamnetin, skimmianine, scopoletin, and melicarpinone. Hence, ML may be used as promising medicinal plant for the development of new functional foods, new generation antidiabetic drugs, as a single entity phytomedicine or in

  3. Asymmetric Cleavage of B-Carotene Yields a Transcriptional Repressor of Retinoid X Receptor and Peroxisome Proliferator-Activated Receptor Responses

    Technology Transfer Automated Retrieval System (TEKTRAN)

    B-Carotene and its metabolites exert a broad range of effects, in part by regulating transcriptional responses through specific nuclear receptor activation. Symmetric cleavage of B-carotene can yield 9-cis retinoic acid (9-cisRA), the natural ligand for the nuclear receptor RXR, the obligate heterod...

  4. Kynurenine pathway metabolites are associated with hippocampal activity during autobiographical memory recall in patients with depression.

    PubMed

    Young, Kymberly D; Drevets, Wayne C; Dantzer, Robert; Teague, T Kent; Bodurka, Jerzy; Savitz, Jonathan

    2016-08-01

    Inflammation-related changes in the concentrations of inflammatory mediators such as c-reactive protein (CRP), interleukin 1β (IL-1), and IL-6 as well as kynurenine metabolites are associated with major depressive disorder (MDD) and affect depressive behavior, cognition, and hippocampal plasticity in animal models. We previously reported that the ratios of kynurenic acid (KynA) to the neurotoxic metabolites, 3-hydroxykynurenine (3HK) and quinolinic acid (QA), were positively correlated with hippocampal volume in depression. The hippocampus is critical for autobiographical memory (AM) recall which is impaired in MDD. Here we tested whether the ratios, KynA/3HK and KynA/QA were associated with AM recall performance as well as hippocampal activity during AM recall. Thirty-five unmedicated depressed participants and 25 healthy controls (HCs) underwent fMRI scanning while recalling emotionally-valenced AMs and provided serum samples for the quantification of kynurenine metabolites, CRP, and cytokines (IL-1 receptor antagonist - IL-1RA; IL-6, tumor necrosis factor alpha - TNF, interferon gamma -IFN-γ, IL-10). KynA/3HK and KynA/QA were lower in the MDD group relative to the HCs. The concentrations of the CRP and the cytokines did not differ significantly between the HCs and the MDD group. Depressed individuals recalled fewer specific AMs and displayed increased left hippocampal activity during the recall of positive and negative memories. KynA/3HK was inversely associated with left hippocampal activity during specific AM recall in the MDD group. Further, KynA/QA was positively correlated with percent negative specific memories recalled in the MDD group and showed a non-significant trend toward a positive correlation with percent positive specific memories recalled in HCs. In contrast, neither CRP nor the cytokines were significantly associated with AM recall or activity of the hippocampus during AM recall. Conceivably, an imbalance in levels of KynA versus QA

  5. Baicalin, a metabolite of baicalein with antiviral activity against dengue virus

    PubMed Central

    Moghaddam, Ehsan; Teoh, Boon-Teong; Sam, Sing-Sin; Lani, Rafidah; Hassandarvish, Pouya; Chik, Zamri; Yueh, Andrew; Abubakar, Sazaly; Zandi, Keivan

    2014-01-01

    Baicalin, a flavonoid derived from Scutellaria baicalensis, is the main metabolite of baicalein released following administration in different animal models and human. We previously reported the antiviral activity of baicalein against dengue virus (DENV). Here, we examined the anti-DENV properties of baicalin in vitro, and described the inhibitory potentials of baicalin at different steps of DENV-2 (NGC strain) replication. Our in vitro antiviral experiments showed that baicalin inhibited virus replication at IC50 = 13.5 ± 0.08 μg/ml with SI = 21.5 following virus internalization by Vero cells. Baicalin exhibited virucidal activity against DENV-2 extracellular particles at IC50 = 8.74 ± 0.08 μg/ml and showed anti-adsorption effect with IC50 = 18.07 ± 0.2 μg/ml. Our findings showed that baicalin as the main metabolite of baicalein exerting in vitro anti-DENV activity. Further investigations on baicalein and baicalin to deduce its antiviral therapeutic effects are warranted. PMID:24965553

  6. Synergistic activation of retinoic acid (RA)-responsive genes and induction of embryonal carcinoma cell differentiation by an RA receptor {alpha} (RAR{alpha})-, RAR{beta}-, or RAR{gamma}-selective ligand in combination with retinoid Z receptor-specific ligand

    SciTech Connect

    Roy, B.; Taneja, R.; Chambon, P.

    1995-12-01

    This research indicates thatn retinoic acid receptor (RAR)-retinoid X receptor (RXR) heterodimers activate transcription of RA-responsive genes and induce cell differentiation of P19 and F9 cells in a ligand-dependent manner. 43 refs., 4 figs., 2 tabs.

  7. Antifungal, Phytotoxic, and Cytotoxic Activities of Metabolites from Epichloë bromicola, a Fungus Obtained from Elymus tangutorum Grass.

    PubMed

    Song, Qiu-Yan; Nan, Zhi-Biao; Gao, Kun; Song, Hui; Tian, Pei; Zhang, Xing-Xu; Li, Chun-Jie; Xu, Wen-Bo; Li, Xiu-Zhang

    2015-10-14

    The development of high-quality herbage is an important aspect of animal husbandry. Inoculating beneficial fungi onto inferior grass is a feasible strategy for producing new varieties of high-quality herbage. Epichloë bromicola is a candidate fungus that is isolated from Elymus tangutorum. A total of 17 metabolites, 1-17, were obtained from E. bromicola, and their biological activities were assayed. Metabolite 1 exhibited antifungal activities against Alternaria alternata, Fusarium avenaceum, Bipolaris sorokiniana, and Curvularia lunata. EC50 values ranged from 0.7 to 5.3 μM, which were better than the positive control, chlorothalonil. Metabolite 8 displayed obvious phytotoxic effects toward Lolium perenne and Poa crymophila seedlings, and it was as active as glyphosate. None of these isolated metabolites displayed cytotoxicity against Madin-Darby bovine kidney cells. The IC50 values were greater than 100 μM, and the metabolites increased the growth of the cells at a concentration of 12.5 μM. The bioassay indicated that E. bromicola may be a beneficial fungus for producing new varieties of herbage with various resistances. Additionally, metabolite 7, 3-(2'-(4″-hydroxyphenyl)acetoxy)-2S-methylpropanoic acid, is a new natural product, and its stereochemistry was determined by means of optical rotation computation and chemical reactions. PMID:26395226

  8. Retinoic acid-induced IgG production in TLR-activated human primary B cells involves ULK1-mediated autophagy

    PubMed Central

    Eriksen, Agnete Bratsberg; Torgersen, Maria Lyngaas; Holm, Kristine Lillebø; Abrahamsen, Greger; Spurkland, Anne; Moskaug, Jan Øivind; Simonsen, Anne; Blomhoff, Heidi Kiil

    2015-01-01

    In the present study we have established a vital role of autophagy in retinoic acid (RA)-induced differentiation of toll-like receptor (TLR)-stimulated human B cells into Ig-secreting cells. Thus, RA enhanced autophagy in TLR9- and CD180-stimulated peripheral blood B cells, as revealed by increased levels of the autophagosomal marker LC3B-II, enhanced colocalization between LC3B and the lysosomal marker Lyso-ID, by a larger percentage of cells with more than 5 characteristic LC3B puncta, and by the concomitant reduction in the level of SQSTM1/p62. Furthermore, RA induced expression of the autophagy-inducing protein ULK1 at the transcriptional level, in a process that required the retinoic acid receptor RAR. By inhibiting autophagy with specific inhibitors or by knocking down ULK1 by siRNA, the RA-stimulated IgG production in TLR9- and CD180-mediated cells was markedly reduced. We propose that the identified prominent role of autophagy in RA-mediated IgG-production in normal human B cells provides a novel mechanism whereby vitamin A exerts its important functions in the immune system. PMID:25749095

  9. Understanding the interactions between metabolites isolated from Achyrocline satureioides in relation to its antibacterial activity.

    PubMed

    Joray, Mariana Belén; Palacios, Sara María; Carpinella, María Cecilia

    2013-02-15

    As part of our ongoing research on the antibacterial activity of Achyrocline satureioides, this study seeks to better understand the interactions between the metabolites isolated from this plant. For this purpose, the combined effect of 23-methyl-6-O-desmethylauricepyrone (1), quercetin (2) and 3-O-methylquercetin (3), obtained through bioguided fractionation from A. satureioides ethanol extract, was evaluated against Staphylococcus aureus and Escherichia coli. In first place, the antibacterial effect of the combination of flavonols 2 and 3 was assessed, as these showed individual effectiveness lower than or equal to that of the fraction from which they were obtained. When the flavonols were applied together at concentrations below their minimum inhibitory concentration (MIC) values, a synergistic effect (FICI<0.30) against S. aureus was observed. In addition, compounds 2 and 3 in combination reduced 1000 times the MIC of compound 1, showing a clear synergistic interaction (FICI<0.15) in treatments against the Gram (+) bacterium. The most active combination against E. coli showed an additive interaction (FICI<0.62) between the three assayed compounds 1-3. These results indicated the existence of concerted action between these metabolites, evidence of the importance of the synergistic interactions between the components of plant-derived extracts for the control of pathogenic bacteria. PMID:23207251

  10. Urinary metabolites of isorhynchophylline in rats and their neuroprotective activities in the HT22 cell assay

    PubMed Central

    Chen, Fangfang; Qi, Wen; Sun, Jiahong; Simpkins, James W.; Yuan, Dan

    2015-01-01

    Isorhynchophylline is one of the major alkaloids from the Uncaria hook possessing the effects of lowered blood pressure, vasodilatation and protection against ischemia-induced neuronal damage. However, the metabolic pathway of isorhynchophylline has not been fully reported yet. In this paper, the metabolism of isorhynchophylline was investigated in rats. Five metabolites were isolated by using solvent extraction and repeated chromatographic methods, and identified by spectroscopic methods including UV, MS, NMR and CD experiments. Three new compounds were identified as 5-oxoisorhynchophyllic acid-22-O-β-D-glucuronide (M1), 17-O-demethyl-16,17-dihydro isorhynchophylline (M2) and 5-oxoisorhynchophyllic acid (M4) together with two known compounds isorhynchophylline (M0) and rhynchophylline (M3). Possible metabolic pathways of isorhynchophylline are proposed. Furthermore, the activity assay for all the metabolites showed that isorhynchophylline (M0) exhibited potent neuroprotective effects against glutamate-induced HT22 cell death. However, little or weak neuroprotective activities were observed for M1–M4. Our present study is important to further understand its metabolic fate and disposition in humans. PMID:24910000

  11. In vitro metabolism of pyripyropene A and ACAT inhibitory activity of its metabolites.

    PubMed

    Matsuda, Daisuke; Ohshiro, Taichi; Ohtawa, Masaki; Yamazaki, Hiroyuki; Nagamitsu, Tohru; Tomoda, Hiroshi

    2015-01-01

    Pyripyropene A (PPPA, 1) of fungal origin, a selective inhibitor of acyl-CoA:cholesterol acyltransferase 2 (ACAT2), proved orally active in atherogenic mouse models. The in vitro metabolites of 1 in liver microsomes and plasma of human, rabbit, rat and mouse were analyzed by ultra fast liquid chromatography and liquid chromatography/tandem mass spectrometry. In the liver microsomes from all species, successive hydrolysis occurred at the 1-O-acetyl residue, then at the 11-O-acetyl residue of 1, while the 7-O-acetyl residue was resistant to hydrolysis. Furthermore, dehydrogenation of the newly generated 11-alcoholic hydroxyl residue occurred in human and mouse-liver microsomes, while oxidation of the pyridine ring occurred in human and rabbit liver microsomes. On the other hand, hydrolysis of the 7-O-acetyl residue proceeded only in the mouse plasma. These data indicated that the in vitro metabolic profiles of 1 have subtle differences among animal species. All of the PPPA metabolites observed in liver microsomes and plasma markedly decreased ACAT2 inhibitory activity. These findings will help us to synthesize new PPPA derivatives more effective in in vivo study than 1. PMID:25005817

  12. Urinary metabolites of isorhynchophylline in rats and their neuroprotective activities in the HT22 cell assay.

    PubMed

    Chen, Fangfang; Qi, Wen; Sun, Jiahong; Simpkins, James W; Yuan, Dan

    2014-09-01

    Isorhynchophylline is one of the major alkaloids from the Uncaria hook possessing the effects of lowered blood pressure, vasodilatation and protection against ischemia-induced neuronal damage. However, the metabolic pathway of isorhynchophylline has not been fully reported yet. In this paper, the metabolism of isorhynchophylline was investigated in rats. Five metabolites were isolated by using solvent extraction and repeated chromatographic methods, and identified by spectroscopic methods including UV, MS, NMR and CD experiments. Three new compounds were identified as 5-oxoisorhynchophyllic acid-22-O-β-D-glucuronide (M1), 17-O-demethyl-16,17-dihydro isorhynchophylline (M2) and 5-oxoisorhynchophyllic acid (M4) together with two known compounds isorhynchophylline (M0) and rhynchophylline (M3). Possible metabolic pathways of isorhynchophylline are proposed. Furthermore, the activity assay for all the metabolites showed that isorhynchophylline (M0) exhibited potent neuroprotective effects against glutamate-induced HT22 cell death. However, little or weak neuroprotective activities were observed for M1-M4. Our present study is important to further understand its metabolic fate and disposition in humans. PMID:24910000

  13. The vitamin D3 metabolite-type activity of Solanum malacoxylon.

    PubMed

    Basudde, C D; Humphreys, D J

    1976-01-01

    1. Administration of an aqueous extract of the dried leaves of Solanum malacoxylon (DLSM) to rats causes a rapid hyperphosphataemia and a decrease in plasma alkaline phosphatase activity; the two effects are typical of 1,25(OH)2D3, the hormonally active metabolite of vitamin D3. 2. DLSM, like both vitamin D3 and parathyroid hormone, increases plasma calcium and citrate levels in rats. The effect of DLSM in influencing plasma citrate, and the role of this important metabolite in mineral metabolism is discussed. 3. A decrease of plasma magnesium levels occurs in rats following treatment with DLSM. This decrease, which is associated with a renal loss of this cation, is remarkably similar to that produced by hypervitaminosis D3. 4. Prolonged administration of DLSM to vitamin D deficient rats causes a polyuria, hypercalciuria, hyperphosphaturia, hypermagnesuria, an increase in urinary total hydroxyproline, an increase in plasma total hexosamines, and a corresponding decrease in the bone total hexosamines. These effects, some of which can also be produced by hyperparathyroidism, or following the administration of parathyroid extract (PTE), large doses of vitamin D3, or 1,25(OH)2D3, suggest that DLSM, like the latter compounds, is capable of causing bone mineral mobilization, and the dissolution of bone organic matrix. PMID:212224

  14. Cytochrome P450-dependent eicosapentaenoic acid metabolites are novel BK channel activators.

    PubMed

    Lauterbach, Birgit; Barbosa-Sicard, Eduardo; Wang, Mong-Heng; Honeck, Horst; Kärgel, Eva; Theuer, Jürgen; Schwartzman, Michal L; Haller, Hermann; Luft, Friedrich C; Gollasch, Maik; Schunck, Wolf-Hagen

    2002-02-01

    P450-dependent arachidonic acid (AA) metabolites regulate arterial tone by modulating calcium-activated (BK) potassium channels in vascular smooth muscle cells (VSMC). Because eicosapentaenoic acid (EPA) has been reported to improve vascular function, we tested the hypothesis that P450-dependent epoxygenation of EPA produces alternative vasoactive compounds. We synthesized the 5 regioisomeric epoxyeicosattrienoic acids (EETeTr) and examined them for effects on K(+) currents in rat cerebral artery VSMCs with the patch-clamp technique. 11(R),12(S)-epoxyeicosatrienoic acid (50 nmol/L) was used for comparison and stimulated K(+) currents 6-fold at +60 mV. However, 17(R),18(S)-EETeTr elicited a more than 14-fold increase. 17(S),18(R)-EET and the remaining four regioisomers were inactive. The effect of 17(R),18(S)-EETeTr was blocked by tetraethylammonium but not by 4-aminopyridine. VSMCs expressed P450s 4A1 and 4A3. Recombinant P450 4A1 hydroxylated EPA at C-19 and C-20 and epoxygenated the 17,18-double bond, yielding the R, S- and S, R-enantiomers in a ratio of 64:36. We conclude that 17(R),18(S)-EETeTr represents a novel, potent activator of BK potassium channels. Furthermore, this metabolite can be directly produced in VSMCs. We suggest that 17(R),18(S)-EETeTr may function as an important hyperpolarizing factor, particularly with EPA-rich diets. PMID:11882617

  15. Antimicrobial and Cytotoxic Activity of Extracts of Ferula heuffelii Griseb. ex Heuff. and Its Metabolites.

    PubMed

    Pavlović, Ivan; Petrović, Silvana; Milenković, Marina; Stanojković, Tatjana; Nikolić, Dejan; Krunić, Aleksej; Niketić, Marjan

    2015-10-01

    The antimicrobial and cytotoxic activities of isolates (CHCl3 and MeOH extracts and selected metabolites) obtained from the underground parts of the Balkan endemic plant Ferula heuffelii Griseb. ex Heuff. were assessed. The CHCl3 and MeOH extracts exhibited moderate antimicrobial activity, being more pronounced against Gram-positive than Gram-negative bacteria, especially against Staphylococcus aureus (MIC=12.5 μg/ml for both extracts) and Micrococcus luteus (MIC=50 and 12.5 μg/ml, resp.). Among the tested metabolites, (6E)-1-(2,4-dihydroxyphenyl)-3,7,11-trimethyl-3-vinyldodeca-6,10-dien-1-one (2) and (2S*,3R*)-2-[(3E)-4,8-dimethylnona-3,7-dien-1-yl]-2,3-dihydro-7-hydroxy-2,3-dimethylfuro[3,2-c]coumarin (4) demonstrated the best antimicrobial activity. Compounds 2 and 4 both strongly inhibited the growth of M. luteus (MIC=11.2 and 5.2 μM, resp.) and Staphylococcus epidermidis (MIC=22.5 and 10.5 μM, resp.) and compound 2 additionally also the growth of Bacillus subtilis (MIC=11.2 μM). The cytotoxic activity of the isolates was tested against three human cancer cell lines, viz., cervical adenocarcinoma (HeLa), chronic myelogenous leukemia (K562), and breast cancer (MCF-7) cells. The CHCl3 extract exhibited strong cytotoxic activity against all cell lines (IC50 <11.0 μg/ml). All compounds strongly inhibited the growth of the K562 and HeLa cell lines. Compound 4 exhibited also a strong activity against the MCF-7 cell line, comparable to that of cisplatin (IC50 =22.32±1.32 vs. 18.67±0.75μM). PMID:26460563

  16. Direct photoaffinity labeling of cellular retinoic acid-binding protein I (CRABP-I) with all-trans-retinoic acid: identification of amino acids in the ligand binding site.

    PubMed

    Chen, G; Radominska-Pandya, A

    2000-10-17

    Cellular retinoic acid-binding proteins I and II (CRABP-I and -II, respectively) are transport proteins for all-trans-retinoic acid (RA), an active metabolite of vitamin A (retinol), and have been reported to be directly involved in the metabolism of RA. In this study, direct photoaffinity labeling with [11,12-(3)H]RA was used to identify amino acids comprising the ligand binding site of CRABP-I. Photoaffinity labeling of CRABP-I with [(3)H]RA was light- and concentration-dependent and was protected by unlabeled RA and various retinoids, indicating that the labeling was directed to the RA-binding site. Photolabeled CRABP-I was hydrolyzed with endoproteinase Lys-C to yield radioactive peptides, which were separated by reversed-phase HPLC for analysis by Edman degradation peptide sequencing. This method identified five modified amino acids from five separate HPLC fractions: Trp7, Lys20, Arg29, Lys38, and Trp109. All five amino acids are located within one side of the "barrel" structure in the area indicated by the reported crystal structure as the ligand binding site. This is the first direct identification of specific amino acids in the RA-binding site of CRABPs by photoaffinity labeling. These results provide significant information about the ligand binding site of the CRABP-I molecule in solution. PMID:11027136

  17. Citrus fruits as a treasure trove of active natural metabolites that potentially provide benefits for human health.

    PubMed

    Lv, Xinmiao; Zhao, Siyu; Ning, Zhangchi; Zeng, Honglian; Shu, Yisong; Tao, Ou; Xiao, Cheng; Lu, Cheng; Liu, Yuanyan

    2015-01-01

    Citrus fruits, which are cultivated worldwide, have been recognized as some of the most high-consumption fruits in terms of energy, nutrients and health supplements. What is more, a number of these fruits have been used as traditional medicinal herbs to cure diseases in several Asian countries. Numerous studies have focused on Citrus secondary metabolites as well as bioactivities and have been intended to develop new chemotherapeutic or complementary medicine in recent decades. Citrus-derived secondary metabolites, including flavonoids, alkaloids, limonoids, coumarins, carotenoids, phenolic acids and essential oils, are of vital importance to human health due to their active properties. These characteristics include anti-oxidative, anti-inflammatory, anti-cancer, as well as cardiovascular protective effects, neuroprotective effects, etc. This review summarizes the global distribution and taxonomy, numerous secondary metabolites and bioactivities of Citrus fruits to provide a reference for further study. Flavonoids as characteristic bioactive metabolites in Citrus fruits are mainly introduced. PMID:26705419

  18. Antimicrobial and antiprotozoal activities of secondary metabolites from the fungus Eurotium repens

    PubMed Central

    Gao, Jiangtao; Radwan, Mohamed M.; León, Francisco; Wang, Xiaoning; Jacob, Melissa R.; Tekwani, Babu L.; Khan, Shabana I.; Lupien, Shari; Hill, Robert A.; Dugan, Frank M.; Cutler, Horace G.

    2011-01-01

    In this study, we examined in vitro antibacterial, antifungal, antimalarial, and antileishmanial activities of secondary metabolites (1–8) isolated from the fungus Eurotium repens. All compounds showed mild to moderate antibacterial or antifungal or both activities except 7. The activity of compound 6 was the best of the group tested. The in vitro antimalarial evaluation of these compounds revealed that compounds 1–3, 5, and 6 showed antimalarial activities against both chloroquine-sensitive and chloroquine-resistant strains of Plasmodium falciparum with IC50 values in the range of 1.1–3.0 μg/ml without showing any cytotoxicity to the mammalian cells. Compound 5 displayed the highest antimalarial activity. Antileishmanial activity against Leishmania donovani promastigotes was observed for compounds 1–6 with IC50 values ranging from 6.2 to 23 μg/ml. Antileishmanial activity of compounds 5 and 6 (IC50 values of 7.5 and 6.2 μg/ml, respectively) was more potent than 1–4 (IC50 values ranging from 19–23 μg/ml). Compounds 7 and 8 did not show any antiprotozoal effect. Preliminary structure and activity relationship studies indicated that antibacterial, antifungal, antimalarial, and antileishmanial activities associated with phenol derivates (1–6) seem to be dependent on the number of double bonds in the side chain, which would be important for lead optimization in the future. PMID:23024574

  19. Phenotypic and metabolic investigation of a CSF-1R kinase receptor inhibitor (BLZ945) and its pharmacologically active metabolite.

    PubMed

    Krauser, Joel A; Jin, Yi; Walles, Markus; Pfaar, Ulrike; Sutton, James; Wiesmann, Marion; Graf, Daniel; Pflimlin-Fritschy, Veronique; Wolf, Thierry; Camenisch, Gian; Swart, Piet

    2015-02-01

    1. 4-[2((1R,2R)-2-Hydroxycyclohexylamino)-benzothiazol-6-yloxyl]-pyridine-2-carboxylic acid methylamide (BLZ945) is a small molecule inhibitor of CSF-1R kinase activity within osteoclasts designed to prevent skeletal related events in metastatic disease. Key metabolites were enzymatically and structurally characterized to understand the metabolic fate of BLZ945 and pharmacological implications. The relative intrinsic clearances for metabolites were derived from in vitro studies using human hepatocytes, microsomes and phenotyped with recombinant P450 enzymes. 2. Formation of a pharmacologically active metabolite (M9) was observed in human hepatocytes. The M9 metabolite is a structural isomer (diastereomer) of BLZ945 and is about 4-fold less potent. This isomer was enzymatically formed via P450 oxidation of the BLZ945 hydroxyl group, followed by aldo-keto reduction to the alcohol (M9). 3. Two reaction phenotyping approaches based on fractional clearances were applied to BLZ945 using hepatocytes and liver microsomes. The fraction metabolized (fm) or contribution ratio was determined for each metabolic reaction type (oxidation, glucuronidation or isomerization) as well as for each metabolite. The results quantitatively illustrate contribution ratios of the involved enzymes and pathways, e.g. the isomerization to metabolite M9 accounted for 24% intrinsic clearance in human hepatocytes. In summary, contribution ratios for the Phase I and Phase II pathways can be determined in hepatocytes. PMID:25180976

  20. Mutagenic activity and metabolites in the urine of workers exposed to trinitrotoluene (TNT).

    PubMed

    Ahlborg, G; Einistö, P; Sorsa, M

    1988-05-01

    Urine samples taken after work and after a free weekend from 50 workers employed in various activities in a chemical plant manufacturing explosives were analysed. On the basis of hygienic surveys, the subjects were divided into three categories of exposure to trinitrotoluene (TNT). The urine analyses consisted of gas chromatographic identification of TNT and its two metabolites, 4-ADNT and 2-ADNT, and a determination of the mutagenic activity. Two frame shift detector strains of Salmonella typhimurium were used, TA 98 and TA 98 NR, the latter being deficient in endogenous nitroreductase activity. On the basis of previous results on TNT mutagenicity, no exogeneous metabolic system was used to test the urine concentrates. Both tester strains showed that the mean urinary mutagenic activity was higher in the after work samples than in post weekend samples from the same subjects, showing that bacterial nitroreductase activity was not significantly responsible for the mutagenicity, although the response was higher with strain TA 98 than with TA 98 NR. The interindividual variation in urine mutagenicity was high, however, and the difference between the two sampling times was statistically significant (p less than 0.05) only for the high exposed group (workers in trotyl foundry and sieve house). Correlation between urinary mutagenicity and concentration of TNT in urine was poor; correlation was significant only with the urinary concentration of 4-ADNT. The correlation between urinary TNT and both metabolites was good (p less than 0.001). These results suggest that analysis of 4-ADNT in urine would be a sufficient biological measure for controlling exposure to TNT. PMID:3378017

  1. Biotransformation of Bisphenol AF to Its Major Glucuronide Metabolite Reduces Estrogenic Activity

    PubMed Central

    Yin, Jie; Zhang, Jing; Feng, Yixing; Shao, Bing

    2013-01-01

    Bisphenol AF (BPAF), an endocrine disrupting chemical, can induce estrogenic activity through binding to estrogen receptor (ER). However, the metabolism of BPAF in vivo and the estrogenic activity of its metabolites remain unknown. In the present study, we identified four metabolites including BPAF diglucuronide, BPAF glucuronide (BPAF-G), BPAF glucuronide dehydrated and BPAF sulfate in the urine of Sprague-Dawley (SD) rats. BPAF-G was further characterized by nuclear magnetic resonance (NMR). After treatment with a single dose of BPAF, BPAF was metabolized rapidly to BPAF-G, as detected in the plasma of SD rats. Biotransformation of BPAF to BPAF-G was confirmed with human liver microsomes (HLM), and Vmax of glucuronidation for HLM was 11.6 nmol/min/mg. We also found that BPAF glucuronidation could be mediated through several human recombinant UDP-glucuronosyltransferases (UGTs) including UGT1A1, UGT1A3, UGT1A8, UGT1A9, UGT2B4, UGT2B7, UGT2B15 and UGT2B17, among which UGT2B7 showed the highest efficiency of glucuronidation. To explain the biological function of BPAF biotransformation, the estrogenic activities of BPAF and BPAF-G were evaluated in ER-positive breast cancer T47D and MCF7 cells. BPAF significantly stimulates ER-regulated gene expression and cell proliferation at the dose of 100 nM and 1 μM in breast cancer cells. However, BPAF-G did not show any induction of estrogenic activity at the same dosages, implying that formation of BPAF-G is a potential host defense mechanism against BPAF. Based on our study, biotransformation of BPAF to BPAF-G can eliminate BPAF-induced estrogenic activity, which is therefore considered as reducing the potential threat to human beings. PMID:24349450

  2. Biotransformation of bisphenol AF to its major glucuronide metabolite reduces estrogenic activity.

    PubMed

    Li, Ming; Yang, Yunjia; Yang, Yi; Yin, Jie; Zhang, Jing; Feng, Yixing; Shao, Bing

    2013-01-01

    Bisphenol AF (BPAF), an endocrine disrupting chemical, can induce estrogenic activity through binding to estrogen receptor (ER). However, the metabolism of BPAF in vivo and the estrogenic activity of its metabolites remain unknown. In the present study, we identified four metabolites including BPAF diglucuronide, BPAF glucuronide (BPAF-G), BPAF glucuronide dehydrated and BPAF sulfate in the urine of Sprague-Dawley (SD) rats. BPAF-G was further characterized by nuclear magnetic resonance (NMR). After treatment with a single dose of BPAF, BPAF was metabolized rapidly to BPAF-G, as detected in the plasma of SD rats. Biotransformation of BPAF to BPAF-G was confirmed with human liver microsomes (HLM), and Vmax of glucuronidation for HLM was 11.6 nmol/min/mg. We also found that BPAF glucuronidation could be mediated through several human recombinant UDP-glucuronosyltransferases (UGTs) including UGT1A1, UGT1A3, UGT1A8, UGT1A9, UGT2B4, UGT2B7, UGT2B15 and UGT2B17, among which UGT2B7 showed the highest efficiency of glucuronidation. To explain the biological function of BPAF biotransformation, the estrogenic activities of BPAF and BPAF-G were evaluated in ER-positive breast cancer T47D and MCF7 cells. BPAF significantly stimulates ER-regulated gene expression and cell proliferation at the dose of 100 nM and 1 μM in breast cancer cells. However, BPAF-G did not show any induction of estrogenic activity at the same dosages, implying that formation of BPAF-G is a potential host defense mechanism against BPAF. Based on our study, biotransformation of BPAF to BPAF-G can eliminate BPAF-induced estrogenic activity, which is therefore considered as reducing the potential threat to human beings. PMID:24349450

  3. Isophosphoramide mustard, a metabolite of ifosfamide with activity against murine tumours comparable to cyclophosphamide.

    PubMed Central

    Struck, R. F.; Dykes, D. J.; Corbett, T. H.; Suling, W. J.; Trader, M. W.

    1983-01-01

    Isophosphoramide mustard was synthesized and was found to demonstrate activity essentially comparable to cyclophosphamide and ifosfamide against L1210 and P388 leukaemia. Lewis lung carcinoma, mammary adenocarcinoma 16/C, ovarian sarcoma M5076, and colon tumour 6A, in mice and Yoshida ascitic sarcoma in rats. At doses less than, or equivalent to, the LD10, isophosphoramide mustard retained high activity against cyclophosphamide-resistant L1210 and P388 leukaemias, but was less active against intracerebrally-implanted P388 leukaemia while cyclophosphamide produced a 4 log10 tumour cell reduction. It was also less active (one log10 lower cell kill) than cyclophosphamide against the B16 melonoma. Metabolism studies on ifosfamide in mice identified isophosphoramide mustard in blood. In addition, unchanged drug, carboxyifosfamide, 4-ketoifosfamide, dechloroethyl cyclophosphamide, dechloroethylifosfamide, and alcoifosfamide were identified. The latter 4 metabolites were also identified in urine from an ifosfamide-treated dog. In a simulated in vitro pharmacokinetic experiment against L1210 leukaemia in which drugs were incubated at various concentrations for various times, both 4-hydroxycyclophosphamide and isophosphoramide mustard exhibited significant cytoxicity at concentration times time values of 100-1000 micrograms X min ml-1, while acrolein was significantly cytotoxic at 10 micrograms X min ml-1. Treatment of mice with drug followed by L1210 cells demonstrated a shorter duration of effective levels of cytotoxic activity for isophosphoramide mustard and phosphoramide mustard in comparison with cyclophosphamide and ifosfamide. Isophosphoramide mustard and 2-chloroethylamine, a potential hydrolysis product of isophosphoramide mustard and carboxyifosfamide, were less mutagenic in the standard Ames test than the 2 corresponding metabolites of cyclophosphamide [phosphoramide mustard and bis(2-chloroethyl)amine]. PMID:6821629

  4. Plasma cathepsin D isoforms and their active metabolites increase after myocardial infarction and contribute to plasma renin activity.

    PubMed

    Naseem, R Haris; Hedegard, Wade; Henry, Timothy D; Lessard, Jennifer; Sutter, Kathryn; Katz, Stephen A

    2005-03-01

    Plasma renin activity (PRA) is often found to increase after myocardial infarction (MI). Elevated PRA may contribute to increased myocardial angiotensin II that is responsible for maladaptive remodeling of the myocardium after MI. We hypothesized that MI would also result in cardiac release of cathepsin D, a ubiquitous lysosomal enzyme with high renin sequence homology. Cathepsin D release from damaged myocardial tissue could contribute to angiotensin formation by acting as an enzymatic alternate to renin. We assessed circulating renin and cathepsin D from both control and MI patient plasma (7-20 hours after MI) using shallow gradient focusing that allowed for independent measurement of both enzymes. Cathepsin D was increased significantly in the plasma after MI (P < 0.001). Furthermore, circulating active cathepsin D metabolites were also significantly elevated after MI (P < 0.04), and contained the majority of cathepsin D activity in plasma. Spiking control plasma with cathepsin D resulted in a variable but significant (P = 0.005) increase in PRA using a clinical assay. We conclude that 7-20 hours after MI, plasma cathepsin D is significantly elevated and most of the active enzymatic activity is circulating as plasma metabolites. Circulating cathepsin D can falsely increase clinical PRA determinations, and may also provide an alternative angiotensin formation pathway after MI. PMID:15739123

  5. Symphonia globulifera, a widespread source of complex metabolites with potent biological activities.

    PubMed

    Fromentin, Yann; Cottet, Kevin; Kritsanida, Marina; Michel, Sylvie; Gaboriaud-Kolar, Nicolas; Lallemand, Marie-Christine

    2015-01-01

    Symphonia globulifera has been widely used in traditional medicine and has therefore been subjected to several phytochemical studies in the American and African continents. Interestingly, some disparities have been observed concerning its metabolic profile. Several phytochemical studies of S. globulifera have led to the identification of more than 40 compounds, including several polycyclic polyprenylated acylphloroglucinols. Biological evaluations have pointed out the promising biological activities of these secondary metabolites, mostly as antiparasitic or antimicrobial, confirming the traditional use of this plant. The purpose of this review is to describe the natural occurrence, botanical aspects, ethnomedicinal use, structure, and biogenesis, as well as biological activities of compounds isolated from this species according to their provenance. PMID:25590372

  6. Bioactive Metabolites from Chaetomium aureum: Structure Elucidation and Inhibition of the Hsp90 Machine Chaperoning Activity

    PubMed Central

    Kabbaj, Fatima Zahra; Lu, Su; Faouzi, My El Abbés; Meddah, Bouchra; Proksch, Peter; Cherrah, Yahya; Altenbach, Hans-Josef; Aly, Amal H.; Chadli, Ahmed; Debbab, Abdessamad

    2014-01-01

    Chemical investigation of the EtOAc extract of the fungus Chaetomium aureum, an endophyte of the Moroccan medicinal plant Thymelaea lythroides, afforded one new resorcinol derivative named chaetorcinol, together with five known metabolites. The structures of the isolated compounds were determined on the basis of one- and two-dimensional NMR spectroscopy and high-resolution mass spectrometry as well as by comparison with the literature. All compounds were tested for their activity towards the Hsp90 chaperoning machine in vitro using the progesterone receptor (PR) and rabbit reticulocyte lysate (RRL). Among the isolated compounds, only sclerotiorin efficiently inhibited the Hsp90 machine chaperoning activity. However, sclerotiorin showed no cytotoxic effect on breast cancer Hs578T, MDA-MB-231 and prostate cancer LNCaP cell lines. Interestingly, deacetylation of sclerotiorin increased its cytotoxicity toward the tested cell lines over a period of 48h. PMID:25482429

  7. Triphenyl phosphate-induced developmental toxicity in zebrafish: Potential role of the retinoic acid receptor

    PubMed Central

    Isales, Gregory M.; Hipszer, Rachel A.; Raftery, Tara D.; Chen, Albert; Stapleton, Heather M.; Volz, David C.

    2015-01-01

    Using zebrafish as a model, we previously reported that developmental exposure to triphenyl phosphate (TPP) – a high-production volume organophosphate-based flame retardant – results in dioxin-like cardiac looping impairments that are independent of the aryl hydrocarbon receptor. Using a pharmacologic approach, the objective of this study was to investigate the potential role of retinoic acid receptor (RAR) – a nuclear receptor that regulates vertebrate heart morphogenesis – in mediating TPP-induced developmental toxicity in zebrafish. We first revealed that static exposure of zebrafish from 5-72 hours post-fertilization (hpf) to TPP in the presence of non-toxic concentrations of an RAR antagonist (BMS493) significantly enhanced TPP-induced toxicity (relative to TPP alone), even though identical non-toxic BMS493 concentrations mitigated retinoic acid (RA)-induced toxicity. BMS493-mediated enhancement of TPP toxicity was not a result of differential TPP uptake or metabolism, as internal embryonic doses of TPP and diphenyl phosphate (DPP) – a primary TPP metabolite - were not different in the presence or absence of BMS493. Using real-time PCR, we then quantified the relative change in expression of cytochrome P450 26a1 (cyp26a1) – a major target gene for RA-induced RAR activation in zebrafish – and found that RA and TPP exposure resulted in a ∼5-fold increase and decrease in cyp26a1 expression, respectively, relative to vehicle-exposed embryos. To address whether TPP may interact with human RARs, we then exposed Chinese hamster ovary cells stably transfected with chimeric human RARα-, RARβ-, or RARγ to TPP in the presence of RA, and found that TPP significantly inhibited RA-induced luciferase activity in a concentration-dependent manner. Overall, our findings suggest that zebrafish RARs may be involved in mediating TPP-induced developmental toxicity, a mechanism of action that may have relevance to humans. PMID:25725299

  8. In-stream attenuation of neuro-active pharmaceuticals and their metabolites

    USGS Publications Warehouse

    Writer, Jeffrey; Antweiler, Ronald C.; Ferrar, Imma; Ryan, Joseph N.; Thurman, Michael

    2013-01-01

    In-stream attenuation was determined for 14 neuro-active pharmaceuticals and associated metabolites. Lagrangian sampling, which follows a parcel of water as it moves downstream, was used to link hydrological and chemical transformation processes. Wastewater loading of neuro-active compounds varied considerably over a span of several hours, and thus a sampling regime was used to verify that the Lagrangian parcel was being sampled and a mechanism was developed to correct measured concentrations if it was not. In-stream attenuation over the 5.4-km evaluated reach could be modeled as pseudo-first-order decay for 11 of the 14 evaluated neuro-active pharmaceutical compounds, illustrating the capacity of streams to reduce conveyance of neuro-active compounds downstream. Fluoxetine and N-desmethyl citalopram were the most rapidly attenuated compounds (t1/2 = 3.6 ± 0.3 h, 4.0 ± 0.2 h, respectively). Lamotrigine, 10,11,-dihydro-10,11,-dihydroxy-carbamazepine, and carbamazepine were the most persistent (t1/2 = 12 ± 2.0 h, 12 ± 2.6 h, 21 ± 4.5 h, respectively). Parent compounds (e.g., buproprion, carbamazepine, lamotrigine) generally were more persistent relative to their metabolites. Several compounds (citalopram, venlafaxine, O-desmethyl-venlafaxine) were not attenuated. It was postulated that the primary mechanism of removal for these compounds was interaction with bed sediments and stream biofilms, based on measured concentrations in stream biofilms and a column experiment using stream sediments.

  9. In-stream attenuation of neuro-active pharmaceuticals and their metabolites.

    PubMed

    Writer, Jeffrey H; Antweiler, Ronald C; Ferrer, Imma; Ryan, Joseph N; Thurman, E Michael

    2013-09-01

    In-stream attenuation was determined for 14 neuro-active pharmaceuticals and associated metabolites. Lagrangian sampling, which follows a parcel of water as it moves downstream, was used to link hydrological and chemical transformation processes. Wastewater loading of neuro-active compounds varied considerably over a span of several hours, and thus a sampling regime was used to verify that the Lagrangian parcel was being sampled and a mechanism was developed to correct measured concentrations if it was not. In-stream attenuation over the 5.4-km evaluated reach could be modeled as pseudo-first-order decay for 11 of the 14 evaluated neuro-active pharmaceutical compounds, illustrating the capacity of streams to reduce conveyance of neuro-active compounds downstream. Fluoxetine and N-desmethyl citalopram were the most rapidly attenuated compounds (t1/2 = 3.6 ± 0.3 h, 4.0 ± 0.2 h, respectively). Lamotrigine, 10,11,-dihydro-10,11,-dihydroxy-carbamazepine, and carbamazepine were the most persistent (t1/2 = 12 ± 2.0 h, 12 ± 2.6 h, 21 ± 4.5 h, respectively). Parent compounds (e.g., buproprion, carbamazepine, lamotrigine) generally were more persistent relative to their metabolites. Several compounds (citalopram, venlafaxine, O-desmethyl-venlafaxine) were not attenuated. It was postulated that the primary mechanism of removal for these compounds was interaction with bed sediments and stream biofilms, based on measured concentrations in stream biofilms and a column experiment using stream sediments. PMID:23952127

  10. Continuing hunt for endophytic actinomycetes as a source of novel biologically active metabolites.

    PubMed

    Masand, Meeta; Jose, Polpass Arul; Menghani, Ekta; Jebakumar, Solomon Robinson David

    2015-12-01

    Drug-resistant pathogens and persistent agrochemicals mount the detrimental threats against human health and welfare. Exploitation of beneficial microorganisms and their metabolic inventions is most promising way to tackle these two problems. Since the successive discoveries of penicillin and streptomycin in 1940s, numerous biologically active metabolites have been discovered from different microorganisms, especially actinomycetes. In recent years, actinomycetes that inhabit unexplored environments have received significant attention due to their broad diversity and distinctive metabolic potential with medical, agricultural and industrial importance. In this scenario, endophytic actinomycetes that inhabit living tissues of plants are emerging as a potential source of novel bioactive compounds for the discovery of drug leads. Also, endophytic actinomycetes are considered as bio-inoculants to improve crop performance through organic farming practices. Further efforts on exploring the endophytic actinomycetes associated with the plants warrant the likelihood of discovering new taxa and their metabolites with novel chemical structures and biotechnological importance. This mini-review highlights the recent achievements in isolation of endophytic actinomycetes and an assortment of bioactive compounds. PMID:26410426

  11. Atrazine and its main metabolites alter the locomotor activity of larval zebrafish (Danio rerio).

    PubMed

    Liu, Zhenzhen; Wang, Yueyi; Zhu, Zhihong; Yang, Enlu; Feng, Xiayan; Fu, Zhengwei; Jin, Yuanxiang

    2016-04-01

    Atrazine (ATZ) and its main chlorometabolites, i.e., diaminochlorotriazine (DACT), deisopropylatrazine (DIP), and deethylatrazine (DE), have been widely detected in aquatic systems near agricultural fields. However, their possible effects on aquatic animals are still not fully understood. In this study, it was observed that several developmental endpoints such as the heart beat, hatchability, and morphological abnormalities were influenced by ATZ and its metabolites in different developmental stages. In addition, after 5 days of exposure to 30, 100, 300 μg L(-1) ATZ and its main chlorometabolites, the swimming behaviors of larval zebrafish were significantly disturbed, and the acetylcholinesterase (AChE) activities were consistently inhibited. Our results also demonstrate that ATZ and its main chlorometabolites are neuroendocrine disruptors that impact the expression of neurotoxicity-related genes such as Ache, Gap43, Gfap, Syn2a, Shha, Mbp, Elavl3, Nestin and Ngn1 in early developmental stages of zebrafish. According to our results, it is possible that not only ATZ but also its metabolites (DACT, DIP and DE) have the same or even more toxic effects on different endpoints of the early developmental stages of zebrafish. PMID:26803580

  12. Anthocyanins and their gut metabolites reduce the adhesion of monocyte to TNFα-activated endothelial cells at physiologically relevant concentrations.

    PubMed

    Krga, Irena; Monfoulet, Laurent-Emmanuel; Konic-Ristic, Aleksandra; Mercier, Sylvie; Glibetic, Maria; Morand, Christine; Milenkovic, Dragan

    2016-06-01

    An increasing number of evidence suggests a protective role of dietary anthocyanins against cardiovascular diseases. Anthocyanins' extensive metabolism indicates that their metabolites could be responsible for the protective effects associated with consumption of anthocyanin-rich foods. The aim of this work was to investigate the effect of plasma anthocyanins and their metabolites on the adhesion of monocytes to TNFα-activated endothelial cells and on the expression of genes encoding cell adhesion molecules. Human umbilical vein endothelial cells (HUVECs) were exposed to circulating anthocyanins: cyanidin-3-arabinoside, cyanidin-3-galactoside, cyanidin-3-glucoside, delphinidin-3-glucoside, peonidin-3-glucoside, anthocyanin degradation product: 4-hydroxybenzaldehyde, or to their gut metabolites: protocatechuic, vanillic, ferulic and hippuric acid, at physiologically-relevant concentrations (0.1-2 μM) and time of exposure. Both anthocyanins and gut metabolites decreased the adhesion of monocytes to HUVECs, with a magnitude ranging from 18.1% to 47%. The mixture of anthocyanins and that of gut metabolites also reduced monocyte adhesion. However, no significant effect on the expression of genes encoding E-selectin, ICAM1 and VCAM1 was observed, suggesting that other molecular targets are involved in the observed effect. In conclusion, this study showed the potency of anthocyanins and their gut metabolites to modulate the adhesion of monocytes to endothelial cells, the initial step in atherosclerosis development, under physiologically-relevant conditions. PMID:26873533

  13. Linking diet, physical activity, cardiorespiratory fitness and obesity to serum metabolite networks: findings from a population-based study

    PubMed Central

    Floegel, A; Wientzek, A; Bachlechner, U; Jacobs, S; Drogan, D; Prehn, C; Adamski, J; Krumsiek, J; Schulze, M B; Pischon, T; Boeing, H

    2014-01-01

    Objective: It is not yet resolved how lifestyle factors and intermediate phenotypes interrelate with metabolic pathways. We aimed to investigate the associations between diet, physical activity, cardiorespiratory fitness and obesity with serum metabolite networks in a population-based study. Methods: The present study included 2380 participants of a randomly drawn subcohort of the European Prospective Investigation into Cancer and Nutrition-Potsdam. Targeted metabolomics was used to measure 127 serum metabolites. Additional data were available including anthropometric measurements, dietary assessment including intake of whole-grain bread, coffee and cake and cookies by food frequency questionnaire, and objectively measured physical activity energy expenditure and cardiorespiratory fitness in a subsample of 100 participants. In a data-driven approach, Gaussian graphical modeling was used to draw metabolite networks and depict relevant associations between exposures and serum metabolites. In addition, the relationship of different exposure metabolite networks was estimated. Results: In the serum metabolite network, the different metabolite classes could be separated. There was a big group of phospholipids and acylcarnitines, a group of amino acids and C6-sugar. Amino acids were particularly positively associated with cardiorespiratory fitness and physical activity. C6-sugar and acylcarnitines were positively associated with obesity and inversely with intake of whole-grain bread. Phospholipids showed opposite associations with obesity and coffee intake. Metabolite networks of coffee intake and obesity were strongly inversely correlated (body mass index (BMI): r=−0.57 and waist circumference: r=−0.59). A strong positive correlation was observed between metabolite networks of BMI and waist circumference (r=0.99), as well as the metabolite networks of cake and cookie intake with cardiorespiratory fitness and intake of whole-grain bread (r=0.52 and r=0

  14. Benzene's metabolites alter c-MYB activity via reactive oxygen species in HD3 cells

    SciTech Connect

    Wan, Joanne; Winn, Louise M. . E-mail: winnl@queensu.ca

    2007-07-15

    Benzene is a known leukemogen that is metabolized to form reactive intermediates and reactive oxygen species (ROS). The c-Myb oncoprotein is a transcription factor that has a critical role in hematopoiesis. c-Myb transcript and protein have been overexpressed in a number of leukemias and cancers. Given c-Myb's role in hematopoiesis and leukemias, it is hypothesized that benzene interferes with the c-Myb signaling pathway and that this involves ROS. To investigate our hypothesis, we evaluated whether benzene, 1,4-benzoquinone, hydroquinone, phenol, and catechol generated ROS in chicken erythroblast HD3 cells, as measured by 5-(and-6)-chloromethyl-2',7'-dichlorodihydrofluorescein diacetate (DCFDA) and dihydrorhodamine-123 (DHR-123), and whether the addition of 100 U/ml of the antioxidating enzyme superoxide dismutase (SOD) could prevent ROS generation. Reduced to oxidized glutathione ratios (GSH:GSSG) were also assessed as well as hydroquinone and benzoquinone's effects on c-Myb protein levels and activation of a transiently transfected reporter construct. Finally we attempted to abrogate benzene metabolite mediated increases in c-Myb activity with the use of SOD. We found that benzoquinone, hydroquinone, and catechol increased DCFDA fluorescence, increased DHR-123 fluorescence, decreased GSH:GSSG ratios, and increased reporter construct expression after 24 h of exposure. SOD was able to prevent DCFDA fluorescence and c-Myb activity caused by benzoquinone and hydroquinone only. These results are consistent with other studies, which suggest metabolite differences in benzene-mediated toxicity. More importantly, this study supports the hypothesis that benzene may mediate its toxicity through ROS-mediated alterations in the c-Myb signaling pathway.

  15. The antitumor activity study of ginsenosides and metabolites in lung cancer cell

    PubMed Central

    Xu, Feng-Yuan; Shang, Wen-Qing; Yu, Jia-Jun; Sun, Qian; Li, Ming-Qing; Sun, Jian-Song

    2016-01-01

    Ginseng and its components exert various biological effects, including antioxidant, anti-carcinogenic, anti-mutagenic, and antitumor activity. Ginsenosides are the main biological components of ginseng. Protopanaxadiol (PPD) and protopanaxatriol (PPT) are two metabolites of ginsenosides. However, the difference between these compounds in anti-lung cancer is unclear. The present study aimed to evaluate the antitumor activity of PPD, PPT, Ginsenosides-Rg3 (G-Rg3) and Ginsenosides-Rh2 (G-Rh2) in lung cancer cell. After treatment with cisplatin, PPD, PPT, G-Rg3 or G-Rh2, the viability, apoptosis level and invasiveness of lung cell lines (A549 cell, a lung adenocarcinoma cell line and SK-MES-1 cell, a lung squamous cell line) in vitro were analyzed by Cell Counting Kit-8 (CCK8), Annexin V/PI apoptosis and Matrigel invasion assays, respectively. Here we found that all these compounds led to significant decreases of viability and invasiveness and an obvious increase of apoptosis of A549 and SK-MES-1 cells. Among these, the viability of SK-MES-1 cell treated with PPT was decreased to 66.8%, and this effect was closest to Cisplatin. G-Rg3 had the highest stimulatory effect on apoptosis, and PTT had the highest inhibitory effect on cell invasiveness in A549 and SK-MES-1 cells. These results indicate that both ginsenosides and two metabolites have antitumor activity on lung cancer cell in vitro. However, PPT is more powerful for inhibiting the viability and invasiveness of lung cancer cell, especially lung squamous cell. G-Rg3 has the best pro-apoptosis effects. This study provides a scientific basis for potential therapeutic strategies targeted to lung cancer by further structure modification. PMID:27186294

  16. Cysteamine, the natural metabolite of pantetheinase, shows specific activity against Plasmodium.

    PubMed

    Min-Oo, Gundula; Ayi, Kodjo; Bongfen, Silayuv E; Tam, Mifong; Radovanovic, Irena; Gauthier, Susan; Santiago, Helton; Rothfuchs, Antonio Gigliotti; Roffê, Ester; Sher, Alan; Mullick, Alaka; Fortin, Anny; Stevenson, Mary M; Kain, Kevin C; Gros, Philippe

    2010-08-01

    In mice, loss of pantetheinase activity causes susceptibility to infection with Plasmodium chabaudi AS. Treatment of mice with the pantetheinase metabolite cysteamine reduces blood-stage replication of P. chabaudi and significantly increases survival. Similarly, a short exposure of Plasmodium to cysteamine ex vivo is sufficient to suppress parasite infectivity in vivo. This effect of cysteamine is specific and not observed with a related thiol (dimercaptosuccinic acid) or with the pantethine precursor of cysteamine. Also, cysteamine does not protect against infection with the parasite Trypanosoma cruzi or the fungal pathogen Candida albicans, suggesting cysteamine acts directly against the parasite and does not modulate host inflammatory response. Cysteamine exposure also blocks replication of P. falciparum in vitro; moreover, these treated parasites show higher levels of intact hemoglobin. This study highlights the in vivo action of cysteamine against Plasmodium and provides further evidence for the involvement of pantetheinase in host response to this infection. PMID:20219464

  17. Cysteamine, the natural metabolite of pantetheinase, shows specific activity against Plasmodium

    PubMed Central

    Min-Oo, Gundula; Ayi, Kodjo; Bongfen, Silayuv E.; Tam, Mifong; Radovanovic, Irena; Gauthier, Susan; Santiago, Helton; Rothfuchs, Antonio Gigliotti; Roffê, Ester; Sher, Alan; Mullick, Alaka; Fortin, Anny; Stevenson, Mary M.; Kain, Kevin C.; Gros, Philippe

    2016-01-01

    In mice, loss of pantetheinase activity causes susceptibility to infection with Plasmodium chabaudi AS. Treatment of mice with the pantetheinase metabolite cysteamine reduces blood-stage replication of P. chabaudi and significantly increases survival. Similarly, a short exposure of Plasmodium to cysteamine ex vivo is sufficient to suppress parasite infectivity in vivo. This effect of cysteamine is specific and not observed with a related thiol (dimercaptosuccinic acid) or with the pantethine precursor of cysteamine. Also, cysteamine does not protect against infection with the parasite Trypanosoma cruzi or the fungal pathogen Candida albicans, suggesting cysteamine acts directly against the parasite and does not modulate host inflammatory response. Cysteamine exposure also blocks replication of P. falciparum in vitro; moreover, these treated parasites show higher levels of intact hemoglobin. This study highlights the in vivo action of cysteamine against Plasmodium and provides further evidence for the involvement of pantetheinase in host response to this infection. PMID:20219464

  18. Evaluation of in vitro antiprotozoal activity of Ajuga laxmannii and its secondary metabolites.

    PubMed

    Atay, Irem; Kirmizibekmez, Hasan; Kaiser, Marcel; Akaydin, Galip; Yesilada, Erdem; Tasdemir, Deniz

    2016-09-01

    Context Some Ajuga L. (Lamiaceae) species are traditionally used for the treatment of malaria, as well as fever, which is a common symptom of many parasitic diseases. Objective In the continuation of our studies on the identification of antiprotozoal secondary metabolites of Turkish Lamiaceae species, we have investigated the aerial parts of Ajuga laxmannii. Materials and methods The aerial parts of A. laxmannii were extracted with MeOH. The H2O subextract was subjected to polyamide, C18-MPLC and SiO2 CCs to yield eight metabolites. The structures of the isolates were elucidated by NMR spectroscopy and MS analyses. The extract, subextracts as well as the isolates were tested for their in vitro antiprotozoal activities against Plasmodium falciparum, Trypanasoma brucei rhodesiense, T. cruzi and Leishmania donovani at concentrations of 90-0.123 μg/mL. Results Two iridoid glycosides harpagide (1) and 8-O-acetylharpagide (2), three o-coumaric acid derivatives cis-melilotoside (3), trans-melilotoside (4) and dihydromelilotoside (5), two phenylethanoid glycosides verbascoside (6) and galactosylmartynoside (7) and a flavone-C-glycoside, isoorientin (8) were isolated. Many compounds showed moderate to good antiparasitic activity, with isoorientin (8) displaying the most significant antimalarial potential (an IC50 value of 9.7 μg/mL). Discussion and conclusion This is the first report on the antiprotozoal evaluation of A. laxmannii extracts and isolates. Furthermore, isoorientin and dihydromelilotoside are being reported for the first time from the genus Ajuga. PMID:26734766

  19. Metabolite fingerprinting of pennycress (Thlaspi arvense L.) embryos to assess active pathways during oil synthesis

    PubMed Central

    Tsogtbaatar, Enkhtuul; Cocuron, Jean-Christophe; Sonera, Marcos Corchado; Alonso, Ana Paula

    2015-01-01

    Pennycress (Thlaspi arvense L.), a plant naturalized to North America, accumulates high levels of erucic acid in its seeds, which makes it a promising biodiesel and industrial crop. The main carbon sinks in pennycress embryos were found to be proteins, fatty acids, and cell wall, which respectively represented 38.5, 33.2, and 27.0% of the biomass at 21 days after pollination. Erucic acid reached a maximum of 36% of the total fatty acids. Together these results indicate that total oil and erucic acid contents could be increased to boost the economic competitiveness of this crop. Understanding the biochemical basis of oil synthesis in pennycress embryos is therefore timely and relevant to guide future breeding and/or metabolic engineering efforts. For this purpose, a combination of metabolomics approaches was conducted to assess the active biochemical pathways during oil synthesis. First, gas chromatography–mass spectrometry (GC-MS) profiling of intracellular metabolites highlighted three main families of compounds: organic acids, amino acids, and sugars/sugar alcohols. Secondly, these intermediates were quantified in developing pennycress embryos by liquid chromatography–tandem mass spectrometry (LC-MS/MS) in multiple reaction monitoring mode. Finally, partitional clustering analysis grouped the intracellular metabolites that shared a similar pattern of accumulation over time into eight clusters. This study underlined that: (i) sucrose might be stored rather than cleaved into hexoses; (ii) glucose and glutamine would be the main sources of carbon and nitrogen, respectively; and (iii) glycolysis, the oxidative pentose phosphate pathway, the tricarboxylic acid cycle, and the Calvin cycle were active in developing pennycress embryos. PMID:25711705

  20. Metabolite fingerprinting of pennycress (Thlaspi arvense L.) embryos to assess active pathways during oil synthesis.

    PubMed

    Tsogtbaatar, Enkhtuul; Cocuron, Jean-Christophe; Sonera, Marcos Corchado; Alonso, Ana Paula

    2015-07-01

    Pennycress (Thlaspi arvense L.), a plant naturalized to North America, accumulates high levels of erucic acid in its seeds, which makes it a promising biodiesel and industrial crop. The main carbon sinks in pennycress embryos were found to be proteins, fatty acids, and cell wall, which respectively represented 38.5, 33.2, and 27.0% of the biomass at 21 days after pollination. Erucic acid reached a maximum of 36% of the total fatty acids. Together these results indicate that total oil and erucic acid contents could be increased to boost the economic competitiveness of this crop. Understanding the biochemical basis of oil synthesis in pennycress embryos is therefore timely and relevant to guide future breeding and/or metabolic engineering efforts. For this purpose, a combination of metabolomics approaches was conducted to assess the active biochemical pathways during oil synthesis. First, gas chromatography-mass spectrometry (GC-MS) profiling of intracellular metabolites highlighted three main families of compounds: organic acids, amino acids, and sugars/sugar alcohols. Secondly, these intermediates were quantified in developing pennycress embryos by liquid chromatography-tandem mass spectrometry (LC-MS/MS) in multiple reaction monitoring mode. Finally, partitional clustering analysis grouped the intracellular metabolites that shared a similar pattern of accumulation over time into eight clusters. This study underlined that: (i) sucrose might be stored rather than cleaved into hexoses; (ii) glucose and glutamine would be the main sources of carbon and nitrogen, respectively; and (iii) glycolysis, the oxidative pentose phosphate pathway, the tricarboxylic acid cycle, and the Calvin cycle were active in developing pennycress embryos. PMID:25711705

  1. Assessment of the Potential Biological Activity of Low Molecular Weight Metabolites of Freshwater Macrophytes with QSAR

    PubMed Central

    Fedorova, Elena V.; Krylova, Julia V.

    2016-01-01

    The paper focuses on the assessment of the spectrum of biological activities (antineoplastic, anti-inflammatory, antifungal, and antibacterial) with PASS (Prediction of Activity Spectra for Substances) for the major components of three macrophytes widespread in the Holarctic species of freshwater, emergent macrophyte with floating leaves, Nuphar lutea (L.) Sm., and two species of submergent macrophyte groups, Ceratophyllum demersum L. and Potamogeton obtusifolius (Mert. et Koch), for the discovery of their ecological and pharmacological potential. The predicted probability of anti-inflammatory or antineoplastic activities above 0.8 was observed for twenty compounds. The same compounds were also characterized by high probability of antifungal and antibacterial activity. Six metabolites, namely, hexanal, pentadecanal, tetradecanoic acid, dibutyl phthalate, hexadecanoic acid, and manool, were a part of the major components of all three studied plants, indicating their high ecological significance and a certain universalism in their use by various species of water plants for the implementation of ecological and biochemical functions. This report underlines the role of identified compounds not only as important components in regulation of biochemical and metabolic pathways and processes in aquatic ecological systems, but also as potential pharmacological agents in the fight against different diseases. PMID:27200207

  2. Antiproliferative, Antibacterial and Antifungal Activity of the Lichen Xanthoria parietina and Its Secondary Metabolite Parietin

    PubMed Central

    Basile, Adriana; Rigano, Daniela; Loppi, Stefano; Di Santi, Annalisa; Nebbioso, Angela; Sorbo, Sergio; Conte, Barbara; Paoli, Luca; De Ruberto, Francesca; Molinari, Anna Maria; Altucci, Lucia; Bontempo, Paola

    2015-01-01

    Lichens are valuable natural resources used for centuries throughout the world as medicine, food, fodder, perfume, spices and dyes, as well as for other miscellaneous purposes. This study investigates the antiproliferative, antibacterial and antifungal activity of the acetone extract of the lichen Xanthoria parietina (Linnaeus) Theodor Fries and its major secondary metabolite, parietin. The extract and parietin were tested for antimicrobial activity against nine American Type Culture Collection standard and clinically isolated bacterial strains, and three fungal strains. Both showed strong antibacterial activity against all bacterial strains and matched clinical isolates, particularly against Staphylococcus aureus from standard and clinical sources. Among the fungi tested, Rhizoctonia solani was the most sensitive. The antiproliferative effects of the extract and parietin were also investigated in human breast cancer cells. The extract inhibited proliferation and induced apoptosis, both effects being accompanied by modulation of expression of cell cycle regulating genes such as p16, p27, cyclin D1 and cyclin A. It also mediated apoptosis by activating extrinsic and intrinsic cell death pathways, modulating Tumor Necrosis Factor-related apoptosis-inducing ligand (TRAIL) and B-cell lymphoma 2 (Bcl-2), and inducing Bcl-2-associated agonist of cell death (BAD) phosphorylation. Our results indicate that Xanthoria parietina is a major potential source of antimicrobial and anticancer substances. PMID:25860944

  3. Antiproliferative, antibacterial and antifungal activity of the lichen Xanthoria parietina and its secondary metabolite parietin.

    PubMed

    Basile, Adriana; Rigano, Daniela; Loppi, Stefano; Di Santi, Annalisa; Nebbioso, Angela; Sorbo, Sergio; Conte, Barbara; Paoli, Luca; De Ruberto, Francesca; Molinari, Anna Maria; Altucci, Lucia; Bontempo, Paola

    2015-01-01

    Lichens are valuable natural resources used for centuries throughout the world as medicine, food, fodder, perfume, spices and dyes, as well as for other miscellaneous purposes. This study investigates the antiproliferative, antibacterial and antifungal activity of the acetone extract of the lichen Xanthoria parietina (Linnaeus) Theodor Fries and its major secondary metabolite, parietin. The extract and parietin were tested for antimicrobial activity against nine American Type Culture Collection standard and clinically isolated bacterial strains, and three fungal strains. Both showed strong antibacterial activity against all bacterial strains and matched clinical isolates, particularly against Staphylococcus aureus from standard and clinical sources. Among the fungi tested, Rhizoctonia solani was the most sensitive. The antiproliferative effects of the extract and parietin were also investigated in human breast cancer cells. The extract inhibited proliferation and induced apoptosis, both effects being accompanied by modulation of expression of cell cycle regulating genes such as p16, p27, cyclin D1 and cyclin A. It also mediated apoptosis by activating extrinsic and intrinsic cell death pathways, modulating Tumor Necrosis Factor-related apoptosis-inducing ligand (TRAIL) and B-cell lymphoma 2 (Bcl-2), and inducing Bcl-2-associated agonist of cell death (BAD) phosphorylation. Our results indicate that Xanthoria parietina is a major potential source of antimicrobial and anticancer substances. PMID:25860944

  4. Assessment of the Potential Biological Activity of Low Molecular Weight Metabolites of Freshwater Macrophytes with QSAR.

    PubMed

    Kurashov, Evgeny A; Fedorova, Elena V; Krylova, Julia V; Mitrukova, Galina G

    2016-01-01

    The paper focuses on the assessment of the spectrum of biological activities (antineoplastic, anti-inflammatory, antifungal, and antibacterial) with PASS (Prediction of Activity Spectra for Substances) for the major components of three macrophytes widespread in the Holarctic species of freshwater, emergent macrophyte with floating leaves, Nuphar lutea (L.) Sm., and two species of submergent macrophyte groups, Ceratophyllum demersum L. and Potamogeton obtusifolius (Mert. et Koch), for the discovery of their ecological and pharmacological potential. The predicted probability of anti-inflammatory or antineoplastic activities above 0.8 was observed for twenty compounds. The same compounds were also characterized by high probability of antifungal and antibacterial activity. Six metabolites, namely, hexanal, pentadecanal, tetradecanoic acid, dibutyl phthalate, hexadecanoic acid, and manool, were a part of the major components of all three studied plants, indicating their high ecological significance and a certain universalism in their use by various species of water plants for the implementation of ecological and biochemical functions. This report underlines the role of identified compounds not only as important components in regulation of biochemical and metabolic pathways and processes in aquatic ecological systems, but also as potential pharmacological agents in the fight against different diseases. PMID:27200207

  5. A Practical Strategy to Discover New Antitumor Compounds by Activating Silent Metabolite Production in Fungi by Diethyl Sulphate Mutagenesis

    PubMed Central

    Fang, Shi-Ming; Wu, Chang-Jing; Li, Chang-Wei; Cui, Cheng-Bin

    2014-01-01

    Many fungal biosynthetic pathways are silent in standard culture conditions, and activation of the silent pathways may enable access to new metabolites with antitumor activities. The aim of the present study was to develop a practical strategy for microbial chemists to access silent metabolites in fungi. We demonstrated this strategy using a marine-derived fungus Penicillium purpurogenum G59 and a modified diethyl sulphate mutagenesis procedure. Using this strategy, we discovered four new antitumor compounds named penicimutanolone (1), penicimutanin A (2), penicimutanin B (3), and penicimutatin (4). Structures of the new compounds were elucidated by spectroscopic methods, especially extensive 2D NMR analysis. Antitumor activities were assayed by the MTT method using human cancer cell lines. Bioassays and HPLC-photodiode array detector (PDAD)-UV and HPLC-electron spray ionization (ESI)-MS analyses were used to estimate the activated secondary metabolite production. Compounds 2 and 3 had novel structures, and 1 was a new compound belonging to a class of very rare natural products from which only four members are so far known. Compounds 1–3 inhibited several human cancer cell lines with IC50 values lower than 20 μM, and 4 inhibited the cell lines to some extent. These results demonstrated the effectiveness of this strategy to discover new compounds by activating silent fungal metabolic pathways. These discoveries provide rationale for the increased use of chemical mutagenesis strategies in silent fungal metabolite studies. PMID:24681631

  6. Structural Characterization of a Therapeutic Anti-Methamphetamine Antibody Fragment: Oligomerization and Binding of Active Metabolites

    PubMed Central

    Gokulan, Kuppan; Varughese, Kottayil I.

    2013-01-01

    Vaccines and monoclonal antibodies (mAb) for treatment of (+)-methamphetamine (METH) abuse are in late stage preclinical and early clinical trial phases, respectively. These immunotherapies work as pharmacokinetic antagonists, sequestering METH and its metabolites away from sites of action in the brain and reduce the rewarding and toxic effects of the drug. A key aspect of these immunotherapy strategies is the understanding of the subtle molecular interactions important for generating antibodies with high affinity and specificity for METH. We previously determined crystal structures of a high affinity anti-METH therapeutic single chain antibody fragment (scFv6H4, KD = 10 nM) in complex with METH and the (+) stereoisomer of 3,4-methylenedioxymethamphetamine (MDMA, or “ecstasy”). Here we report the crystal structure of scFv6H4 in homo-trimeric unbound (apo) form (2.60Å), as well as monomeric forms in complex with two active metabolites; (+)-amphetamine (AMP, 2.38Å) and (+)-4-hydroxy methamphetamine (p-OH-METH, 2.33Å). The apo structure forms a trimer in the crystal lattice and it results in the formation of an intermolecular composite beta-sheet with a three-fold symmetry. We were also able to structurally characterize the coordination of the His-tags with Ni2+. Two of the histidine residues of each C-terminal His-tag interact with Ni2+ in an octahedral geometry. In the apo state the CDR loops of scFv6H4 form an open conformation of the binding pocket. Upon ligand binding, the CDR loops adopt a closed formation, encasing the drug almost completely. The structural information reported here elucidates key molecular interactions important in anti-methamphetamine abuse immunotherapy. PMID:24349338

  7. Integrated semi-physiological pharmacokinetic model for both sunitinib and its active metabolite SU12662

    PubMed Central

    Yu, Huixin; Steeghs, Neeltje; Kloth, Jacqueline S L; de Wit, Djoeke; van Hasselt, J G Coen; van Erp, Nielka P; Beijnen, Jos H; Schellens, Jan H M; Mathijssen, Ron H J; Huitema, Alwin D R

    2015-01-01

    Aims Previously published pharmacokinetic (PK) models for sunitinib and its active metabolite SU12662 were based on a limited dataset or lacked important elements such as correlations between sunitinib and its metabolite. The current study aimed to develop an improved PK model that circumvented these limitations and to prove the utility of the PK model in treatment optimization in clinical practice. Methods One thousand two hundred and five plasma samples from 70 cancer patients were collected from three PK studies with sunitinib and SU12662. A semi-physiological PK model for sunitinib and SU12662 was developed incorporating pre-systemic metabolism using non-linear mixed effects modelling (nonmem). Allometric scaling based on body weight was applied. The final model was used for simulation of the PK of different treatment regimens. Results Sunitinib and SU12662 PK were best described by a one and two compartment model, respectively. Introduction of pre-systemic formation of SU12662 strongly improved model fit, compared with solely systemic metabolism. The clearance of sunitinib and SU12662 was estimated at 35.7 (relative standard error (RSE) 5.7%) l h−1 and 17.1 (RSE 7.4%) l h−1, respectively for 70 kg patients. Correlation coefficients were estimated between inter-individual variability of both clearances, both volumes of distribution and between clearance and volume of distribution of SU12662 as 0.53, 0.48 and 0.45, respectively. Simulation of the PK model predicted correctly the ratio of patients who did not reach proposed PK targets for efficacy. Conclusions A semi-physiological PK model for sunitinib and SU12662 in cancer patients was presented including pre-systemic metabolism. The model was superior to previous PK models in many aspects. PMID:25393890

  8. Metabolism of 20(S)-Ginsenoside Rg₂ by Rat Liver Microsomes: Bioactivation to SIRT1-Activating Metabolites.

    PubMed

    Ma, Li-Yuan; Zhou, Qi-Le; Yang, Xin-Bao; Wang, Hong-Ping; Yang, Xiu-Wei

    2016-01-01

    20(S)-Ginsenoside Rg₂ (1) has recently become a hot research topic due to its potent bioactivities and abundance in natural sources such as the roots, rhizomes and stems-leaves of Panax ginseng. However, due to the lack of studies on systematic metabolic profiles, the prospects for new drug development of 1 are still difficult to predict, which has become a huge obstacle for its safe clinical use. To solve this problem, investigation of the metabolic profiles of 1 in rat liver microsomes was first carried out. To identify metabolites, a strategy of combined analyses based on prepared metabolites by column chromatography and ultra-performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry (UPLC-Q-TOF/MS) was performed. As a result, four metabolites M1-M4, including a rare new compound named ginsenotransmetin A (M1), were isolated and the structures were confirmed by spectroscopic analyses. A series of metabolites of 1, MA-MG, were also tentatively identified by UPLC-Q-TOF/MS in rat liver microsomal incubate of 1. Partial metabolic pathways were proposed. Among them, 1 and its metabolites M1, M3 and M4 were discovered for the first time to be activators of SIRT1. The SIRT1 activating effects of the metabolite M1 was comparable to those of 1, while the most interesting SIRT1 activatory effects of M3 and M4 were higher than that of 1 and comparable with that of resveratrol, a positive SIRT1 activator. These results indicate that microsome-dependent metabolism may represent a bioactivation pathway for 1. This study is the first to report the metabolic profiles of 1 in vitro, and the results provide an experimental foundation to better understand the in vivo metabolic fate of 1. PMID:27294899

  9. Anti-apoptotic role of retinoic acid in the inner ear of noise-exposed mice

    SciTech Connect

    Ahn, Joong Ho; Kang, Hun Hee; Kim, Young-Jin; Chung, Jong Woo . E-mail: jwchung@amc.seoul.kr

    2005-09-23

    Exposure to loud noise can induce temporary or permanent hearing loss, and acoustic trauma is the major cause of hearing impairment in industrial nations. However, the mechanisms underlying the death of hair cells after acoustic trauma remain unclear. In addition to its involvement in cellular stress and apoptosis, the c-Jun N-terminal kinase (JNK), a member of the mitogen-activated protein kinase family, is involved in cell survival, transformation, embryonic morphogenesis, and differentiation. JNK is primarily activated by various environmental stresses including noise, and the phenotypic result appears be to cell death. All-trans retinoic acid (ATRA) is an active metabolite of vitamin A that regulates a wide range of biological processes, including cell proliferation, differentiation, and morphogenesis. We evaluated the role of ATRA in preserving hearing in mice exposed to noise that can induce permanent hearing loss. Mice fed with ATRA before and during 3 consecutive days of noise exposure had a more preserved hearing threshold than mice fed sesame oil or saline. Histological and TUNEL staining of the cochlea showed significantly enhanced preservation of the organ of Corti, including outer hair cells and relatively low apoptotic nuclei, in mice-fed ATRA than in mice-fed sesame oil or saline. Phospho-JNK immunohistochemistry showed that ATRA inhibited the activation of JNK. These results suggest that ATRA has an anti-apoptotic effect on cochleae exposed to noise.

  10. Cytotoxic, Antiangiogenic and Antitelomerase Activity of Glucosyl- and Acyl- Resveratrol Prodrugs and Resveratrol Sulfate Metabolites.

    PubMed

    Falomir, Eva; Lucas, Ricardo; Peñalver, Pablo; Martí-Centelles, Rosa; Dupont, Alexia; Zafra-Gómez, Alberto; Carda, Miguel; Morales, Juan C

    2016-07-15

    Resveratrol (RES) is a natural polyphenol with relevant and varied biological activity. However, its low bioavailability and rapid metabolism to its glucuronate and sulfate conjugates has opened a debate on the mechanisms underlying its bioactivity. RES prodrugs are being developed to overcome these problems. We have synthesized a series of RES prodrugs and RES sulfate metabolites (RES-S) and evaluated their biological activities. RES glucosylated prodrugs (RES-Glc) were more cytotoxic in HT-29 and MCF-7 cells than RES itself whereas RES-S showed similar or higher cytotoxicity than RES. VEGF production was decreased by RES-Glc, and RES-disulfate (RES-diS) diminished it even more than RES. Finally, RES-Glc and RES-diS inhibited hTERT gene expression to a higher extent than RES. In conclusion, resveratrol prodrugs are promising candidates as anticancer drugs. In addition, RES-S showed distinct biological activity, thus indicating they are not simply RES reservoirs. PMID:27147200

  11. Antifungal activity of metabolites of the endophytic fungus Trichoderma brevicompactum from garlic

    PubMed Central

    Shentu, Xuping; Zhan, Xiaohuan; Ma, Zheng; Yu, Xiaoping; Zhang, Chuanxi

    2014-01-01

    The endophytic fungus strain 0248, isolated from garlic, was identified as Trichoderma brevicompactum based on morphological characteristics and the nucleotide sequences of ITS1-5.8S- ITS2 and tef1. The bioactive compound T2 was isolated from the culture extracts of this fungus by bioactivity-guided fractionation and identified as 4β-acetoxy-12,13- epoxy-Δ9-trichothecene (trichodermin) by spectral analysis and mass spectrometry. Trichodermin has a marked inhibitory activity on Rhizoctonia solani, with an EC50 of 0.25 μgmL−1. Strong inhibition by trichodermin was also found for Botrytis cinerea, with an EC50 of 2.02 μgmL−1. However, a relatively poor inhibitory effect was observed for trichodermin against Colletotrichum lindemuthianum (EC50 = 25.60 μgmL−1). Compared with the positive control Carbendazim, trichodermin showed a strong antifungal activity on the above phytopathogens. There is little known about endophytes from garlic. This paper studied in detail the identification of endophytic T. brevicompactum from garlic and the characterization of its active metabolite trichodermin. PMID:24948941

  12. Effect of Competition on the Production and Activity of Secondary Metabolites in Aspergillus species

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Secondary metabolites are of intense interest to humans due to their pharmaceutical and/or toxic properties. Aspergillus species secrete these metabolites by themselves and in the presence of other fungal species. Here, we have performed co-cultivation competition assays among different Aspergillu...

  13. EFFECTS OF METHOPRENE, ITS METABOLITES, AND BREAKDOWN PRODUCTS ON RETINOID-ACTIVATED PATHWAYS IN TRANSFECTED CELL LINES

    EPA Science Inventory

    Methoprene is a terpene-based insecticide designed to act as an agonist of insect juvenile hormone, which is essential for the transition from larval to adult forms in some metamorphic insects. Recent evidence suggests that a methoprene metabolite, methoprene acid, activates a ve...

  14. Increased active metabolite formation explains the greater platelet inhibition with prasugrel compared to high-dose clopidogrel.

    PubMed

    Payne, Christopher D; Li, Ying Grace; Small, David S; Ernest, C Steven; Farid, Nagy A; Jakubowski, Joseph A; Brandt, John T; Salazar, Daniel E; Winters, Kenneth J

    2007-11-01

    Prasugrel pharmacodynamics and pharmacokinetics after a 60-mg loading dose (LD) and daily 10-mg maintenance doses (MD) were compared in a 3-way crossover study to clopidogrel 600-mg/75-mg and 300-mg/75-mg LD/MD in 41 healthy, aspirin-free subjects. Each LD was followed by 7 days of daily MD and a 14-day washout period. Inhibition of platelet aggregation (IPA) was assessed by turbidometric aggregometry (20 and 5 microM ADP). Prasugrel 60-mg achieved higher mean IPA (54%) 30 minutes post-LD than clopidogrel 300-mg (3%) or 600-mg (6%) (P < 0.001) and greater IPA by 1 hour (82%) and 2 hours (91%) than the 6-hour IPA for clopidogrel 300-mg (51%) or 600-mg (69%) (P < 0.01). During MD, IPA for prasugrel 10-mg (78%) exceeded that of clopidogrel (300-mg/75-mg, 56%; 600-mg/75-mg, 52%; P < 0.001). Active metabolite area under the concentration-time curve (AUC0-tlast) after prasugrel 60-mg (594 ng.hr/mL) was 2.2 times that after clopidogrel 600-mg. Prasugrel active metabolite AUC0-tlast was consistent with dose-proportionality from 10-mg to 60-mg, while clopidogrel active metabolite AUC0-tlast exhibited saturable absorption and/or metabolism. In conclusion, greater exposure to prasugrel's active metabolite results in faster onset, higher levels, and less variability of platelet inhibition compared with high-dose clopidogrel in healthy subjects. PMID:18030066

  15. Endoxifen, the active metabolite of tamoxifen, inhibits cloned hERG potassium channels.

    PubMed

    Chae, Yun Ju; Lee, Keon Jin; Lee, Hong Joon; Sung, Ki-Wug; Choi, Jin-Sung; Lee, Eun Hui; Hahn, Sang June

    2015-04-01

    The effects of tamoxifen, and its active metabolite endoxifen (4-hydroxy-N-desmethyl-tamoxifen), on hERG currents stably expressed in HEK cells were investigated using the whole-cell patch-clamp technique and an immunoblot assay. Tamoxifen and endoxifen inhibited hERG tail currents at -50mV in a concentration-dependent manner with IC50 values of 1.2 and 1.6μM, respectively. The steady-state activation curve of the hERG currents was shifted to the hyperpolarizing direction in the presence of endoxifen. The voltage-dependent inhibition of hERG currents by endoxifen increased steeply in the voltage range of channel activation. The inhibition by endoxifen displayed a shallow voltage dependence (δ=0.18) in the full activation voltage range. A fast application of endoxifen induced a reversible block of hERG tail currents during repolarization in a concentration-dependent manner, which suggested an interaction with the open state of the channel. Endoxifen also decreased the hERG current elicited by a 5s depolarizing pulse to +60mV to inactivate the hERG currents, suggesting an interaction with the activated (open and/or inactivated) states of the channels. Tamoxifen and endoxifen inhibited the hERG channel protein trafficking to the plasma membrane in a concentration-dependent manner with endoxifen being more potent than tamoxifen. These results indicated that tamoxifen and endoxifen inhibited the hERG current by direct channel blockage and by the disruption of channel trafficking to the plasma membrane in a concentration-dependent manner. A therapeutic concentration of endoxifen inhibited the hERG current by preferentially interacting with the activated (open and/or inactivated) states of the channel. PMID:25680947

  16. Matrine cooperates with all-trans retinoic acid on differentiation induction of all-trans retinoic acid-resistant acute promyelocytic leukemia cells (NB4-LR1): possible mechanisms.

    PubMed

    Wu, Dijiong; Shao, Keding; Sun, Jie; Zhu, Fuyun; Ye, Baodong; Liu, Tingting; Shen, Yiping; Huang, He; Zhou, Yuhong

    2014-03-01

    Retinoic acid resistance results in refractory disease, and recovery in acute promyelocytic leukemia remains a challenge in clinical practice, with no ideal chemotherapeutic drug currently available. Here we report on the effect of an active compound of Sophora flavescens called matrine (0.1 mmol/L) combined with all-trans retinoic acid (1 µmol/L) in alleviating retinoic acid resistance in acute promyelocytic leukemia-derived NB4-LR1 cells by differentiation induction, as can be seen by an induced morphology change, increased CD11b expression, and nitro blue tetrazolium reduction activity, and a decreased expression of the promyelocytic leukemia-retinoic acid receptor α fusion gene and protein product. We further explored the probable mechanism of how matrine promotes the recovery of differentiation ability in NB4-LR1 cells when exposed to all-trans retinoic acid. We observed that the combination of all-trans retinoic acid and matrine can increase the level of cyclic adenosine monophosphate and protein kinase A activity, reduce telomerase activity, and downregulate the protein expression of topoisomerase II beta in NB4-LR1 cells. The results of this study suggest the possible clinical utility of matrine in the treatment of retinoic acid-resistant acute promyelocytic leukemia. PMID:24619838

  17. Arsenic Trioxide (ATO) cooperates with All Trans Retinoic Acid (ATRA) to enhance MAPK activation and differentiation in Human Myeloblastic Leukemia (HL-60) cells

    PubMed Central

    Nayak, Satyaprakash; Shen, Miaoqing; Varner, Jeffrey D.; Yen, Andrew

    2016-01-01

    Arsenic trioxide (ATO) synergistically promotes retinoic acid (RA)-induced differentiation of HL-60 myeloblastic leukemia cells, a PML-RARα negative cell line. In PML-RARα positive myeloid leukemia cells, ATO is known to cause degradation of PML-RARα with subsequent induced myeloid differentiation. We find now that ATO by itself does not cause differentiation of the PML-RARα negative HL-60 cells, but enhances RA’s capability to cause differentiation. RA-induced differentiation of HL-60 cells is known to be propelled by an induced hyperactive/persistent MAPK signal. ATO augmented RA induced RAF/MEK/ERK axis signaling and expression of CD11b, an integrin receptor that is a myeloid differentiation marker. p47PHOX, a component of the respiratory burst machinery and inducible oxidative metabolism, functional differentiation marker were also enhanced. However, ATO did not enhance RA-induced CD38 expression, an early cell surface differentiation marker. ATO enhanced RA-induced population growth retardation without evidence of apoptosis or an enhanced G1/0 growth arrest. But compared to RA, ATO plus RA showed reduced pAKT, suggesting that an overall biosynthetic/metabolic retardation was seminal to the apparent enhanced growth retardation due to ATO. In sum, our results indicate that ATO can augment action of RA in causing differentiation of myeloid leukemia cells through promoting MAPK signaling and independent of PML-RARα. PMID:20615082

  18. Activation of Notch1 inhibits medial edge epithelium apoptosis in all-trans retinoic acid-induced cleft palate in mice.

    PubMed

    Zhang, Yadong; Dong, Shiyi; Wang, Weicai; Wang, Jianning; Wang, Miao; Chen, Mu; Hou, Jinsong; Huang, Hongzhang

    2016-08-26

    Administration of all-trans retinoic acid (atRA) on E12.0 (embryonic day 12.0) leads to failure of medial edge epithelium (MEE) disappearance and cleft palate. However, the molecular mechanism underlying the relationship between atRA and MEE remains to be identified. In this study, atRA (200 mg/kg) administered by gavage induced a 75% incidence of cleft palate in C57BL/6 mice. Notch1 was up-regulated in MEE cells in the atRA-treated group compared with the controls at E15.0, together with reduced apoptosis and elevated proliferation. Next, we investigated the mechanisms underlying atRA, Notch1 and MEE degradation in palate organ culture. Our results revealed that down-regulation of Notch1 partially rescued the inhibition of atRA-induced palate fusion. Molecular analysis indicated that atRA increased the expression of Notch1 and Rbpj and decreased the expression of P21. In addition, depletion of Notch1 expression decreased the expression of Rbpj and increased the expression of P21. Moreover, inhibition of Rbpj expression partially reversed atRA-induced MEE persistence and increased P21 expression. These findings demonstrate that atRA inhibits MEE degradation, which in turn induces a cleft palate, possibly through the Notch1/RBPjk/P21 signaling pathway. PMID:27343556

  19. Novel retinoic acid receptor ligands in Xenopus embryos.

    PubMed Central

    Blumberg, B; Bolado, J; Derguini, F; Craig, A G; Moreno, T A; Chakravarti, D; Heyman, R A; Buck, J; Evans, R M

    1996-01-01

    Retinoids are a large family of natural and synthetic compounds related to vitamin A that have pleiotropic effects on body physiology, reproduction, immunity, and embryonic development. The diverse activities of retinoids are primarily mediated by two families of nuclear retinoic acid receptors, the RARs and RXRs. Retinoic acids are thought to be the only natural ligands for these receptors and are widely assumed to be the active principle of vitamin A. However, during an unbiased, bioactivity-guided fractionation of Xenopus embryos, we were unable to detect significant levels of all-trans or 9-cis retinoic acids. Instead, we found that the major bioactive retinoid in the Xenopus egg and early embryo is 4-oxoretinaldehyde, which is capable of binding to and transactivating RARs. In addition to its inherent activity, 4-oxoretinaldehyde appears to be a metabolic precursor of two other RAR ligands, 4-oxoretinoic acid and 4-oxoretinol. The remarkable increase in activity of retinaldehyde and retinol as a consequence of 4-oxo derivatization suggests that this metabolic step could serve a critical regulatory function during embryogenesis. Images Fig. 1 Fig. 4 PMID:8643496

  20. Anticancer Activities of Protopanaxadiol- and Protopanaxatriol-Type Ginsenosides and Their Metabolites

    PubMed Central

    Chen, Xiao-Jia; Zhang, Xiao-Jing; Shui, Yan-Mei; Wan, Jian-Bo

    2016-01-01

    Recently, most anticancer drugs are derived from natural resources such as marine, microbial, and botanical sources, but the low success rates of chemotherapies and the development of multidrug resistance emphasize the importance of discovering new compounds that are both safe and effective against cancer. Ginseng types, including Asian ginseng, American ginseng, and notoginseng, have been used traditionally to treat various diseases, due to their immunomodulatory, neuroprotective, antioxidative, and antitumor activities. Accumulating reports have shown that ginsenosides, the major active component of ginseng, were helpful for tumor treatment. 20(S)-Protopanaxadiol (PDS) and 20(S)-protopanaxatriol saponins (PTS) are two characteristic types of triterpenoid saponins in ginsenosides. PTS holds capacity to interfere with crucial metabolism, while PDS could affect cell cycle distribution and prodeath signaling. This review aims at providing an overview of PTS and PDS, as well as their metabolites, regarding their different anticancer effects with the proposal that these compounds might be potent additions to the current chemotherapeutic strategy against cancer. PMID:27446225

  1. Microbial communication leading to the activation of silent fungal secondary metabolite gene clusters

    PubMed Central

    Netzker, Tina; Fischer, Juliane; Weber, Jakob; Mattern, Derek J.; König, Claudia C.; Valiante, Vito; Schroeckh, Volker; Brakhage, Axel A.

    2015-01-01

    Microorganisms form diverse multispecies communities in various ecosystems. The high abundance of fungal and bacterial species in these consortia results in specific communication between the microorganisms. A key role in this communication is played by secondary metabolites (SMs), which are also called natural products. Recently, it was shown that interspecies “talk” between microorganisms represents a physiological trigger to activate silent gene clusters leading to the formation of novel SMs by the involved species. This review focuses on mixed microbial cultivation, mainly between bacteria and fungi, with a special emphasis on the induced formation of fungal SMs in co-cultures. In addition, the role of chromatin remodeling in the induction is examined, and methodical perspectives for the analysis of natural products are presented. As an example for an intermicrobial interaction elucidated at the molecular level, we discuss the specific interaction between the filamentous fungi Aspergillus nidulans and Aspergillus fumigatus with the soil bacterium Streptomyces rapamycinicus, which provides an excellent model system to enlighten molecular concepts behind regulatory mechanisms and will pave the way to a novel avenue of drug discovery through targeted activation of silent SM gene clusters through co-cultivations of microorganisms. PMID:25941517

  2. Anticancer Activities of Protopanaxadiol- and Protopanaxatriol-Type Ginsenosides and Their Metabolites.

    PubMed

    Chen, Xiao-Jia; Zhang, Xiao-Jing; Shui, Yan-Mei; Wan, Jian-Bo; Gao, Jian-Li

    2016-01-01

    Recently, most anticancer drugs are derived from natural resources such as marine, microbial, and botanical sources, but the low success rates of chemotherapies and the development of multidrug resistance emphasize the importance of discovering new compounds that are both safe and effective against cancer. Ginseng types, including Asian ginseng, American ginseng, and notoginseng, have been used traditionally to treat various diseases, due to their immunomodulatory, neuroprotective, antioxidative, and antitumor activities. Accumulating reports have shown that ginsenosides, the major active component of ginseng, were helpful for tumor treatment. 20(S)-Protopanaxadiol (PDS) and 20(S)-protopanaxatriol saponins (PTS) are two characteristic types of triterpenoid saponins in ginsenosides. PTS holds capacity to interfere with crucial metabolism, while PDS could affect cell cycle distribution and prodeath signaling. This review aims at providing an overview of PTS and PDS, as well as their metabolites, regarding their different anticancer effects with the proposal that these compounds might be potent additions to the current chemotherapeutic strategy against cancer. PMID:27446225

  3. Solid Lipid Nanoparticles Loaded with Retinoic Acid and Lauric Acid as an Alternative for Topical Treatment of Acne Vulgaris.

    PubMed

    Silva, Elton Luiz; Carneiro, Guilherme; De Araújo, Lidiane Advíncula; Trindade, Mariana de Jesus Vaz; Yoshida, Maria Irene; Oréfice, Rodrigo Lambert; Farias, Luis de Macêdo; De Carvalho, Maria Auxiliadora Roque; Dos Santos, Simone Gonçalves; Goulart, Gisele Assis Castro; Alves, Ricardo José; Ferreira, Lucas Antônio Miranda

    2015-01-01

    Topical therapy is the first choice for the treatment of mild to moderate acne and all-trans retinoic acid is one of the most used drugs. The combination of retinoids and antimicrobials is an innovative approach for acne therapy. Recently, lauric acid, a saturated fatty acid, has shown strong antimicrobial activity against Propionibacterium acnes. However, topical application of retinoic acid is followed by high incidence of side-effects, including erythema and irritation. Solid lipid nanoparticles represent an alternative to overcome these side-effects. This work aims to develop solid lipid nanoparticles loaded with retinoic acid and lauric acid and evaluate their antibacterial activity. The influence of lipophilic stearylamine on the characteristics of solid lipid nanoparticles was investigated. Solid lipid nanoparticles were characterized for size, zeta potential, encapsulation efficiency, differential scanning calorimetry and X-ray diffraction. The in vitro inhibitory activity of retinoic acid-lauric acid-loaded solid lipid nanoparticles was evaluated against Propionibacterium acnes, Staphylococcus aureus and Staphylococcus epidermidis. High encapsulation efficiency was obtained at initial time (94 ± 7% and 100 ± 4% for retinoic acid and lauric acid, respectively) and it was demonstrated that lauric acid-loaded-solid lipid nanoparticles provided the incorporation of retinoic acid. However, the presence of stearylamine is necessary to ensure stability of encapsulation. Moreover, retinoic acid-lauric acid-loaded solid lipid nanoparticles showed growth inhibitory activity against Staphylococcus epidermidis, Propionibacterium acnes and Staphylococcus aureus, representing an interesting alternative for the topical therapy of acne vulgaris. PMID:26328443

  4. 9-cis-retinoic acid in combination with retinal pigment epithelium induces apoptosis in cultured retinal explants only during early postnatal development.

    PubMed

    Söderpalm, A K; Karlsson, J; Caffé, A R; vanVeen, T

    1999-12-10

    Retinoic acid is one of the active metabolites of vitamin A and has profound effects on the development of the CNS including retina. Previously, we have shown that rod-specific apoptosis is induced in retinal explants from neonatal mice by exposure to 9-cis-retinoic acid (9CRA) when the retinal pigment epithelium (RPE) is present. In explants lacking RPE, it instead has a differentiation-promoting effect seen as an accelerated opsin expression on postnatal day 3. To investigate the long-term effect of 9CRA exposure, we have explanted retinas from neonatal C3H mice with or without RPE attached and placed in organ culture. After 19 or 48 h in culture or 7, 8 or 13 days in culture, the explants were either fixed for histochemical examination or frozen for assay of DEVDase activity. We found that long-term exposure to 9CRA caused a decrease in the number of cell layers in the outer nuclear layer (ONL) only in explants with the RPE attached. When explants with RPE attached were exposed to 9CRA only during the second postnatal week, neither an increase in DEVDase activity, TUNEL-positive cells, nor a decrease in cell layers of the ONL could be demonstrated, indicating that the retina was insensitive to the apoptosis-inducing effect of 9CRA after the first postnatal week. The absence of RPE in control explants resulted in a higher number of rosettes and the extrusion of cells into the subretinal space. PMID:10611516

  5. Interferon regulatory factor-1 binds c-Cbl, enhances mitogen activated protein kinase signaling and promotes retinoic acid-induced differentiation of HL-60 human myelo-monoblastic leukemia cells

    PubMed Central

    Shen, Miaoqing; Bunaciu, Rodica P.; Congleton, Johanna; Jensen, Holly A.; Sayam, Lavanya G.; Varner, Jeffrey D.; Yen, Andrew

    2014-01-01

    All-trans retinoic acid (RA) and interferons (IFNs) have efficacy in treating certain leukemias and lymphomas, respectively, motivating interest in their mechanism of action to improve therapy. Both RA and IFNs induce interferon regulatory factor-1 (IRF-1). We find that in HL-60 myeloblastic leukemia cells which undergo mitogen activated protien kinase (MAPK)-dependent myeloid differentiation in response to RA, IRF-1 propels differentiation. RA induces MAPK-dependent expression of IRF-1. IRF-1 binds c-Cbl, a MAPK related adaptor. Ectopic IRF-1 expression causes CD38 expression and activation of the Raf/MEK/ERK axis, and enhances RA-induced differentiation by augmenting CD38, CD11b, respiratory burst and G0 arrest. Ectopic IRF-1 expression also decreases the activity of aldehyde dehydrogenase 1, a stem cell marker, and enhances RA-induced ALDH1 down-regulation. Interestingly, expression of aryl hydrocarbon receptor (AhR), which is RA-induced and known to down-regulate Oct4 and drive RA-induced differentiation, also enhances IRF-1 expression. The data are consistent with a model whereby IRF-1 acts downstream of RA and AhR to enhance Raf/MEK/ERK activation and propel differentiation. PMID:21740303

  6. Retinoic acid induces Sertoli cell paracrine signals for spermatogonia differentiation but cell autonomously drives spermatocyte meiosis

    PubMed Central

    Raverdeau, Mathilde; Gely-Pernot, Aurore; Féret, Betty; Dennefeld, Christine; Benoit, Gérard; Davidson, Irwin; Chambon, Pierre; Mark, Manuel; Ghyselinck, Norbert B.

    2012-01-01

    Direct evidence for a role of endogenous retinoic acid (RA), the active metabolite of vitamin A in the initial differentiation and meiotic entry of spermatogonia, and thus in the initiation of spermatogenesis is still lacking. RA is synthesized by dedicated enzymes, the retinaldehyde dehydrogenases (RALDH), and binds to and activates nuclear RA receptors (RARA, RARB, and RARG) either within the RA-synthesizing cells or in the neighboring cells. In the present study, we have used a combination of somatic genetic ablations and pharmacological approaches in vivo to show that during the first, prepubertal, spermatogenic cycle (i) RALDH-dependent synthesis of RA by Sertoli cells (SC), the supporting cells of the germ cell (GC) lineage, is indispensable to initiate differentiation of A aligned into A1 spermatogonia; (ii) RARA in SC mediates the effects of RA, possibly through activating Mafb expression, a gene whose Drosophila homolog is mandatory to GC differentiation; (iii) RA synthesized by premeiotic spermatocytes cell autonomously induces meiotic initiation through controlling the RAR-dependent expression of Stra8. Furthermore, we show that RA of SC origin is no longer necessary for the subsequent spermatogenic cycles but essential to spermiation. Altogether, our data establish that the effects of RA in vivo on spermatogonia differentiation are indirect, via SC, but direct on meiotic initiation in spermatocytes, supporting thereby the notion that, contrary to the situation in the female, RA is necessary to induce meiosis in the male. PMID:23012458

  7. The role of Zic transcription factors in regulating hindbrain retinoic acid signaling

    PubMed Central

    2013-01-01

    Background The reiterated architecture of cranial motor neurons aligns with the segmented structure of the embryonic vertebrate hindbrain. Anterior-posterior identity of cranial motor neurons depends, in part, on retinoic acid signaling levels. The early vertebrate embryo maintains a balance between retinoic acid synthetic and degradative zones on the basis of reciprocal expression domains of the retinoic acid synthesis gene aldhehyde dehydrogenase 1a2 (aldh1a2) posteriorly and the oxidative gene cytochrome p450 type 26a1 (cyp26a1) in the forebrain, midbrain, and anterior hindbrain. Results This manuscript investigates the role of zinc finger of the cerebellum (zic) transcription factors in regulating levels of retinoic acid and differentiation of cranial motor neurons. Depletion of zebrafish Zic2a and Zic2b results in a strong downregulation of aldh1a2 expression and a concomitant reduction in activity of a retinoid-dependent transgene. The vagal motor neuron phenotype caused by loss of Zic2a/2b mimics a depletion of Aldh1a2 and is rescued by exogenously supplied retinoic acid. Conclusion Zic transcription factors function in patterning hindbrain motor neurons through their regulation of embryonic retinoic acid signaling. PMID:23937294

  8. Retinoic acid amide inhibits JAK/STAT pathway in lung cancer which leads to apoptosis.

    PubMed

    Li, Hong-Xing; Zhao, Wei; Shi, Yan; Li, Ya-Na; Zhang, Lian-Shuang; Zhang, Hong-Qin; Wang, Dong

    2015-11-01

    Small cell lung cancer (SCLC) accounts for 12 to 16% of lung neoplasms and has a high rate of metastasis. The present study demonstrates the antiproliferative effect of retinoic acid amide in vitro and in vivo against human lung cancer cells. The results from MTT assay showed a significant growth inhibition of six tested lung cancer cell lines and inhibition of clonogenic growth at 30 μM. Retinoic acid amide also leads to G2/M-phase cell cycle arrest and apoptosis of lung cancer cells. It caused inhibition of JAK2, STAT3, and STAT5, increased the level of p21WAF1, and decreased cyclin A, cyclin B1, and Bcl-XL expression. Retinoic acid amide exhibited a synergistic effect on antiproliferative effects of methotrexate in lung cancer cells. In lung tumor xenografts, the tumor volume was decreased by 82.4% compared to controls. The retinoic acid amide-treated tumors showed inhibition of JAK2/STAT3 activation and Bcl-XL expression. There was also increase in expression of caspase-3 and caspase-9 in tumors on treatment with retinoic acid amide. Thus, retinoic acid amide exhibits promising antiproliferative effects against human lung cancer cells in vitro and in vivo and enhances the antiproliferative effect of methotrexate. PMID:26044560

  9. Retinoic acid-primed human dendritic cells inhibit Th9 cells and induce Th1/Th17 cell differentiation.

    PubMed

    Rampal, Ritika; Awasthi, Amit; Ahuja, Vineet

    2016-07-01

    All-trans-retinoic acid plays a central role in mucosal immunity, where it promotes its synthesis by up-regulating CD103 expression on dendritic cells, induces gut tropic (α4β7(+) and CCR9(+)) T cells, and inhibits Th1/Th17 differentiation. Recently, murine studies have highlighted the proinflammatory role of retinoic acid in maintaining inflammation under a variety of pathologic conditions. However, as a result of limited human data, we investigated the effect of retinoic acid on human dendritic cells and CD4(+) T cell responses in the presence of polarizing (Th1/Th9/Th17) and inflammatory (LPS-induced dendritic cells) conditions. We report a novel role of retinoic acid in an inflammatory setup, where retinoic acid-primed dendritic cells (retinoic acid-monocyte-derived dendritic cells) up-regulated CCR9(+)T cells, which were observed to express high levels of IFN-γ in the presence of Th1/Th17 conditions. Retinoic acid-monocyte-derived dendritic cells, under Th17 conditions, also favored the induction of IL-17(+) T cells. Furthermore, in the presence of TGF-β1 and IL-4, retinoic acid-monocyte-derived dendritic cells inhibited IL-9 and induced IFN-γ expression on T cells. Experiments with naïve CD4(+) T cells, activated in the presence of Th1/Th17 conditions and absence of DCs, indicated that retinoic acid inhibited IFN-γ and IL-17 expression on T cells. These data revealed that in the face of inflammatory conditions, retinoic acid, in contrast from its anti-inflammatory role, could maintain or aggravate the intestinal inflammation. PMID:26980802

  10. Functional imaging of focal brain activation in conscious rats: impact of [(14)C]glucose metabolite spreading and release.

    PubMed

    Cruz, Nancy F; Ball, Kelly K; Dienel, Gerald A

    2007-11-15

    Labeled glucose and its analogs are widely used in imaging and metabolic studies of brain function, astrocyte-neuron interactions, and neurotransmission. Metabolite shuttling among astrocytes and neurons is essential for cell-cell transfer of neurotransmitter precursors and supply and elimination of energy metabolites, but dispersion and release of labeled compounds from activated tissue would reduce signal registration in metabolic labeling studies, causing underestimation of focal functional activation. Processes and pathways involved in metabolite trafficking and release were therefore assessed in the auditory pathway of conscious rats. Unilateral monotonic stimulation increased glucose utilization (CMR(glc)) in tonotopic bands in the activated inferior colliculus by 35-85% compared with contralateral tissue when assayed with [(14)C]deoxyglucose (DG), whereas only 20-30% increases were registered with [1- or 6-(14)C]glucose. Tonotopic bands were not evident with [1-(14)C]glucose unless assayed during halothane anesthesia or pretreatment with probenecid but were detectable with [6-(14)C]glucose. Extracellular lactate levels transiently doubled during acoustic stimulation, so metabolite spreading was assessed by microinfusion of [(14)C]tracers into the inferior colliculus. The volume of tissue labeled by [1-(14)C]glucose exceeded that by [(14)C]DG by 3.2- and 1.4-fold during rest and acoustic activation, respectively. During activation, the tissue volume labeled by U-(14)C-labeled glutamine and lactate rose, whereas that by glucose fell 50% and that by DG was unchanged. Dispersion of [1-(14)C]glucose and its metabolites during rest was also reduced 50% by preinfusion of gap junction blockers. To summarize, during brain activation focal CMR(glc) is underestimated with labeled glucose because of decarboxylation reactions, spreading within tissue and via the astrocyte syncytium, and release from activated tissue. These findings help explain the fall in CMR(O2)/CMR

  11. Colon cancer chemopreventive effects of baicalein, an active enteric microbiome metabolite from baicalin

    PubMed Central

    WANG, CHONG-ZHI; ZHANG, CHUN-FENG; CHEN, LINA; ANDERSON, SAMANTHA; LU, FANG; YUAN, CHUN-SU

    2015-01-01

    Baicalin is a major constituent of Scutellaria baicalensis, which is a commonly used herbal medicine in many Asian countries. After oral ingestion, intestinal micro-biota metabolism may change parent compound's structure and its biological activities. However, whether baicalin can be metabolized by enteric microbiota and the related anti-cancer activity is not clear. In this study, using human enteric microbiome incubation and HPLC analysis, we observed that baicalin can be quickly converted to baicalein. We compared the antiproliferative effects of baicalin and baicalein using a panel of human cancer cell lines, including three human colorectal cancer (CRC) cell lines. In vitro antiproliferative effects on CRC cells were verified using an in vivo xenograft nude mouse model. Baicalin showed limited antiproliferative effects on some of these cancer cell lines. Baicalein, however, showed significant antiproliferative effects in all the tested cancer cell lines, especially on HCT-116 human colorectal cancer cells. In vivo antitumor results supported our in vitro data. We demonstrated that baicalein exerts potent S phase cell cycle arrest and pro-apoptotic effects in HCT-116 cells. Baicalein induced the activation of caspase 3 and 9. The in silico modeling suggested that baicalein forms hydrogen bonds with residues Ser251 and Asp253 at the active site of caspase 3, while interactions with residues Leu227 and Asp228 in caspase 9 through its hydroxyl groups. Data from this study suggested that baicalein is a potent anticancer metabolite derived from S. baicalensis. Enteric microbiota play a key role in the colon cancer chemoprevention of S. baicalensis. PMID:26398706

  12. Unexpected hormonal activity of a catechol equine estrogen metabolite reveals reversible glutathione conjugation

    PubMed Central

    Peng, Kuan-Wei; Chang, Minsun; Wang, Yue-Ting; Wang, Zhican; Qin, Zhihui; Bolton, Judy L.; Thatcher, Gregory R. J.

    2010-01-01

    4-Hydroxyequilenin (4-OHEN) is a major phase I metabolite of the equine estrogens present in widely prescribed hormone replacement formulations. 4-OHEN is autoxidized to an electrophilic o-quinone that has been shown to redox cycle, generating ROS, and to covalently modify proteins and DNA and thus potentially to act as a chemical carcinogen. To establish the ability of 4-OHEN to act as a hormonal carcinogen at the estrogen receptor (ER), estrogen responsive gene expression and proliferation were studied in ER(+) breast cancer cells. Recruitment by 4-OHEN of ER to estrogen responsive elements (ERE) of DNA in MCF-7 cells was also studied and observed. 4-OHEN was a potent estrogen, with additional weak activity associated with binding to the arylhydrocarbon receptor (AhR). The potency of 4-OHEN towards classical ERα mediated activity was unexpected given the reported rapid autoxidation and trapping of the resultant quinone by GSH. Addition of thiols to cell cultures did not attenuate the estrogenic activity of 4-OHEN and pre-formed thiol conjugates added to cell incubations only marginally reduced ERE-luciferase induction. On reaction of the 4OHEN-GSH conjugate with NADPH, 4-OHEN was observed to be regenerated at a rate dependent upon NADPH concentration, indicating that intracellular non-enzymatic and enzymatic regeneration of 4-OHEN accounts for the observed estrogenic activity of 4-OHEN. 4-OHEN is therefore capable of inducing chemical and hormonal pathways that may contribute to estrogen-dependent carcinogenesis, and trapping by cellular thiols does not provide a mechanism of termination of these pathways. PMID:20540524

  13. 3D-QSAR Studies on a Series of Dihydroorotate Dehydrogenase Inhibitors: Analogues of the Active Metabolite of Leflunomide

    PubMed Central

    Li, Shun-Lai; He, Mao-Yu; Du, Hong-Guang

    2011-01-01

    The active metabolite of the novel immunosuppressive agent leflunomide has been shown to inhibit the enzyme dihydroorotate dehydrogenase (DHODH). This enzyme catalyzes the fourth step in de novo pyrimidine biosynthesis. Self-organizing molecular field analysis (SOMFA), a simple three-dimensional quantitative structure-activity relationship (3D-QSAR) method is used to study the correlation between the molecular properties and the biological activities of a series of analogues of the active metabolite. The statistical results, cross-validated rCV2 (0.664) and non cross-validated r2 (0.687), show a good predictive ability. The final SOMFA model provides a better understanding of DHODH inhibitor-enzyme interactions, and may be useful for further modification and improvement of inhibitors of this important enzyme. PMID:21686163

  14. Synthesis of new optically active propargylic fluorides and application to the enantioselective synthesis of monofluorinated analogues of fatty acid metabolites.

    PubMed

    Prakesch, M; Grée, D; Grée, R

    2001-05-01

    A new approach to obtain optically active unsaturated or polyunsaturated systems with a single fluorine atom in an allylic or propargylic position is reported. Central to this strategy is the high regio- and stereocontrol observed during the fluorination of propargylic alcohols allowing a short and efficient synthesis of 1. Further, simple functional group transformations gave the enals 2 and 3. These three key intermediates were used for the preparation of optically active monofluorinated analogues of fatty acid metabolites. PMID:11325281

  15. A fish-feeding laboratory bioassay to assess the antipredatory activity of secondary metabolites from the tissues of marine organisms.

    PubMed

    Marty, Micah J; Pawlik, Joseph R

    2015-01-01

    Marine chemical ecology is a young discipline, having emerged from the collaboration of natural products chemists and marine ecologists in the 1980s with the goal of examining the ecological functions of secondary metabolites from the tissues of marine organisms. The result has been a progression of protocols that have increasingly refined the ecological relevance of the experimental approach. Here we present the most up-to-date version of a fish-feeding laboratory bioassay that enables investigators to assess the antipredatory activity of secondary metabolites from the tissues of marine organisms. Organic metabolites of all polarities are exhaustively extracted from the tissue of the target organism and reconstituted at natural concentrations in a nutritionally appropriate food matrix. Experimental food pellets are presented to a generalist predator in laboratory feeding assays to assess the antipredatory activity of the extract. The procedure described herein uses the bluehead, Thalassoma bifasciatum, to test the palatability of Caribbean marine invertebrates; however, the design may be readily adapted to other systems. Results obtained using this laboratory assay are an important prelude to field experiments that rely on the feeding responses of a full complement of potential predators. Additionally, this bioassay can be used to direct the isolation of feeding-deterrent metabolites through bioassay-guided fractionation. This feeding bioassay has advanced our understanding of the factors that control the distribution and abundance of marine invertebrates on Caribbean coral reefs and may inform investigations in diverse fields of inquiry, including pharmacology, biotechnology, and evolutionary ecology. PMID:25650625

  16. The ex vivo antiplatelet activation potential of fruit phenolic metabolite hippuric acid.

    PubMed

    Santhakumar, Abishek Bommannan; Stanley, Roger; Singh, Indu

    2015-08-01

    Polyphenol-rich fruit and vegetable intake has been associated with reduction in platelet hyperactivity, a significant contributor to thrombus formation. This study was undertaken to investigate the possible role of hippuric acid, a predominant metabolite of plant cyclic polyols, phenolic acids and polyphenols, in reduction of platelet activation-related thrombogenesis. Fasting blood samples were collected from 13 healthy subjects to analyse the effect of varying concentrations of hippuric acid (100 μM, 200 μM, 500 μM, 1 mM and 2 mM) on activation-dependant platelet surface-marker expression. Procaspase activating compound-1 (PAC-1) and P-selectin/CD62P monoclonal antibodies were used to evaluate platelet activation-related conformational changes and α-granule release respectively using flow cytometry. Platelets were stimulated ex vivo via the P2Y1/P2Y12- adenosine diphosphate (ADP) pathway of platelet activation. Hippuric acid at a concentration of 1 mM and 2 mM significantly reduced P-selectin/CD62P expression (p = 0.03 and p < 0.001 respectively) induced by ADP. Hippuric acid at 2 mM concentration also inhibited PAC-1 activation-dependant antibody expression (p = 0.03). High ex vivo concentrations of hippuric acid can therefore significantly reduce P-selectin and PAC-1 expression thus reducing platelet activation and clotting potential. However, although up to 11 mM of hippuric acid can be excreted in the urine per day following consumption of fruit, hippuric acid is actively excreted with a recorded Cmax for hippuric acid in human plasma at 250-300 μM. This is lower than the blood concentration of 1-2 mM shown to be bioactive in this research. The contribution of hippuric acid to the protective effects of fruit and vegetable intake against vascular disorders by the pathways measured is therefore low but could be synergistic with lowered doses of antiplatelet drugs and help reduce risk of thrombosis in current antiplatelet drug sensitive populations. PMID

  17. Modulation of ultraviolet light-, ethyl methanesulfonate-, and 7,12-dimethylbenz(A)anthracene-induced unscheduled DNA synthesis by retinol and retinoic acid in the primary rat hepatocyte

    SciTech Connect

    Budroe, J.D.; Shaddock, J.G.; Casciano, D.A.

    1987-01-01

    The effects of retinol and retinoic acid on unscheduled DNA synthesis (UDS) in primary Sprague-Dawley rat hepatocytes were studied in the presence and absence of know chemical and physical mutagens. Neither retinol or retinoic acid caused a significant increase in UDS over solvent control at concentrations ranging from 1 ..mu..M to 50 ..mu..M. Retinol and retinoic acid did not significantly affect 200..mu..g/mL ethyl methanesulfonate (EMS)- or 32 J/m/sup 2/ ultraviolet light (UV)-induced UDS at concentrations ranging from 1..mu..M to 50 ..mu..M. In contrast, retinol and retinoic acid significantly inhibited 2.5 ..mu..g/mL and 5.0 ..mu..g/mL 7,12-dimethyl-benz(a)-anthracene(DMBA)-induced UDS at concentrations of 1..mu..M or greater. Retinol-and retinoic acid-induced hepatocytotoxicity was studied in vitro using lactate dehydrogenase (LDH) release as an indicator of cytoxicity. Neither retinol nor retinoic acid caused significant increases in LDH release over solvent control 3 hours after treatment, whereas retinol caused a biologically significant increase in LDH release 24 hours posttreatment at concentrations of 50 ..mu..M and 100 ..mu..M. These data suggest that nontoxic concentrations of retinol and retinoic acid do not inhibit the DNA excision repair process but apparently affect the effective DNA adduct load due to the ultimate species of DMBA metabolite responsible for hepatocellular DNA damage.

  18. Modulation of ultraviolet light-, ethyl methanesulfonate-, and 7,12-dimethylbenz(a)anthracene-induced unscheduled DNA synthesis by retinol and retinoic acid in the primary rat hepatocyte

    SciTech Connect

    Budroe, J.D.; Shaddock, J.G.; Casciano, D.A.

    1987-01-01

    The effects of retinol and retinoic acid on unscheduled DNA synthesis (UDS) in primary Sprague-Dawley rat hepatocytes were studied in the presence and absence of known chemical and physical mutagens. Neither retinol nor retinoic acid caused a significant increase in UDS over solvent control at concentrations ranging from 1 microM to 50 microM. Retinol and retinoic acid did not significantly affect 200 micrograms/mL ethyl methanesulfonate(EMS)- or 32 J/m2 ultraviolet light(UV)-induced UDS at concentrations ranging from 1 microM to 50 microM. In contrast, retinol and retinoic acid significantly inhibited 2.5 micrograms/mL and 5.0 micrograms/mL 7,12-dimethyl-benz(a)anthracene(DMBA)-induced UDS at concentrations of 1 microM or greater. Retinol- and retinoic acid-induced hepatocytotoxicity was studied in vitro using lactate dehydrogenase (LDH) release as an indicator of cytoxicity. Neither retinol nor retinoic acid caused significant increases in LDH release over solvent control 3 hours after treatment, whereas retinol caused a biologically significant increase in LDH release 24 hours posttreatment at concentrations of 50 microM and 100 microM. These data suggest that nontoxic concentrations of retinol and retinoic acid do not inhibit the DNA excision repair process but apparently affect the effective DNA adduct load due to the ultimate species of DMBA metabolite responsible for hepatocellular DNA damage.

  19. Molecular structure of antihypertensive drug perindopril, its active metabolite perindoprilat and impurity F

    NASA Astrophysics Data System (ADS)

    Remko, M.; Bojarska, J.; Ježko, P.; Maniukiewicz, W.; Olczak, A.

    2013-03-01

    The molecular structure of the antihypertensive drug perindopril (2S,3aS,7aS)-1-[(2S)-2-[[(2S)-1-ethoxy-1-oxopentan-2-yl]amino]propanoyl]-2,3,3a,4,5,6,7,7a-octahydroindole-2 carboxylic acid), its active metabolite perindoprilat ((2S,3aS,7aS)-1-[(2S)-2-[[(2S)-1-carboxybutyl]amino]propanoyl]-2,3,3a,4,5,6,7,7a-octahydroindole-2-carboxylic acid), and impurity F (ethyl (2S)-2-((3S,5aS,9aS,10aS)-3-methyl-1,4-dioxodecahydropyrazino[1,2-a]indol-2(1H)-yl) pentanoate) has been investigated using B3LYP/6-31g(d) and B3LYP/6-311+g(d,p) model chemistry. It has been found that solid state conformations of perindoprilat occur close to, but not actually at minima on the computed gas-phase potential energy surfaces. Both, neutral and zwitterionic structures of perindopril and perindoprilat have been investigated. Relative stability of individual ionized species of this drug has been determined. Water has a remarkable effect on the geometry of the perindopril species studied.

  20. Imaging of Endogenous Metabolites of Plant Leaves by Mass Spectrometry Based on Laser Activated Electron Tunneling

    PubMed Central

    Huang, Lulu; Tang, Xuemei; Zhang, Wenyang; Jiang, Ruowei; Chen, Disong; Zhang, Juan; Zhong, Hongying

    2016-01-01

    A new mass spectrometric imaging approach based on laser activated electron tunneling (LAET) was described and applied to analysis of endogenous metabolites of plant leaves. LAET is an electron-directed soft ionization technique. Compressed thin films of semiconductor nanoparticles of bismuth cobalt zinc oxide were placed on the sample plate for proof-of-principle demonstration because they can not only absorb ultraviolet laser but also have high electron mobility. Upon laser irradiation, electrons are excited from valence bands to conduction bands. With appropriate kinetic energies, photoexcited electrons can tunnel away from the barrier and eventually be captured by charge deficient atoms present in neutral molecules. Resultant unpaired electron subsequently initiates specific chemical bond cleavage and generates ions that can be detected in negative ion mode of the mass spectrometer. LAET avoids the co-crystallization process of routinely used organic matrix materials with analyzes in MALDI (matrix assisted-laser desorption ionization) analysis. Thus uneven distribution of crystals with different sizes and shapes as well as background peaks in the low mass range resulting from matrix molecules is eliminated. Advantages of LAET imaging technique include not only improved spatial resolution but also photoelectron capture dissociation which produces predictable fragment ions. PMID:27053227

  1. Imaging of Endogenous Metabolites of Plant Leaves by Mass Spectrometry Based on Laser Activated Electron Tunneling

    NASA Astrophysics Data System (ADS)

    Huang, Lulu; Tang, Xuemei; Zhang, Wenyang; Jiang, Ruowei; Chen, Disong; Zhang, Juan; Zhong, Hongying

    2016-04-01

    A new mass spectrometric imaging approach based on laser activated electron tunneling (LAET) was described and applied to analysis of endogenous metabolites of plant leaves. LAET is an electron-directed soft ionization technique. Compressed thin films of semiconductor nanoparticles of bismuth cobalt zinc oxide were placed on the sample plate for proof-of-principle demonstration because they can not only absorb ultraviolet laser but also have high electron mobility. Upon laser irradiation, electrons are excited from valence bands to conduction bands. With appropriate kinetic energies, photoexcited electrons can tunnel away from the barrier and eventually be captured by charge deficient atoms present in neutral molecules. Resultant unpaired electron subsequently initiates specific chemical bond cleavage and generates ions that can be detected in negative ion mode of the mass spectrometer. LAET avoids the co-crystallization process of routinely used organic matrix materials with analyzes in MALDI (matrix assisted-laser desorption ionization) analysis. Thus uneven distribution of crystals with different sizes and shapes as well as background peaks in the low mass range resulting from matrix molecules is eliminated. Advantages of LAET imaging technique include not only improved spatial resolution but also photoelectron capture dissociation which produces predictable fragment ions.

  2. Imaging of Endogenous Metabolites of Plant Leaves by Mass Spectrometry Based on Laser Activated Electron Tunneling.

    PubMed

    Huang, Lulu; Tang, Xuemei; Zhang, Wenyang; Jiang, Ruowei; Chen, Disong; Zhang, Juan; Zhong, Hongying

    2016-01-01

    A new mass spectrometric imaging approach based on laser activated electron tunneling (LAET) was described and applied to analysis of endogenous metabolites of plant leaves. LAET is an electron-directed soft ionization technique. Compressed thin films of semiconductor nanoparticles of bismuth cobalt zinc oxide were placed on the sample plate for proof-of-principle demonstration because they can not only absorb ultraviolet laser but also have high electron mobility. Upon laser irradiation, electrons are excited from valence bands to conduction bands. With appropriate kinetic energies, photoexcited electrons can tunnel away from the barrier and eventually be captured by charge deficient atoms present in neutral molecules. Resultant unpaired electron subsequently initiates specific chemical bond cleavage and generates ions that can be detected in negative ion mode of the mass spectrometer. LAET avoids the co-crystallization process of routinely used organic matrix materials with analyzes in MALDI (matrix assisted-laser desorption ionization) analysis. Thus uneven distribution of crystals with different sizes and shapes as well as background peaks in the low mass range resulting from matrix molecules is eliminated. Advantages of LAET imaging technique include not only improved spatial resolution but also photoelectron capture dissociation which produces predictable fragment ions. PMID:27053227

  3. Exploring the chemodiversity and biological activities of the secondary metabolites from the marine fungus Neosartorya pseudofischeri.

    PubMed

    Liang, Wan-Ling; Le, Xiu; Li, Hou-Jin; Yang, Xiang-Ling; Chen, Jun-Xiong; Xu, Jun; Liu, Huan-Liang; Wang, Lai-You; Wang, Kun-Teng; Hu, Kun-Chao; Yang, De-Po; Lan, Wen-Jian

    2014-11-01

    The production of fungal metabolites can be remarkably influenced by various cultivation parameters. To explore the biosynthetic potentials of the marine fungus, Neosartorya pseudofischeri, which was isolated from the inner tissue of starfish Acanthaster planci, glycerol-peptone-yeast extract (GlyPY) and glucose-peptone-yeast extract (GluPY) media were used to culture this fungus. When cultured in GlyPY medium, this fungus produced two novel diketopiperazines, neosartins A and B (1 and 2), together with six biogenetically-related known diketopiperazines,1,2,3,4-tetrahydro-2, 3-dimethyl-1,4-dioxopyrazino[1,2-a]indole (3), 1,2,3,4-tetrahydro-2-methyl-3-methylen e-1,4-dioxopyrazino[1,2-a]indole (4), 1,2,3,4-tetrahydro-2-methyl-1,3,4-trioxopyrazino[1,2-a] indole (5), 6-acetylbis(methylthio)gliotoxin (10), bisdethiobis(methylthio)gliotoxin (11), didehydrobisdethiobis(methylthio)gliotoxin (12) and N-methyl-1H-indole-2-carboxamide (6). However, a novel tetracyclic-fused alkaloid, neosartin C (14), a meroterpenoid, pyripyropene A (15), gliotoxin (7) and five known gliotoxin analogues, acetylgliotoxin (8), reduced gliotoxin (9), 6-acetylbis(methylthio)gliotoxin (10), bisdethiobis(methylthio) gliotoxin (11) and bis-N-norgliovictin (13), were obtained when grown in glucose-containing medium (GluPY medium). This is the first report of compounds 3, 4, 6, 9, 10 and 12 as naturally occurring. Their structures were determined mainly by MS, 1D and 2D NMR data. The possible biosynthetic pathways of gliotoxin-related analogues and neosartin C were proposed. The antibacterial activity of compounds 2-14 and the cytotoxic activity of compounds 4, 5 and 7-13 were evaluated. Their structure-activity relationships are also preliminarily discussed. PMID:25421322

  4. Exploring the Chemodiversity and Biological Activities of the Secondary Metabolites from the Marine Fungus Neosartorya pseudofischeri

    PubMed Central

    Liang, Wan-Ling; Le, Xiu; Li, Hou-Jin; Yang, Xiang-Ling; Chen, Jun-Xiong; Xu, Jun; Liu, Huan-Liang; Wang, Lai-You; Wang, Kun-Teng; Hu, Kun-Chao; Yang, De-Po; Lan, Wen-Jian

    2014-01-01

    The production of fungal metabolites can be remarkably influenced by various cultivation parameters. To explore the biosynthetic potentials of the marine fungus, Neosartorya pseudofischeri, which was isolated from the inner tissue of starfish Acanthaster planci, glycerol-peptone-yeast extract (GlyPY) and glucose-peptone-yeast extract (GluPY) media were used to culture this fungus. When cultured in GlyPY medium, this fungus produced two novel diketopiperazines, neosartins A and B (1 and 2), together with six biogenetically-related known diketopiperazines,1,2,3,4-tetrahydro-2,3-dimethyl-1,4-dioxopyrazino[1,2-a]indole (3), 1,2,3,4-tetrahydro-2-methyl-3-methylene-1,4-dioxopyrazino[1,2-a]indole (4), 1,2,3,4-tetrahydro-2-methyl-1,3,4-trioxopyrazino[1,2-a] indole (5), 6-acetylbis(methylthio)gliotoxin (10), bisdethiobis(methylthio)gliotoxin (11), didehydrobisdethiobis(methylthio)gliotoxin (12) and N-methyl-1H-indole-2-carboxamide (6). However, a novel tetracyclic-fused alkaloid, neosartin C (14), a meroterpenoid, pyripyropene A (15), gliotoxin (7) and five known gliotoxin analogues, acetylgliotoxin (8), reduced gliotoxin (9), 6-acetylbis(methylthio)gliotoxin (10), bisdethiobis(methylthio) gliotoxin (11) and bis-N-norgliovictin (13), were obtained when grown in glucose-containing medium (GluPY medium). This is the first report of compounds 3, 4, 6, 9, 10 and 12 as naturally occurring. Their structures were determined mainly by MS, 1D and 2D NMR data. The possible biosynthetic pathways of gliotoxin-related analogues and neosartin C were proposed. The antibacterial activity of compounds 2–14 and the cytotoxic activity of compounds 4, 5 and 7–13 were evaluated. Their structure-activity relationships are also preliminarily discussed. PMID:25421322

  5. Aspirin's Active Metabolite Salicylic Acid Targets High Mobility Group Box 1 to Modulate Inflammatory Responses.

    PubMed

    Choi, Hyong Woo; Tian, Miaoying; Song, Fei; Venereau, Emilie; Preti, Alessandro; Park, Sang-Wook; Hamilton, Keith; Swapna, G V T; Manohar, Murli; Moreau, Magali; Agresti, Alessandra; Gorzanelli, Andrea; De Marchis, Francesco; Wang, Huang; Antonyak, Marc; Micikas, Robert J; Gentile, Daniel R; Cerione, Richard A; Schroeder, Frank C; Montelione, Gaetano T; Bianchi, Marco E; Klessig, Daniel F

    2015-01-01

    Salicylic acid (SA) and its derivatives have been used for millennia to reduce pain, fever and inflammation. In addition, prophylactic use of acetylsalicylic acid, commonly known as aspirin, reduces the risk of heart attack, stroke and certain cancers. Because aspirin is rapidly de-acetylated by esterases in human plasma, much of aspirin's bioactivity can be attributed to its primary metabolite, SA. Here we demonstrate that human high mobility group box 1 (HMGB1) is a novel SA-binding protein. SA-binding sites on HMGB1 were identified in the HMG-box domains by nuclear magnetic resonance (NMR) spectroscopic studies and confirmed by mutational analysis. Extracellular HMGB1 is a damage-associated molecular pattern molecule (DAMP), with multiple redox states. SA suppresses both the chemoattractant activity of fully reduced HMGB1 and the increased expression of proinflammatory cytokine genes and cyclooxygenase 2 (COX-2) induced by disulfide HMGB1. Natural and synthetic SA derivatives with greater potency for inhibition of HMGB1 were identified, providing proof-of-concept that new molecules with high efficacy against sterile inflammation are attainable. An HMGB1 protein mutated in one of the SA-binding sites identified by NMR chemical shift perturbation studies retained chemoattractant activity, but lost binding of and inhibition by SA and its derivatives, thereby firmly establishing that SA binding to HMGB1 directly suppresses its proinflammatory activities. Identification of HMGB1 as a pharmacological target of SA/aspirin provides new insights into the mechanisms of action of one of the world's longest and most used natural and synthetic drugs. It may also provide an explanation for the protective effects of low-dose aspirin usage. PMID:26101955

  6. Cytotoxicity and characterization of an active metabolite of benzamide riboside, a novel inhibitor of IMP dehydrogenase.

    PubMed

    Gharehbaghi, K; Paull, K D; Kelley, J A; Barchi, J J; Marquez, V E; Cooney, D A; Monks, A; Scudiero, D; Krohn, K; Jayaram, H N

    1994-03-15

    Benzamide riboside exhibits significant cytotoxicity against a variety of human tumor cells in culture. On the basis of metabolic studies, the primary target of this drug's action appears to be IMP dehydrogenase (IMPDH). Incubation of human myelogenous leukemia K562 cells with an IC50 concentration of benzamide riboside resulted in an expansion of IMP pools (5.9-fold), with a parallel reduction in the concentration of GMP (90%), GDP (63%), GTP (55%) and dGTP (40%). On kinetic grounds, it was deduced that benzamide riboside (whose Ki versus IMPDH is 6.4 mM, while that of its 5'-monophosphate is 3.9 mM) or its 5'-monophosphate were unlikely to be responsible for inhibition of this target enzyme, IMPDH, since only micromolar concentrations of benzamide riboside were needed to exert potent inhibition of tumor-cell growth. Studies on the metabolism of this C-nucleoside have revealed the presence of a new peak eluting in the nucleoside diphosphate area on HPLC. Treatment of this peak with venom phosphodiesterase degraded it and concurrently nullified its inhibitory activity versus IMPDH; alkaline phosphatase, on the other hand, totally failed to digest the anabolite. These results suggest that the metabolite in question is the phosphodiester, benzamide adenine dinucleotide (BAD). Evidence that the inhibitor was an analog of NAD, wherein the nicotinamide moiety has been replaced by benzamide, was provided by both NMR and mass spectrometric analysis and confirmed by enzymatic synthesis. Further insight into the nature of the active principle was obtained from kinetic studies, which established that BAD competitively inhibited NAD utilization by partially purified IMPDH from K562 cells with a Ki of 0.118 microM. In concert, these studies establish that benzamide riboside exhibits potent antiproliferative activity by inhibiting IMPDH through BAD. PMID:7907081

  7. Electrochemiluminescent Arrays for Cytochrome P450-Activated Genotoxicity Screening. DNA Damage from Benzo[a]pyrene Metabolites

    PubMed Central

    Hvastkovs, Eli G.; So, Minjeong; Krishnan, Sadagopan; Bajrami, Besnik; Tarun, Maricar; Jansson, Ingela; Schenkman, John B.; Rusling, James F.

    2007-01-01

    Arrays suitable for genotoxicity screening are reported that generate metabolites from cytochrome P450 enzymes (CYPs) in thin-film spots. Array spots containing DNA, various human cyt P450s, and electrochemiluminescence (ECL) generating metallopolymer [Ru(bpy)2PVP10]2+ were exposed to H2O2 to activate the enzymes. ECL from all spots was visualized simultaneously using a CCD camera. Using benzo[a]pyrene as a test substrate, enzyme activity for producing DNA damage in the arrays was found in the order CYP1B1 > CYP1A2 > CYP1A1 > CYP2E1 > myoglobin, the same as the order of their metabolic activity. Thus, these arrays estimate the relative propensity of different enzymes to produce genotoxic metabolites. This is the first demonstration of ECL arrays for high-throughput in vitro genotoxicity screening. PMID:17261025

  8. Effects of 3-O-methyldopa, L-3,4-dihydroxyphenylalanine metabolite, on locomotor activity and dopamine turnover in rats.

    PubMed

    Onzawa, Yoritaka; Kimura, Yasuhiro; Uzuhashi, Kengo; Shirasuna, Megumi; Hirosawa, Tasuku; Taogoshi, Takanori; Kihira, Kenji

    2012-01-01

    It has been well known that 3-O-methyldopa (3-OMD) is a metabolite of L-3,4-dihydroxyphenylalanine (L-DOPA) formed by catechol O-methyltransferase (COMT), and 3-OMD blood level often reaches higher than physiological level in Parkinson's disease (PD) patients receiving long term L-DOPA therapy. However, the physiological role of 3-OMD has not been well understood. Therefore, in order to clarify the effects of 3-OMD on physiological function, we examined the behavioral alteration in rats based on locomotor activity, and measured dopamine (DA) and its metabolites levels in rats at the same time after 3-OMD subchronic administration. The study results showed that repeated administrations of 3-OMD increased its blood and the striatum tissue levels in those rats, and decreased locomotor activity in a dose dependent manner. Although 3-OMD subchronic administration showed no significant change in DA level in the striatum, DA metabolite levels, such as 3,4-dihydroxyphenylacetic acid (DOPAC), 3-methoxytyramine (3-MT), and homovanillic acid (HVA) were significantly decreased. After 3-OMD washout period (7 d), locomotor activity and DA turnover in those rats returned to normal levels. Furthermore, locomotor activity and DA turnover decreased by 3-OMD administration were recovered to normal level by acute L-DOPA administration. These results suggested that 3-OMD affect to locomotor activity via DA neuron system. In conclusion, 3-OMD itself may have a disadvantage in PD patients receiving L-DOPA therapy. PMID:22863920

  9. [Secondary metabolites, lethality and antimicrobial activity of extracts from three corals and three marine mollusks from Sucre, Venezuela].

    PubMed

    Ordaz, Gabriel; D'Armas, Haydelba; Yáñez, Dayanis; Hernández, Juan; Camacho, Angel

    2010-06-01

    The study of biochemical activity of extracts obtained from marine organisms is gaining interest as some have proved to have efficient health or industrial applications. To evaluate lethality and antimicrobial activities, some chemical tests were performed on crude extracts of the octocorals Eunicea sp., Muricea sp. and Pseudopterogorgia acerosa and the mollusks Pteria colymbus, Phyllonotus pomum and Chicoreus brevifrons, collected in Venezuelan waters. The presence of secondary metabolites like alkaloids, unsaturated sterols and pentacyclic triterpenes in all invertebrates, was evidenced. Additionally, sesquiterpenlactones, saponins, tannins, cyanogenic and cardiotonic glycosides were also detected in some octocoral extracts, suggesting that biosynthesis of these metabolites is typical in this group. From the lethality bioassays, all extracts resulted lethal to Artemia salina (LC50<1000 microg/ml) with an increased of lethal activity with exposition time. P. pomum extract showed the highest lethality rate (LC50=46.8 microg/ml). Compared to the octocorals, mollusks extracts displayed more activity and a greater action spectrum against different bacterial strains, whereas octocorals also inhibited some fungi strains growth. Staphylococcus aureus was the most susceptible to the antimicrobial power of the extracts (66.7%), whereas Pseudomonas aeruginosa, Candida albicans and Aspergillus niger were not affected. The antibiosis shown by marine organisms extracts indicates that some of their biosynthesized metabolites are physiologically active, and may have possible cytotoxic potential or as a source of antibiotic components. PMID:20527468

  10. Direction of estradiol metabolism as a control of its hormonal action--uterotrophic activity of estradiol metabolites.

    PubMed

    Martucci, C; Fishman, J

    1977-12-01

    The uterotrophic activities of the catechol metabolites of estradiol 2-hydroxyestrone, 2-methoxyestrone and 2-hydroxyestradiol were measured under conditions of continuous administration of sc implanted paraffin pellets. The activity of these estrogens was compared to that of estradiol-17beta and its other principal metabolites estrone, estriol and 15alpha-hydroxyestriol (estetrol). The major catechol estrogens, 2-hydroxyestrone and 2-methoxyestrone, and the pregnancy metabolite, 15alpha-hydroxyestriol, exhibited no uterotrophic activity. The minor catecholestrogen, 2-hydroxyestradiol, showed some activity whose character was different from that exhibited by implants of estradiol, estrone and estriol all of which were equipotent uterotrophic agents. Implants of 2-hydroxyestrone in the presence of estradiol or estriol pellets did not diminish the response to the latter indicating that the 2-hydroxyestrone is not antiestrogenic under these conditions. It is concluded that the direction of estradiol metabolism can have a profound influence on the expression of peripheral hormonal activity with hydroxylation at C-2 terminating and hydroxylation at C-16 extending it. PMID:590186

  11. The HER2 inhibitor TAK165 Sensitizes Human Acute Myeloid Leukemia Cells to Retinoic Acid-Induced Myeloid Differentiation by activating MEK/ERK mediated RARα/STAT1 axis.

    PubMed

    Shao, Xuejing; Liu, Yujia; Li, Yangling; Xian, Miao; Zhou, Qian; Yang, Bo; Ying, Meidan; He, Qiaojun

    2016-01-01

    The success of all-trans retinoic acid (ATRA) in differentiation therapy for patients with acute promyelocytic leukemia (APL) highly encourages researches to apply this therapy to other types of acute myeloid leukemia (AML). However, AML, with the exception of APL, fails to respond to differentiation therapy. Therefore, research strategies to further sensitize cells to retinoids and to extend the range of AMLs that respond to retinoids beyond APLs are urgently needed. In this study, we showed that TAK165, a HER2 inhibitor, exhibited a strong synergy with ATRA to promote AML cell differentiation. We observed that TAK165 sensitized the AML cells to ATRA-induced cell growth inhibition, G0/G1 phase arrest, CD11b expression, mature morphologic changes, NBT reduction and myeloid regulator expression. Unexpectedly, HER2 pathway might not be essential for TAK165-enhanced differentiation when combined with ATRA, while the enhanced differentiation was dependent on the activation of the RARα/STAT1 axis. Furthermore, the MEK/ERK cascade regulated the activation of STAT1. Taken together, our study is the first to evaluate the synergy of TAK165 and ATRA in AML cell differentiation and to assess new opportunities for the combination of TAK165 and ATRA as a promising approach for future differentiation therapy. PMID:27074819

  12. The HER2 inhibitor TAK165 Sensitizes Human Acute Myeloid Leukemia Cells to Retinoic Acid-Induced Myeloid Differentiation by activating MEK/ERK mediated RARα/STAT1 axis

    PubMed Central

    Shao, Xuejing; Liu, Yujia; Li, Yangling; Xian, Miao; Zhou, Qian; Yang, Bo; Ying, Meidan; He, Qiaojun

    2016-01-01

    The success of all-trans retinoic acid (ATRA) in differentiation therapy for patients with acute promyelocytic leukemia (APL) highly encourages researches to apply this therapy to other types of acute myeloid leukemia (AML). However, AML, with the exception of APL, fails to respond to differentiation therapy. Therefore, research strategies to further sensitize cells to retinoids and to extend the range of AMLs that respond to retinoids beyond APLs are urgently needed. In this study, we showed that TAK165, a HER2 inhibitor, exhibited a strong synergy with ATRA to promote AML cell differentiation. We observed that TAK165 sensitized the AML cells to ATRA-induced cell growth inhibition, G0/G1 phase arrest, CD11b expression, mature morphologic changes, NBT reduction and myeloid regulator expression. Unexpectedly, HER2 pathway might not be essential for TAK165-enhanced differentiation when combined with ATRA, while the enhanced differentiation was dependent on the activation of the RARα/STAT1 axis. Furthermore, the MEK/ERK cascade regulated the activation of STAT1. Taken together, our study is the first to evaluate the synergy of TAK165 and ATRA in AML cell differentiation and to assess new opportunities for the combination of TAK165 and ATRA as a promising approach for future differentiation therapy. PMID:27074819

  13. CSF biomarkers of monocyte activation and chemotaxis correlate with magnetic resonance spectroscopy metabolites during chronic HIV disease.

    PubMed

    Anderson, Albert M; Fennema-Notestine, Christine; Umlauf, Anya; Taylor, Michael J; Clifford, David B; Marra, Christina M; Collier, Ann C; Gelman, Benjamin B; McArthur, Justin C; McCutchan, J Allen; Simpson, David M; Morgello, Susan; Grant, Igor; Letendre, Scott L

    2015-10-01

    Human immunodeficiency virus (HIV)-associated neurocognitive disorders (HAND) persist despite combination antiretroviral therapy (cART), supporting the need to better understand HIV neuropathogenesis. Magnetic resonance spectroscopy (MRS) of the brain has demonstrated abnormalities in HIV-infected individuals despite cART. We examined the associations between MRS metabolites and selected cerebrospinal fluid (CSF) biomarkers reflecting monocyte/macrophage activation and chemotaxis. A multicenter cross-sectional study involving five sites in the USA was conducted. The following CSF biomarkers were measured: soluble CD14 (sCD14), monocyte chemotactic protein-1 (MCP-1), interferon inducible protein 10 (IP-10), and stromal cell-derived growth factor 1 alpha (SDF-1α). The following MRS metabolites were measured from basal ganglia (BG), frontal white matter (FWM), and frontal gray matter (FGM): N-acetylaspartate (NAA), myo-inositol (MI), choline (Cho), and creatine (Cr). CSF biomarkers were compared to absolute MRS metabolites as well as metabolite/Cr ratios using linear regression. Eighty-three HIV-infected individuals were included, 78 % on cART and 37 % with HAND. The most robust positive correlations were between MCP-1 and Cho in BG (R (2) 0.179, p < 0.001) as well as MCP-1 and MI in FWM (R (2) 0.137, p = 0.002). Higher Cr levels in FWM were associated with MCP-1 (R (2) 0. 075, p = 0.01) and IP-10 (R (2) 0.106, p = 0.003). Comparing biomarkers to MRS metabolite/Cr ratios impacted some relationships, e.g., higher sCD14 levels were associated with lower Cho/Cr ratios in FGM (R (2) 0.224, p < 0.001), although higher MCP-1 levels remained associated with Cho/Cr in BG. These findings provide evidence that monocyte activation and chemotaxis continue to contribute to HIV-associated brain abnormalities in cART-treated individuals. PMID:26069183

  14. Activation of Dormant Secondary Metabolite Production by Introducing Neomycin Resistance into the Deep-Sea Fungus, Aspergillus versicolor ZBY-3

    PubMed Central

    Dong, Yuan; Cui, Cheng-Bin; Li, Chang-Wei; Hua, Wei; Wu, Chang-Jing; Zhu, Tian-Jiao; Gu, Qian-Qun

    2014-01-01

    A new ultrasound-mediated approach has been developed to introduce neomycin-resistance to activate silent pathways for secondary metabolite production in a bio-inactive, deep-sea fungus, Aspergillus versicolor ZBY-3. Upon treatment of the ZBY-3 spores with a high concentration of neomycin by proper ultrasound irradiation, a total of 30 mutants were obtained by single colony isolation. The acquired resistance of the mutants to neomycin was confirmed by a resistance test. In contrast to the ZBY-3 strain, the EtOAc extracts of 22 of the 30 mutants inhibited the human cancer K562 cells, indicating that these mutants acquired a capability to produce antitumor metabolites. HPLC-photodiode array detector (PDAD)-UV and HPLC-electron spray ionization (ESI)-MS analyses of the EtOAc extracts of seven bioactive mutants and the ZBY-3 strain indicated that diverse secondary metabolites have been newly produced in the mutant extracts in contrast to the ZBY-3 extract. The followed isolation and characterization demonstrated that six metabolites, cyclo(d-Pro-d-Phe) (1), cyclo(d-Tyr-d-Pro) (2), phenethyl 5-oxo-l-prolinate (3), cyclo(l-Ile-l-Pro) (4), cyclo(l-Leu-l-Pro) (5) and 3β,5α,9α-trihydroxy-(22E,24R)-ergosta-7,22-dien-6-one (6), were newly produced by the mutant u2n2h3-3 compared to the parent ZBY-3 strain. Compound 3 was a new compound; 2 was isolated from a natural source for the first time, and all of these compounds were also not yet found in the metabolites of other A. versicolor strains. Compounds 1–6 inhibited the K562 cells, with inhibition rates of 54.6% (1), 72.9% (2), 23.5% (3), 29.6% (4), 30.9% (5) and 51.1% (6) at 100 μg/mL, and inhibited also other human cancer HL-60, BGC-823 and HeLa cells, to some extent. The present study demonstrated the effectiveness of the ultrasound-mediated approach to activate silent metabolite production in fungi by introducing acquired resistance to aminoglycosides and its potential for discovering new compounds from silent

  15. Relationship between binding affinities to cellular retinoic acid-binding protein and biological potency of a new series of retinoids.

    PubMed

    Sani, B P; Dawson, M I; Hobbs, P D; Chan, R L; Schiff, L J

    1984-01-01

    Binding affinities of a new and unusual series of retinoic acid analogues to cellular retinoic acid-binding protein, a possible mediator of their biological function in the control of differentiation and tumorigenesis, and to serum albumin, their plasma transport protein, were determined. Also, biological activity of these retinoids in the reversal of keratinization in hamster tracheal organ cultures was assessed and compared with their binding affinities. Analogues that possessed high biological activity showed high binding efficiency to cellular retinoic acid-binding protein. Those that were biologically less active were poor binders to the binding protein. Three retinoids, 4657-57, 3920-59, and 4445-75, which showed 90 to 100% binding efficiency of that of retinoic acid for cellular retinoic acid-binding protein expressed high biological activity detectable in the range of 10(-10) M as against 10(-11) M for retinoic acid. The correlation noticed in these two activities not only enhances the confidence in the two assay procedures but also paves the way for design and development of potential chemopreventive agents. No apparent differences were observed in the binding affinities of the retinoids to binding proteins of a normal tissue or a tumor tissue. No correlation existed between the binding affinities of these retinoids to serum albumin and their biological activity. Structure-activity relationships of the retinoids in relation to their binding affinities and biological activities have been discussed. PMID:6317169

  16. Constituents within pulp mill effluent deplete retinoid stores in white sucker and bind to rainbow trout retinoic acid receptors and retinoid X receptors.

    PubMed

    Alsop, Derek; Hewitt, Mark; Kohli, Mohan; Brown, Scott; Van Der Kraak, Glen

    2003-12-01

    Wild female and male white sucker (Catostomus commersoni) inhabiting an area receiving pulp mill effluent had reduced hepatic levels of retinol, didehydroretinol, retinyl esters, and didehydroretinyl esters, while vitamin E levels were unaffected. This disruption of the retinoid system led us to test methanol and dichloromethane extracts from the effluent of 11 pulp mills from across Canada for their ability to bind to rainbow trout (Oncorhynchus mykiss) retinoic acid receptors (RARs) from the gill and retinoid X receptors (RXRs) from the liver. Concentrated extracts of the final effluent from 6 of the 11 pulp mills were able to displace greater than 25% of the receptor-bound [3H]all-trans retinoic acid (RA) or [3H]9-cis RA from trout RARs and RXRs, respectively. The ability of the extracts to displace retinoic acid did not appear to be linked to the pulping or treatment processes. Moreover, extracts with the greatest activity came from thermomechanical mills, suggesting the compounds may originate from the wood furnish. In addition, extracts prepared from wood furnish (wood chips: white spruce [50%], lodgepole pine [47%], and balsam fir [3%]) from one mill were able to displace [3H]RA from the RARs and RXRs. The 4-hydroxy RA, a metabolite of RA that has been shown to be generated in greater quantities in fish exposed to P450-inducing xenobiotics, was able to displace [3H]all-trans RA from trout RARs as effectively as unlabeled all-trans RA. These results suggest that pulp mill effluent may impact the retinoid system of fish at multiple sites, either by decreasing hepatic retinoid stores or through contributing additional ligands (from the wood furnish) that can bind to RA receptors. PMID:14713038

  17. Pre-systemic elimination of tilidine: localization and consequences for the formation of the active metabolite nortilidine.

    PubMed

    Eichbaum, Christine; Mathes, Kristin; Burhenne, Jürgen; Markert, Christoph; Blank, Antje; Mikus, Gerd

    2015-02-01

    The therapeutic activity of tilidine, an opioid analgesic, is mainly related to its active metabolite nortilidine. Nortilidine formation mainly occurs during the high intestinal first-pass metabolism of tilidine by N-demethylation. Elimination of the active nortilidine to the inactive bisnortilidine is also mediated by N-demethylation and is supposed to take place in the liver, probably at a smaller rate. The aim of this study was the investigation of the pre-systemic elimination of tilidine using grapefruit juice (GFJ) as an intestinal CYP3A4 inhibitor and efavirenz (EFV) as a CYP3A4 activator. A randomized, open, placebo-controlled, cross-over study was conducted in 12 healthy volunteers using 100 mg tilidine solution p.o., regular strength GFJ 250 mL (3 times at 12-hr intervals) and EFV 400 mg (12 hr before tilidine administration). Tilidine, nortilidine and bisnortilidine in plasma and urine were quantified by a validated LC/MS/MS analysis. GFJ did not change any pharmacokinetic parameter of tilidine and its metabolites, which suggests that intestinal CYP3A4 does not contribute to the first-pass metabolism of tilidine. No effect of EFV on the pharmacokinetics of the active nortilidine was observed except a significant reduction of the terminal elimination half-life by 15%. Overall elimination (renal and metabolic clearances) was unaffected by every treatment. CYP3A4 does not seem to play a major role in tilidine first-pass and overall metabolism. Other unknown metabolites and their enzymes responsible for their formation have to be investigated as they account for the majority of renally excreted metabolites. PMID:25223231

  18. Not flavone-8-acetic acid (FAA) but its murine metabolite 6-OH-FAA exhibits remarkable antivascular activities in vitro.

    PubMed

    Pham, Minh Hien; Dauzonne, Daniel; Chabot, Guy G

    2016-06-01

    Flavone-8-acetic acid (FAA) has been proved to be a potent vascular-disrupting agent in mice. Unfortunately, FAA did not produce any anticancer activity in clinical trials. Previously, we had reported that FAA is metabolized by mouse microsomes into six metabolites, whereas it was poorly metabolized by human microsomes, with fewer metabolites formed in lesser amounts. Especially, 6-OH-FAA was not formed by human microsomes. In this work, two major available metabolites, 4'-OH-FAA and 6-OH-FAA, were tested and compared with the parent compound FAA for their potential antivascular activities in vitro. The ability of the products to induce morphological changes, disrupt preformed capillaries of EA.hy926 endothelial cells and inhibit tubulin polymerization in vitro was assessed. The action mechanism was determined using the RhoA and Rac1 inhibitors. At 25 µg/ml, 6-OH-FAA induced morphological changes and membrane blebbing, whereas 300 µg/ml of FAA and 4'-OH-FAA slightly changed the morphology without inducing membrane blebbing. At 300 µg/ml, 6-OH-FAA produced morphological changes that were 2.1-6.9-fold greater than that produced by FAA and 4'-OH-FAA, an effect that was consistent with its much greater inhibitory effect on tubulin polymerization compared with FAA and 4'-OH-FAA. 6-OH-FAA significantly disrupted the EA.hy926 cell capillaries. 6-OH-FAA activities were prevented in EA.hy926 cells pretreated with RhoA, but not Rac1, inhibitor. In this short communication we report for the first time that, in vitro, 6-OH-FAA, a mouse-specific FAA metabolite, exhibits significantly stronger antivascular activities compared with FAA and 4'-OH-FAA, which are mediated through the RhoA kinase pathway. PMID:26901071

  19. Population pharmacokinetic modeling of oxcarbazepine active metabolite in Chinese patients with epilepsy.

    PubMed

    Yu, Yunli; Zhang, Quanying; Xu, Wenjun; Lv, Chengzhe; Hao, Gang

    2016-08-01

    The aim of the study was to develop a population pharmacokinetic (PPK) model of oxcarbazepine and optimize the treatment of oxcarbazepine in Chinese patients with epilepsy. A total of 108 oxcarbazepine therapeutic drug monitoring samples from 78 patients with epilepsy were collected in this study. The pharmacologically active metabolite 10,11-dihydro-10-hydrocarbamazepine (MHD) was used as the analytical target for monitoring therapy of oxcarbazepine. Patients' clinical data were retrospectively collected. The PPK model for MHD was developed using Phoenix NLME 1.2 with a non-linear mixed-effect model. MHD pharmacokinetics obeys a one-compartment model with first-order absorption and elimination. The effect of age, gender, red blood cell count, red blood cell specific volume, hemoglobin (HGB), alanine aminotransferase (ALT), aspartate aminotransferase (AST), blood urea nitrogen (BUN), and serum creatine were analyzed. Bootstrap and data splitting were used simultaneously to validate the final PPK models. The mean values of volume of distribution and clearance of MHD in the patients were 14.2 L and 2.38 L h(-1), respectively. BUN and HGB influenced the MHD volume of distribution according to the following equation: V = tvV × (BUN/4.76)(-0.007) × (HGB/140)(-0.001) × e (ηV) . The MHD clearance was dependent on ALT and gender as follows: CL = tvCL × (ALT/30)(0.181) × (gender) × 1.083 × e (ηCL). The final PPK model was demonstrated to be suitable and effective and it can be used to evaluate the pharmacokinetic parameters of MHD in Chinese patients with epilepsy and to choose an optimal dosage regimen of oxcarbazepine on the basis of these parameters. PMID:25700977

  20. Antinociceptive activity of extracts and secondary metabolites from wild growing and micropropagated plants of Renealmia alpinia

    PubMed Central

    Gómez-Betancur, Isabel; Cortés, Natalie; Benjumea, Dora; Osorio, Edison; León, Francisco; Cutler, Stephen J.

    2015-01-01

    Ethnopharmacological relevance Renealmia alpinia is native to the American continent and can be found from Mexico to Brazil, and in the Caribbean islands. It is known as “matandrea” in Colombia, and it has been commonly used in traditional medicine to treat painful diseases and ailments. Based on its traditional uses, it is of interest to evaluate the pharmacologic effects of this plant and its secondary metabolites. Materials and methods Methanol and aqueous extracts of wild and micropropagated R. alpinia (leaves) were obtained and chemically compared by High Performance Thin Layer Chromatography (HPTLC). The antinociceptive activity of these extracts was examined using an in vivo assay (Siegmund test). Additionally, the dichloromethane extract of R. alpinia was fractionated and pure compounds were isolated by chromatographic methods. The structure elucidation of isolated compounds was performed by NMR experiments and spectroscopic techniques and comparison with the literature data. Purified compounds were evaluated for their in vitro binding affinity for opioids and cannabinoids receptors. Results The dichloromethane extract of the plant’s aerial part afforded sinostrobin (1), naringenin 7,4′-dimethyl ether (2), 2′,6′-dihydroxy-4′-methoxychalcone (3), 4-methoxy-6-(2-phenylethenyl)-2H-pyran-2-one (4), naringenin 7-methyl ether (5) and 3,5-heptanediol, 1,7-diphenyl (6), which were isolated using chromatographic methods. Their chemical structures were established by physical and spectroscopic techniques. The antinociceptive effects observed in mice by extracts of wild and micropropagated plants were similar. The compounds isolated from R. alpinia do not show affinity to opioid or cannabinoid receptors. Conclusion Aqueous and methanol extracts of R. alpinia provide antinociceptive and analgesic effects in an in vivo model. These results contribute additional insight as to why this plant is traditionally used for pain management. Also, this is the first

  1. Tissue accumulation kinetics of ciclesonide-active metabolite and budesonide in mice.

    PubMed

    Mårs, Ulla; d'Argy, Roland; Hallbeck, Karin; Miller-Larsson, Anna; Edsbäcker, Staffan

    2013-06-01

    Inhaled corticosteroids (ICS) are mainstay treatment of asthma and chronic obstructive pulmonary disease. However, highly lipophilic ICS accumulate in systemic tissues, which may lead to adverse systemic effects. The accumulation of a new, highly lipophilic ICS, ciclesonide and its active metabolite (des-CIC) has not yet been reported. Here, we have compared tissue accumulation of des-CIC and an ICS of a moderate lipophilicity, budesonide (BUD), after 14 days of once-daily treatment in mice. Single, three or 14 daily doses of [(3) H]-des-CIC or [(3) H]-BUD were administered subcutaneously to male CD1 albino mice, which were killed at 4 hr, 24 hr or 5 days after the last dose. Distribution of tissue concentration of radioactivity was studied by quantitative whole-body autoradiography. Pattern of radioactivity distribution across most tissues was similar for both corticosteroids after a single as well as after repeated dosing. However, tissue concentration of radioactivity differed between des-CIC and BUD. After a single dose, concentrations of radioactivity for both corticosteroids were low for most tissues but increased over 14 days of daily dosing. The tissue radioactivity of des-CIC at 24 hr and 5 days after the 14th dose was 2-3 times higher than that of BUD in majority of tissues. Tissue accumulation, assessed as concentration of tissue radioactivity 5 days after the 14th versus 3rd dose, showed an average ratio of 5.2 for des-CIC and 2.7 for BUD (p < 0.0001). In conclusion, des-CIC accumulated significantly more than BUD. Systemic accumulation may lead to increased risk of adverse systemic side effects during long-term therapy. PMID:23256845

  2. Pharmacokinetics, safety and tolerability of triflusal and its main active metabolite HTB in healthy Chinese subjects.

    PubMed

    Wang, M; Zhang, Q; Huang, M; Zong, S; Hua, W; Zhou, W

    2014-05-01

    Triflusal presents comparable antiplatelet activity to aspirin while presenting a more favourable safety profile, and is used in the treatment of thrombosis. The study aimed to evaluate the pharmacokinetics and safety of triflusal and its major metabolite 2-(hydroxyl)-4-(trifluoromethyl)- benzoic acid (HTB) in healthy Chinese subjects.30 healthy subjects were recruited in this randomized, single-center, and open-label, parallel, single ascending doses (300, 600, 900 mg) and multiple doses (600 mg, once daily for 7 days) study. Plasma samples were analyzed with a validated liquid chromatography tandem mass spectrometry (LC/MS/MS) method. Safety was assessed by adverse events, ECG, laboratory testing, and vital signs.Triflusal was safe and well tolerated. After single-dose administration, triflusal was rapidly absorbed with a mean Tmax of 0.55-0.92 h and a mean t1/2 kel of 0.35-0.65 h, HTB was absorbed with a mean Tmax of 2.35-3.03 h and a mean t1/2 kel of 52.5-65.57 h. Cmax and AUC for triflusal and HTB were approximately dose proportional over the 300-900 mg dose range. In the steady state, the accumulation index (R) indicated that the exposure of triflusal increased slightly with repeated dosing, and the exposure of HTB increased obviously. 3 adverse events certainly related to the investigational drugs occurred in the multiple-dose phase.Following oral dosing under fasting condition, triflusal is promptly absorbed and rapidly depleted from the systemic circulation. HTB is quickly generated from triflusal and slowly eliminated. Triflusal accumulates slightly in the body. HTB plasma concentration builds up progressively toward steady-state. PMID:24105106

  3. All-trans retinoic acid ameliorates glycemic control in diabetic mice via modulating pancreatic islet production of vascular endothelial growth factor-A.

    PubMed

    Chien, Chiao-Yun; Yuan, Tze-An; Cho, Candy Hsin-Hua; Chang, Fang-Pei; Mao, Wan-Yu; Wu, Ruei-Ren; Lee, Hsuan-Shu; Shen, Chia-Ning

    2016-09-01

    Patients with type 1 diabetes mellitus are associated with impairment in vitamin A metabolism. This study evaluated whether treatment with retinoic acid, the biologically active metabolite of vitamin A, can ameliorate diabetes. All-trans retinoic acid (atRA) was used to treat streptozotocin (STZ)-induced diabetic mice which revealed atRA administration ameliorated blood glucose levels of diabetic mice. This hyperglycemic amelioration was accompanied by an increase in the amount of β cells co-expressed Pdx1 and insulin and by restoration of the vascular laminin expression. The atRA-induced production of vascular endothelial growth factor-A from the pancreatic islets was possibly the key factor that mediated the restoration of islet vascularity and recovery of β-cell mass. Furthermore, the combination of islet transplantation and atRA administration significantly rescued hyperglycemia in diabetic mice. These findings suggest that vitamin A derivatives can potentially be used as a supplementary treatment to improve diabetes management and glycemic control. PMID:27381866

  4. Retinoic acid modulates rat Ito cell proliferation, collagen, and transforming growth factor beta production.

    PubMed Central

    Davis, B H; Kramer, R T; Davidson, N O

    1990-01-01

    Recent studies suggest that vitamin A plays an inhibitory role with respect to "activation" of the hepatic Ito cell, a likely effector of hepatic fibrogenesis. Ito cell "activation" during fibrogenesis is characterized by a decrease in intracellular vitamin A and an increase in cellular proliferation and collagen production. To explore the hypothesis that retinoids have the capacity to diminish Ito cell activation, cultured Ito cells were exposed to retinoic acid and its effects assessed on three key features: cell proliferation, collagen protein production and mRNA abundance, and transforming growth factor beta protein production. Retinoic acid was 100-1,000X more potent than retinol with respect to inhibition of Ito cell proliferation. Interstitial collagen and transforming growth factor beta production were also reduced by 10(-6) M retinoic acid. The relative abundance of type I collagen mRNA however, was not significantly altered. By contrast, retinoic acid administration to rats caused a marked reduction in the abundance of type I collagen mRNA in both total hepatic and purified Ito cell RNA. The relative abundance of rat hepatic fibronectin or apolipoprotein E mRNA was not significantly altered. These studies demonstrate that retinoic acid can differentially modulate several key features of hepatic fibrogenesis in vitro and in vivo. Images PMID:2254460

  5. Retinol Promotes In Vitro Growth of Proximal Colon Organoids through a Retinoic Acid-Independent Mechanism

    PubMed Central

    Nibe, Yoichi; Akiyama, Shintaro; Matsumoto, Yuka; Nozaki, Kengo; Fukuda, Masayoshi; Hayashi, Ayumi; Mizutani, Tomohiro; Oshima, Shigeru; Watanabe, Mamoru; Nakamura, Tetsuya

    2016-01-01

    Retinol (ROL), the alcohol form of vitamin A, is known to control cell fate decision of various types of stem cells in the form of its active metabolite, retinoic acid (RA). However, little is known about whether ROL has regulatory effects on colonic stem cells. We examined in this study the effect of ROL on the growth of murine normal colonic cells cultured as organoids. As genes involved in RA synthesis from ROL were differentially expressed along the length of the colon, we tested the effect of ROL on proximal and distal colon organoids separately. We found that organoid forming efficiency and the expression level of Lgr5, a marker gene for colonic stem cells were significantly enhanced by ROL in the proximal colon organoids, but not in the distal ones. Interestingly, neither retinaldehyde (RAL), an intermediate product of the ROL-RA pathway, nor RA exhibited growth promoting effects on the proximal colon organoids, suggesting that ROL-dependent growth enhancement in organoids involves an RA-independent mechanism. This was confirmed by the observation that an inhibitor for RA-mediated gene transcription did not abrogate the effect of ROL on organoids. This novel role of ROL in stem cell maintenance in the proximal colon provides insights into the mechanism of region-specific regulation for colonic stem cell maintenance. PMID:27564706

  6. Retinoic acid induces nuclear accumulation of Raf1 during differentiation of HL-60 cells

    SciTech Connect

    Smith, James; Bunaciu, Rodica P.; Reiterer, Gudrun; Coder, David; George, Thaddeus; Asaly, Michael; Yen, Andrew

    2009-08-01

    All trans-retinoic acid (RA) is a standard therapeutic agent used in differentiation induction therapy treatment of acute promyelocytic leukemia (APL). RA and its metabolites use a diverse set of signal transduction pathways during the differentiation program. In addition to the direct transcriptional targets of the nuclear RAR and RXR receptors, signals derived from membrane receptors and the Raf-MEK-ERK pathway are required. Raf1 phosphorylation and the prolonged activation of Raf1 persisting during the entire differentiation process are required for RA-dependent differentiation of HL-60 cells. Here we identify a nuclear redistribution of Raf1 during the RA-induced differentiation of HL-60 cells. In addition, the nuclear accumulation of Raf1 correlates with an increase in Raf1 phosphorylated at serine 621. The serine 621 phosphorylated Raf1 is predominantly localized in the nucleus. The RA-dependent nuclear accumulation of Raf1 suggests a novel nuclear role for Raf1 during the differentiation process.

  7. The retinoic acid receptor agonist Am80 increases hippocampal ADAM10 in aged SAMP8 mice.

    PubMed

    Kitaoka, Kazuyoshi; Shimizu, Noriyuki; Ono, Koji; Chikahisa, Sachiko; Nakagomi, Madoka; Shudo, Koichi; Ishimura, Kazunori; Séi, Hiroyoshi; Yoshizaki, Kazuo

    2013-09-01

    The retinoic acid (RA, a vitamin A metabolite) receptor (RAR) is a transcription factor. Vitamin A/RA administration improves the Alzheimer's disease (AD)- and age-related attenuation of memory/learning in mouse models. Recently, a disintegrin and metalloproteinase domain-containing protein 10 (ADAM10) was identified as a key molecule in RA-mediated anti-AD mechanisms. We investigated the effect of chronic administration of the RAR agonist Am80 (tamibarotene) on ADAM10 expression in senescence-accelerated mice (SAMP8). Moreover, we estimated changes in the expression of the amyloid precursor protein (APP), amyloid beta (Aβ), and hairy/enhancer of split (Hes), which are mediated by ADAM10. Spatial working memory and the levels of a hippocampal proliferation marker (Ki67) were also assessed in these mice. ADAM10 mRNA and protein expression was significantly reduced in the hippocampus of 13-month-old SAMP8 mice; their expression improved significantly after Am80 administration. Further, after Am80 administration, the expression levels of Hes5 and Ki67 were restored and the deterioration of working memory was suppressed, whereas APP and Aβ levels remained unchanged. Our results suggest that Am80 administration effectively improves dementia by activating the hippocampal ADAM10-Notch-Hes5 proliferative pathway. PMID:23624141

  8. Importance of ALDH1A enzymes in determining human testicular retinoic acid concentrations

    PubMed Central

    Arnold, Samuel L.; Kent, Travis; Hogarth, Cathryn A.; Schlatt, Stefan; Prasad, Bhagwat; Haenisch, Michael; Walsh, Thomas; Muller, Charles H.; Griswold, Michael D.; Amory, John K.; Isoherranen, Nina

    2015-01-01

    Retinoic acid (RA), the active metabolite of vitamin A, is required for spermatogenesis and many other biological processes. RA formation requires irreversible oxidation of retinal to RA by aldehyde dehydrogenase enzymes of the 1A family (ALDH1A). While ALDH1A1, ALDH1A2, and ALDH1A3 all form RA, the expression pattern and relative contribution of these enzymes to RA formation in the testis is unknown. In this study, novel methods to measure ALDH1A protein levels and intrinsic RA formation were used to accurately predict RA formation velocities in individual human testis samples and an association between RA formation and intratesticular RA concentrations was observed. The distinct localization of ALDH1A in the testis suggests a specific role for each enzyme in controlling RA formation. ALDH1A1 was found in Sertoli cells, while only ALDH1A2 was found in spermatogonia, spermatids, and spermatocytes. In the absence of cellular retinol binding protein (CRBP)1, ALDH1A1 was predicted to be the main contributor to intratesticular RA formation, but when CRBP1 was present, ALDH1A2 was predicted to be equally important in RA formation as ALDH1A1. This study provides a comprehensive novel methodology to evaluate RA homeostasis in human tissues and provides insight to how the individual ALDH1A enzymes mediate RA concentrations in specific cell types. PMID:25502770

  9. Regulation of human tonsillar T-cell proliferation by the active metabolite of vitamin D3.

    PubMed Central

    Nunn, J D; Katz, D R; Barker, S; Fraher, L J; Hewison, M; Hendy, G N; O'Riordan, J L

    1986-01-01

    We have examined the effects of 1,25(OH)2D3 on T-cell populations isolated by buoyant density and E rosetting from human tonsils. Cell proliferation was assessed by measuring the incorporation of 125iododeoxyuridine; interleukin-2 (IL-2) production was measured using an IL-2-dependent cell line, and the number of 1,25(OH)2D3 receptors was measured by whole-cell nuclear association assay. At a concentration of 10(-7) M, 1,25(OH)2D3 inhibited mitogen-induced T-cell proliferation in all E+ T-cell populations. This effect was more pronounced in the cells from the intermediate and high density layers and was reflected both in cell proliferative responses and in relative IL-2 synthesis. By adding the 1,25(OH)2D3 during the course of the mitogen assay, we demonstrated that activation of the T cell precedes the 1,25(OH)2D3-mediated inhibition. Cells that had been preincubated with mitogen in the presence of the 1,25(OH)2D3 were refractory to further stimulation by mitogens. Receptors for 1,25(OH)2D3 could not be detected in unstimulated T cells. However, activation led to the expression of high-affinity receptors for 1,25(OH)2D3. Co-incubation of the cells with mitogen and 1,25(OH)2D3 increased the number of receptors compared with mitogen alone. The effects provide further evidence for the hypothesis that 1,25(OH)2D3 is an important potential modulator of the immune system through its action on T cells. Taking our observations in conjunction with the known capacity of monocytes to hydroxylate the precursor metabolite (and thus synthesize the active form of cholecalciferol), the results support the suggestion that 1,25(OH)2D3 plays a role as a local mediator of mononuclear phagocyte-T cell interaction in human lymphomedullary tissues. PMID:3026959

  10. Trace Amine-Associated Receptor 1 (TAAR1) is Activated by Amiodarone Metabolites

    PubMed Central

    Snead, Aaron N.; Miyakawa, Motonori; Tan, Edwin S.; Scanlan, Thomas S.

    2012-01-01

    Amiodarone (Cordarone, Wyeth-Ayerst Pharmaceuticals) is a clinically available drug used to treat a wide variety of cardiac arrhythmias. We report here the synthesis and characterization of a panel of potential amiodarone metabolites that have significant structural similarity to thyroid hormone and its metabolites the iodothyronamines. Several of these amiodarone derivatives act as specific agonists of the G protein-coupled receptor (GPCR) trace amine-associated receptor 1 (TAAR1). This result demonstrates a novel molecular target for amiodarone derivatives with potential clinical significance. PMID:18752950

  11. [Detection of fungal metabolites showing toxic activity through Artemia salina bioassay].

    PubMed

    González, Ana María; Presa, Maximiliano; Latorre, María Gabriela; Lurá, María Cristina

    2007-03-01

    The aim of this study was to detect toxic metabolites from fungi contaminating food and medicinal herbs by applying the toxicity assay to Artemia salina. According to toxicity percentages, the extracts were classified as nontoxic (NT), slightly toxic (ST), toxic (T) and highly toxic (HT). Those classified as T and HT were assayed for mycotoxins. Only 6 out of 71 strains were found to be T (8.5%) for A. salina. Penicillium brevicompactum Dierckx, isolated from sausages, was found to be HT, mainly due to the presence of ochratoxin A and two other unidentified metabolites. PMID:17592895

  12. Cadmium induces retinoic acid signaling by regulating retinoic acid metabolic gene expression.

    PubMed

    Cui, Yuxia; Freedman, Jonathan H

    2009-09-11

    The transition metal cadmium is an environmental teratogen. In addition, cadmium and retinoic acid can act synergistically to induce forelimb malformations. The molecular mechanism underlying the teratogenicity of cadmium and the synergistic effect with retinoic acid has not been addressed. An evolutionarily conserved gene, beta,beta-carotene 15,15'-monooxygenase (BCMO), which is involved in retinoic acid biosynthesis, was studied in both Caenorhabditis elegans and murine Hepa 1-6 cells. In C. elegans, bcmo-1 was expressed in the intestine and was cadmium inducible. Similarly, in Hepa 1-6 cells, Bcmo1 was induced by cadmium. Retinoic acid-mediated signaling increased after 24-h exposures to 5 and 10 microm cadmium in Hepa 1-6 cells. Examination of gene expression demonstrated that the induction of retinoic acid signaling by cadmium may be mediated by overexpression of Bcmo1. Furthermore, cadmium inhibited the expression of Cyp26a1 and Cyp26b1, which are involved in retinoic acid degradation. These results indicate that cadmium-induced teratogenicity may be due to the ability of the metal to increase the levels of retinoic acid by disrupting the expression of retinoic acid-metabolizing genes. PMID:19556237

  13. Cadmium Induces Retinoic Acid Signaling by Regulating Retinoic Acid Metabolic Gene Expression*

    PubMed Central

    Cui, Yuxia; Freedman, Jonathan H.

    2009-01-01

    The transition metal cadmium is an environmental teratogen. In addition, cadmium and retinoic acid can act synergistically to induce forelimb malformations. The molecular mechanism underlying the teratogenicity of cadmium and the synergistic effect with retinoic acid has not been addressed. An evolutionarily conserved gene, β,β-carotene 15,15′-monooxygenase (BCMO), which is involved in retinoic acid biosynthesis, was studied in both Caenorhabditis elegans and murine Hepa 1–6 cells. In C. elegans, bcmo-1 was expressed in the intestine and was cadmium inducible. Similarly, in Hepa 1–6 cells, Bcmo1 was induced by cadmium. Retinoic acid-mediated signaling increased after 24-h exposures to 5 and 10 μm cadmium in Hepa 1–6 cells. Examination of gene expression demonstrated that the induction of retinoic acid signaling by cadmium may be mediated by overexpression of Bcmo1. Furthermore, cadmium inhibited the expression of Cyp26a1 and Cyp26b1, which are involved in retinoic acid degradation. These results indicate that cadmium-induced teratogenicity may be due to the ability of the metal to increase the levels of retinoic acid by disrupting the expression of retinoic acid-metabolizing genes. PMID:19556237

  14. Non-targeted Metabolite Profiling and Scavenging Activity Unveil the Nutraceutical Potential of Psyllium (Plantago ovata Forsk).

    PubMed

    Patel, Manish K; Mishra, Avinash; Jha, Bhavanath

    2016-01-01

    Non-targeted metabolomics implies that psyllium (Plantago ovata) is a rich source of natural antioxidants, PUFAs (ω-3 and ω-6 fatty acids) and essential and sulfur-rich amino acids, as recommended by the FAO for human health. Psyllium contains phenolics and flavonoids that possess reducing capacity and reactive oxygen species (ROS) scavenging activities. In leaves, seeds, and husks, about 76, 78, 58% polyunsaturated, 21, 15, 20% saturated, and 3, 7, 22% monounsaturated fatty acids were found, respectively. A range of FAs (C12 to C24) was detected in psyllium and among different plant parts, a high content of the nutritive indicators ω-3 alpha-linolenic acid CPS (57%) and ω-6 linoleic acid CPS (18%) was detected in leaves. Similarly, total content of phenolics and the essential amino acid valine were also detected utmost in leaves followed by sulfur-rich amino acids and flavonoids. In total, 36 different metabolites were identified in psyllium, out of which 26 (13 each) metabolites were detected in leaves and seeds, whereas the remaining 10 were found in the husk. Most of the metabolites are natural antioxidants, phenolics, flavonoids, or alkaloids and can be used as nutrient supplements. Moreover, these metabolites have been reported to have several pharmaceutical applications, including anti-cancer activity. Natural plant ROS scavengers, saponins, were also detected. Based on metabolomic data, the probable presence of a flavonoid biosynthesis pathway was inferred, which provides useful insight for metabolic engineering in the future. Non-targeted metabolomics, antioxidants and scavenging activities reveal the nutraceutical potential of the plant and also suggest that psyllium leaves can be used as a green salad as a dietary supplement to daily food. PMID:27092153

  15. Non-targeted Metabolite Profiling and Scavenging Activity Unveil the Nutraceutical Potential of Psyllium (Plantago ovata Forsk)

    PubMed Central

    Patel, Manish K.; Mishra, Avinash; Jha, Bhavanath

    2016-01-01

    Non-targeted metabolomics implies that psyllium (Plantago ovata) is a rich source of natural antioxidants, PUFAs (ω-3 and ω-6 fatty acids) and essential and sulfur-rich amino acids, as recommended by the FAO for human health. Psyllium contains phenolics and flavonoids that possess reducing capacity and reactive oxygen species (ROS) scavenging activities. In leaves, seeds, and husks, about 76, 78, 58% polyunsaturated, 21, 15, 20% saturated, and 3, 7, 22% monounsaturated fatty acids were found, respectively. A range of FAs (C12 to C24) was detected in psyllium and among different plant parts, a high content of the nutritive indicators ω-3 alpha-linolenic acid (57%) and ω-6 linoleic acid (18%) was detected in leaves. Similarly, total content of phenolics and the essential amino acid valine were also detected utmost in leaves followed by sulfur-rich amino acids and flavonoids. In total, 36 different metabolites were identified in psyllium, out of which 26 (13 each) metabolites were detected in leaves and seeds, whereas the remaining 10 were found in the husk. Most of the metabolites are natural antioxidants, phenolics, flavonoids, or alkaloids and can be used as nutrient supplements. Moreover, these metabolites have been reported to have several pharmaceutical applications, including anti-cancer activity. Natural plant ROS scavengers, saponins, were also detected. Based on metabolomic data, the probable presence of a flavonoid biosynthesis pathway was inferred, which provides useful insight for metabolic engineering in the future. Non-targeted metabolomics, antioxidants and scavenging activities reveal the nutraceutical potential of the plant and also suggest that psyllium leaves can be used as a green salad as a dietary supplement to daily food. PMID:27092153

  16. Cellular Metabolic Activity and the Oxygen and Hydrogen Stable Isotope Composition of Intracellular Water and Metabolites

    NASA Astrophysics Data System (ADS)

    Kreuzer-Martin, H. W.; Hegg, E. L.

    2008-12-01

    biomass of Bacillus subtilis, a Gram-positive bacterium, showed the same pattern. Rapidly-dividing cells derived fewer of their O and H atoms from environmental water than did more slowly-growing cells and spores. To test whether a eukaryotic cell, surrounded by only a membrane, would also maintain an isotopic gradient and a detectable percentage of metabolic water, we applied our approach to cultured rat fibroblasts. Preliminary results showed that approximately 50% of the O and H atoms in exponentially growing cells were derived from metabolic activity. In quiescent cells, metabolic activity generated approximately 25% of the O and H atoms in intracellular water. Thus far, the data we have obtained is consistent with the following model: (1) Intracellular water is composed of water that diffuses in from the extracellular environment and water that is created as a result of metabolic activity. (2) The relative amounts of environmental and metabolic water inside a cell are a function of the cell's metabolic activity. (3) The oxygen and hydrogen isotope ratios of cellular metabolites are a function of those of intracellular water, and therefore reflect the metabolic activity of the cell at the time of biosynthesis.

  17. Loss-of-Function of HtrA1 Abrogates All-Trans Retinoic Acid-Induced Osteogenic Differentiation of Mouse Adipose-Derived Stromal Cells Through Deficiencies in p70S6K Activation.

    PubMed

    Glanz, Stephan; Mirsaidi, Ali; López-Fagundo, Cristina; Filliat, Gladys; Tiaden, André N; Richards, Peter J

    2016-05-01

    All-trans retinoic acid (ATRA) is a potent inducer of osteogenic differentiation in mouse adipose-derived stromal cells (mASCs), although the underlying mechanisms responsible for its mode of action have yet to be completely elucidated. High temperature requirement protease A1 (HtrA1) is a newly recognized modulator of human multipotent stromal cell (MSC) osteogenesis and as such, may play a role in regulating ATRA-dependent osteogenic differentiation of mASCs. In this study, we assessed the influence of small interfering RNA (siRNA)-induced repression of HtrA1 production on mASC osteogenesis and examined its effects on ATRA-mediated mammalian target of rapamycin (mTOR) signaling. Inhibition of HtrA1 production in osteogenic mASCs resulted in a significant reduction of alkaline phosphatase activity and mineralized matrix formation. Western blot analyses revealed the rapid activation of Akt (Ser473) and p70S6K (Thr389) in ATRA-treated mASCs, and that levels of phosphorylated p70S6K were noticeably reduced in HtrA1-deficient mASCs. Further studies using mTOR inhibitor rapamycin and siRNA specific for the p70S6K gene Rps6kb1 confirmed ATRA-mediated mASC osteogenesis as being dependent on p70S6K activation. Finally, transfection of cells with a constitutively active rapamycin-resistant p70S6K mutant could restore the mineralizing capacity of HtrA1-deficient mASCs. These findings therefore lend further support for HtrA1 as a positive mediator of MSC osteogenesis and provide new insights into the molecular mode of action of ATRA in regulating mASC lineage commitment. PMID:26950191

  18. All-Trans Retinoic Acid Induces TGF-β2 in Intestinal Epithelial Cells via RhoA- and p38α MAPK-Mediated Activation of the Transcription Factor ATF2

    PubMed Central

    Namachivayam, Kopperuncholan; MohanKumar, Krishnan; Arbach, Dima; Jagadeeswaran, Ramasamy; Jain, Sunil K.; Natarajan, Viswanathan; Mehta, Dolly; Jankov, Robert P.; Maheshwari, Akhil

    2015-01-01

    Objective We have shown previously that preterm infants are at risk of necrotizing enterocolitis (NEC), an inflammatory bowel necrosis typically seen in infants born prior to 32 weeks’ gestation, because of the developmental deficiency of transforming growth factor (TGF)-β2 in the intestine. The present study was designed to investigate all-trans retinoic acid (atRA) as an inducer of TGF-β2 in intestinal epithelial cells (IECs) and to elucidate the involved signaling mechanisms. Methods AtRA effects on intestinal epithelium were investigated using IEC6 cells. TGF-β2 expression was measured using reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) and Western blots. Signaling pathways were investigated using Western blots, transiently-transfected/transduced cells, kinase arrays, chromatin immunoprecipitation, and selective small molecule inhibitors. Results AtRA-treatment of IEC6 cells selectively increased TGF-β2 mRNA and protein expression in a time- and dose-dependent fashion, and increased the activity of the TGF-β2 promoter. AtRA effects were mediated via RhoA GTPase, Rho-associated, coiled-coil-containing protein kinase 1 (ROCK1), p38α MAPK, and activating transcription factor (ATF)-2. AtRA increased phospho-ATF2 binding to the TGF-β2 promoter and increased histone H2B acetylation in the TGF-β2 nucleosome, which is typically associated with transcriptional activation. Conclusions AtRA induces TGF-β2 expression in IECs via RhoA- and p38α MAPK-mediated activation of the transcription factor ATF2. Further studies are needed to investigate the role of atRA as a protective/therapeutic agent in gut mucosal inflammation. PMID:26225425

  19. Mixture toxicity of the antiviral drug Tamiflu((R)) (oseltamivir ethylester) and its active metabolite oseltamivir acid.

    PubMed

    Escher, Beate I; Bramaz, Nadine; Lienert, Judit; Neuwoehner, Judith; Straub, Jürg Oliver

    2010-02-18

    Tamiflu (oseltamivir ethylester) is an antiviral agent for the treatment of influenza A and B. The pro-drug Tamiflu is converted in the human body to the pharmacologically active metabolite, oseltamivir acid, with a yield of 75%. Oseltamivir acid is indirectly photodegradable and slowly biodegradable in sewage works and sediment/water systems. A previous environmental risk assessment has concluded that there is no bioaccumulation potential of either of the compounds. However, little was known about the ecotoxicity of the metabolite. Ester hydrolysis typically reduces the hydrophobicity and thus the toxicity of a compound. In this case, a zwitterionic, but overall neutral species is formed from the charged parent compound. If the speciation and predicted partitioning into biological membranes is considered, the metabolite may have a relevant contribution to the overall toxicity. These theoretical considerations triggered a study to investigate the toxicity of oseltamivir acid (OA), alone and in binary mixtures with its parent compound oseltamivir ethylester (OE). OE and OA were found to be baseline toxicants in the bioluminescence inhibition test with Vibrio fischeri. Their mixture effect lay between predictions for concentration addition and independent action for the mixture ratio excreted in urine and nine additional mixture ratios of OE and OA. In contrast, OE was an order of magnitude more toxic than OA towards algae, with a more pronounced effect when the direct inhibition of photosystem II was used as toxicity endpoint opposed to the 24h growth rate endpoint. The binary mixtures in this assay yielded experimental mixture effects that agreed with predictions for independent action. This is consistent with the finding that OE exhibits slightly enhanced toxicity, while OA acts as baseline toxicant. Therefore, with respect to mixture classification, the two compounds can be considered as acting according to different modes of toxic action, although there are

  20. Three conazoles increase hepatic microsomal retinoic acid metabolism and decrease mouse hepatic retinoic acid levels in vivo

    SciTech Connect

    Chen, P.-J.; Padgett, William T.; Moore, Tanya; Winnik, Witold; Lambert, Guy R.; Thai, Sheau-Fung; Hester, Susan D.; Nesnow, Stephen

    2009-01-15

    Conazoles are fungicides used in agriculture and as pharmaceuticals. In a previous toxicogenomic study of triazole-containing conazoles we found gene expression changes consistent with the alteration of the metabolism of all trans-retinoic acid (atRA), a vitamin A metabolite with cancer-preventative properties (Ward et al., Toxicol. Pathol. 2006; 34:863-78). The goals of this study were to examine effects of propiconazole, triadimefon, and myclobutanil, three triazole-containing conazoles, on the microsomal metabolism of atRA, the associated hepatic cytochrome P450 (P450) enzyme(s) involved in atRA metabolism, and their effects on hepatic atRA levels in vivo. The in vitro metabolism of atRA was quantitatively measured in liver microsomes from male CD-1 mice following four daily intraperitoneal injections of propiconazole (210 mg/kg/d), triadimefon (257 mg/kg/d) or myclobutanil (270 mg/kg/d). The formation of both 4-hydroxy-atRA and 4-oxo-atRA were significantly increased by all three conazoles. Propiconazole-induced microsomes possessed slightly greater metabolizing activities compared to myclobutanil-induced microsomes. Both propiconazole and triadimefon treatment induced greater formation of 4-hydroxy-atRA compared to myclobutanil treatment. Chemical and immuno-inhibition metabolism studies suggested that Cyp26a1, Cyp2b, and Cyp3a, but not Cyp1a1 proteins were involved in atRA metabolism. Cyp2b10/20 and Cyp3a11 genes were significantly over-expressed in the livers of both triadimefon- and propiconazole-treated mice while Cyp26a1, Cyp2c65 and Cyp1a2 genes were over-expressed in the livers of either triadimefon- or propiconazole-treated mice, and Cyp2b10/20 and Cyp3a13 genes were over-expressed in the livers of myclobutanil-treated mice. Western blot analyses indicated conazole induced-increases in Cyp2b and Cyp3a proteins. All three conazoles decreased hepatic atRA tissue levels ranging from 45-67%. The possible implications of these changes in hepatic atRA levels

  1. Integrated circuit-based electrochemical sensor for spatially resolved detection of redox-active metabolites in biofilms

    PubMed Central

    Bellin, Daniel L.; Sakhtah, Hassan; Rosenstein, Jacob K.; Levine, Peter M.; Thimot, Jordan; Emmett, Kevin; Dietrich, Lars E. P.; Shepard, Kenneth L.

    2014-01-01

    Despite advances in monitoring spatiotemporal expression patterns of genes and proteins with fluorescent probes, direct detection of metabolites and small molecules remains challenging. A technique for spatially resolved detection of small molecules would benefit the study of redox-active metabolites produced by microbial biofilms, which can drastically affect colony development. Here we present an integrated circuit-based electrochemical sensing platform featuring an array of working electrodes and parallel potentiostat channels. “Images” over a 3.25 × 0.9 mm area can be captured with a diffusion-limited spatial resolution of 750 μm. We demonstrate that square wave voltammetry can be used to detect, identify, and quantify (for concentrations as low as 2.6 μM) four distinct redox-active metabolites called phenazines. We characterize phenazine production in both wild-type and mutant Pseudomonas aeruginosa PA14 colony biofilms, and find correlations with fluorescent reporter imaging of phenazine biosynthetic gene expression. PMID:24510163

  2. Metabolites from Aspergillus fumigatus, an endophytic fungus associated with Melia azedarach, and their antifungal, antifeedant, and toxic activities.

    PubMed

    Li, Xiao-Jun; Zhang, Qiang; Zhang, An-Ling; Gao, Jin-Ming

    2012-04-01

    Thirty-nine fungal metabolites 1-39, including two new alkaloids, 12β-hydroxy-13α-methoxyverruculogen TR-2 (6) and 3-hydroxyfumiquinazoline A (16), were isolated from the fermentation broth of Aspergillus fumigatus LN-4, an endophytic fungus isolated from the stem bark of Melia azedarach. Their structures were elucidated on the basis of detailed spectroscopic analysis (mass spectrometry and one- and two-dimensional NMR experiments) and by comparison of their NMR data with those reported in the literature. These isolated compounds were evaluated for in vitro antifungal activities against some phytopathogenic fungi, toxicity against brine shrimps, and antifeedant activities against armyworm larvae (Mythimna separata Walker). Among them, sixteen compounds showed potent antifungal activities against phytopathogenic fungi (Botrytis cinerea, Alternaria solani, Alternaria alternata, Colletotrichum gloeosporioides, Fusarium solani, Fusarium oxysporum f. sp. niveum, Fusarium oxysporum f. sp. vasinfectum, and Gibberella saubinettii), and four of them, 12β-hydroxy-13α-methoxyverruculogen TR-2 (6), fumitremorgin B (7), verruculogen (8), and helvolic acid (39), exhibited antifungal activities with MIC values of 6.25-50 μg/mL, which were comparable to the two positive controls carbendazim and hymexazol. In addition, of eighteen that exerted moderate lethality toward brine shrimps, compounds 7 and 8 both showed significant toxicities with median lethal concentration (LC(50)) values of 13.6 and 15.8 μg/mL, respectively. Furthermore, among nine metabolites that were found to possess antifeedant activity against armyworm larvae, compounds 7 and 8 gave the best activity with antifeedant indexes (AFI) of 50.0% and 55.0%, respectively. Structure-activity relationships of the metabolites were also discussed. PMID:22409377

  3. Spectrofluorimetric determination of 3-methylflavone-8-carboxylic acid, the main active metabolite of flavoxate hydrochloride in human urine

    NASA Astrophysics Data System (ADS)

    Zaazaa, Hala E.; Mohamed, Afaf O.; Hawwam, Maha A.; Abdelkawy, Mohamed

    2015-01-01

    A simple, sensitive and selective spectrofluorimetric method has been developed for the determination of 3-methylflavone-8-carboxylic acid as the main active metabolite of flavoxate hydrochloride in human urine. The proposed method was based on the measurement of the native fluorescence of the metabolite in methanol at an emission wavelength 390 nm, upon excitation at 338 nm. Moreover, the urinary excretion pattern has been calculated using the proposed method. Taking the advantage that 3-methylflavone-8-carboxylic acid is also the alkaline degradate, the proposed method was applied to in vitro determination of flavoxate hydrochloride in tablets dosage form via the measurement of its corresponding degradate. The method was validated in accordance with the ICH requirements and statistically compared to the official method with no significant difference in performance.

  4. Contrasting Roles For All-Trans Retinoic Acid in TGF-ß-mediated Induction of Foxp3 and Il10 Genes in Developing Regulatory T Cells

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Extrathymic induction of regulatory T cells (Treg) is essential to the regulation of effector T cell responses in the periphery. TGF-ß has been shown to induce Foxp3-expressing Tregs both in vitro and in vivo. More recently, the vitamin A metabolite, all-trans retinoic acid (at-RA), has been found t...

  5. Solving the Jigsaw Puzzle of Wound-Healing Potato Cultivars: Metabolite Profiling and Antioxidant Activity of Polar Extracts

    PubMed Central

    2015-01-01

    Potato (Solanum tuberosum L.) is a worldwide food staple, but substantial waste accompanies the cultivation of this crop due to wounding of the outer skin and subsequent unfavorable healing conditions. Motivated by both economic and nutritional considerations, this metabolite profiling study aims to improve understanding of closing layer and wound periderm formation and guide the development of new methods to ensure faster and more complete healing after skin breakage. The polar metabolites of wound-healing tissues from four potato cultivars with differing patterns of tuber skin russeting (Norkotah Russet, Atlantic, Chipeta, and Yukon Gold) were analyzed at three and seven days after wounding, during suberized closing layer formation and nascent wound periderm development, respectively. The polar extracts were assessed using LC-MS and NMR spectroscopic methods, including multivariate analysis and tentative identification of 22 of the 24 biomarkers that discriminate among the cultivars at a given wound-healing time point or between developmental stages. Differences among the metabolites that could be identified from NMR- and MS-derived biomarkers highlight the strengths and limitations of each method, also demonstrating the complementarity of these approaches in terms of assembling a complete molecular picture of the tissue extracts. Both methods revealed that differences among the cultivar metabolite profiles diminish as healing proceeds during the period following wounding. The biomarkers included polyphenolic amines, flavonoid glycosides, phenolic acids and glycoalkaloids. Because wound healing is associated with oxidative stress, the free radical scavenging activities of the extracts from different cultivars were measured at each wounding time point, revealing significantly higher scavenging activity of the Yukon Gold periderm especially after 7 days of wounding. PMID:24998264

  6. Active Oxygen Metabolites and Thromboxane in Phorbol Myristate Acetate Toxicity to the Isolated, Perfused Rat Lung.

    NASA Astrophysics Data System (ADS)

    Carpenter, Laurie Jean

    When administered intravenously or intratracheally to rats, rabbits and sheep, phorbol myristate acetate (PMA) produces changes in lung morphology and function are similar to those seen in humans with the adult respiratory distress syndrome (ARDS). Therefore, it is thought that information about the mechanism of ARDS development can be gained from experiments using PMA-treated animals. Currently, the mechanisms by which PMA causes pneumotoxicity are unknown. Results from other studies in rabbits and in isolated, perfused rabbit lungs suggest that PMA-induced lung injury is mediated by active oxygen species from neutrophils (PMN), whereas studies in sheep and rats suggest that PMN are not required for the toxic response. The role of PMN, active oxygen metabolites and thromboxane (TxA_2) in PMA-induced injury to isolated, perfused rat lungs (IPLs) was examined in this thesis. To determine whether PMN were required for PMA to produce toxicity to the IPL, lungs were perfused for 30 min with buffer containing various concentrations of PMA (in the presence or absence of PMN). When concentrations >=q57 ng/ml were added to medium devoid of added PMN, perfusion pressure and lung weight increased. When a concentration of PMA (14-28 ng/ml) that did not by itself cause lungs to accumulate fluid was added to the perfusion medium containing PMN (1 x 10 ^8), perfusion pressure increased, and lungs accumulated fluid. These results indicate that high concentrations of PMA produce lung injury which is independent of PMN, whereas injury induced by lower concentrations is PMN-dependent. To examine whether active oxygen species were involved in mediating lung injury induced by PMA and PMN, lungs were coperfused with the oxygen radical scavengers SOD and/or catalase. Coperfusion with either or both of these enzymes totally protected lungs against injury caused by PMN and PMA. These results suggest that active oxygen species (the hydroxyl radical in particular), mediate lung injury in

  7. Tissue distribution study of columbianadin and its active metabolite columbianetin in rats.

    PubMed

    Zhang, You-Bo; Yang, Xiu-Wei

    2016-02-01

    Columbianadin, one of the main bioactive constituents of the roots of Angelica pubescens Maxim. f. biserrata Shan et Yuan, has been found to possess obvious pharmacological effects in previous studies. In this study, a valid and sensitive reverse-phase high-performance liquid chromatography (RP-HPLC) method was established and validated for the determination of columbianadin (CBN) and its active metabolite columbianetin (CBT) in rat tissue samples. Sample separation was performed on an RP-HPLC column using a mobile phase of MeOH-H2 O (75:25, v/v) at a flow rate of 1.0 mL/min. The UV absorbance of the samples was measured at the wavelength 325 nm. The calibration curves for CBN were linear over the ranges of 0.5-20 µg/g for brain, testes and muscle, 1.0-10.0 µg/g for stomach and intestine, and 0.2-20.0 µg/g for heart, liver, spleen, lung and kidney. The calibration curves for CBT were linear over the ranges of 0.5-25 µg/g for stomach and intestine, and 0.1-10.0 µg/g for heart, liver, spleen, lung and kidney. The analysis method was successfully applied to a tissue distribution study of CBN and CBT after intravenous administration of CBN to rats. The results of this study indicated that CBN could be detected in all of the selected tissues after i.v. administration. CBN was distributed to rat tissues rapidly and could be metabolized to CBT in most detected tissues. Of the detected tissues, heart had the highest uptake of CBN, which suggested that heart might be one of the main target tissues of CBN. Concentrations of CBT were obviously higher in the digestive system than in other assayed tissues. The information provided by this research is very useful for gaining knowledge of the capacities of CBN and CBT to access different tissues. PMID:26115176

  8. Mitogen-activated protein kinases regulate expression of neuronal nitric oxide synthase and neurite outgrowth via non-classical retinoic acid receptor signaling in human neuroblastoma SH-SY5Y cells.

    PubMed

    Fujibayashi, Tatsuya; Kurauchi, Yuki; Hisatsune, Akinori; Seki, Takahiro; Shudo, Koichi; Katsuki, Hiroshi

    2015-10-01

    We have previously shown that retinoic acid receptor (RAR) stimulation by an agonist Am80 recruits nitric oxide-dependent signaling via increased expression of neuronal nitric oxide synthase (nNOS) in rat midbrain slice cultures. Using neuroblastoma SH-SY5Y cells, here we investigated the mechanisms of RAR-induced nNOS expression, together with relationship between nNOS expression and neurite outgrowth. Am80 promoted neurite outgrowth, which was attenuated by inhibitors of phosphoinositide 3-kinase (PI3K; LY294002), c-Jun N-terminal kinase (JNK; SP600125) and p38 mitogen-activated protein kinase (p38 MAPK; SB203580). A selective nNOS inhibitor 3-bromo-nitroindazole also suppressed Am80-induced neurite outgrowth. Am80-induced increase in nNOS protein expression was attenuated by LY294002, SP600125 and SB203580, whereas increase in nNOS mRNA expression was attenuated only by LY294002. Am80-induced activation of JNK and p38 MAPK was blocked by LY294002, suggesting that these kinases acted downstream of PI3K. We also confirmed that DAX1, a nuclear receptor reported to regulate nNOS expression, was up-regulated in response to Am80. siRNA-mediated knockdown of DAX1 abrogated Am80-induced nNOS expression and neurite outgrowth. These results reveal for the first time that nNOS expression is crucial for RAR-mediated neurite outgrowth, and that non-genomic signaling such as JNK and p38 MAPK is involved in RAR-mediated nNOS expression. PMID:26422672

  9. Keratinocyte lipid fluidity under the influence of cholesterols, hydrocortisones, "active lipid", tocopherol and retinoic acid--a fluorescence polarization study with regard to physiological and pathophysiological epidermopoiesis and its therapeutic accessibility.

    PubMed

    Bonnekoh, B; Daefler, S; Krueger, G R; Mahrle, G

    1991-01-01

    Lipid fluidity of freshly isolated human (H) and guinea pig (GP) keratinocytes (K) was determined as the reciprocal of diphenylhexatriene (DPH) fluorescence polarization (P-value), the temperature being kept at 25 degrees C and cell density standardized to 550,000 per ml (level of statistical significance a less than 0.05). An experimental model involving short-term incubations (2.5 hours, 37 degrees C) of GPK in 1% ethanolic lipid solutions (15 mg lipid agent per ml ethanol) was set up to investigate accumulation a) of cholesterol due to terminal differentiation of keratinocytes and b) of cholesteryl sulfate due to the lack of steroid sulfatase activity in recessive X-linked ichthyosis (RXLI). In comparison to the control including 1% ethanol (P = 0.291 +/- 0.004), significant rigidifying effects were demonstrated for cholesteryl hemisuccinate (0.331 +/- 0.005) and cholesteryl sulfate (0.310 +/- 0.002). Correspondingly, a significant increase of the P-value was also induced by cholesteryl hemisuccinate in HK. Rigidification of GPK by a preincubation with cholesteryl sulfate (P = 0.306 +/- 0.002) could be antagonized by a subsequent short-term incubation with "active lipid (mixture 721)" (0.285 +/- 0.003, a less than 0.05) which may be relevant for future therapeutic strategies in RXLI. Other steran molecules such as hydrocortisone-21-hemisuccinate or hydrocortisone acetate did not affect lipid fluidity. With regard to the therapeutic potency of retinoids in epidermopoietic disorders, incubations of HK with all-trans-retinoic-acid were compared to those with also lipophilic vitamin E, i.e. d-alpha-tocopherol, for 2.5 hours at 37 degrees C using 1% DMSO as a solvent.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:1893078

  10. Peroxisome Proliferator-Activated Receptor Activation is Associated with Altered Plasma One-Carbon Metabolites and B-Vitamin Status in Rats

    PubMed Central

    Lysne, Vegard; Strand, Elin; Svingen, Gard F. T.; Bjørndal, Bodil; Pedersen, Eva R.; Midttun, Øivind; Olsen, Thomas; Ueland, Per M.; Berge, Rolf K.; Nygård, Ottar

    2016-01-01

    Plasma concentrations of metabolites along the choline oxidation pathway have been linked to increased risk of major lifestyle diseases, and peroxisome proliferator-activated receptors (PPARs) have been suggested to be involved in the regulation of key enzymes along this pathway. In this study, we investigated the effect of PPAR activation on circulating and urinary one-carbon metabolites as well as markers of B-vitamin status. Male Wistar rats (n = 20) received for 50 weeks either a high-fat control diet or a high-fat diet with tetradecylthioacetic acid (TTA), a modified fatty acid and pan-PPAR agonist with high affinity towards PPARα. Hepatic gene expression of PPARα, PPARβ/δ and the enzymes involved in the choline oxidation pathway were analyzed and concentrations of metabolites were analyzed in plasma and urine. TTA treatment altered most biomarkers, and the largest effect sizes were observed for plasma concentrations of dimethylglycine, nicotinamide, methylnicotinamide, methylmalonic acid and pyridoxal, which were all higher in the TTA group (all p < 0.01). Hepatic Pparα mRNA was increased after TTA treatment, but genes of the choline oxidation pathway were not affected. Long-term TTA treatment was associated with pronounced alterations on the plasma and urinary concentrations of metabolites related to one-carbon metabolism and B-vitamin status in rats. PMID:26742069

  11. Peroxisome Proliferator-Activated Receptor Activation is Associated with Altered Plasma One-Carbon Metabolites and B-Vitamin Status in Rats.

    PubMed

    Lysne, Vegard; Strand, Elin; Svingen, Gard F T; Bjørndal, Bodil; Pedersen, Eva R; Midttun, Øivind; Olsen, Thomas; Ueland, Per M; Berge, Rolf K; Nygård, Ottar

    2016-01-01

    Plasma concentrations of metabolites along the choline oxidation pathway have been linked to increased risk of major lifestyle diseases, and peroxisome proliferator-activated receptors (PPARs) have been suggested to be involved in the regulation of key enzymes along this pathway. In this study, we investigated the effect of PPAR activation on circulating and urinary one-carbon metabolites as well as markers of B-vitamin status. Male Wistar rats (n = 20) received for 50 weeks either a high-fat control diet or a high-fat diet with tetradecylthioacetic acid (TTA), a modified fatty acid and pan-PPAR agonist with high affinity towards PPARα. Hepatic gene expression of PPARα, PPARβ/δ and the enzymes involved in the choline oxidation pathway were analyzed and concentrations of metabolites were analyzed in plasma and urine. TTA treatment altered most biomarkers, and the largest effect sizes were observed for plasma concentrations of dimethylglycine, nicotinamide, methylnicotinamide, methylmalonic acid and pyridoxal, which were all higher in the TTA group (all p < 0.01). Hepatic Pparα mRNA was increased after TTA treatment, but genes of the choline oxidation pathway were not affected. Long-term TTA treatment was associated with pronounced alterations on the plasma and urinary concentrations of metabolites related to one-carbon metabolism and B-vitamin status in rats. PMID:26742069

  12. Altered retinoic acid signalling underpins dentition evolution.

    PubMed

    Gibert, Yann; Samarut, Eric; Pasco-Viel, Emmanuel; Bernard, Laure; Borday-Birraux, Véronique; Sadier, Alexa; Labbé, Catherine; Viriot, Laurent; Laudet, Vincent

    2015-03-01

    Small variations in signalling pathways have been linked to phenotypic diversity and speciation. In vertebrates, teeth represent a reservoir of adaptive morphological structures that are prone to evolutionary change. Cyprinid fish display an impressive diversity in tooth number, but the signals that generate such diversity are unknown. Here, we show that retinoic acid (RA) availability influences tooth number size in Cyprinids. Heterozygous adult zebrafish heterozygous for the cyp26b1 mutant that encodes an enzyme able to degrade RA possess an extra tooth in the ventral row. Expression analysis of pharyngeal mesenchyme markers such as dlx2a and lhx6 shows lateral, anterior and dorsal expansion of these markers in RA-treated embryos, whereas the expression of the dental epithelium markers dlx2b and dlx3b is unchanged. Our analysis suggests that changes in RA signalling play an important role in the diversification of teeth in Cyprinids. Our work illustrates that through subtle changes in the expression of rate-limiting enzymes, the RA pathway is an active player of tooth evolution in fish. PMID:25652838

  13. Altered retinoic acid signalling underpins dentition evolution

    PubMed Central

    Gibert, Yann; Samarut, Eric; Pasco-Viel, Emmanuel; Bernard, Laure; Borday-Birraux, Véronique; Sadier, Alexa; Labbé, Catherine; Viriot, Laurent; Laudet, Vincent

    2015-01-01

    Small variations in signalling pathways have been linked to phenotypic diversity and speciation. In vertebrates, teeth represent a reservoir of adaptive morphological structures that are prone to evolutionary change. Cyprinid fish display an impressive diversity in tooth number, but the signals that generate such diversity are unknown. Here, we show that retinoic acid (RA) availability influences tooth number size in Cyprinids. Heterozygous adult zebrafish heterozygous for the cyp26b1 mutant that encodes an enzyme able to degrade RA possess an extra tooth in the ventral row. Expression analysis of pharyngeal mesenchyme markers such as dlx2a and lhx6 shows lateral, anterior and dorsal expansion of these markers in RA-treated embryos, whereas the expression of the dental epithelium markers dlx2b and dlx3b is unchanged. Our analysis suggests that changes in RA signalling play an important role in the diversification of teeth in Cyprinids. Our work illustrates that through subtle changes in the expression of rate-limiting enzymes, the RA pathway is an active player of tooth evolution in fish. PMID:25652838

  14. Characterisation of metabolites of the putative cancer chemopreventive agent quercetin and their effect on cyclo-oxygenase activity

    PubMed Central

    Jones, D J L; Lamb, J H; Verschoyle, R D; Howells, L M; Butterworth, M; Lim, C K; Ferry, D; Farmer, P B; Gescher, A J

    2004-01-01

    Quercetin (3,5,7,3′,4′-pentahydroxyflavone) is a flavone with putative ability to prevent cancer and cardiovascular diseases. Its metabolism was evaluated in rats and human. Rats received quercetin via the intravenous (i.v.) route and metabolites were isolated from the plasma, urine and bile. Analysis was by high-performance liquid chromatography and confirmation of species identity was achieved by mass spectrometry. Quercetin and isorhamnetin, the 3′-O-methyl analogue, were found in both the plasma and urine. In addition, several polar peaks were characterised as sulphated and glucuronidated conjugates of quercetin and isorhamnetin. Extension of the metabolism studies to a cancer patient who had received quercetin as an i.v. bolus showed that (Quercetin removed) isorhamnetin and quercetin 3′-O-sulphate were major plasma metabolites. As a catechol, quercetin can potentially be converted to a quinone and subsequently conjugated with glutathione (GSH). Oxidation of quercetin with mushroom tyrosinase in the presence of GSH furnished GSH conjugates of quercetin, two mono- and one bis-substituted conjugates. However, these species were not found in biomatrices in rats treated with quercetin. As cyclo-oxygenase-2 (COX-2) expression is mechanistically linked to carcinogenesis, we examined whether quercetin and its metabolites can inhibit COX-2 in a human colorectal cancer cell line (HCA-7). Isorhamnetin and its 4′-isomer tamarixetin were potent inhibitors, reflected in a 90% decrease in prostaglandin E-2 (PGE-2) levels, a marker of COX-2 activity. Quercetin was less effective, with a 50% decline. Quercetin 3- and 7-O-sulphate had no effect on PGE-2. The results indicate that quercetin may exert its pharmacological effects, at least in part, via its metabolites. PMID:15292928

  15. Reduced photoinhibition under low irradiance enhanced Kacip Fatimah (Labisia pumila Benth) secondary metabolites, phenyl alanine lyase and antioxidant activity.

    PubMed

    Ibrahim, Mohd Hafiz; Jaafar, Hawa Z E

    2012-01-01

    A randomized complete block design experiment was designed to characterize the relationship between production of total flavonoids and phenolics, anthocyanin, photosynthesis, maximum efficiency of photosystem II (Fv/Fm), electron transfer rate (Fm/Fo), phenyl alanine lyase activity (PAL) and antioxidant (DPPH) in Labisia pumila var. alata, under four levels of irradiance (225, 500, 625 and 900 μmol/m(2)/s) for 16 weeks. As irradiance levels increased from 225 to 900 μmol/m(2)/s, the production of plant secondary metabolites (total flavonoids, phenolics and antocyanin) was found to decrease steadily. Production of total flavonoids and phenolics reached their peaks under 225 followed by 500, 625 and 900 μmol/m(2)/s irradiances. Significant positive correlation of production of total phenolics, flavonoids and antocyanin content with Fv/Fm, Fm/Fo and photosynthesis indicated up-regulation of carbon-based secondary metabolites (CBSM) under reduced photoinhibition on the under low light levels condition. At the lowest irradiance levels, Labisia pumila extracts also exhibited a significantly higher antioxidant activity (DPPH) than under high irradiance. The improved antioxidative activity under low light levels might be due to high availability of total flavonoids, phenolics and anthocyanin content in the plant extract. It was also found that an increase in the production of CBSM was due to high PAL activity under low light, probably signifying more availability of phenylalanine (Phe) under this condition. PMID:22754297

  16. Reduced Photoinhibition under Low Irradiance Enhanced Kacip Fatimah (Labisia pumila Benth) Secondary Metabolites, Phenyl Alanine Lyase and Antioxidant Activity

    PubMed Central

    Ibrahim, Mohd Hafiz; Jaafar, Hawa Z.E.

    2012-01-01

    A randomized complete block design experiment was designed to characterize the relationship between production of total flavonoids and phenolics, anthocyanin, photosynthesis, maximum efficiency of photosystem II (Fv/Fm), electron transfer rate (Fm/Fo), phenyl alanine lyase activity (PAL) and antioxidant (DPPH) in Labisia pumila var. alata, under four levels of irradiance (225, 500, 625 and 900 μmol/m2/s) for 16 weeks. As irradiance levels increased from 225 to 900 μmol/m2/s, the production of plant secondary metabolites (total flavonoids, phenolics and antocyanin) was found to decrease steadily. Production of total flavonoids and phenolics reached their peaks under 225 followed by 500, 625 and 900 μmol/m2/s irradiances. Significant positive correlation of production of total phenolics, flavonoids and antocyanin content with Fv/Fm, Fm/Fo and photosynthesis indicated up-regulation of carbon-based secondary metabolites (CBSM) under reduced photoinhibition on the under low light levels condition. At the lowest irradiance levels, Labisia pumila extracts also exhibited a significantly higher antioxidant activity (DPPH) than under high irradiance. The improved antioxidative activity under low light levels might be due to high availability of total flavonoids, phenolics and anthocyanin content in the plant extract. It was also found that an increase in the production of CBSM was due to high PAL activity under low light, probably signifying more availability of phenylalanine (Phe) under this condition. PMID:22754297

  17. Anti-onchocerca Metabolites from Cyperus articulatus: Isolation, In Vitro Activity and In Silico 'Drug-Likeness'.

    PubMed

    Metuge, Jonathan Alunge; Babiaka, Smith B; Mbah, James A; Ntie-Kang, Fidele; Ayimele, Godfred A; Cho-Ngwa, Fidelis

    2014-08-01

    The aims of this investigation were to isolate active ingredients from the roots/rhizomes of Cyperus articulatus used as herbal medicine in Cameroon for the treatment of human onchocerciasis and to assess the efficacy of the metabolites on the Onchocerca worm. The antifilarial activity was evaluated in vitro on microfilariae (Mfs) and adult worms of the bovine derived Onchocerca ochengi, a close relative of Onchocerca volvulus. Cytotoxicity was assessed in vitro on monkey kidney epithelial cells. The structures of the active compounds were determined using spectroscopic methods and their drug-likeness evaluated using Lipinski parameters. Two secondary metabolites, AMJ1 [containing mustakone (1) as the major component] and linoleic acid or (9Z,12Z)-octadeca-9,12-dienoic acid (2) were isolated. Both compounds were found to kill both the microfilariae and adult worms of O. ochengi in a dose dependent manner. The IC50s for AMJ1 were 15.7 µg/mL for Mfs, 17.4 µg/mL for adult males and 21.9 µg/mL for adult female worms while for linoleic acid the values were, 15.7 µg/mL for Mfs, 31.0 µg/mL for adult males and 44.2 µg/mL for adult females. The present report provides the first ever evidence of the anti-Onchocerca efficacy of AMJ1 and linoleic acid. Thus, these secondary metabolites may provide a lead for design and development of new antifilarial agents. PMID:25089243

  18. Effectiveness of clopidogrel dose escalation to normalize active metabolite exposure and antiplatelet effects in CYP2C19 poor metabolizers.

    PubMed

    Horenstein, Richard B; Madabushi, Rajnikanth; Zineh, Issam; Yerges-Armstrong, Laura M; Peer, Cody J; Schuck, Robert N; Figg, William Douglas; Shuldiner, Alan R; Pacanowski, Michael A

    2014-08-01

    Carriers of two copies of the loss-of-function CYP2C19*2 variant convert less clopidogrel into its active metabolite, resulting in diminished antiplatelet responses and higher cardiovascular event rates. To evaluate whether increasing the daily clopidogrel dose in poor metabolizers (PM) overcomes the effect of the CYP2C19 * 2 variant, we enrolled 18 healthy participants in a genotype-stratified, multi-dose, three-period, fixed-sequence crossover study. Six participants with the *1/*1 extensive (EM), *1/*2 intermediate (IM), and *2/*2 poor metabolizer genotypes each received 75 mg, 150 mg, and 300 mg each for 8 days. In each period, maximal platelet aggregation 4 hours post-dose (MPA4) and active metabolite area under the curve (AUC) differed among genotype groups (P < .05 for all). At day 8, PMs needed 300 mg daily and IMs needed 150 mg daily to attain a similar MPA4 as EMs on the 75 mg dose (32.6%, 33.2%, 31.3%, respectively). Similarly, PMs needed 300 mg daily to achieve active metabolite concentrations that were similar to EMs on 75 mg (AUC 37.7 and 33.5 ng h/mL, respectively). These results suggest that quadrupling the usual clopidogrel dose might be necessary to overcome the effect of poor CYP2C19 metabolism. PMID:24710841

  19. Multiple modes of inhibition of human cytochrome P450 2J2 by dronedarone, amiodarone and their active metabolites.

    PubMed

    Karkhanis, Aneesh; Lam, Hui Yuan; Venkatesan, Gopalakrishnan; Koh, Siew Kwan; Chai, Christina Li Lin; Zhou, Lei; Hong, Yanjun; Kojodjojo, Pipin; Chan, Eric Chun Yong

    2016-05-01

    Dronedarone, a multiple ion channel blocker is prescribed for the treatment of paroxysmal and persistent atrial fibrillation. While dronedarone does not precipitate toxicities like its predecessor amiodarone, its clinical use has been associated with idiosyncratic hepatic and cardiac adverse effects and drug-drug interactions (DDIs). As dronedarone is a potent mechanism-based inactivator of CYP3A4 and CYP3A5, a question arose if it exerts a similar inhibitory effect on CYP2J2, a prominent cardiac CYP450 enzyme. In this study, we demonstrated that CYP2J2 is reversibly inhibited by dronedarone (Ki=0.034μM), amiodarone (Ki=4.8μM) and their respective pharmacologically active metabolites namely N-desbutyldronedarone (NDBD) (Ki=0.55μM) and N-desethylamiodarone (NDEA) (Ki=7.4μM). Moreover, time-, concentration- and NADPH-dependent irreversible inactivation of CYP2J2 was investigated where inactivation kinetic parameters (KI, kinact) and partition ratio (r) of dronedarone (0.05μM, 0.034min(-1), 3.3), amiodarone (0.21μM, 0.015min(-1), 20.7) and NDBD (0.48μM, 0.024min(-1), 21.7) were observed except for NDEA. The absence of the characteristic Soret peak, lack of recovery of CYP2J2 activity upon dialysis, and biotransformation of dronedarone and NDBD to quinone-oxime reactive metabolites further confirmed the irreversible inactivation of CYP2J2 by dronedarone and NDBD is via the covalent adduction of CYP2J2. Our novel findings illuminate the possible mechanisms of DDIs and cardiac adverse effects due to both reversible inhibition and irreversible inactivation of CYP2J2 by dronedarone, amiodarone and their active metabolites. PMID:26972388

  20. Reactive metabolites and agranulocytosis.

    PubMed

    Uetrecht, J P

    1996-01-01

    Central to most hypotheses of the mechanism of idiosyncratic drug-induced blood dyscrasias is the involvement of reactive metabolites. In view of the reactive nature of the majority of such metabolites, it is likely that they are formed by, or in close proximity to the blood cells affected. The major oxidative system of neutrophils generates hypochlorous acid. We have demonstrated that the drugs associated with the highest incidence of agranulocytosis are oxidized to reactive metabolites by hypochlorous acid and/or activated neutrophils. There are many mechanisms by which such reactive metabolites could induce agranulocytosis. In the case of aminopyrine-induced agranulocytosis, most cases appear to involve drug-dependent anti-neutrophil antibodies, and these are likely to be induced by cell membrane antigens modified by the reactive metabolite of aminopyrine. The target of agranulocytosis associated with many other drugs is usually neutrophil precursors and may involve cytotoxicity or a cell-mediated immune reaction induced by a reactive metabolite. In the case of aplastic anaemia, there is evidence in some cases for involvement of cytotoxic T cells, which could either be induced by metabolites generated by neutrophils, or more likely, by reactive metabolites generated by stem cells. PMID:8987247

  1. In vivo anti-inflammatory activity of some naturally occurring O- and N-prenyl secondary metabolites.

    PubMed

    Epifano, Francesco; Genovese, Salvatore; Fiorito, Serena; della Loggia, Roberto; Tubaro, Aurelia; Sosa, Silvio

    2014-01-01

    A series of O- and N-prenyl secondary metabolites of insect, fungal, and plant origin have been evaluated for their topical anti-inflammatory activity using the Croton oil ear test in mice as a model of acute inflammation. Some of the tested compounds revealed an effect (ID50 = 0.31 divided by 0.56 micromol/cm2) comparable with that of the reference non-steroidal anti-inflammatory drug indomethacin (ID50 = 0.23 micromol/cm2). PMID:24660470

  2. Prostaglandin endoperoxide synthetase and the activation of benzo(a)pyrene to reactive metabolites in vivo in guinea pigs

    SciTech Connect

    Garattini, E.; Coccia, P.; Romano, M.; Jiritano, L.; Noseda, A.; Salmona, M.

    1984-11-01

    The role of prostaglandin endoperoxide synthetase in the in vivo activation of benzo(a)pyrene to reactive metabolites capable of interacting irreversibly with cellular macromolecules was studied in guinea pig liver, lung, kidney, spleen, small intestine, colon, and brain. DNA and protein covalent binding experiments were made after systemic administration of acetylsalicylic acid (200 mg/kg) followed by radiolabeled benzo(a)pyrene (4 microgram/kg). Results are compared with a control situation in which the prostaglandin endoperoxide synthetase inhibitor (acetylsalicylic acid) was not administered. No decrease in the level of DNA or protein benzo(a)pyrene-derived covalent binding was observed in any of the tissues studied.

  3. Transthyretin Binding Heterogeneity and Anti-amyloidogenic Activity of Natural Polyphenols and Their Metabolites.

    PubMed

    Florio, Paola; Folli, Claudia; Cianci, Michele; Del Rio, Daniele; Zanotti, Giuseppe; Berni, Rodolfo

    2015-12-11

    Transthyretin (TTR) is an amyloidogenic protein, the amyloidogenic potential of which is enhanced by a number of specific point mutations. The ability to inhibit TTR fibrillogenesis is known for several classes of compounds, including natural polyphenols, which protect the native state of TTR by specifically interacting with its thyroxine binding sites. Comparative analyses of the interaction and of the ability to protect the TTR native state for polyphenols, both stilbenoids and flavonoids, and some of their main metabolites have been carried out. A main finding of this investigation was the highly preferential binding of resveratrol and thyroxine, both characterized by negative binding cooperativity, to distinct sites in TTR, consistent with the data of x-ray analysis of TTR in complex with both ligands. Although revealing the ability of the two thyroxine binding sites of TTR to discriminate between different ligands, this feature has allowed us to evaluate the interactions of polyphenols with both resveratrol and thyroxine preferential binding sites, by using resveratrol and radiolabeled T4 as probes. Among flavonoids, genistein and apigenin were able to effectively displace resveratrol from its preferential binding site, whereas genistein also showed the ability to interact, albeit weakly, with the preferential thyroxine binding site. Several glucuronidated polyphenol metabolites did not exhibit significant competition for resveratrol and thyroxine preferential binding sites and lacked the ability to stabilize TTR. However, resveratrol-3-O-sulfate was able to significantly protect the protein native state. A rationale for the in vitro properties found for polyphenol metabolites was provided by x-ray analysis of their complexes with TTR. PMID:26468275

  4. Plant Polyphenols and Oxidative Metabolites of the Herbal Alkenylbenzene Methyleugenol Suppress Histone Deacetylase Activity in Human Colon Carcinoma Cells

    PubMed Central

    Groh, Isabel Anna Maria; Chen, Chen; Lüske, Claudia; Cartus, Alexander Thomas; Esselen, Melanie

    2013-01-01

    Evidence has been provided that diet and environmental factors directly influence epigenetic mechanisms associated with cancer development in humans. The inhibition of histone deacetylase (HDAC) activity and the disruption of the HDAC complex have been recognized as a potent strategy for cancer therapy and chemoprevention. In the present study, we investigated whether selected plant constituents affect HDAC activity or HDAC1 protein status in the human colon carcinoma cell line HT29. The polyphenols (−)-epigallocatechin-3-gallate (EGCG) and genistein (GEN) as well as two oxidative methyleugenol (ME) metabolites were shown to inhibit HDAC activity in intact HT29 cells. Concomitantly, a significant decrease of the HDAC1 protein level was observed after incubation with EGCG and GEN, whereas the investigated ME metabolites did not affect HDAC1 protein status. In conclusion, dietary compounds were found to possess promising HDAC-inhibitory properties, contributing to epigenetic alterations in colon tumor cells, which should be taken into account in further risk/benefit assessments of polyphenols and alkenylbenzenes. PMID:23476753

  5. Magnolia Extract, Magnolol, and Metabolites: Activation of Cannabinoid CB2 Receptors and Blockade of the Related GPR55

    PubMed Central

    2012-01-01

    The bark of Magnolia officinalis is used in Asian traditional medicine for the treatment of anxiety, sleeping disorders, and allergic diseases. We found that the extract and its main bioactive constituents, magnolol and honokiol, can activate cannabinoid (CB) receptors. In cAMP accumulation studies, magnolol behaved as a partial agonist (EC50 = 3.28 μM) with selectivity for the CB2 subtype, while honokiol was less potent showing full agonistic activity at CB1 and antagonistic properties at CB2. We subsequently synthesized the major metabolites of magnolol and found that tetrahydromagnolol (7) was 19-fold more potent than magnolol (EC50 CB2 = 0.170 μM) exhibiting high selectivity versus CB1. Additionally, 7 behaved as an antagonist at GPR55, a CB-related orphan receptor (KB = 13.3 μM, β-arrestin translocation assay). Magnolol and its metabolites may contribute to the biological activities of Magnolia extract via the observed mechanisms of action. Furthermore, the biphenylic compound magnolol provides a simple novel lead structure for the development of agonists for CB receptors and antagonists for the related GPR55. PMID:24900561

  6. Plant polyphenols and oxidative metabolites of the herbal alkenylbenzene methyleugenol suppress histone deacetylase activity in human colon carcinoma cells.

    PubMed

    Groh, Isabel Anna Maria; Chen, Chen; Lüske, Claudia; Cartus, Alexander Thomas; Esselen, Melanie

    2013-01-01

    Evidence has been provided that diet and environmental factors directly influence epigenetic mechanisms associated with cancer development in humans. The inhibition of histone deacetylase (HDAC) activity and the disruption of the HDAC complex have been recognized as a potent strategy for cancer therapy and chemoprevention. In the present study, we investigated whether selected plant constituents affect HDAC activity or HDAC1 protein status in the human colon carcinoma cell line HT29. The polyphenols (-)-epigallocatechin-3-gallate (EGCG) and genistein (GEN) as well as two oxidative methyleugenol (ME) metabolites were shown to inhibit HDAC activity in intact HT29 cells. Concomitantly, a significant decrease of the HDAC1 protein level was observed after incubation with EGCG and GEN, whereas the investigated ME metabolites did not affect HDAC1 protein status. In conclusion, dietary compounds were found to possess promising HDAC-inhibitory properties, contributing to epigenetic alterations in colon tumor cells, which should be taken into account in further risk/benefit assessments of polyphenols and alkenylbenzenes. PMID:23476753

  7. Top-down Targeted Metabolomics Reveals a Sulfur-Containing Metabolite with Inhibitory Activity against Angiotensin-Converting Enzyme in Asparagus officinalis.

    PubMed

    Nakabayashi, Ryo; Yang, Zhigang; Nishizawa, Tomoko; Mori, Tetsuya; Saito, Kazuki

    2015-05-22

    The discovery of bioactive natural compounds containing sulfur, which is crucial for inhibitory activity against angiotensin-converting enzyme (ACE), is a challenging task in metabolomics. Herein, a new S-containing metabolite, asparaptine (1), was discovered in the spears of Asparagus officinalis by targeted metabolomics using mass spectrometry for S-containing metabolites. The contribution ratio (2.2%) to the IC50 value in the crude extract showed that asparaptine (1) is a new ACE inhibitor. PMID:25922884

  8. Excretion of tectoridin metabolites in rat urine and bile orally administrated at different dosages and their inhibitory activity against aldose reductase.

    PubMed

    Qu, Jialin; Wu, Zhizhen; Gao, Jie; Wen, Hao; Wang, Tao; Yuan, Dan

    2014-12-01

    This study investigated the urinary and biliary excretion of tectoridin, a major active isoflavonoid found in the flowers of Pueraria thomsonii Benth. and the rhizomes of Belamcanda chinensis (L.) DC. Using UHPLC/Q-TOFMS, seven glucuronides and/or sulfated metabolites and four Phase I metabolites were simultaneously quantified in rat urine after oral administration of tectoridin at 100 and 200 mg/kg. Over a 72-h period, 14.2% and 14.7% of the tectoridin were excreted as eleven metabolites in urine, among which, two major metabolites tectorigenin-7-O-β-D-glucuronide (Te-7G) and tectorigenin accounted for 5.5-5.5% and 4.3-4.4%. Furthermore, the cumulative excretion of four glucuronides and sulfated metabolites in bile accounted for 7.3% and 3.9% of the dose within 60 h, among which, Te-7G and tectorigenin-7-O-glucuronide-4'-O-sulfate (Te-7G-4'S) accounted for 2.3-3.0% and 1.4-3.9%, respectively. The results indicate that the urine was the primary elimination route, and glucuronidation after deglycosylation at C-7 position was the major metabolic pathway of tectoridin in vivo. Moreover, the inhibitory activities of tectoridin and its five metabolites on rat lens aldose reductase were confirmed (IC₅₀: 1.4-15.5 μM), whereas irisolidone-7-O-glucuronide (Ir-7G) and irisolidone showed little activity. PMID:25256063

  9. Activity and characterization of secondary metabolites produced by a new microorganism for control of plant diseases.

    PubMed

    Ko, Wen-Hsiung; Tsou, Yi-Jung; Lin, Mei-Ju; Chern, Lih-Ling

    2010-09-30

    Microorganisms capable of utilizing vegetable tissues for growth in soils were isolated and their vegetable broth cultures were individually sprayed directly on leaves to test their ability to control Phytophthora blight of bell pepper caused by Phytophthora capsici. Liquid culture of Streptomyces strain TKA-5, a previously undescribed species obtained in this study, displayed several desirable disease control characteristics in nature, including high potency, long lasting and ability to control also black leaf spot of spoon cabbage caused by Alternaria brassicicolca. The extract was fungicidal to P. capsici but fungistatic to A. brassicicola. It was stable at high temperature and high pH. However, after exposure to pH 2 for 24h, the extract was no longer inhibitory to P. capsici although it was still strongly inhibitory to A. brassicicola. After treatment with cation or anion exchange resins, the extract lost its inhibitory effect against P. capsici but not A. brassicicola. The results suggest that the extract contained two different kinds of inhibitory metabolites, one against P. capsici with both positive and negative charges on its molecule and another against A. brassicicola with no charges on its molecule. The inhibitory metabolites were soluble in ethanol or methanol but not in water, ether or chloroform. They were dialyzable in the membrane tubing with molecular weight cut-off of 10,000, 1000 or 500 but not 100, indicating that the inhibitors have a molecular weight between 500 and 100. Results also showed that both inhibitors are not proteins. PMID:20580869

  10. Activation of the Silent Secondary Metabolite Production by Introducing Neomycin-Resistance in a Marine-Derived Penicillium purpurogenum G59

    PubMed Central

    Wu, Chang-Jing; Yi, Le; Cui, Cheng-Bin; Li, Chang-Wei; Wang, Nan; Han, Xiao

    2015-01-01

    Introduction of neomycin-resistance into a marine-derived, wild-type Penicillium purpurogenum G59 resulted in activation of silent biosynthetic pathways for the secondary metabolite production. Upon treatment of G59 spores with neomycin and dimethyl sulfoxide (DMSO), a total of 56 mutants were obtained by single colony isolation. The acquired resistance of mutants to neomycin was testified by the resistance test. In contrast to the G59 strain, the EtOAc extracts of 28 mutants inhibited the human cancer K562 cells, indicating that the 28 mutants have acquired the capability to produce bioactive metabolites. HPLC-photodiode array detector (PDAD)-UV and HPLC-electron spray ionization (ESI)-MS analyses further indicated that diverse secondary metabolites have been newly produced in the bioactive mutant extracts. Followed isolation and characterization demonstrated that five bioactive secondary metabolites, curvularin (1), citrinin (2), penicitrinone A (3), erythro-23-O-methylneocyclocitrinol (4) and 22E-7α-methoxy-5α,6α-epoxyergosta-8(14),22-dien-3β-ol (5), were newly produced by a mutant, 4-30, compared to the G59 strain. All 1–5 were also not yet found in the secondary metabolites of other wild type P. purpurogenum strains. Compounds 1–5 inhibited human cancer K562, HL-60, HeLa and BGC-823 cells to varying extents. Both present bioassays and chemical investigations demonstrated that the introduction of neomycin-resistance into the marine-derived fungal G59 strain could activate silent secondary metabolite production. The present work not only extended the previous DMSO-mediated method for introducing drug-resistance in fungi both in DMSO concentrations and antibiotics, but also additionally exemplified effectiveness of this method for activating silent fungal secondary metabolites. This method could be applied to other fungal isolates to elicit their metabolic potentials to investigate secondary metabolites from silent biosynthetic pathways. PMID:25913704

  11. Isolation and Identification of Twelve Metabolites of Isocorynoxeine in Rat Urine and their Neuroprotective Activities in HT22 Cell Assay

    PubMed Central

    Qi, Wen; Chen, Fangfang; Sun, Jiahong; Simpkins, James W.; Yuan, Dan

    2015-01-01

    Isocorynoxeine, one of the major alkaloids from Uncaria Hook, shows the effects of lowering blood pressure, vasodilatation, and protection against ischemia-induced neuronal damage. In this paper, the metabolism of isocorynoxeine was investigated in rats. Twelve metabolites and the parent drug were isolated by using solvent extraction and repeated chromatographic methods, and determined by spectroscopic methods including UV, MS, NMR, and CD experiments. Seven new compounds were identified as 11-hydroxyisocorynoxeine, 5-oxoisocorynoxeinic acid-22-O-β-D-glucuronide, 10-hydroxyisocorynoxeine, 17-O-demethyl-16,17-dihydro-5-oxoisocorynoxeine, 5-oxoisocorynoxeinic acid, 21-hydroxy-5-oxoisocorynoxeine, and oxireno[18,19]-5-oxoisocorynoxeine, together with six known compounds identified as isocorynoxeine, 18,19-dehydrocorynoxinic acid, 18,19-dehydrocorynoxinic acid B, corynoxeine, isocorynoxeine-N-oxide, and corynoxeine-N-oxide. Possible metabolic pathways of isocorynoxeine are proposed. Furthermore, the activity assay for the parent drug and some of its metabolites showed that isocorynoxeine exhibited a significant neuroprotective effect against glutamate-induced HT22 cell death at the maximum concentration. However, little or weak neuroprotective activities were observed for M-3, M-6, M-7, and M-10. Our present study is important to further understand their metabolic fate and disposition in humans. PMID:25519834

  12. Isolation and identification of twelve metabolites of isocorynoxeine in rat urine and their neuroprotective activities in HT22 cell assay.

    PubMed

    Qi, Wen; Chen, Fangfang; Sun, Jiahong; Simpkins, James W; Yuan, Dan

    2015-01-01

    Isocorynoxeine, one of the major alkaloids from Uncaria Hook, shows the effects of lowering blood pressure, vasodilatation, and protection against ischemia-induced neuronal damage. In this paper, the metabolism of isocorynoxeine was investigated in rats. Twelve metabolites and the parent drug were isolated by using solvent extraction and repeated chromatographic methods, and determined by spectroscopic methods including UV, MS, NMR, and CD experiments. Seven new compounds were identified as 11-hydroxyisocorynoxeine, 5-oxoisocorynoxeinic acid-22-O-β-D-glucuronide, 10-hydroxyisocorynoxeine, 17-O-demethyl-16,17-dihydro-5-oxoisocorynoxeine, 5-oxoisocorynoxeinic acid, 21-hydroxy-5-oxoisocorynoxeine, and oxireno[18, 19]-5-oxoisocorynoxeine, together with six known compounds identified as isocorynoxeine, 18,19-dehydrocorynoxinic acid, 18,19-dehydrocorynoxinic acid B, corynoxeine, isocorynoxeine-N-oxide, and corynoxeine-N-oxide. Possible metabolic pathways of isocorynoxeine are proposed. Furthermore, the activity assay for the parent drug and some of its metabolites showed that isocorynoxeine exhibited a significant neuroprotective effect against glutamate-induced HT22 cell death at the maximum concentration. However, little or weak neuroprotective activities were observed for M-3, M-6, M-7, and M-10. Our present study is important to further understand their metabolic fate and disposition in humans. PMID:25519834

  13. Anti-phytopathogenic activity of sporothriolide, a metabolite from endophyte Nodulisporium sp. A21 in Ginkgo biloba.

    PubMed

    Cao, Ling-Ling; Zhang, Ying-Ying; Liu, Ying-Jie; Yang, Ting-Ting; Zhang, Jin-Long; Zhang, Zheng-Guang; Shen, Li; Liu, Jun-Yan; Ye, Yong-Hao

    2016-05-01

    Phytopathogenic fungi such as Rhizoctonia solani and Sclerotinia sclerotiorum caused multiple plant diseases resulting in severe loss of crop production. Increasing documents endorsed that endophytes are a striking resource pool for numerous metabolites with various bioactivities such as anti-fungal. Here we reported the characterization and anti-phytopathogenic activity of sporothriolide, a metabolite produced by Nodulisporium sp. A21-an endophytic fungus in the leaves of Ginkgo biloba. Among the total twenty-five endophytic fungi isolated from the healthy leaves of G. biloba, the fermentation broth (FB) of the strain A21 was found potently inhibitory activity against R. solani and S. sclerotiorum using mycelia growth inhibition method. A21 was then identified as Nodulisporium sp., the asexual stage of Hypoxylon sp., by microscopic examination and ITS rDNA sequence data comparison. Under the bioassay-guided fractionation, sporothriolide was isolated from the petroleum ether extract of the FB of A21, whose structure was established by integrated interpretation of HR-ESI-MS and (1)H- and (13)C-NMR. Furthermore, the crystal structure of sporothriolide was first reported. In addition, sporothriolide was validated to be potently antifungal against R. solani, S. sclerotiorum and inhibit conidium germination of Magnaporthe oryzae in vitro and in vivo, indicating that it could be used as a lead compound for new fungicide development. PMID:27017876

  14. Mass spectrometry-based metabolite profiling and antioxidant activity of Aloe vera ( Aloe barbadensis Miller) in different growth stages.

    PubMed

    Lee, Sarah; Do, Seon-Gil; Kim, Sun Yeou; Kim, Jinwan; Jin, Yoojeong; Lee, Choong Hwan

    2012-11-14

    Metabolite profiling of four different-sized Aloe vera plants was performed using gas chromatography-ion trap-mass spectrometry (GC-IT-MS) and ultra performance liquid chromatography-quadrupole-time of flight-mass spectrometry (UPLC-Q-TOF-MS) with multivariate analysis. Amino acids, sugars, and organic acids related to growth and development were identified by sizes. In particular, the relative contents of glucose, fructose, alanine, valine, and aspartic acid increased gradually as the size of the aloe increased. Anthraquinone derivatives such as 7-hydroxy-8-O-methylaloin, 7-hydroxyaloin A, and 6'-malonylnataloins A and B increased gradually, whereas chromone derivatives decreased continuously as the size of the aloe increased. The A30 aloe (size = 20-30 cm) with relatively high contents of aloins A and B, was suggested to have antioxidant components showing the highest antioxidant activity among the four different sizes of aloe. These data suggested that MS-based metabolomic approaches can illuminate metabolite changes associated with growth and development and can explain their change of antioxidant activity. PMID:23050594

  15. Mechanistic toxicodynamic model for receptor-mediated toxicity of diazoxon, the active metabolite of diazinon, in Daphnia magna.

    PubMed

    Kretschmann, Andreas; Ashauer, Roman; Hitzfeld, Kristina; Spaak, Piet; Hollender, Juliane; Escher, Beate I

    2011-06-01

    The organothiophosphate diazinon inhibits the target site acetylcholinesterase only after activation to its metabolite diazoxon. Commonly, the toxicity of xenobiotics toward aquatic organisms is expressed as a function of the external concentration and the resulting effect on the individual level after fixed exposure times. This approach does not account for the time dependency of internal processes such as uptake, metabolism, and interaction of the toxicant with the target site. Here, we develop a mechanistic toxicodynamic model for Daphnia magna and diazoxon, which accounts for the inhibition of the internal target site acetylcholinesterase and its link to the observable effect, immobilization, and mortality. The model was parametrized by experiments performed in vitro with the active metabolite diazoxon on enzyme extracts and in vivo with the parent compound diazinon. The mechanism of acetylcholinesterase inhibition was shown to occur irreversibly in two steps via formation of a reversible enzyme-inhibitor complex. The corresponding kinetic parameters revealed a very high sensitivity of acetylcholinesterase from D. magna toward diazoxon, which corresponds well with the high toxicity of diazinon toward this species. Recovery of enzyme activity but no recovery from immobilization was observed after in vivo exposure to diazinon. The toxicodynamic model combining all in vitro and in vivo parameters was successfully applied to describe the time course of immobilization in dependence of acetylcholinesterase activity during exposure to diazinon. The threshold value for enzyme activity below which immobilization set in amounted to 40% of the control activity. Furthermore, the model enabled the prediction of the time-dependent diazoxon concentration directly present at the target site. PMID:21539304

  16. IDO1 Metabolites Activate β-catenin Signaling to Promote Cancer Cell Proliferation and Colon Tumorigenesis in Mice

    PubMed Central

    Thaker, Ameet I.; Rao, M Suprada; Bishnupuri, Kumar S.; Kerr, Thomas A; Foster, Lynne; Marinshaw, Jeffrey M.; Newberry, Rodney D.; Stenson, William F.; Ciorba, Matthew A

    2013-01-01

    BACKGROUND & AIMS Indoleamine 2,3 dioxygenase-1 (IDO1) catabolizes tryptophan along the kynurenine pathway. Though IDO1 is expressed in inflamed and neoplastic epithelial cells of the colon, its role in colon tumorigenesis is not well understood. We used genetic and pharmacologic approaches to manipulate IDO1 activity in mice with colitis-associated cancer and human colon cancer cell lines. METHODS C57Bl6 wild type (control), IDO1−/−, Rag1−/−, Rag1/IDO1 double knockout mice were exposed to azoxymethane and dextran sodium sulfate (DSS) to induce colitis and tumorigenesis. Colitis severity was assessed by measurements of disease activity, cytokine levels and histologic analysis. In vitro experiments were conducted using HCT116 and HT29 human colon cancer cells. 1-methyl tryptophan and small interfering RNA were used to inhibit IDO1. Kynurenine pathway metabolites were used to simulate IDO1 activity. RESULTS C57Bl6 mice given pharmacologic inhibitors of IDO1 and IDO1−/− mice had lower tumor burdens and reduced proliferation in the neoplastic epithelium following administration of DSS and azoxymethane than control mice. These reductions were also observed in Rag1/IDO1 double knockout mice compared to Rag1−/− mice (which lack mature adaptive immunity). In human colon cancer cells, blockade of IDO1 activity reduced nuclear and activated β-catenin, transcription of its target genes (cyclin D1 and Axin2), and ultimately proliferation. Exogenous administration of IDO1 pathway metabolites kynurenine and quinolinic acid led to activation of β-catenin and proliferation of human colon cancer cells, and increased tumor growth in mice. CONCLUSIONS IDO1, which catabolizes tryptophan, promotes colitis-associated tumorigenesis in mice, independent of its ability to limit T-cell mediated immune surveillance. The epithelial cell-autonomous survival advantage provided by IDO1 to colon epithelial cells indicate its potential as a therapeutic target. PMID:23669411

  17. 20(S)-protopanaxadiol, an active ginseng metabolite, exhibits strong antidepressant-like effects in animal tests.

    PubMed

    Xu, Changjiang; Teng, Jijun; Chen, Weidong; Ge, Qiang; Yang, Zhiqi; Yu, Chunying; Yang, Zirong; Jia, William

    2010-12-01

    Ginseng has been used for mood adjustment in traditional Chinese medicine for thousands of years. Our previous study has shown that, total ginsenosides, the major pharmacologically functional ingredients of ginseng, possess antidepressant activity. In the present study, we hypothesized that an intestinal metabolite of ginseng, 20(S)-protopanaxadiol (code name S111), as a post metabolism compound (PMC) of ingested ginsenosides, may be responsible for the antidepressant activity of ginseng. To test this hypothesis, antidepressant-like activity of orally given S111 was measured in animal tests including tail suspension test, forced swimming test and rat olfactory bulbectomy depression model. In all those tests, S111 demonstrated antidepressant-like activity as potent as fluoxetine. S111 treated bulbectomy animals had higher levels of monoamine neurotransmitters in the brain and in vitro reuptake assay showed that S111 had a mild inhibitory effect. Furthermore, S111 but not fluoxetine significantly reduced brain oxidative stress and down-regulated serum corticosterone concentration in bulbectomy animals. No disturbance to central nervous system (CNS) normal functions were found in S111 treated animals. These results suggest that the ginseng active metabolite S111 is a potential antidepressant. Since the monoamine reuptake activity of this compound is rather weak, it remains to be investigated whether its antidepressant-like effect is by mechanisms that are different from current antidepressants. Furthermore, this study has demonstrated that post metabolism compounds (PMCs) of herb medicines such as S111 may be a novel source for drug discovery from medicinal herbs. PMID:20647027

  18. Allocation of secondary metabolites, photosynthetic capacity, and antioxidant activity of Kacip Fatimah (Labisia pumila Benth) in response to CO2 and light intensity.

    PubMed

    Ibrahim, Mohd Hafiz; Jaafar, Hawa Z E; Karimi, Ehsan; Ghasemzadeh, Ali

    2014-01-01

    A split plot 3 by 4 experiment was designed to investigate and distinguish the relationships among production of secondary metabolites, soluble sugar, phenylalanine ammonia lyase (PAL; EC 4.3.1.5) activity, leaf gas exchange, chlorophyll content, antioxidant activity (DPPH), and lipid peroxidation under three levels of CO2 (400, 800, and 1200 μ mol/mol) and four levels of light intensity (225, 500, 625, and 900 μ mol/m(2)/s) over 15 weeks in Labisia pumila. The production of plant secondary metabolites, sugar, chlorophyll content, antioxidant activity, and malondialdehyde content was influenced by the interactions between CO2 and irradiance. The highest accumulation of secondary metabolites, sugar, maliondialdehyde, and DPPH activity was observed under CO2 at 1200 μ mol/mol + light intensity at 225 μ mol/m(2)/s. Meanwhile, at 400 μ mol/mol CO2 + 900 μ mol/m(2)/s light intensity the production of chlorophyll and maliondialdehyde content was the highest. As CO2 levels increased from 400 to 1200 μ mol/mol the photosynthesis, stomatal conductance, f v /f m (maximum efficiency of photosystem II), and PAL activity were enhanced. The production of secondary metabolites displayed a significant negative relationship with maliondialdehyde indicating lowered oxidative stress under high CO2 and low irradiance improved the production of plant secondary metabolites that simultaneously enhanced the antioxidant activity (DPPH), thus improving the medicinal value of Labisia pumila under this condition. PMID:24683336

  19. Allocation of Secondary Metabolites, Photosynthetic Capacity, and Antioxidant Activity of Kacip Fatimah (Labisia pumila Benth) in Response to CO2 and Light Intensity

    PubMed Central

    Jaafar, Hawa Z. E.; Karimi, Ehsan; Ghasemzadeh, Ali

    2014-01-01

    A split plot 3 by 4 experiment was designed to investigate and distinguish the relationships among production of secondary metabolites, soluble sugar, phenylalanine ammonia lyase (PAL; EC 4.3.1.5) activity, leaf gas exchange, chlorophyll content, antioxidant activity (DPPH), and lipid peroxidation under three levels of CO2 (400, 800, and 1200 μmol/mol) and four levels of light intensity (225, 500, 625, and 900 μmol/m2/s) over 15 weeks in Labisia pumila. The production of plant secondary metabolites, sugar, chlorophyll content, antioxidant activity, and malondialdehyde content was influenced by the interactions between CO2 and irradiance. The highest accumulation of secondary metabolites, sugar, maliondialdehyde, and DPPH activity was observed under CO2 at 1200 μmol/mol + light intensity at 225 μmol/m2/s. Meanwhile, at 400 μmol/mol CO2 + 900 μmol/m2/s light intensity the production of chlorophyll and maliondialdehyde content was the highest. As CO2 levels increased from 400 to 1200 μmol/mol the photosynthesis, stomatal conductance, fv/fm (maximum efficiency of photosystem II), and PAL activity were enhanced. The production of secondary metabolites displayed a significant negative relationship with maliondialdehyde indicating lowered oxidative stress under high CO2 and low irradiance improved the production of plant secondary metabolites that simultaneously enhanced the antioxidant activity (DPPH), thus improving the medicinal value of Labisia pumila under this condition. PMID:24683336

  20. Metabolic activation of tris(2,3-dibromopropyl)phosphate to reactive intermediates. II. Covalent binding, reactive metabolite formation, and differential metabolite-specific DNA damage in vivo.

    PubMed

    Pearson, P G; Omichinski, J G; Holme, J A; McClanahan, R H; Brunborg, G; Søderlund, E J; Dybing, E; Nelson, S D

    1993-02-01

    Analogs of tris(2,3-dibromopropyl)phosphate (Tris-BP) either labeled at specific positions with carbon-14 and phosphorus-32 or dual-labeled with both deuterium and tritium were administered to male Wistar rats at a nephrotoxic dose of 360 mumol/kg. The covalent binding of Tris-BP metabolites to hepatic, renal, and testicular proteins was determined after 9 and 24 hr, and plasma concentrations of bis(2,3-dibromopropyl)-phosphate (Bis-BP) formed metabolically from Tris-BP were measured at intervals throughout the initial 9-hr postdosing period. The covalent binding of 14C-Tris-BP metabolites in the kidney (2495 +/- 404 pmol/mg protein) was greater than that in the liver (476 +/- 123 pmol/mg protein) or testes (94 +/- 11 pmol/mg protein); the extent of renal covalent protein binding of Tris-BP metabolites was decreased by 82 and 84% when deuterium was substituted at carbon-2 and carbon-3, respectively. Substitution of Tris-BP with deuterium at carbon-2 or carbon-3 also decreased the mean area under the curve for Bis-BP plasma concentration by 48 and 57%, respectively. The mechanism of Tris-BP-induced renal and hepatic DNA damage was evaluated in Wistar rats by an automated alkaline elution procedure after the administration of analogs of Tris-BP or Bis-BP labeled at specific positions with deuterium. Renal DNA damage was decreased when Tris-BP was substituted with deuterium at either carbon-2 or carbon-3; the magnitude of the change correlated with both a decrease in the area under the Bis-BP plasma curve and a decrease in renal covalent binding of Tris-BP metabolites for each of the deuterated analogs. In marked contrast, analogs of Bis-BP labeled with deuterium at carbon-2 or carbon-3 did not show a decrease in the severity of renal DNA damage compared to unlabeled Bis-BP. On the basis of these observations a metabolic scheme for hepatic P-450-mediated oxidation at either carbon-2 or carbon-3 of Tris-BP affording Bis-BP by two alternate pathways that are susceptible

  1. The Oxidized Linoleic Acid Metabolite-Cytochrome P450 System is Active in Biopsies from Patients with Inflammatory Dental Pain

    PubMed Central

    Ruparel, Shivani; Hargreaves, Kenneth M.; Eskander, Michael; Rowan, Spencer; de Almeida, Jose F.A.; Roman, Linda; Henry, Michael A.

    2013-01-01

    Endogenous TRPV1 agonists such as oxidized linoleic acid metabolites (OLAMs) and the enzymes releasing them [e.g., cytochrome P450 (CYP)], are up-regulated following inflammation in the rat. However, it is not known if such agonists are elevated in human inflammatory pain conditions. Since TRPV1 is expressed in human dental pulp nociceptors, we hypothesized that OLAM-CYP machinery is active in this tissue type and is increased under painful inflammatory conditions such as irreversible pulpitis (IP). The aim of this study was to compare CYP expression and linoleic acid (LA) metabolism in normal versus inflamed human dental pulp. Our data showed that exogenous LA metabolism was significantly increased in IP tissues compared to normal tissues and that pretreatment with a CYP inhibitor, ketoconazole, significantly inhibited LA metabolism. Additionally, extracts obtained from LA-treated inflamed tissues, evoked significant inward currents in TG neurons, and were blocked by pretreatment with the TRPV1 antagonist, IRTX. Moreover, extracts obtained from ketoconazole-pretreated inflamed tissues significantly reduced inward currents in TG neurons. These data suggest that LA metabolites produced in human inflamed tissues act as TRPV1 agonists and that the metabolite production can be targeted by CYP inhibition. In addition, immunohistochemical analysis of two CYP isoforms, CYP2J and CYP3A1, were shown to be predominately expressed in immune cells infiltrating the inflamed dental pulp, emphasizing the paracrine role of CYP enzymes in OLAM regulation. Collectively, our data indicates that the machinery responsible for OLAM production is up-regulated during inflammation and can be targeted to develop potential analgesics for inflammatory-induced dental pain. PMID:23867730

  2. A retinoic acid receptor-specific element controls the retinoic acid receptor-beta promoter.

    PubMed

    Hoffmann, B; Lehmann, J M; Zhang, X K; Hermann, T; Husmann, M; Graupner, G; Pfahl, M

    1990-11-01

    The morphogen retinoic acid (RA) regulates gene transcription by interacting with specific nuclear receptors that recognize DNA sequences near responsive promoters. While much has recently been learned about the nuclear receptor proteins, little is known about the genes that are directly regulated by RA and their cis-acting response elements recognized by these receptors. Here we have analyzed the RA receptor-beta (RAR beta) gene promoter that is controlled by RA. We find that a RA-responsive element (RARE) is located adjacent to the TATA box. The RARE shows a direct repeat symmetry which is essential for its function. While thyroid hormone-responsive elements can also function as RAR response elements, we show here that this RARE is activated by endogenous RARs and RAR beta, but cannot be regulated by thyroid hormone receptors and other known nuclear receptors. In addition, we find that RAR gamma is a poor activator of this RARE. However, the response element is bound with high affinity by both RAR beta and RAR gamma as well as by thyroid hormone receptors. Thus, interaction between specific response elements and receptors is insufficient for gene activation. PMID:2177841

  3. Retinoic acid differentially affects in vitro proliferation, differentiation and mineralization of two fish bone-derived cell lines: different gene expression of nuclear receptors and ECM proteins.

    PubMed

    Fernández, Ignacio; Tiago, Daniel M; Laizé, Vincent; Leonor Cancela, M; Gisbert, Enric

    2014-03-01

    Retinoic acid (RA), the main active metabolite of vitamin A, regulates vertebrate morphogenesis through signaling pathways not yet fully understood. Such process involves the specific activation of retinoic acid and retinoid X receptors (RARs and RXRs), which are nuclear receptors of the steroid/thyroid hormone receptor superfamily. Teleost fish are suitable models to study vertebrate development, such as skeletogenesis. Cell systems capable of in vitro mineralization have been developed for several fish species and may provide new insights into the specific cellular and molecular events related to vitamin A activity in bone, complementary to in vivo studies. This work aims at investigating the in vitro effects of RA (0.5 and 12.5 μM) on proliferation, differentiation and extracellular matrix (ECM) mineralization of two gilthead seabream bone-derived cell lines (VSa13 and VSa16), and at identifying molecular targets of its action through gene expression analysis. RA induced phenotypic changes and cellular proliferation was inhibited in both cell lines in a cell type-dependent manner (36-59% in VSa13 and 17-46% in VSa16 cells). While RA stimulated mineral deposition in VSa13 cell cultures (50-62% stimulation), it inhibited the mineralization of extracellular matrix in VSa16 cells (11-57% inhibition). Expression of hormone receptor genes (rars and rxrs), and extracellular matrix-related genes such as matrix and bone Gla proteins (mgp and bglap), osteopontin (spp1) and type I collagen (col1a1) were differentially regulated upon exposure to RA in proliferating, differentiating and mineralizing cultures of VSa13 and VSa16 cells. Altogether, our results show: (i) RA affects proliferative and mineralogenic activities in two fish skeletal cell types and (ii) that during phenotype transitions, specific RA nuclear receptors and bone-related genes are differentially expressed in a cell type-dependent manner. PMID:24291400

  4. Current results on biological activities of lichen secondary metabolites: a review.

    PubMed

    Molnár, Katalin; Farkas, Edit

    2010-01-01

    Lichens are symbiotic organisms of fungi and algae or cyanobacteria. Lichen-forming fungi synthesize a great variety of secondary metabolites, many of which are unique. Developments in analytical techniques and experimental methods have resulted in the identification of about 1050 lichen substances (including those found in cultures). In addition to their role in lichen chemotaxonomy and systematics, lichen secondary compounds have several possible biological roles, including photoprotection against intense radiation, as well as allelochemical, antiviral, antitumor, antibacterial, antiherbivore, and antioxidant action. These compounds are also important factors in metal homeostasis and pollution tolerance of lichen thalli. Although our knowledge of the contribution of these extracellular products to the success of the lichen symbiosis has increased significantly in the last decades, their biotic and abiotic roles have not been entirely explored. PMID:20469633

  5. Protopanaxadiol, an Active Ginseng Metabolite, Significantly Enhances the Effects of Fluorouracil on Colon Cancer

    PubMed Central

    Wang, Chong-Zhi; Zhang, Zhiyu; Wan, Jin-Yi; Zhang, Chun-Feng; Anderson, Samantha; He, Xin; Yu, Chunhao; He, Tong-Chuan; Qi, Lian-Wen; Yuan, Chun-Su

    2015-01-01

    In this study, we evaluated the effects of protopanaxadiol (PPD), a gut microbiome induced ginseng metabolite, in increasing the anticancer effects of a chemotherapeutic agent fluorouracil (5-FU) on colorectal cancer. An in vitro HCT-116 colorectal cancer cell proliferation test was conducted to observe the effects of PPD, 5-FU and their co-administration and the related mechanisms of action. Then, an in vivo xenografted athymic mouse model was used to confirm the in vitro data. Our results showed that the human gut microbiome converted ginsenoside compound K to PPD as a metabolite. PPD and 5-FU significantly inhibited HCT-116 cell proliferation in a concentration-dependent manner (both p < 0.01), and the effects of 5-FU were very significantly enhanced by combined treatment with PPD (p < 0.01). Cell cycle evaluation demonstrated that 5-FU markedly induced the cancer cell S phase arrest, while PPD increased arrest in G1 phase. Compared to the control, 5-FU and PPD increased apoptosis, and their co-administration significantly increased the number of apoptotic cells (p < 0.01). Using bioluminescence imaging, in vivo data revealed that 5-FU significantly reduced the tumor growth up to Day 20 (p < 0.05). PPD and 5-FU co-administration very significantly reduced the tumor size in a dose-related manner (p < 0.01 compared to the 5-FU alone). The quantification of the tumor size and weight changes for 43 days supported the in vivo imaging data. Our results demonstrated that the co-administration of PPD and 5-FU significantly inhibited the tumor growth, indicating that PPD significantly enhanced the anticancer action of 5-FU, a commonly used chemotherapeutic agent. PPD may have a clinical value in 5-FU’s cancer therapeutics. PMID:25625815

  6. Anti-inflammatory and anti-hyperalgesic effect of all-trans retinoic acid in carrageenan-induced paw edema in Wistar rats: Involvement of peroxisome proliferator-activated receptor-β/δ receptors

    PubMed Central

    Gill, Navneet; Bijjem, Krishna Reddy V.; Sharma, Pyare L.

    2013-01-01

    Objective: In this study, we investigated the role of peroxisome proliferator-activated receptors (PPAR)-β/δ receptors in carrageenan-induced inflammation and in the anti-inflammatory effects of all-trans retinoic acid (ATRA). Materials and Methods: The λ-carrageenan (0.1 ml of 1% w/v) was injected into intra-plantar (i.pl.) region of the hind paw to produce acute inflammation. Paw volume was measured by using the mercury plethysmography. Further, mechanical and thermal hyperalgesia (TH) were assessed by using the dynamic plantar aesthesiometer and plantar test apparatus, respectively. In addition, markers of oxido-nitrosative stress were assessed spectrophotometrically in the hind paw tissue 5 h post-carrageenan. Results: An i.pl injection of carrageenan has produced a marked mechanical hyperalgesia (MH) and TH in ipsilateral paw, which was associated with significant elevated oxido-nitrosative stress. Treatment with ATRA (5 mg/kg/p.o/4 days) and GW0742, a selective PPAR-β/δ receptor agonist (0.1 mg/kg/i.p/4 days), significantly decreased the paw volume, mechanical and TH as compared to vehicle control. Administration of GSK0660, selective PPAR-β/δ receptor antagonist, at a dose of (0.3 mg/kg/i.p/4 days), did not produce a significant effect on carrageenan-induced paw edema, MH and TH. However, co-administration of GSK0660 (0.3 mg/kg/i.p/4 days) along with both ATRA (5 mg/kg/p.o/4 days) and GW0742 (0.1 mg/kg/i.p/4 days), significantly reverse the decreased paw edema, MH, and TH. These observed ameliorative effects on inflammatory pain symptoms are correlated with the extent of reduction of oxido-nitrosative stress. Conclusion: From above findings, it can be concluded that ATRA exerts anti-inflammatory and anti-hyperalgesic effect, possibly through activation of PPAR-β/δ and subsequent reduction of oxido-nitrosative stress. PMID:23833373

  7. Retinoic Acid Prevents Disruption of Blood-Spinal Cord Barrier by Inducing Autophagic Flux After Spinal Cord Injury.

    PubMed

    Zhou, Yulong; Zheng, Binbin; Ye, Libing; Zhang, Hongyu; Zhu, Sipin; Zheng, Xiaomeng; Xia, Qinghai; He, Zili; Wang, Qingqing; Xiao, Jian; Xu, Huazi

    2016-04-01

    Spinal cord injury (SCI) induces the disruption of the blood-spinal cord barrier (BSCB), which leads to infiltration of blood cells, inflammatory responses and neuronal cell death, with subsequent development of spinal cord secondary damage. Recent reports pointed to an important role of retinoic acid (RA), the active metabolite of the vitamin A, in the induction of the blood-brain barrier (BBB) during human and mouse development, however, it is unknown whether RA plays a role in maintaining BSCB integrity under the pathological conditions such as SCI. In this study, we investigated the BSCB protective role of RA both in vivo and in vitro and demonstrated that autophagy are involved in the BSCB protective effect of RA. Our data show that RA attenuated BSCB permeability and also attenuated the loss of tight junction molecules such as P120, β-catenin, Occludin and Claudin5 after injury in vivo as well as in brain microvascular endothelial cells. In addition, RA administration improved functional recovery of the rat model of trauma. We also found that RA could significantly increase the expression of LC3-II and decrease the expression of p62 both in vivo and in vitro. Furthermore, combining RA with the autophagy inhibitor chloroquine (CQ) partially abolished its protective effect on the BSCB and exacerbated the loss of tight junctions. Together, our studies indicate that RA improved functional recovery in part by the prevention of BSCB disruption via the activation of autophagic flux after SCI. PMID:26582233

  8. Antitumor Activity of Hierridin B, a Cyanobacterial Secondary Metabolite Found in both Filamentous and Unicellular Marine Strains

    PubMed Central

    Ramos, Vitor; Pereira, Alban R.; Fernandes, Virgínia C.; Domingues, Valentina F.; Gerwick, William H.; Vasconcelos, Vitor M.; Martins, Rosário

    2013-01-01

    Cyanobacteria are widely recognized as a valuable source of bioactive metabolites. The majority of such compounds have been isolated from so-called complex cyanobacteria, such as filamentous or colonial forms, which usually display a larger number of biosynthetic gene clusters in their genomes, when compared to free-living unicellular forms. Nevertheless, picocyanobacteria are also known to have potential to produce bioactive natural products. Here, we report the isolation of hierridin B from the marine picocyanobacterium Cyanobium sp. LEGE 06113. This compound had previously been isolated from the filamentous epiphytic cyanobacterium Phormidium ectocarpi SAG 60.90, and had been shown to possess antiplasmodial activity. A phylogenetic analysis of the 16S rRNA gene from both strains confirmed that these cyanobacteria derive from different evolutionary lineages. We further investigated the biological activity of hierridin B, and tested its cytotoxicity towards a panel of human cancer cell lines; it showed selective cytotoxicity towards HT-29 colon adenocarcinoma cells. PMID:23922738

  9. Role of retinoic acid metabolizing cytochrome P450s, CYP26, in inflammation and cancer

    PubMed Central

    Stevison, Faith; Jing, Jing; Tripathy, Sasmita; Isoherranen, Nina

    2016-01-01

    Vitamin A (retinol) and its active metabolite, all-trans-retinoic acid (atRA), play critical roles in regulating the differentiation, growth and migration of immune cells. Similarly, as critical signaling molecules in the regulation of the cell cycle, retinoids are important in cancers. Concentrations of atRA are tightly regulated in tissues, predominantly by the availability of retinol, synthesis of atRA by ALDH1A enzymes and metabolism and clearance of atRA by CYP26 enzymes. The ALDH1A and CYP26 enzymes are expressed in several cell types in the immune system and in cancer cells. In the immune system the ALDH1A and CYP26 enzymes appear to modulate RA concentrations. Consequently, alterations in the activity of ALDH1A and CYP26 enzymes are expected to change disease outcomes in inflammation. There is increasing evidence from various disease models of intestinal and skin inflammation that treatment with atRA has a positive effect on disease markers. However, whether aberrant atRA concentrations or atRA synthesis and metabolism play a role in inflammatory disease development and progression is not well understood. In cancers, especially in acute promyelocytic leukemia and neuroblastoma, increasing intracellular concentrations of atRA appears to provide clinical benefit. Inhibition of the CYP26 enzymes to increase atRA concentrations and combat therapy resistance has been pursued as a drug target in these cancers. This chapter covers the current knowledge of how atRA and retinol regulate the immune system and inflammation, how retinol and atRA metabolism is altered in inflammation and cancer and what roles atRA metabolizing enzymes have in immune responses and cancers. PMID:26233912

  10. Effects of Secondary Plant Metabolites on Microbial Populations: Changes in Community Structure and Metabolic Activity in Contaminated Environments.

    PubMed

    Musilova, Lucie; Ridl, Jakub; Polivkova, Marketa; Macek, Tomas; Uhlik, Ondrej

    2016-01-01

    Secondary plant metabolites (SPMEs) play an important role in plant survival in the environment and serve to establish ecological relationships between plants and other organisms. Communication between plants and microorganisms via SPMEs contained in root exudates or derived from litter decomposition is an example of this phenomenon. In this review, the general aspects of rhizodeposition together with the significance of terpenes and phenolic compounds are discussed in detail. We focus specifically on the effect of SPMEs on microbial community structure and metabolic activity in environments contaminated by polychlorinated biphenyls (PCBs) and polyaromatic hydrocarbons (PAHs). Furthermore, a section is devoted to a complex effect of plants and/or their metabolites contained in litter on bioremediation of contaminated sites. New insights are introduced from a study evaluating the effects of SPMEs derived during decomposition of grapefruit peel, lemon peel, and pears on bacterial communities and their ability to degrade PCBs in a long-term contaminated soil. The presented review supports the "secondary compound hypothesis" and demonstrates the potential of SPMEs for increasing the effectiveness of bioremediation processes. PMID:27483244

  11. Light-induced biochemical variations in secondary metabolite production and antioxidant activity in callus cultures of Stevia rebaudiana (Bert).

    PubMed

    Ahmad, Naveed; Rab, Abdur; Ahmad, Nisar

    2016-01-01

    Stevia rebaudiana (S. rebaudiana) is a very important species with worldwide medicinal and commercial uses. Light is one of the major elicitors that fluctuate morphogenic potential and biochemical responses. In the present study, we investigated the effect of various spectral lights on biomass accumulation and secondary metabolite production in callus cultures of S. rebaudiana. Leaf explants were placed on Murashige and Skoog (MS) medium and exposed to various spectral lights. 6-Benzyle adenine (BA) and 2, 4-dichlorophenoxy acetic acid (2, 4-D; 2.0 mgl(-1)) were used for callus induction. The control light (16/8h) produced optimum callogenic response (92.73%) than other colored lights. Compared to other colored lights, control grown cultures displayed maximum biomass accumulation (5.78 gl(-1)) during a prolonged log phase at the 18th day of growth kinetics. Cultures grown under blue light enhanced total phenolic content (TPC; 102.32 μg/g DW), total flavonoid content (TFC; 22.07 μg/g DW) and total antioxidant capacity (TAC; 11.63 μg/g DW). On the contrary, green and red lights improved reducing power assay (RPA; 0.71Fe(II)g(-1) DW) and DPPH-radical scavenging activity (DRSA; 80%). Herein, we concluded that the utilization of colored lights is a promising strategy for enhanced production of antioxidant secondary metabolites in callus cultures of S. rebaudiana. PMID:26688290

  12. Effects of Secondary Plant Metabolites on Microbial Populations: Changes in Community Structure and Metabolic Activity in Contaminated Environments

    PubMed Central

    Musilova, Lucie; Ridl, Jakub; Polivkova, Marketa; Macek, Tomas; Uhlik, Ondrej

    2016-01-01

    Secondary plant metabolites (SPMEs) play an important role in plant survival in the environment and serve to establish ecological relationships between plants and other organisms. Communication between plants and microorganisms via SPMEs contained in root exudates or derived from litter decomposition is an example of this phenomenon. In this review, the general aspects of rhizodeposition together with the significance of terpenes and phenolic compounds are discussed in detail. We focus specifically on the effect of SPMEs on microbial community structure and metabolic activity in environments contaminated by polychlorinated biphenyls (PCBs) and polyaromatic hydrocarbons (PAHs). Furthermore, a section is devoted to a complex effect of plants and/or their metabolites contained in litter on bioremediation of contaminated sites. New insights are introduced from a study evaluating the effects of SPMEs derived during decomposition of grapefruit peel, lemon peel, and pears on bacterial communities and their ability to degrade PCBs in a long-term contaminated soil. The presented review supports the “secondary compound hypothesis” and demonstrates the potential of SPMEs for increasing the effectiveness of bioremediation processes. PMID:27483244

  13. Observation of an Unusual Electronically Distorted Semiquinone Radical of PCB Metabolites in the Active Site of Prostaglandin H Synthase-2

    PubMed Central

    Wangpradit, Orarat; Moman, Edelmiro; Nolan, Kevin B.; Buettner, Garry R.; Robertson, Larry W.; Luthe, Gregor

    2013-01-01

    The activation of the metabolites of airborne polychlorinated biphenyls (PCBs) into highly reactive radicals is of fundamental importance. We found that human recombinant prostaglandin H synthase-2 (hPGHS-2) biotransforms dihydroxy-PCBs, such as 4-chlorobiphenyl-2′,5′-hydroquinone (4-CB-2′,5′H2Q), into semiquinone radicals via one-electron oxidation. Using electron paramagnetic resonance (EPR) spectroscopy, we observed the formation of the symmetric quartet spectrum (1:3:3:1 by area) of 4-chlorobiphenyl-2′,5′-semiquinone radical (4-CB-2′,5′-SQ•−) from 4-CB-2′,5′H2Q. This spectrum changed to an asymmetric spectrum with time: the change can be explained as the overlap of two different semiquinone radical species. Hindered rotation of the 4-CB-2′,5′-SQ•− appears not to be a major factor for the change in lineshape because increasing the viscosity of the medium with glycerol produced no significant change in lineshape. Introduction of a fluorine, which increases the steric hindrance for rotation of the dihydroxy-PCB studied, also produced no significant changes. An in silico molecular docking model of 4-CB-2′,5′H2Q in the peroxidase site of hPGHS-2 together with ab initio quantum mechanical studies indicate that the close proximity of a negatively charged carboxylic acid in the peroxidase active site may be responsible for the observed perturbation in the spectrum. This study provides new insights into the formation of semiquinones from PCB metabolites and underscores the potential role of PGHS-2 in the metabolic activation of PCBs. PMID:20843536

  14. Identification of Tazarotenic Acid as the First Xenobiotic Substrate of Human Retinoic Acid Hydroxylase CYP26A1 and CYP26B1.

    PubMed

    Foti, Robert S; Isoherranen, Nina; Zelter, Alex; Dickmann, Leslie J; Buttrick, Brian R; Diaz, Philippe; Douguet, Dominique

    2016-05-01

    Cytochrome P450 (CYP) 26A1 and 26B1 are heme-containing enzymes responsible for metabolizing all-trans retinoic acid (at-RA). No crystal structures have been solved, and therefore homology models that provide structural information are extremely valuable for the development of inhibitors of cytochrome P450 family 26 (CYP26). The objectives of this study were to use homology models of CYP26A1 and CYP26B1 to characterize substrate binding characteristics, to compare structural aspects of their active sites, and to support the role of CYP26 in the metabolism of xenobiotics. Each model was verified by dockingat-RA in the active site and comparing the results to known metabolic profiles ofat-RA. The models were then used to predict the metabolic sites of tazarotenic acid with results verified by in vitro metabolite identification experiments. The CYP26A1 and CYP26B1 homology models predicted that the benzothiopyranyl moiety of tazarotenic acid would be oriented toward the heme of each enzyme and suggested that tazarotenic acid would be a substrate of CYP26A1 and CYP26B1. Metabolite identification experiments indicated that CYP26A1 and CYP26B1 oxidatively metabolized tazarotenic acid on the predicted moiety, with in vitro rates of metabolite formation by CYP26A1 and CYP26B1 being the highest across a panel of enzymes. Molecular analysis of the active sites estimated the active-site volumes of CYP26A1 and CYP26B1 to be 918 Å(3)and 977 Å(3), respectively. Overall, the homology models presented herein describe the enzyme characteristics leading to the metabolism of tazarotenic acid by CYP26A1 and CYP26B1 and support a potential role for the CYP26 enzymes in the metabolism of xenobiotics. PMID:26937021

  15. Effects of the microbial secondary metabolite benzothiazole on the nutritional physiology and enzyme activities of Bradysia odoriphaga (Diptera: Sciaridae).

    PubMed

    Zhao, Yunhe; Xu, Chunmei; Wang, Qiuhong; Wei, Yan; Liu, Feng; Xu, Shuangyu; Zhang, Zhengqun; Mu, Wei

    2016-05-01

    Bradysia odoriphaga (Diptera: Sciaridae) is the major pest that damages Chinese chive production. As a volatile compound derived from microbial secondary metabolites, benzothiazole has been determined to possess fumigant activity against B. odoriphaga. However, the mechanism of action of benzothiazole is not well understood. In the present study, fourth-instar larvae of B. odoriphaga were exposed to LC10 and LC30 of benzothiazole. Sublethal concentrations (LC10 and LC30) of benzothiazole significantly reduced the food consumption of the larvae on the second day after treatment (2 DAT). However, there were no significant changes in pupal weight among the different treatments. We also measured the protein, lipid, carbohydrate, and trehalose contents and the digestive enzyme activities of the larvae, and the results suggest that benzothiazole reduced the nutrient accumulation and decreased the digestive enzyme activities of B. odoriphaga. In addition, the activity of glutathione S-transferase was significantly decreased at 6h after treatment with benzothiazole, whereas general esterase activities were significantly increased at 6 and 24h after treatment. The results of this study indicate that benzothiazole interferes in the normal food consumption and digestion process by decreasing the activities of digestive enzymes. These results provide valuable information for understanding the toxicity of benzothiazole and for exploring volatile compound for the control of this pest. PMID:27017881

  16. Histopathology, enzyme activities, and PAH metabolites in English sole collected near coastal pulp mills

    SciTech Connect

    Brand, D.G.

    1995-12-31

    The bottom-feeding flatfish, English sole (Pleuronectes vetulus), is widely distributed along the B.C. Pacific coast and fulfills a majority of the requirements as a sentinel species for environmental effects monitoring programs. Studies involving the use of histopathological, biochemical, and chemical tools with English sole collected near the vicinity of B.C. pulp mills are currently being conducted. Analysis, to date, has revealed idiopathic liver lesions to be strongly dependent on location of capture with a prevalence of 30% preneoplastic and neoplastic lesions found in fish collected near pulp mills. All fish residing near pulp mills show hepatocellular hemosiderosis, an iron storage disorder. The mixed-function oxidizing enzyme, EROD, was found to be induced in fish collected near pulp mills. However, the levels of conjugating enzymes, GST and UDP-GT, were found to be unchanged when compared with reference fish. PAH metabolites, measured as FACs in bile, are also present in English sole collected from the mill sites and the conjugated derivatives are presently being identified by HPLC/ES-MS techniques, The relationships between these observations will be discussed.

  17. mu Opioid receptor-mediated G-protein activation by heroin metabolites: evidence for greater efficacy of 6-monoacetylmorphine compared with morphine.

    PubMed

    Selley, D E; Cao, C C; Sexton, T; Schwegel, J A; Martin, T J; Childers, S R

    2001-08-15

    The efficacy of heroin metabolites for the stimulation of mu opioid receptor-mediated G-protein activation was investigated using agonist-stimulated [(35)S]guanosine-5'-O-(gamma-thio)-triphosphate binding. In rat thalamic membranes, heroin and its primary metabolite, 6-monoacetylmorphine (6-MAM), were more efficacious than morphine or morphine-6-beta D-glucuronide. This increased efficacy was not due to increased action of heroin and 6-MAM at delta receptors, as determined by competitive antagonism by naloxone, lack of antagonism by naltrindole, and competitive partial antagonism with morphine. In agreement with this interpretation, the same relative efficacy profile of heroin and its metabolites was observed at the cloned human mu opioid receptor expressed in C6 glioma cells. Moreover, these efficacy differences were GDP-dependent in a manner consistent with accepted mechanisms of receptor-mediated G-protein activation. The activity of heroin was attributed to in vitro deacetylation to 6-MAM, as confirmed by HPLC analysis. These results indicate that the heroin metabolite 6-MAM possesses higher efficacy than other heroin metabolites at mu opioid receptors, which may contribute to the higher efficacy of heroin compared with morphine in certain behavioral paradigms in vivo. PMID:11448454

  18. A third human retinoic acid receptor, hRAR-. gamma

    SciTech Connect

    Krust, A.; Kastner, Ph.; Petkovich, M.; Zelent, A.; Chambon, P. )

    1989-07-01

    Retinoic acid receptors (RARs) are retinoic acid (RA)-inducible enhancer factors belonging to the superfamily of steroid/thyroid nuclear receptors. The authors have previously characterized two human RAR (hRAR-{alpha} and hRAR-{beta}) cDNAs and have recently cloned their murine cognates (mRAR-{alpha} and mRAR-{beta}) together with a third RAR (mRAR-{gamma}) whose RNA was detected predominantly in skin, a well-known target for RA. mRAR-{gamma} cDNA was used here to clone its human counterpart (hRAR-{gamma}) from a T47D breast cancer cell cDNA library. Using a transient transfection assay in HeLa cells and a reporter gene harboring a synthetic RA responsive element, they demonstrate that hRAR-{gamma} cDNA indeed encodes a RA-inducible transcriptional trans-activator. Interestingly, comparisons of the amino acid sequences of all six human and mouse RARs indicate that the interspecies conservation of a given member of the RAR subfamily (either {alpha}, {beta}, or {gamma}) is much higher than the conservation of all three receptors within a given species. These observations indicate that RAR-{alpha}, -{beta}, and -{gamma} may perform specific functions. They show also that hRAR-{gamma} RNA is the predominant RAR RNA species in human skin, which suggests that hRAR-{gamma} mediates some of the retinoid effects in this tissue.

  19. Retinoic acid-induced neural differentiation of embryonal carcinoma cells.

    PubMed Central

    Jones-Villeneuve, E M; Rudnicki, M A; Harris, J F; McBurney, M W

    1983-01-01

    We have previously shown that the P19 line of embryonal carcinoma cells develops into neurons, astroglia, and fibroblasts after aggregation and exposure to retinoic acid. The neurons were initially identified by their morphology and by the presence of neurofilaments within their cytoplasm. We have more fully documented the neuronal nature of these cells by showing that their cell surfaces display tetanus toxin receptors, a neuronal cell marker, and that choline acetyl-transferase and acetyl cholinesterase activities appear coordinately in neuron-containing cultures. Several days before the appearance of neurons, there is a marked decrease in the amount of an embryonal carcinoma surface antigen, and at the same time there is a substantial decrease in the volumes of individual cells. Various retinoids were able to induce the development of neurons in cultures of aggregated P19 cells, but it did not appear that polyamine metabolism was involved in the effect. We have isolated a mutant clone which does not differentiate in the presence of any of the drugs which are normally effective in inducing differentiation of P19 cells. This mutant and others may help to elucidate the chain of events triggered by retinoic acid and other differentiation-inducing drugs. Images PMID:6656766

  20. Retinoic Acid and Affective Disorders: The Evidence for an Association

    PubMed Central

    Bremner, J Douglas; Shearer, Kirsty; McCaffery, Peter

    2011-01-01

    Objective Isotretinoin (13-cis-retinoic acid, or 13-cis-RA) (Accutane), approved by the FDA for the treatment of acne, carries a black box warning related to the risk of depression, suicide, and psychosis. Retinoic acid (RA), the active form of vitamin A, regulates gene expression in the brain, and isotretinoin is its 13-cis isomer. Retinoids represent a group of compounds derived from vitamin A that perform a large variety of functions in many systems, in particular the CNS, and abnormal retinoid levels can have neurological effects. Although infrequent, proper recognition and treatment of psychiatric side effects in acne patients is critical given the risk of death and disability. This paper reviews the evidence for a relationship between isotretinoin, depression and suicidality. Data Sources Evidence examined includes: 1) case reports; 2) temporal association between onset of depression and exposure to the drug; 3) challenge-rechallenge cases; 4) class effect (other compounds in the same class, like vitamin A, having similar neuropsychiatric effects); 5) dose response; and 6) biologically plausible mechanisms. Study Selection All papers in the literature related to isotretinoin, depression and suicide were reviewed, as well as papers related to class effect, dose response, and biological plausibility. Data Extraction Information from individual articles in the literature was extracted. Data Synthesis The literature reviewed is consistent with an association between isotretinoin administration, depression and suicide in some individuals. Conclusions The relationship between isotretinoin and depression may have implications for a greater understanding of the neurobiology of affective disorders. PMID:21903028

  1. Thuringiensin: A Thermostable Secondary Metabolite from Bacillus thuringiensis with Insecticidal Activity against a Wide Range of Insects

    PubMed Central

    Liu, Xiaoyan; Ruan, Lifang; Peng, Donghai; Li, Lin; Sun, Ming; Yu, Ziniu

    2014-01-01

    Thuringiensin (Thu), also known as β-exotoxin, is a thermostable secondary metabolite secreted by Bacillus thuringiensis. It has insecticidal activity against a wide range of insects, including species belonging to the orders Diptera, Coleoptera, Lepidoptera, Hymenoptera, Orthoptera, and Isoptera, and several nematode species. The chemical formula of Thu is C22H32O19N5P, and it is composed of adenosine, glucose, phosphoric acid, and gluconic diacid. In contrast to the more frequently studied insecticidal crystal protein, Thu is not a protein but a small molecule oligosaccharide. In this review, a detailed and updated description of the characteristics, structure, insecticidal mechanism, separation and purification technology, and genetic determinants of Thu is provided. PMID:25068925

  2. Retinoic Acid Improves Morphology of Cultured Peritoneal Mesothelial Cells from Patients Undergoing Dialysis

    PubMed Central

    Retana, Carmen; Sanchez, Elsa I.; Gonzalez, Sirenia; Perez-Lopez, Alejandro; Cruz, Armando; Lagunas-Munoz, Jesus; Alfaro-Cruz, Carmen; Vital-Flores, Socorro; Reyes, José L.

    2013-01-01

    Patients undergoing continuous ambulatory peritoneal dialysis are classified according to their peritoneal permeability as low transporter (low solute permeability) or High transporter (high solute permeability). Factors that determine the differences in permeability between them have not been fully disclosed. We investigated morphological features of cultured human peritoneal mesothelial cells from low or high transporter patients and its response to All trans retinoic Acid (ATRA, vitamin A active metabolite), as compared to non-uremic human peritoneal mesothelial cells. Control cells were isolated from human omentum. High or low transporter cells were obtained from dialysis effluents. Cells were cultured in media containing ATRA (0, 50, 100 or 200 nM). We studied length and distribution of microvilli and cilia (scanning electron microscopy), epithelial (cytokeratin, claudin-1, ZO-1 and occludin) and mesenchymal (vimentin and α-smooth muscle actin) transition markers by immunofluorescence and Western blot, and transforming growth factor β1 expression by Western blot. Low and high transporter exhibited hypertrophic cells, reduction in claudin-1, occludin and ZO-1 expression, cytokeratin and vimentin disorganization and positive α-smooth muscle actin label. Vimentin, α-smooth muscle actin and transforming growth factor- β1 were overexpressed in low transporter. Ciliated cells were diminished in low and high transporters. Microvilli number and length were severely reduced in high transporter. ATRA reduced hypertrophic cells number in low transporter. It also improved cytokeratin and vimentin organization, decreased vimentin and α-smooth muscle actin expression, and increased claudin 1, occludin and ZO-1 expression, in low and high transporter. In low transporter, ATRA reduced transforming growth factor-β1 expression. ATRA augmented percentage of ciliated cells in low and high transporter. It also augmented cilia length in high transporter. Alterations in

  3. Evaluation of the pharmacokinetics of oral amitriptyline and its active metabolite nortriptyline in fed and fasted Greyhound dogs.

    PubMed

    Norkus, C; Rankin, D; KuKanich, B

    2015-12-01

    This study reports the pharmacokinetics of oral amitriptyline and its active metabolite nortriptyline in Greyhound dogs. Five healthy Greyhound dogs were enrolled in a randomized crossover design. A single oral dose of amitriptyline hydrochloride (actual mean dose 8.1 per kg) was administered to fasted or fed dogs. Blood samples were collected at predetermined times from 0 to 24 h after administration, and plasma drug concentrations were measured by liquid chromatography with mass spectrometry. Noncompartmental pharmacokinetic analyses were performed. Two dogs in the fasted group vomited following amitriptyline administration and were excluded from analysis. The range of amitriptyline CMAX for the remaining fasted dogs (n = 3) was 22.8-64.5 ng/mL compared to 30.6-127 ng/mL for the fed dogs (n = 5). The range of the amitriptyline AUCINF for the three fasted dogs was 167-720 h·ng/mL compared to 287-1146 h·ng/mL for fed dogs. The relative bioavailability of amitriptyline in fasted dogs compared to fed dogs was 69-91% (n = 3). The exposure of the active metabolite nortriptyline was correlated to amitriptyline exposure (R(2)  = 0.84). Due to pharmacokinetic variability and the small number of dogs completing this study, further studies are needed assessing the impact of feeding on oral amitriptyline pharmacokinetics. Amitriptyline may be more likely to cause vomiting in fasted dogs. PMID:25989225

  4. Gene Expression Profiling Elucidates a Specific Role for RARγ in the Retinoic Acid Induced Differentiation of F9 Teratocarcinoma Stem Cells

    PubMed Central

    Su, Dan; Gudas, Lorraine J

    2010-01-01

    The biological effects of all-trans-retinoic acid (RA), a major active metabolite of retinol, are mainly mediated through its interactions with retinoic acid receptor (RARs α, β, γ) and retinoid X receptor (RXRs α, β, γ) heterodimers. RAR/RXR heterodimers activate transcription by binding to RA-response elements (RAREs or RXREs) in the promoters of primary target genes. Murine F9 teratocarcinoma stem cells have been widely used as a model for cellular differentiation and RA signaling during embryonic development. We identified and characterized genes that are differentially expressed in F9 wild type (Wt) and F9 RAR γ−/− cells, with and without RA treatment, through the use of oligonucleotide based microarrays. Our data indicate that RARγ, in the absence of exogenous RA, modulates gene expression. Genes such as Sfrp2, Tie1, Fbp2, Emp1, and Emp3 exhibited higher transcript levels in RA treated Wt, RARα−/− and RARβ2−/− lines than in RA-treated RARγ−/− cells, and represent specific RARγ targets. Other genes, such as Runx1, were expressed at lower levels in both F9 RARβ2−/− and RARγ−/− cell lines then in F9 Wt and RARα−/−. Genes specifically induced by RA at 6h with the protein synthesis inhibitor cycloheximide in F9 Wt, but not in RARγ−/− cells, included Hoxa3, Hoxa5, Gas1, Cyp26a1, Sfrp2, Fbp2, and Emp1. These genes represent specific primary RARγ targets in F9 cells. Several genes in the Wnt signaling pathway were regulated by RARγ. Delineation of the receptor specific actions of RA with respect to cell proliferation and differentiation should result in more effective therapies with this drug. PMID:18164278

  5. Retinoic Acid Modulates Interferon-γ Production by Hepatic Natural Killer T Cells via Phosphatase 2A and the Extracellular Signal-Regulated Kinase Pathway

    PubMed Central

    Chang, Heng-Kwei

    2015-01-01

    Retinoic acid (RA), an active metabolite converted from vitamin A, plays an active role in immune function, such as defending against infections and immune regulation. Although RA affects various types of immune cells, including antigen-presenting cells, B lymphocytes, and T lymphocytes, whether it affects natural killer T (NKT) cells remain unknown. In this study, we found that RA decreased interferon (IFN)-γ production by activated NKT cells through T-cell receptor (TCR) and CD28. We also found that RA reduced extracellular signal-regulated kinase (ERK) phosphorylation, but increased phosphatase 2A (PP2A) activity in TCR/CD28-stimulated NKT cells. The increased PP2A activity, at least partly, contributed to the reduction of ERK phosphorylation. Since inhibition of ERK activation decreases IFN-γ production by TCR/CD28-stimulated NKT cells, RA may downregulate IFN-γ production by TCR/CD28-stimulated NKT cells through the PP2A-ERK pathway. Our results demonstrated a novel function of RA in modulating the IFN-γ expression by activated NKT cells. PMID:25343668

  6. In Vitro Transformation of Chlorinated Parabens by the Liver S9 Fraction: Kinetics, Metabolite Identification, and Aryl Hydrocarbon Receptor Agonist Activity.

    PubMed

    Terasaki, Masanori; Wada, Takeshi; Nagashima, Satoshi; Makino, Masakazu; Yasukawa, Hiro

    2016-01-01

    We investigated the kinetics of in vitro transformation of a dichlorinated propyl paraben (2-propyl 3,5-dichloro-4-hydroxybenzoate; Cl2PP) by the rat liver S9 fraction and assessed the aryl hydrocarbon receptor (AhR) agonist activity of the metabolite products identified in HPLC and GC/MS analysis and by metabolite syntheses. The results indicated that the chlorination of Cl2PP reduced its degradation rate by approximately 40-fold. Two hydroxylated metabolite products showed AhR agonist activity of up to 39% of that of the parent Cl2PP when assessed in a yeast (YCM3) reporter gene assay. The determination of the metabolic properties of paraben bioaccumulation presented here provides further information on the value of risk assessments of chlorinated parabens as a means to ensure human health and environmental safety. PMID:27250800

  7. The combination of glutamate receptor antagonist MK-801 with tamoxifen and its active metabolites potentiates their antiproliferative activity in mouse melanoma K1735-M2 cells

    SciTech Connect

    Ribeiro, Mariana P.C.; Nunes-Correia, Isabel; Santos, Armanda E.; Custódio, José B.A.

    2014-02-15

    Recent reports suggest that N-methyl-D-aspartate receptor (NMDAR) blockade by MK-801 decreases tumor growth. Thus, we investigated whether other ionotropic glutamate receptor (iGluR) antagonists were also able to modulate the proliferation of melanoma cells. On the other hand, the antiestrogen tamoxifen (TAM) decreases the proliferation of melanoma cells, and is included in combined therapies for melanoma. As the efficacy of TAM is limited by its metabolism, we investigated the effects of the NMDAR antagonist MK-801 in combination with TAM and its active metabolites, 4-hydroxytamoxifen (OHTAM) and endoxifen (EDX). The NMDAR blockers MK-801 and memantine decreased mouse melanoma K1735-M2 cell proliferation. In contrast, the NMDAR competitive antagonist APV and the AMPA and kainate receptor antagonist NBQX did not affect cell proliferation, suggesting that among the iGluR antagonists only the NMDAR channel blockers inhibit melanoma cell proliferation. The combination of antiestrogens with MK-801 potentiated their individual effects on cell biomass due to diminished cell proliferation, since it decreased the cell number and DNA synthesis without increasing cell death. Importantly, TAM metabolites combined with MK-801 promoted cell cycle arrest in G1. Therefore, the data obtained suggest that the activity of MK-801 and antiestrogens in K1735-M2 cells is greatly enhanced when used in combination. - Highlights: • MK-801 and memantine decrease melanoma cell proliferation. • The combination of MK-801 with antiestrogens inhibits melanoma cell proliferation. • These combinations greatly enhance the effects of the compounds individually. • MK-801 combined with tamoxifen active metabolites induces cell cycle arrest in G1. • The combination of MK-801 and antiestrogens is an innovative strategy for melanoma.

  8. In silico Identification of Ergosterol as a Novel Fungal Metabolite Enhancing RuBisCO Activity in Lycopersicum esculentum.

    PubMed

    Mitra, Joyeeta; Narad, Priyanka; Sengupta, Abhishek; Sharma, P D; Paul, P K

    2016-09-01

    RuBisCO (EC 4.1.1.39), a key enzyme found in stroma of chloroplast, is important for fixing atmospheric CO2 in plants. Alterations in the activity of RuBisCO could influence photosynthetic yield. Therefore, to understand the activity of the protein, knowledge about its structure is pertinent. Though the structure of Nicotiana RuBisCO has been modeled, the structure of tomato RuBisCO is still unknown. RuBisCO extracted from chloroplasts of tomato leaves was subjected to MALDI-TOF-TOF followed by Mascot Search. The protein sequence based on gene identification numbers was subjected to in silico model construction, characterization and docking studies. The primary structure analysis revealed that protein was stable, neutral, hydrophilic and has an acidic pI. The result though indicates a 90 % homology with other members of Solanaceae but differs from the structure of Arabidopsis RuBisCO. Different ligands were docked to assess the activity of RuBisCO against these metabolite components. Out of the number of modulators tested, ergosterol had the maximum affinity (E = -248.08) with RuBisCO. Ergosterol is a major cell wall component of fungi and has not been reported to be naturally found in plants. It is a known immune elicitor in plants. The current study throws light on its role in affecting RuBisCO activity in plants, thereby bringing changes in the photosynthetic rate. PMID:26253718

  9. Developmental expression of retinoic acid receptors (RARs)

    PubMed Central

    Dollé, Pascal

    2009-01-01

    Here, I review the developmental expression features of genes encoding the retinoic acid receptors (RARs) and the 'retinoid X' or rexinoid receptors (RXRs). The first detailed expression studies were performed in the mouse over two decades ago, following the cloning of the murine Rar genes. These studies revealed complex expression features at all stages of post-implantation development, one receptor gene (Rara) showing widespread expression, the two others (Rarb and Rarg) with highly regionalized and/or cell type-specific expression in both neural and non-neural tissues. Rxr genes also have either widespread (Rxra, Rxrb), or highly-restricted (Rxrg) expression patterns. Studies performed in zebrafish and Xenopus demonstrated expression of Rar and Rxr genes (both maternal and zygotic), at early pre-gastrulation stages. The eventual characterization of specific enzymes involved in the synthesis of retinoic acid (retinol/retinaldehyde dehydrogenases), or the triggering of its catabolism (CYP26 cytochrome P450s), all of them showing differential expression patterns, led to a clearer understanding of the phenomenons regulated by retinoic acid signaling during development. Functional studies involving targeted gene disruptions in the mouse, and additional approaches such as dominant negative receptor expression in other models, have pinpointed the specific, versus partly redundant, roles of the RARs and RXRs in many developing organ systems. These pleiotropic roles are summarized hereafter in relationship to the receptors’ expression patterns. PMID:19471585

  10. Retinoic acid regulates embryonic development of mammalian submandibular salivary glands.

    PubMed

    Wright, Diana M; Buenger, Deanna E; Abashev, Timur M; Lindeman, Robert P; Ding, Jixiang; Sandell, Lisa L

    2015-11-01

    Organogenesis is orchestrated by cell and tissue interactions mediated by molecular signals. Identification of relevant signals, and the tissues that generate and receive them, are important goals of developmental research. Here, we demonstrate that Retinoic Acid (RA) is a critical signaling molecule important for morphogenesis of mammalian submandibular salivary glands (SMG). By examining late stage RA deficient embryos of Rdh10 mutant mice we show that SMG development requires RA in a dose-dependent manner. Additionally, we find that active RA signaling occurs in SMG tissues, arising earlier than any other known marker of SMG development and persisting throughout gland morphogenesis. At the initial bud stage of development, we find RA production occurs in SMG mesenchyme, while RA signaling occurs in epithelium. We also demonstrate active RA signaling occurs in glands cultured ex vivo, and treatment with an inhibitor of RA signaling blocks growth and branching. Together these data identify RA signaling as a direct regulator of SMG organogenesis. PMID:26278034

  11. The active metabolite of prasugrel inhibits ADP-stimulated thrombo-inflammatory markers of platelet activation: Influence of other blood cells, calcium, and aspirin.

    PubMed

    Frelinger, Andrew L; Jakubowski, Joseph A; Li, Youfu; Barnard, Marc R; Fox, Marsha L; Linden, Matthew D; Sugidachi, Atsuhiro; Winters, Kenneth J; Furman, Mark I; Michelson, Alan D

    2007-07-01

    The novel thienopyridine prodrug prasugrel, a platelet P2Y(12) ADP receptor antagonist, requires in vivo metabolism for activity. Although pharmacological data have been collected on the effects of prasugrel on platelet aggregation, there are few data on the direct effects of the prasugrel's active metabolite, R-138727, on other aspects of platelet function. Here we examined the effects of R-138727 on thrombo-inflammatory markers of platelet activation, and the possible modulatory effects of other blood cells, calcium, and aspirin. Blood (PPACK or citrate anticoagulated) from healthy donors pre- and post-aspirin was incubated with R-138727 and the response to ADP assessed in whole blood or platelet-rich plasma (PRP) by aggregometry and flow cytometric analysis of leukocyte-platelet aggregates, platelet surface P-selectin, and GPIIb-IIIa activation. Low-micromolar concentrations of R-138727 resulted in a rapid and consistent inhibition of these ADP-stimulated thrombo-inflammatory markers. These rapid kinetics required physiological calcium levels, but were largely unaffected by aspirin. Lower IC(50) values in whole blood relative to PRP suggested that other blood cells affect ADP-induced platelet activation and hence the net inhibition by R-138727. R-138727 did not inhibit P2Y(12)-mediated ADP-induced shape change, even at concentrations that completely inhibited platelet aggregation, confirming the specificity of R-138727 for P2Y(12). In conclusion, R-138727, the active metabolite of prasugrel, results in rapid, potent, consistent, and selective inhibition of P2Y(12)-mediated up-regulation of thrombo-inflammatory markers of platelet activation. This inhibition is enhanced in the presence other blood cells and calcium, but not aspirin. PMID:17598013

  12. Identification and characterization of a functional retinoic acid/thyroid hormone-response element upstream of the human insulin gene enhancer.

    PubMed Central

    Clark, A R; Wilson, M E; London, N J; James, R F; Docherty, K

    1995-01-01

    A deletion analysis of the human insulin gene extending to 2 kb upstream of the transcription start site provided evidence of regulatory sequences located upstream of the insulin-linked polymorphic region (ILPR). Within this ILPR-distal region is a sequence (Ink, for insulin kilobase upstream) which contains three potential nuclear hormone-receptor half-sites, closely matching the consensus sequence AGGTCA. These sequences are arranged as a palindromic element with zero spacing over-lapping a direct repeat with 2 bp spacing. The Ink sequence was used in electrophoretic mobility-shift assays within nuclear extracts from COS-7 cells overexpressing the vitamin D, thyroid hormone or retinoic acid receptors, or from an insulin-expressing hamster cell line, HIT-T15. These studies suggest that the insulin-expressing cell line contains thyroid hormone and retinoic acid receptors at least, and that these receptors are able to recognize the Ink sequence. Three copies of the Ink sequence were placed upstream of the thymidine kinase promoter and firefly luciferase reporter gene. In COS-7 cells expressing the appropriate nuclear hormone receptor, this construct was responsive to both thyroid hormone (18-fold) and all-trans-retinoic acid (31-fold). In HIT-T15 cells the same construct responded to all-trans-retinoic acid, but not to thyroid hormone. Within the context of a 2 kb insulin gene fragment, the Ink sequence was shown to be activated by retinoic acid and by the retinoic acid receptor, but acted as a negative element in the presence of both retinoic acid and the retinoic acid receptor. Mutagenesis studies demonstrated that the palindromic sequence was important for the retinoic acid response, and for binding of complexes containing retinoic acid receptor. In human islets of Langerhans, retinoic acid was shown to stimulate insulin mRNA levels. These results demonstrate that a functional nuclear hormone-receptor-response element is located upstream of the human ILPR. As

  13. PreQ0 base, an unusual metabolite with anti-cancer activity from Streptomyces qinglanensis 172205.

    PubMed

    Xu, Dongbo; Ma, Min; Liu, Yongfeng; Zhou, Ting; Wang, Kexin; Deng, Zixin; Hong, Kui

    2015-01-01

    PreQ0 base (7-cyano-7-deazaguanine, compound 1) is the biosynthetic precursor of queuosine-tRNA and important synthetic intermediate for bioactive compounds. It was obtained for the first time as a new natural product from a mangrove actinomycete Streptomyces qinglanensis 172205, during the course of searching for anti-cancer compounds from marine microbes. PreQ0 base showed anti-HeLa (IC50 = 62.0 μg/ml) and anti-HepG2 (IC50 = 80.6 μg/ml) activities, higher cytotoxicity than the positive control, fluorouracil. Furthermore, it exhibited weak α -glucosidase inhibitory activity, but no obvious antimicrobial and Aβ1-42 fibrillation inhibitory activities. In silico analysis of the genome of the strain 172205 revealed a putative biosynthetic gene cluster directing the biosynthesis of PreQ0 base. The gene cluster only contained three Open Reading Frames (ORFs), queC, queD and queE. The absence of the key gene queF encoding 7-cyano-7-deazaguanine reductase catalyzing PreQ0 base to PreQ1 base suggested that the strain only has the capacity of accumulation of PreQ0 base as a metabolite, consistent with our observation in chemical identification. PMID:25353335

  14. Activation of p53 with Ilimaquinone and Ethylsmenoquinone, Marine Sponge Metabolites, Induces Apoptosis and Autophagy in Colon Cancer Cells

    PubMed Central

    Lee, Hyun-Young; Chung, Kyu Jin; Hwang, In Hyun; Gwak, Jungsuk; Park, Seoyoung; Ju, Bong Gun; Yun, Eunju; Kim, Dong-Eun; Chung, Young-Hwa; Na, MinKyun; Song, Gyu-Yong; Oh, Sangtaek

    2015-01-01

    The tumor suppressor, p53, plays an essential role in the cellular response to stress through regulating the expression of genes involved in cell cycle arrest, apoptosis and autophagy. Here, we used a cell-based reporter system for the detection of p53 response transcription to identify the marine sponge metabolites, ilimaquinone and ethylsmenoquinone, as activators of the p53 pathway. We demonstrated that ilimaquinone and ethylsmenoquinone efficiently stabilize the p53 protein through promotion of p53 phosphorylation at Ser15 in both HCT116 and RKO colon cancer cells. Moreover, both compounds upregulate the expression of p21WAF1/CIP1, a p53-dependent gene, and suppress proliferation of colon cancer cells. In addition, ilimaquinone and ethylsmenoquinone induced G2/M cell cycle arrest and increased caspase-3 cleavage and the population of cells that positively stained with Annexin V-FITC, both of which are typical biochemical markers of apoptosis. Furthermore, autophagy was elicited by both compounds, as indicated by microtubule-associated protein 1 light chain 3 (LC3) puncta formations and LC3-II turnover in HCT116 cells. Our findings suggest that ilimaquinone and ethylsmenoquinone exert their anti-cancer activity by activation of the p53 pathway and may have significant potential as chemo-preventive and therapeutic agents for human colon cancer. PMID:25603347

  15. Microbial transformation of ginsenoside-Rg₁ by Absidia coerulea and the reversal activity of the metabolites towards multi-drug resistant tumor cells.

    PubMed

    Liu, Xin; Qiao, Lirui; Xie, Dan; Zhang, Yi; Zou, Jianhua; Chen, Xiaoguang; Dai, Jungui

    2011-12-01

    Biotransformation of ginsenoside-Rg₁ (1) by the fungus Absidia coerulea AS 3.2462 yielded five metabolites (2-6). On the basis of spectroscopic data analyses, the metabolites were identified as ginsenoside-F₁ (2), 6α,12β-dihydroxydammar-3-one-20(S)-O-β-D-glucopyranoside (3), 3-oxo-20(S)-protopanaxatriol (4), 3-oxo-7β-hydroxy-20(S)-protopanaxatriol (5), and 3-oxo-7β,15α-dihydroxy-20(S)-protopanaxatriol (6), respectively. Among them, 5 and 6 are new compounds. These results indicated that Absidia coerulea AS 3.2462 could catalyze the specific C-3 dehydrogenation of derivatives of ginsenoside-Rg₁, as well as hydroxylation at the 7β and 15α positions. Metabolites 2, 4 and 5 exhibited moderate reversal activity towards A549/taxol MDR tumor cells in vitro. PMID:21946057

  16. Effects of licochalcon A on the pharmacokinetics of losartan and its active metabolite, EXP-3174, in rats.

    PubMed

    Choi, J S; Choi, J S; Choi, D H

    2013-11-01

    Losartan and licochalcon A interact with cytochrome P-450 (CYP) enzymes and P-glycoprotein (P-gp), and the increase in the use of health supplements may result in licochalcon A being taken concomitantly with losartan to treat or prevent cardiovascular diseases as a combination therapy. The effect of licochalcon A, a natural flavonoid, on the pharmacokinetics of losartan and its active metabolite, EXP-3174, was investigated in rats. Pharmacokinetic parameters of losartan and EXP-3174 were determined after oral administration of losartan (9 mg/kg) to rats in the presence or absence of licochalcon A (0.5, 2.5 and 10 mg/kg). The effect of licochalcon A on P-glycoprotein (P-gp) as well as CYP3A4 and 2C9 activities was also evaluated. Licochalcon A inhibited CYP3A4 and CYP2C9 enzyme activities with 50% inhibition concentrations (IC50) of 2.0 and 0.1 microM, respectively. In addition, licochalcon A significantly enhanced the cellular accumulation of rhodamine-123 in a concentration-dependent manner in MCF-7/ADR cells overexpressing P-gp. The pharmacokinetic parameters of losartan were significantly altered by licochalcon A. Licochalcon A (2.5 mg/kg or 10 mg/kg) increased AUC0-infinity of losartan by 33.4-63.2% and Cmax of losartan by 34.0-62.8%. The total body clearance (CL/F) was significantly decreased (2.5 mg/kg, p < 0.05; 10 mg/kg, p < 0.01) by licochalcon A. Consequently, the absolute bioavailability of losartan in the presence of licochalcon A increased significantly (2.5 mg/kg, p < 0.05; 10 mg/kg, p < 0.01) compared to that in the control group. The relative bioavailability (R.B.) of losartan was 1.15- to 1.63-fold greater than that of the control group. However, there was no significant change in Tmax and t1/2 of losartan in the presence of licochalcon A. Licochalcon A (10 mg/kg) increased the AUC0-infinity of EXP-3174 but this was not significant. Furthermore, concurrent use of licochalcon A (10 mg/kg) significantly decreased the metabolite-parent AUC ratio (M

  17. The combination of glutamate receptor antagonist MK-801 with tamoxifen and its active metabolites potentiates their antiproliferative activity in mouse melanoma K1735-M2 cells.

    PubMed

    Ribeiro, Mariana P C; Nunes-Correia, Isabel; Santos, Armanda E; Custódio, José B A

    2014-02-15

    Recent reports suggest that N-methyl-d-aspartate receptor (NMDAR) blockade by MK-801 decreases tumor growth. Thus, we investigated whether other ionotropic glutamate receptor (iGluR) antagonists were also able to modulate the proliferation of melanoma cells. On the other hand, the antiestrogen tamoxifen (TAM) decreases the proliferation of melanoma cells, and is included in combined therapies for melanoma. As the efficacy of TAM is limited by its metabolism, we investigated the effects of the NMDAR antagonist MK-801 in combination with TAM and its active metabolites, 4-hydroxytamoxifen (OHTAM) and endoxifen (EDX). The NMDAR blockers MK-801 and memantine decreased mouse melanoma K1735-M2 cell proliferation. In contrast, the NMDAR competitive antagonist APV and the AMPA and kainate receptor antagonist NBQX did not affect cell proliferation, suggesting that among the iGluR antagonists only the NMDAR channel blockers inhibit melanoma cell proliferation. The combination of antiestrogens with MK-801 potentiated their individual effects on cell biomass due to diminished cell proliferation, since it decreased the cell number and DNA synthesis without increasing cell death. Importantly, TAM metabolites combined with MK-801 promoted cell cycle arrest in G1. Therefore, the data obtained suggest that the activity of MK-801 and antiestrogens in K1735-M2 cells is greatly enhanced when used in combination. PMID:24240127

  18. Antioxidant activity and metabolite profile of quercetion in vitamin E depleted rats

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Dietary antioxidants interact in a dynamic fashion, including recycling and sparing one another, to decrease oxidative stress. Limited information is available regarding the interrelationships in vivo between quercetin and vitamin E. We investigated the antioxidant activity and metabolism of querc...

  19. Human myeloblastic leukemia cells (HL-60) express a membrane receptor for estrogen that signals and modulates retinoic acid-induced cell differentiation

    SciTech Connect

    Kauss, M. Ariel; Reiterer, Gudrun; Bunaciu, Rodica P.; Yen, Andrew

    2008-10-01

    Estrogen receptors are historically perceived as nuclear ligand activated transcription factors. An estrogen receptor has now been found localized to the plasma membrane of human myeloblastic leukemia cells (HL-60). Its expression occurs throughout the cell cycle, progressively increasing as cells mature from G{sub 1} to S to G{sub 2}/M. To ascertain that the receptor functioned, the effect of ligands, including a non-internalizable estradiol-BSA conjugate and tamoxifen, an antagonist of nuclear estrogen receptor function, were tested. The ligands caused activation of the ERK MAPK pathway. They also modulated the effect of retinoic acid, an inducer of MAPK dependent terminal differentiation along the myeloid lineage in these cells. In particular the ligands inhibited retinoic acid-induced inducible oxidative metabolism, a functional marker of terminal myeloid cell differentiation. To a lesser degree they also diminished retinoic acid-induced earlier markers of cell differentiation, namely CD38 and CD11b. However, they did not regulate retinoic acid-induced G{sub 0} cell cycle arrest. There is thus a membrane localized estrogen receptor in HL-60 myeloblastic leukemia cells that can cause ERK activation and modulates the response of these cells to retinoic acid, indicating crosstalk between the membrane estrogen and retinoic acid evoked pathways relevant to propulsion of cell differentiation.

  20. Human myeloblastic leukemia cells (HL-60) express a membrane receptor for estrogen that signals and modulates retinoic acid-induced cell differentiation.

    PubMed

    Kauss, M Ariel; Reiterer, Gudrun; Bunaciu, Rodica P; Yen, Andrew

    2008-10-01

    Estrogen receptors are historically perceived as nuclear ligand activated transcription factors. An estrogen receptor has now been found localized to the plasma membrane of human myeloblastic leukemia cells (HL-60). Its expression occurs throughout the cell cycle, progressively increasing as cells mature from G(1) to S to G(2)/M. To ascertain that the receptor functioned, the effect of ligands, including a non-internalizable estradiol-BSA conjugate and tamoxifen, an antagonist of nuclear estrogen receptor function, were tested. The ligands caused activation of the ERK MAPK pathway. They also modulated the effect of retinoic acid, an inducer of MAPK dependent terminal differentiation along the myeloid lineage in these cells. In particular the ligands inhibited retinoic acid-induced inducible oxidative metabolism, a functional marker of terminal myeloid cell differentiation. To a lesser degree they also diminished retinoic acid-induced earlier markers of cell differentiation, namely CD38 and CD11b. However, they did not regulate retinoic acid-induced G(0) cell cycle arrest. There is thus a membrane localized estrogen receptor in HL-60 myeloblastic leukemia cells that can cause ERK activation and modulates the response of these cells to retinoic acid, indicating crosstalk between the membrane estrogen and retinoic acid evoked pathways relevant to propulsion of cell differentiation. PMID:18692045

  1. Prostaglandin D2 metabolites as a biomarker of in vivo mast cell activation in systemic mastocytosis and rheumatoid arthritis.

    PubMed

    Cho, Catherine; Nguyen, Anna; Bryant, Katherine J; O'Neill, Sean G; McNeil, H Patrick

    2016-03-01

    Mast cells (MCs) participate in diseases such as systemic mastocytosis (SM) and allergic conditions. Less well understood is the role of MCs in non-allergic inflammatory disorders like rheumatoid arthritis (RA). Studying definitive roles for MCs in human diseases has been hampered by the lack of a well-accepted biomarker for monitoring in vivo MC activation. This study aimed to investigate the utility of urinary tetranor PGDM (T-PGDM) as a biomarker of in vivo MC activation in patients with SM, and apply this biomarker to assess MC involvement in relation to RA disease activity. A prospective, cross-sectional cohort study was conducted to measure a major urinary metabolite of prostaglandin D2, T-PGDM. Urine samples were collected from patients with RA (n = 60), SM (n = 17) and healthy normal controls (n = 16) and T-PGDM excretion was determined by enzyme immunoassay as nanograms per milligram of urinary creatinine (ng/mg Cr). Mean urinary T-PGDM excretion was significantly higher (p < 0.01) in patients with SM compared to controls (37.2 vs. 11.5 ng/mg Cr) with 65% of SM patients showing elevated levels. One third of patients with RA had elevated T-PGDM excretion, and the mean level in the RA group (20.0 ng/mg Cr) was significantly higher than controls (p < 0.01). Medications inhibiting cyclooxygenase reduced T-PGDM excretion. Urinary T-PGDM excretion appears promising as a biomarker of in vivo MC activity and elevated levels in 33% of patients with RA provides evidence of MC activation in this disease. PMID:27042302

  2. Lack of metabolic activation and predominant formation of an excreted metabolite of nontoxic platynecine-type pyrrolizidine alkaloids.

    PubMed

    Ruan, Jianqing; Liao, Cangsong; Ye, Yang; Lin, Ge

    2014-01-21

    Pyrrolizidine alkaloid (PA) poisoning is well-known because of the intake of PA-containing plant-derived natural products and PA-contaminated foodstuffs. Based on different structures of the necine bases, PAs are classified into three types: retronecine, otonecine, and platynecine type. The former two type PAs possessing an unsaturated necine base with a 1,2-double bond are hepatotoxic due to the P450-mediated metabolic activation to generate reactive pyrrolic ester, which interacts with cellular macromolecules leading to toxicity. With a saturated necine base, platynecine-type PAs are reported to be nontoxic and their nontoxicity was hypothesized to be due to the absence of metabolic activation; however, the metabolic pathway responsible for their nontoxic nature is largely unknown. In the present study, to prove the absence of metabolic activation in nontoxic platynecine-type PAs, hepatic metabolism of platyphylline (PLA), a representative platynecine-type PA, was investigated and directly compared with the representatives of two toxic types of PAs in parallel. By determining the pyrrolic ester-derived glutathione conjugate, our results confirmed that the major metabolic pathway of PLA did not lead to formation of the reactive pyrrolic ester. More interestingly, having a metabolic rate similar to that of toxic PAs, PLA also underwent oxidative metabolisms mediated by P450s, especially P450 3A4, the same enzyme that catalyzes metabolic activation of two toxic types of PAs. However, the predominant oxidative dehydrogenation pathway of PLA formed a novel metabolite, dehydroplatyphylline carboxylic acid, which was water-soluble, readily excreted, and could not interact with cellular macromolecules. In conclusion, our study confirmed that the saturated necine bases determine the absence of metabolic activation and thus govern the metabolic pathway responsible for the nontoxic nature of platynecine-type PAs. PMID:24308637

  3. Fumigaclavine C, an fungal metabolite, improves experimental colitis in mice via downregulating Th1 cytokine production and matrix metalloproteinase activity.

    PubMed

    Wu, Xue-Feng; Fei, Ming-Jian; Shu, Ren-Geng; Tan, Ren-Xiang; Xu, Qiang

    2005-09-01

    In the present paper, the effect of Fumigaclavine C, a fungal metabolite, on experimental colitis was examined. Fumigaclavine C, when administered intraperitoneally once a day, significantly reduced the weight loss and mortality rate of mice with experimental colitis induced by intrarectally injection of 2, 4, 6-trinitrobenzene sulfonic acid (TNBS). This compound also markedly alleviated the macroscopic and microscopic appearances of colitis. Furthermore, Fumigaclavine C, given both in vivo and in vitro, showed a marked inhibition on the expression of several inflammatory cytokines, including IL-1beta, IL-2, IL-12alpha, IFN-gamma, TNF-alpha as well as MMP-9 in sacral lymph node cells, colonic patch lymphocytes and colitis tissues from the TNBS colitis mice. Meanwhile, the compound caused a dose-dependent reduction in IL-2 and IFN-gamma from the lymphocytes at the protein level and MMP-9 activity. These results suggest that Fumigaclavine C may alleviate experimental colitis mainly via down-regulating the production of Th1 cytokines and the activity of matrix metalloproteinase. PMID:16023606

  4. Effects of Metabolites Produced from (-)-Epigallocatechin Gallate by Rat Intestinal Bacteria on Angiotensin I-Converting Enzyme Activity and Blood Pressure in Spontaneously Hypertensive Rats.

    PubMed

    Takagaki, Akiko; Nanjo, Fumio

    2015-09-23

    Inhibitory activity of angiotensin I-converting enzyme (ACE) was examined with (-)-epigallocatechin gallate (EGCG) metabolites produced by intestinal bacteria, together with tea catechins. All of the metabolites showed ACE inhibitory activities and the order of IC50 was hydroxyphenyl valeric acids > 5-(3,4,5-trihydroxyphenyl)-γ-valerolactone (1) > trihydroxyphenyl 4-hydroxyvaleric acid ≫ dihydroxyphenyl 4-hydroxyvaleric acid ≫ 5-(3,5-dihydroxyphenyl)-γ-valerolactone (2). Among the catechins, galloylated catechins exhibited stronger ACE inhibitory activity than nongalloylated catechins. Furthermore, the effects of a single oral intake of metabolites 1 and 2 on systolic blood pressure (SBP) were examined with spontaneously hypertensive rats (SHR). Significant decreases in SBP were observed between 2 h after oral administration of 1 (150 mg/kg in SHR) and the control group (p = 0.002) and between 4 h after administration of 2 (200 mg/kg in SHR) and the control group (p = 0.044). These results suggest that the two metabolites have hypotensive effects in vivo. PMID:26323573

  5. Distinct binding determinants for 9-cis retinoic acid are located within AF-2 of retinoic acid receptor alpha.

    PubMed Central

    Tate, B F; Allenby, G; Janocha, R; Kazmer, S; Speck, J; Sturzenbecker, L J; Abarzúa, P; Levin, A A; Grippo, J F

    1994-01-01

    Retinoids exert their physiological action by interacting with two families of nuclear receptors, the retinoic acid receptors (RARs) and the retinoid X receptors (RXRs), which regulate gene expression by forming transcriptionally active heterodimeric RAR/RXR or homodimeric RXR/RXR complexes on DNA. Retinoid receptor activity resides in several regions, including the DNA and ligand binding domains, a dimerization interface, and both a ligand-independent (AF-1) and a ligand-dependent (AF-2) transactivation function. While 9-cis retinoic acid (RA) alone is the cognate ligand for the RXRs, both 9-cis RA and all-trans RA (t-RA) compete for binding with high affinity to the RARs. This latter observation suggested to us that the two isomers may interact with a common binding site. Here we report that RAR alpha has two distinct but overlapping binding sites for 9-cis RA and t-RA. Truncation of a human RAR alpha to 419 amino acids yields a receptor that binds both t-RA and 9-cis RA with high affinity, but truncation to amino acid 404 yields a mutant receptor that binds only t-RA with high affinity. Remarkably, this region also defines a C-terminal boundary for AF-2, as addition of amino acids 405 to 419 restores receptor-mediated gene activity to a truncated human RAR alpha lacking this region. It is interesting to speculate that binding of retinoid stereoisomers to unique sites within an RAR may function with AF-2 to cause differential activation of retinoid-responsive gene pathways. Images PMID:8139538

  6. Structure of neprilysin in complex with the active metabolite of sacubitril

    PubMed Central

    Schiering, Nikolaus; D’Arcy, Allan; Villard, Frederic; Ramage, Paul; Logel, Claude; Cumin, Frederic; Ksander, Gary M.; Wiesmann, Christian; Karki, Rajeshri G.; Mogi, Muneto

    2016-01-01

    Sacubitril is an ethyl ester prodrug of LBQ657, the active neprilysin (NEP) inhibitor, and a component of LCZ696 (sacubitril/valsartan). We report herein the three-dimensional structure of LBQ657 in complex with human NEP at 2 Å resolution. The crystal structure unravels the binding mode of the compound occupying the S1, S1’ and S2’ sub-pockets of the active site, consistent with a competitive inhibition mode. An induced fit conformational change upon binding of the P1’-biphenyl moiety of the inhibitor suggests an explanation for its selectivity against structurally homologous zinc metallopeptidases. PMID:27302413

  7. Structure of neprilysin in complex with the active metabolite of sacubitril.

    PubMed

    Schiering, Nikolaus; D'Arcy, Allan; Villard, Frederic; Ramage, Paul; Logel, Claude; Cumin, Frederic; Ksander, Gary M; Wiesmann, Christian; Karki, Rajeshri G; Mogi, Muneto

    2016-01-01

    Sacubitril is an ethyl ester prodrug of LBQ657, the active neprilysin (NEP) inhibitor, and a component of LCZ696 (sacubitril/valsartan). We report herein the three-dimensional structure of LBQ657 in complex with human NEP at 2 Å resolution. The crystal structure unravels the binding mode of the compound occupying the S1, S1' and S2' sub-pockets of the active site, consistent with a competitive inhibition mode. An induced fit conformational change upon binding of the P1'-biphenyl moiety of the inhibitor suggests an explanation for its selectivity against structurally homologous zinc metallopeptidases. PMID:27302413

  8. Cytotoxic activities of ethyl acetate extract and a metabolite from a Monocillium species.

    PubMed

    Khondkar, Proma; Rahman, M Mukhlesur; Islam, Anwarul

    2005-09-01

    The ethyl acetate soluble fraction of a cultural broth of a Monocillium species afforded the isolation of 5-hydroxymethylfurfural. Both the extract and 5-hydroxymethylfurfural showed significant cytotoxic activities in a brine shrimp bioassay and the LC(50) values were found to be 14.96 microg/mL and 23.71 microg/mL, respectively. PMID:16220580

  9. Total synthesis of the cyanobacterial metabolite nostodione A: discovery of its antiparasitic activity against Toxoplasma gondii.

    PubMed

    McNulty, J; Keskar, K; Bordón, C; Yolken, R; Jones-Brando, L

    2014-08-18

    A total synthesis of the cyanobacterial natural product nostodione A is reported involving a convergent, diversity-oriented route. A small assemblage of structural analogues were prepared and their cytotoxicity and anti-invasion activity against the protozoal parasite Toxoplasma gondii is reported for the first time. PMID:24970332

  10. Salicylate, diflunisal and their metabolites inhibit CBP/p300 and exhibit anticancer activity

    PubMed Central

    Shirakawa, Kotaro; Wang, Lan; Man, Na; Maksimoska, Jasna; Sorum, Alexander W; Lim, Hyung W; Lee, Intelly S; Shimazu, Tadahiro; Newman, John C; Schröder, Sebastian; Ott, Melanie; Marmorstein, Ronen; Meier, Jordan; Nimer, Stephen; Verdin, Eric

    2016-01-01

    Salicylate and acetylsalicylic acid are potent and widely used anti-inflammatory drugs. They are thought to exert their therapeutic effects through multiple mechanisms, including the inhibition of cyclo-oxygenases, modulation of NF-κB activity, and direct activation of AMPK. However, the full spectrum of their activities is incompletely understood. Here we show that salicylate specifically inhibits CBP and p300 lysine acetyltransferase activity in vitro by direct competition with acetyl-Coenzyme A at the catalytic site. We used a chemical structure-similarity search to identify another anti-inflammatory drug, diflunisal, that inhibits p300 more potently than salicylate. At concentrations attainable in human plasma after oral administration, both salicylate and diflunisal blocked the acetylation of lysine residues on histone and non-histone proteins in cells. Finally, we found that diflunisal suppressed the growth of p300-dependent leukemia cell lines expressing AML1-ETO fusion protein in vitro and in vivo. These results highlight a novel epigenetic regulatory mechanism of action for salicylate and derivative drugs. DOI: http://dx.doi.org/10.7554/eLife.11156.001 PMID:27244239

  11. Salicylate, diflunisal and their metabolites inhibit CBP/p300 and exhibit anticancer activity.

    PubMed

    Shirakawa, Kotaro; Wang, Lan; Man, Na; Maksimoska, Jasna; Sorum, Alexander W; Lim, Hyung W; Lee, Intelly S; Shimazu, Tadahiro; Newman, John C; Schröder, Sebastian; Ott, Melanie; Marmorstein, Ronen; Meier, Jordan; Nimer, Stephen; Verdin, Eric

    2016-01-01

    Salicylate and acetylsalicylic acid are potent and widely used anti-inflammatory drugs. They are thought to exert their therapeutic effects through multiple mechanisms, including the inhibition of cyclo-oxygenases, modulation of NF-κB activity, and direct activation of AMPK. However, the full spectrum of their activities is incompletely understood. Here we show that salicylate specifically inhibits CBP and p300 lysine acetyltransferase activity in vitro by direct competition with acetyl-Coenzyme A at the catalytic site. We used a chemical structure-similarity search to identify another anti-inflammatory drug, diflunisal, that inhibits p300 more potently than salicylate. At concentrations attainable in human plasma after oral administration, both salicylate and diflunisal blocked the acetylation of lysine residues on histone and non-histone proteins in cells. Finally, we found that diflunisal suppressed the growth of p300-dependent leukemia cell lines expressing AML1-ETO fusion protein in vitro and in vivo. These results highlight a novel epigenetic regulatory mechanism of action for salicylate and derivative drugs. PMID:27244239

  12. In vitro biological activity of secondary metabolites from Seseli rigidum Waldst. et Kit. (Apiaceae).

    PubMed

    Jakovljević, Dragana; Vasić, Sava; Stanković, Milan; Čomić, Ljiljana; Topuzović, Marina

    2015-12-01

    The antioxidant, antimicrobial activity, total phenolic content and flavonoid concentration of Seseli rigidum Waldst. et Kit. were evaluated. Five different extracts of the aboveground plant parts were obtained by extraction with distilled water, methanol, acetone, ethyl acetate and petroleum ether. Total phenols were determined using the Folin-Ciocalteu's reagent, with the highest values obtained in the acetone extract (102.13 mg GAE/g). The concentration of flavonoids, determined by using a spectrophotometric method with aluminum chloride and expressed in terms of rutin equivalent, was also highest in the acetone extracts (291.58 mg RUE/g). The antioxidant activity was determined in vitro using DPPH reagent. The greatest antioxidant activity was expressed in the aqueous extract (46.15 μg/ml). In vitro antimicrobial activities were determined using a microdilution analysis method; minimum inhibitory concentration (MIC) and minimum microbicidal concentration (MMC) were determined. Methanolic extract had the greatest influence on bacilli (MIC at 0.0391 mg/ml), but the best antimicrobial effect had acetone and ethyl acetate extracts considering their broad impact on bacteria. According to our research, S. rigidum can be regarded as promising candidate for natural plant source with high value of biological compounds. PMID:26616372

  13. Antioxidant and anti-acetylcholinesterase activities of extracts and secondary metabolites from Acacia cyanophylla

    PubMed Central

    Ghribia, Lotfi; Ghouilaa, Hatem; Omrib, Amel; Besbesb, Malek; Janneta, Hichem Ben

    2014-01-01

    Objective To investigate the antioxidant potential and anti-acetycholinesterase activity of compounds and extracts from Acacia cyanophylla (A. cyanophylla). Methods Three polyphenolic compounds were isolated from ethyl acetate extract of A. cyanophylla flowers. They have been identified as isosalipurposide 1, quercetin 2 and naringenin 3. Their structures were elucidated by extensive spectroscopic methods including 1D and 2D NMR experiments as well as ES-MS. The prepared extracts and the isolated compounds 1-3 were tested for their antioxidant activity using 1′-1′-diphenylpicrylhydrazyl (DPPH) and 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) scavenging assays and reducing power. They have been also investigated for inhibitory effect against acetylcholinesterase using the microplate assay. Results In the DPPH test, the EtOAc extract of flowers exhibited the highest antioxidant effect (67.26 µg/mL). Isosalipurposide 1 showed a significant antiradical power against DPPH (81.9 µg/mL). All extracts showed a dose-dependent acetylcholinesterase inhibition. In terms of the IC50 value, the butanolic extract (16.03 µg/mL) was the most potent sample. Isosalipurposide 1 was found to be active against AChE with an IC50 value of 52.04 µg/mL. Conclusions The results demonstrated the important antioxidant and anti-acetylcholinesterase activity of pure compounds and extracts from A. cyanophylla. PMID:25183120

  14. Non-invasive monitoring of adrenocortical activity in captive African Penguin (Spheniscus demersus) by measuring faecal glucocorticoid metabolites.

    PubMed

    Ozella, L; Anfossi, L; Di Nardo, F; Pessani, D

    2015-12-01

    Measurement of faecal glucocorticoid metabolites (FGMs) has become a useful and widely-accepted method for the non-invasive evaluation of stress in vertebrates. In this study we assessed the adrenocortical activity of five captive African Penguins (Spheniscus demersus) by means of FGM evaluation following a biological stressor, i.e. capture and immobilization. In addition, we detected individual differences in secretion of FGMs during a stage of the normal biological cycle of penguins, namely the breeding period, without any external or induced causes of stress. Our results showed that FGM concentrations peaked 5.5-8h after the induced stress in all birds, and significantly decreased within 30 h. As predictable, the highest peak of FGMs (6591 ng/g) was reached by the youngest penguin, which was at its first experience with the stressor. This peak was 1.8-2.7-fold higher compared to those of the other animals habituated to the stimulus. For the breeding period, our results revealed that the increase in FGMs compared to ordinary levels, and the peaks of FGMs, varied widely depending on the age and mainly on the reproductive state of the animal. The bird which showed the lowest peak (2518 ng/g) was an old male that was not in a reproductive state at the time of the study. Higher FGM increases and peaks were reached by the two birds which were brooding (male: 5552%, 96,631 ng/g; female: 1438%, 22,846 ng/g) and by the youngest bird (1582%, 39,700 ng/g). The impact of the reproductive state on FGM levels was unexpected compared to that produced by the induced stress. The EIA used in this study to measure FGM levels proved to be a reliable tool for assessing individual and biologically-relevant changes in FGM concentrations in African Penguin. Moreover, this method allowed detection of physiological stress during the breeding period, and identification of individual differences in relation to the reproductive status. The increase in FGM levels as a response to capture and

  15. Secondary metabolites from Vietnamese marine invertebrates with activity against Trypanosoma brucei and T. cruzi.

    PubMed

    Thao, Nguyen Phuong; No, Joo Hwan; Luyen, Bui Thi Thuy; Yang, Gyongseon; Byun, Soo Young; Goo, Junghyun; Kim, Kyung Tae; Cuong, Nguyen Xuan; Nam, Nguyen Hoai; Van Minh, Chau; Schmidt, Thomas J; Kang, Jong Seong; Kim, Young Ho

    2014-01-01

    Marine-derived natural products from invertebrates comprise an extremely diverse and promising source of the compounds from a wide variety of structural classes. This study describes the discovery of five marine natural products with activity against Trypanosoma species by natural product library screening using whole cell in vitro assays. We investigated the anti-trypanosomal activity of the extracts from the soft corals and echinoderms living in Vietnamese seas. Of the samples screened, the methanolic extracts of several marine organisms exhibited potent activities against cultures of Trypanosoma brucei and T. cruzi (EC50 < 5.0 μg/mL). Among the compounds isolated from these extracts, laevigatol B (1) from Lobophytum crassum and L. laevigatum, (24S)-ergost-4-ene-3-one (2) from Sinularia dissecta, astropectenol A (3) from Astropecten polyacanthus, and cholest-8-ene-3β,5α,6β,7α-tetraol (4) from Diadema savignyi showed inhibitory activity against T. brucei with EC50 values ranging from 1.57 ± 0.14 to 14.6 ± 1.36 μM, relative to the positive control, pentamidine (EC50 = 0.015 ± 0.003 μM). Laevigatol B (1) and 5α-cholest-8(14)-ene-3β,7α-diol (5) exhibited also significant inhibitory effects on T. cruzi. The cytotoxic activity of the pure compounds on mammalian cells was also assessed and found to be insignificant in all cases. This is the first report on the inhibitory effects of marine organisms collected in Vietnamese seas against Trypanosoma species responsible for neglected tropical diseases. PMID:24962391

  16. Estrogenic and androgenic activities of TBBA and TBMEPH, metabolites of novel brominated flame retardants, and selected bisphenols, using the XenoScreen XL YES/YAS assay.

    PubMed

    Fic, Anja; Žegura, Bojana; Gramec, Darja; Mašič, Lucija Peterlin

    2014-10-01

    The present study investigated and compared the estrogenic and androgenic activities of the three different classes of environmental pollutants and their metabolites using the XenoScreen XL YES/YAS assay, which has advantages compared with the original YES/YAS protocol. Contrary to the parent brominated flame retardants TBB and TBPH, which demonstrated no or very weak (anti)estrogenic or (anti)androgenic activities, their metabolites, TBBA and TBMEPH, exhibited anti-estrogenic (IC50 for TBBA=31.75 μM and IC50 for TBMEPH=0.265 μM) and anti-androgenic (IC50 for TBBA=73.95 μM and IC50 for TBMEPH=2.92 μM) activities. These results reveal that metabolism can enhance the anti-estrogenic and anti-androgenic effects of these two novel brominated flame retardants. Based on the activities of BPAF, BPF, BPA and MBP, we can conclude that the XenoScreen XL YES/YAS assay gives comparable results to the (anti)estrogenic or (anti)androgenic assays that are reported in the literature. For BPA, it was confirmed previously that the metabolite formed after an ipso-reaction (hydroxycumyl alcohol) exhibited higher estrogenic activity compared with the parent BPA, but this was not confirmed for BPAF and BPF ipso-metabolites, which were not active in the XenoScreen YES/YAS assay. Among the substituted BPA analogues, bis-GMA exhibited weak anti-estrogenic activity, BADGE demonstrated weak anti-estrogenic and anti-androgenic activities (IC50=13.73 μM), and the hydrolysed product BADGE·2H2O demonstrated no (anti)estrogenic or (anti)androgenic activities. PMID:25048928

  17. Secondary metabolites from the unripe pulp of Persea americana and their antimycobacterial activities.

    PubMed

    Lu, Ying-Chen; Chang, Hsun-Shuo; Peng, Chien-Fang; Lin, Chu-Hung; Chen, Ih-Sheng

    2012-12-15

    The fruits of Persea americana (Avocado) are nowadays used as healthy fruits in the world. Bioassay-guided fractionation of the active ethyl acetate soluble fraction has led to the isolation of five new fatty alcohol derivatives, avocadenols A-D (1-4) and avocadoin (5) from the unripe pulp of P. americana, along with 12 known compounds (6-17). These structures were elucidated by spectroscopic analysis. Among the isolates, avocadenol A (1), avocadenol B (2), (2R,4R)-1,2,4-trihydroxynonadecane (6), and (2R,4R)-1,2,4-trihydroxyheptadec-16-ene (7) showed antimycobacterial activity against Mycobacterium tuberculosis H(37)R(V)in vitro, with MIC values of 24.0, 33.8, 24.9, and 35.7 μg/ml, respectively. PMID:22980888

  18. The Effect of the Lunar Cycle on Fecal Cortisol Metabolite Levels and Foraging Ecology of Nocturnally and Diurnally Active Spiny Mice

    PubMed Central

    Dayan, Tamar; Kronfeld-Schor, Noga

    2011-01-01

    We studied stress hormones and foraging of nocturnal Acomys cahirinus and diurnal A. russatus in field populations as well as in two field enclosures populated by both species and two field enclosures with individuals of A. russatus alone. When alone, A. russatus individuals become also nocturnally active. We asked whether nocturnally active A. russatus will respond to moon phase and whether this response will be obtained also in diurnally active individuals. We studied giving-up densities (GUDs) in artificial foraging patches and fecal cortisol metabolite levels. Both species exhibited elevated fecal cortisol metabolite levels and foraged to higher GUDs in full moon nights; thus A. russatus retains physiological response and behavioral patterns that correlate with full moon conditions, as can be expected in nocturnal rodents, in spite of its diurnal activity. The endocrinological and behavioral response of this diurnal species to moon phase reflects its evolutionary heritage. PMID:21829733

  19. Secondary metabolites of Seseli rigidum: Chemical composition plus antioxidant, antimicrobial and cholinesterase inhibition activity.

    PubMed

    Stankov-Jovanović, V P; Ilić, M D; Mitić, V D; Mihajilov-Krstev, T M; Simonović, S R; Nikolić Mandić, S D; Tabet, J C; Cole, R B

    2015-01-01

    Extracts of different polarity obtained from various plant parts (root, leaf, flower and fruit) of Seseli rigidum were studied by different antioxidant assays: DPPH and ABTS radical scavenging activity, by total reducing power method as well as via total content of flavonoids and polyphenols. Essential oils of all plant parts showed weak antioxidant characteristics. The inhibitory concentration range of the tested extracts, against bacteria Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, Bacillus cereus, and fungi Candida albicans and Aspergillus niger was 0.01-1.50 mg/mL and of a microbicidal 0.02-3.00 mg/mL. In the interaction with cholinesterase, all essential oils proved effective as inhibitors. The highest percentage of inhibition versus human and horse cholinesterase was shown by root essential oil (38.20% and 48.30%, respectively) among oils, and root hexane extract (40.56% and 50.65% respectively). Essential oils and volatile components of all plant parts were identified by GC, GC-MS and headspace/GC-MS. Statistical analysis of the ensemble of results showed that the root essential oil composition differed significantly from essential oils of other parts of the plant. Taking into account all of the studied activities, the root hexane extract showed the best overall properties. By means of high performance liquid chromatography coupled to high resolution mass spectrometry, the 30 most abundant constituents were identified in extracts of different polarity. The presence of identified constituents was linked to observed specific biological activities, thus designating compounds potentially responsible for each exhibited activity. PMID:25863020

  20. Complex secondary metabolites from Ludwigia leptocarpa with potent antibacterial and antioxidant activities.

    PubMed

    Mabou, Florence Déclaire; Tamokou, Jean-de-Dieu; Ngnokam, David; Voutquenne-Nazabadioko, Laurence; Kuiate, Jules-Roger; Bag, Prasanta Kumar

    2016-01-01

    Diarrhea continues to be one of the most common causes of morbidity and mortality among infants and children in developing countries. The aim of the present study was to evaluate the antibacterial and antioxidant activities of extracts and compounds from Ludwigia leptocarpa, a plant traditionally used for its vermifugal, anti-dysenteric, and antimicrobial properties. A methanol extract was prepared by maceration of the dried plant and this was successively extracted with ethyl acetate to obtain an EtOAc extract and with n-butanol to obtain an n-BuOH extract. Column chromatography of the EtOAc and n-BuOH extracts was followed by purification of different fractions, leading to the isolation of 10 known compounds. Structures of isolated compounds were assigned on the basis of spectral analysis and by comparison to structures of compounds described in the literature. Antioxidant activity was evaluated using 1,1-diphenyl-2-picrylhydrazyl (DPPH) and gallic acid equivalent antioxidant capacity (GAEAC) assays. Antibacterial activity was assessed with the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) with respect to strains of a Gram-positive bacterium, Staphylococcus aureus (a major cause of community and hospital-associated infection), and Gram-negative multi-drug-resistant bacteria, Vibrio cholerae (a cause of cholera) and Shigella flexneri (a cause of shigellosis). All of the extracts showed different degrees of antioxidant and antibacterial activities. 2β-hydroxyoleanolic acid, (2R,3S,2''S)-3''',4',4''',5,5'',7,7''-heptahydroxy-3,8"-biflavanone, and luteolin-8-C-glucoside displayed the most potent antibacterial and antioxidant properties, and these properties were in some cases equal to or more potent than those of reference drugs. Overall, the present results show that L. leptocarpa has the potential to be a natural source of anti-diarrheal and antioxidant products, so further investigation is warranted. PMID:27431270

  1. Potential role of nuclear receptor ligand all-trans retinoic acids in the treatment of fungal keratitis

    PubMed Central

    Zhou, Hong-Yan; Zhong, Wei; Zhang, Hong; Bi, Miao-Miao; Wang, Shuang; Zhang, Wen-Song

    2015-01-01

    Fungal keratitis (FK) is a worldwide visual impairment disease. This infectious fungus initiates the primary innate immune response and, later the adaptive immune response. The inflammatory process is related to a variety of immune cells, including macrophages, helper T cells, neutrophils, dendritic cells, and Treg cells, and is associated with proinflammatory, chemotactic and regulatory cytokines. All-trans retinoic acids (ATRA) have diverse immunomodulatory actions in a number of inflammatory and autoimmune conditions. These retinoids regulate the transcriptional levels of target genes through the activation of nuclear receptors. Retinoic acid receptor α (RAR α), retinoic acid receptor γ (RAR γ), and retinoid X receptor α (RXR α) are expressed in the cornea and immune cells. This paper summarizes new findings regarding ATRA in immune and inflammatory diseases and analyzes the perspective application of ATRA in FK. PMID:26309886

  2. The Content of Secondary Metabolites and Antioxidant Activity of Wild Strawberry Fruit (Fragaria vesca L.).

    PubMed

    Dyduch-Siemińska, Magdalena; Najda, Agnieszka; Dyduch, Jan; Gantner, Magdalena; Klimek, Kamila

    2015-01-01

    Chemical analyses carried out in 2011-2013 aimed at evaluating the contents of flavonoids, free phenolic acids, tannins, anthocyanins, and antioxidant activity (%) by means of DPPH radical neutralization ability in fresh and air-dried fruits of three wild strawberry cultivars. Examinations revealed differences in contents of biologically active substances determined in raw versus dried material depending on the cultivar. Mean concentrations of flavonoids and tannins were highest in raw fruits of "Baron von Solemacher" cv., which amounted to 1.244 mg·g(-1) and 6.09%, respectively. Fresh fruits of "Regina" cv. were characterized by the highest average content of phenolic acids and anthocyanins: 4.987 mg·g(-1) and 0.636 mg·100 g(-1). The pattern of mean contents of biologically active substances analyzed in air-dried fruits was similar. Significant differences in abilities to neutralize the DPPH radical to diphenylpicrylhydrazine by extracts made of examined wild strawberry fruits were also indicated. PMID:26539306

  3. Secondary metabolites from endophytic Streptomyces aureofaciens CMUAc130 and their antifungal activity.

    PubMed

    Taechowisan, Thongchai; Lu, Chunhua; Shen, Yuemao; Lumyong, Saisamorn

    2005-05-01

    Streptomyces aureofaciens CMUAc130 was isolated from the root tissue of Zingiber officinale Rosc. (Zingiberaceae). It was an antagonist of Colletotrichum musae and Fusarium oxysporum, the causative agents of anthracnose of banana and wilt of wheat, respectively. Evidence for the in vitro antibiosis of S. aureofaciens CMUAc130 was demonstrated by the zone of fungal-growth inhibition. Microscopic observations showed thickness and bulbous structures at the edges of the inhibited fungal hyphae. The culture filtrate and crude extract from this strain were all inhibitory to tested phytopathogenic fungi. The major active ingredients from the culture filtrate of S. aureofaciens CMUAc130 were purified by silica gel-column chromatography and identified to be (i) 5,7-dimethoxy-4-p-methoxylphenylcoumarin and (ii) 5,7-dimethoxy-4-phenylcoumarin by NMR and mass-spectral data, respectively. Bioassay studies showed that compounds (i) and (ii) had antifungal activities against tested fungi, and their MICs were found to be 120 and 150 microg ml(-1), respectively. This is the first report of compounds (i) and (ii) from micro-organisms as active ingredients for the control of phytopathogenic fungi. PMID:15870476

  4. The Content of Secondary Metabolites and Antioxidant Activity of Wild Strawberry Fruit (Fragaria vesca L.)

    PubMed Central

    Dyduch-Siemińska, Magdalena; Najda, Agnieszka; Dyduch, Jan; Gantner, Magdalena; Klimek, Kamila

    2015-01-01

    Chemical analyses carried out in 2011–2013 aimed at evaluating the contents of flavonoids, free phenolic acids, tannins, anthocyanins, and antioxidant activity (%) by means of DPPH radical neutralization ability in fresh and air-dried fruits of three wild strawberry cultivars. Examinations revealed differences in contents of biologically active substances determined in raw versus dried material depending on the cultivar. Mean concentrations of flavonoids and tannins were highest in raw fruits of “Baron von Solemacher” cv., which amounted to 1.244 mg·g−1 and 6.09%, respectively. Fresh fruits of “Regina” cv. were characterized by the highest average content of phenolic acids and anthocyanins: 4.987 mg·g−1 and 0.636 mg·100 g−1. The pattern of mean contents of biologically active substances analyzed in air-dried fruits was similar. Significant differences in abilities to neutralize the DPPH radical to diphenylpicrylhydrazine by extracts made of examined wild strawberry fruits were also indicated. PMID:26539306

  5. Hesperetin and its sulfate and glucuronide metabolites inhibit TNF-α induced human aortic endothelial cell migration and decrease plasminogen activator inhibitor-1 (PAI-1) levels.

    PubMed

    Giménez-Bastida, Juan Antonio; González-Sarrías, Antonio; Vallejo, Fernando; Espín, Juan Carlos; Tomás-Barberán, Francisco A

    2016-01-01

    Epidemiological, clinical and preclinical studies have reported the protection offered by citrus consumption, mainly orange, against cardiovascular diseases, which is primarily mediated by the antiatherogenic and vasculoprotective effects of the flavanone hesperetin-7-O-rutinoside (hesperidin). However, flavanone aglycones or glycosides are not present in the bloodstream but their derived phase-II metabolites could be the actual bioactive molecules. To date, only a few studies have explored the effects of circulating hesperetin-derived metabolites (glucuronides and sulfates) on endothelial cells. Herein, we describe for the first time the effects of hesperetin 3'-O-glucuronide, hesperetin 7-O-glucuronide, hesperetin 3'-O-sulfate, hesperetin 7-O-sulfate and hesperetin on human aortic endothelial cell (HAEC) migration upon pro-inflammatory stimuli as an essential step to angiogenesis. Hesperetin and its derived metabolites, at physiologically relevant concentrations (1-10 μM), significantly attenuated cell migration in the presence of the pro-inflammatory cytokine TNF-α (50 ng mL(-1)), which was accompanied and perhaps mediated by a significant decrease in the levels of the thrombogenic plasminogen activator inhibitor-1 (PAI-1). However, hesperetin metabolites did not counteract the TNF-α-induced production of pro-inflammatory interleukin-6 (IL-6) and IL-8. We also study here for the first time, the metabolism of hesperetin and its derived metabolites by HAEC with and without a pro-inflammatory stimulus. All these results reinforce the concept according to which circulating phase-II hesperetin metabolites are critical molecules contributing to the cardioprotective effects upon consumption of citrus fruits such as orange. PMID:26456097

  6. Have all-trans retinoic acid and arsenic trioxide replaced all-trans retinoic acid and anthracyclines in APL as standard of care.

    PubMed

    Iland, Harry J; Wei, Andrew; Seymour, John F

    2014-03-01

    Until recently, the standard of care in the treatment of APL has involved the combination of all-trans retinoic acid with anthracycline-based chemotherapy during both induction and consolidation. Additionally, the intensity of consolidation chemotherapy has evolved according to a universally accepted relapse-risk stratification algorithm based on the white cell and platelet counts at presentation. That standard of care is being challenged by the increasing incorporation of arsenic trioxide into front-line treatment protocols, based on two complementary observations. The first is the undoubted anti-leukaemic activity of arsenic trioxide as shown in the relapsed and refractory setting, and in the initial management of low- and intermediate-risk patients. The second is an improved understanding of the action of both all-trans retinoic acid and arsenic trioxide in mediating APL cell eradication, with increasing recognition that PML-RARA fusion protein degradation rather than direct induction of terminal differentiation is the primary mechanism for their ability to eliminate leukaemia initiating cells. As a result, we believe the standard of care for initial therapy in APL is shifting towards an all-trans retinoic acid plus arsenic trioxide-based approach, with additional chemotherapy reserved for patients with high-risk disease. PMID:24907016

  7. Chronic oral treatment with 13-cis-retinoic acid (isotretinoin) or all-trans-retinoic acid does not alter depression-like behaviors in rats.

    PubMed

    Ferguson, Sherry A; Cisneros, F Javier; Gough, B; Hanig, Joseph P; Berry, Kimberly J

    2005-10-01

    Oral treatment with the anti-acne drug Accutane (isotretinoin, 13-cis-retinoic acid) has been associated with suicide ideation and depression. Here, depression-like behaviors (i.e., behavioral despair and anhedonia) were quantified in adult Sprague-Dawley rats gavaged daily beginning at postnatal day (PND) 82 with 13-cis-RA (7.5 or 22.5 mg/kg) or all-trans-retinoic acid (10 or 15 mg/kg ). Tested at PND 130-131 in the Forced Swim Test, 7.5 mg/kg 13-cis-RA marginally decreased immobility and slightly increased climb/struggle durations whereas neither all-trans-retinoic acid group differed from controls. Voluntary saccharin solution (0.03%) intake at PND 102-104 and PND 151-153 was not different from controls in any treated group, although all RA-treated groups had lower intakes. Swim speed in a water maze at PND 180 was similar across groups, indicating no RA-induced differences in physical ability. Open field activity was mildly decreased at PND 91 in 7.5 mg/kg-treated males only, but it was within the control range at PND 119, 147, and 175. Thus, at serum levels similar to those in humans receiving the drug, chronic 13-cis-RA treatment did not severely affect depression-like behaviors in rats. These data do not substantiate the hypothesis of 13-cis-RA-induced depression. PMID:16033993

  8. Lipid metabolism enzyme 5-LOX and its metabolite LTB4 are capable of activating transcription factor NF-{kappa}B in hepatoma cells

    SciTech Connect

    Zhao, Yu; Wang, Wenhui; Wang, Qi; Zhang, Xiaodong; Ye, Lihong

    2012-02-24

    Highlights: Black-Right-Pointing-Pointer 5-LOX is able to upregulate expression of NF-{kappa}B p65. Black-Right-Pointing-Pointer 5-LOX enhances nuclear translocation of NF-{kappa}B p65 via increasing p-I{kappa}B-{alpha} level. Black-Right-Pointing-Pointer 5-LOX stimulates transcriptional activity of NF-{kappa}B in hepatoma cells. Black-Right-Pointing-Pointer LTB4 activates transcriptional activity of NF-{kappa}B in hepatoma cells. -- Abstract: The issue that lipid metabolism enzyme and its metabolites regulate transcription factors in cancer cell is not fully understood. In this study, we first report that the lipid metabolism enzyme 5-Lipoxygenase (5-LOX) and its metabolite leukotriene B4 (LTB4) are capable of activating nuclear factor-{kappa}B (NF-{kappa}B) in hepatoma cells. We found that the treatment of MK886 (an inhibitor of 5-LOX) or knockdown of 5-LOX was able to downregulate the expression of NF-{kappa}B p65 at the mRNA level and decreased the phosphorylation level of inhibitor {kappa}B{alpha} (I{kappa}B{alpha}) in the cytoplasm of hepatoma HepG2 or H7402 cells, which resulted in the decrease of the level of nuclear NF-{kappa}B p65. These were confirmed by immunofluorescence staining in HepG2 cell. Moreover, the above treatments were able to decrease the transcriptional activity of NF-{kappa}B in the cells. The LTB4, one of metabolites of 5-LOX, is responsible for 5-LOX-activated NF-{kappa}B in a dose-dependent manner. Thus, we conclude that the lipid metabolism enzyme 5-LOX and its metabolite LTB4 are capable of activating transcription factor NF-{kappa}B in hepatoma cells. Our finding provides new insight into the significance of lipid metabolism in activation of transcription factors in cancer.

  9. Three New and Eleven Known Unusual C25 Steroids: Activated Production of Silent Metabolites in a Marine-Derived Fungus by Chemical Mutagenesis Strategy using Diethyl Sulphate

    PubMed Central

    Xia, Ming-Wen; Cui, Cheng-Bin; Li, Chang-Wei; Wu, Chang-Jing

    2014-01-01

    Three new (1–3) and 11 known (4–14) C25 steroids with an unusual bicyclo[4.4.1]A/B ring system were isolated by tracing newly produced metabolites in the EtOAc extract of an antitumor mutant AD-1-2 obtained by the diethyl sulphate (DES) mutagenesis of a marine-derived Penicillium purpurogenum G59. HPLC-PDAD-UV and HPLC-ESI-MS analyses indicated that the G59 strain did not produce these metabolites and the production of 1–14 in the mutant AD-1-2 extract was caused by the activation of silent metabolites in the original G59 strain by DES mutagenesis. The structures of the new compounds, named antineocyclocitrinols A (1) and B (2) and 23-O-methylantineocyclocitrinol (3), including their absolute configurations were determined by various spectroscopic methods, especially the NMR and Mo2-induced CD analyses. Compounds 1–3 provide the first examples of the C25 bicyclo[4.4.1]A/B ring steroids with the Z-configuration of 20,22-double bond. All of 1–14 weakly inhibited several human cancer cell lines to varying extents. These results provided additional examples for the successful application of the chemical mutagenesis strategy using DES to discover new compounds by activating silent metabolites in fungal isolates and supported also the effectiveness and usefulness of this new strategy. PMID:24633254

  10. A facile reproducible radioimmunoassay of the mixed metabolites of prostaglandins E, suitable for measurement of relative differences of phospholipase/prostaglandin synthetase activity in vivo.

    PubMed

    Fretland, D J; Cammarata, P S

    1984-04-01

    A relatively simple, reproducible, radioimmunoassay for the mixed metabolites of prostaglandins E (U-PGE-M) in rat and human urine is described. Results of the assay of treated versus control urine extracts correlate well with differences expected from treatments known to alter in vivo phospholipase/prostaglandin synthetase activity. Cross-reactivity of heterogeneous metabolite antiserum with 5 available endogenous prostaglandins and a single metabolite was determined and showed little or no cross reaction. Sensitivity, within-assay precision, interassay reproducibility, and parallelism were also determined and found acceptable. Excretion rates of U-PGE-M by rats and humans were determined, and statistically significant differences could be shown, although absolute values were smaller than estimated absolute values obtained from mass-spectrometric measurements of single, purified metabolites. Normal human male excretion rates differed significantly from those of females. Injection of prostaglandin E1 caused a significant rise in U-PGE-M excretion in rats whereas aspirin and indomethacin caused it to fall. U-PGE-M excretion rates of spontaneous hypertensive rats were significantly less than rates of normotensive controls. Adrenalectomy resulted in excretion of significantly larger amounts of U-PGE-M than in normal or sham-operated controls. A screen of clinically active pharmacological agents and hormones gave results consistent with previously published reports. PMID:6427792

  11. Characterization of two water-soluble lignin metabolites with antiproliferative activities from Inonotus obliquus.

    PubMed

    Wang, Qingjie; Mu, Haibo; Zhang, Lin; Dong, Dongqi; Zhang, Wuxia; Duan, Jinyou

    2015-03-01

    The chaga mushroom, Inonotus obliquus has long been recognized as a remedy for cancer, gastritis, ulcers, and tuberculosis of the bones since the 16th century. Herein we reported the identification of two homogenous biological macromolecules, designated as IOW-S-1 and IOW-S-2 with anti-tumor activities from the hot-water extract of I. obliquus. Their molecular weights were determined to be 37.9 and 24.5kDa by high performance gel permeation chromatography (HPGPC) respectively. Chemical and spectral analysis indicated that both IOW-S-1 and IOW-S-2 were predominant in lignin, along with ∼20% carbohydrates. Examination of cytotoxicity showed that these two lignin-carbohydrate complexes induced cell death in a concentration dependent manner, while this apoptosis induction was largely cell-cycle independent. Further investigation demonstrated that IOW-S-1 or IOW-S-2 inhibited the activation of the nuclear transcription factor in cancer cells. These findings implied that soluble lignin derivatives were one of bioactive components in I. obliquus, and further provided insights into the understanding of molecular basis for diverse medicinal and nutritional values of this mushroom. PMID:25583019

  12. Natural phenolic metabolites from endophytic Aspergillus sp. IFB-YXS with antimicrobial activity.

    PubMed

    Zhang, Wenjing; Wei, Wei; Shi, Jing; Chen, Chaojun; Zhao, Guoyan; Jiao, Ruihua; Tan, Renxiang

    2015-07-01

    Prompted by the pressing necessity to conquer phytopathogenic infections, the antimicrobial compounds were characterized with bioassay-guided method from the ethanol extract derived from the solid-substrate fermentation of Aspergillus sp. IFB-YXS, an endophytic fungus residing in the apparently healthy leave of Ginkgo biloba L. The aim of this work was to evaluate the antimicrobial activity and mechanism(s) of these bioactive compounds against phytopathogens. Among the compounds, xanthoascin (1) is significantly inhibitory on the growth of the phytopathogenic bacterium Clavibacter michiganense subsp. Sepedonicus with a minimum inhibitory concentration (MIC) value of 0.31μg/ml, which is more potent than streptomycin (MIC 0.62μg/ml), an antimicrobial drug co-assayed herein as a positive reference. Moreover, terphenyl derivatives 3, 5 and 6 are also found to be active against other phytopathogens including Xanthomonas oryzae pv. oryzae Swings, Xanthomonas oryzae pv. oryzicola Swings, Erwinia amylovora and Pseudomonas syringae pv. lachrymans etc. The antibacterial mechanism of xanthoascin (1) was addressed to change the cellular permeability of the phytopathogens, leading to the remarkable leakage of nucleic acids out of the cytomembrane. The work highlights the possibility that xanthoascin (1), an analogue of xanthocillin which is used to be an approved antibiotic, may find its renewed application as a potent antibacterial agrichemical. This study contributes to the development of new antimicrobial drugs, especially against C. michiganense subsp. Sepedonicus. PMID:26004581

  13. The In Vitro Antimicrobial Activities of Metabolites from Lactobacillus Strains on Candida Species Implicated in Candida Vaginitis

    PubMed Central

    Ogunshe, Adenike A O; Omotoso, Mopelola A; Bello, Victoria B

    2011-01-01

    Background: Research from developing countries, such as Nigeria, on Lactobacillus species in the female urogenital tract and their role as a barrier to vaginal infection is limited. Therefore, the aim of this study was to assess the clinical biotherapeutic potential of indigenous Lactobacillus species. Methods: Antimicrobial metabolites production were characterised using simple and easily reproducible qualitative and quantitative methods. The in vitro inhibitory effect of Lactobacillus antimicrobials on vulvovaginal candidiasis–associated Candida species was investigated using modified agar spot and agar well-diffusion methods. Results: The maximum levels of lactic acid, hydrogen peroxide, and diacetyl from 20 vaginal Lactobacillus strains from diseased subjects were 1.46 mg/L, 1.36 mmol/L, and 1.72 mg/L respectively. From the 4 healthy subjects, the maximum level of lactic acid was 1.08 mg/L; hydrogen peroxide, 1.36 mmol/L; and diacetyl, 0.86 mg/L. The maximum productions of these substances occurred between 72 and 120 hours of incubation. The in vitro antagonistic activities of vaginal L. acidophilus, L. fermentum, L. brevis, L. plantarum, L. casei, L. delbrueckii, and L. jensenii from diseased subjects inhibited a maximum of 5.71% of the 35 Candida species tested, while vaginal L. acidophilus and L. plantarum from healthy subjects inhibited between 57.1% and 68.6% of Candida species in vitro. Conclusion: Antimicrobial-producing lactobacilli can be considered as adjunct biotherapeutic candidates for the treatment of vulvovaginal candidiasis. PMID:22589669

  14. Bioaccessible (poly)phenol metabolites from raspberry protect neural cells from oxidative stress and attenuate microglia activation.

    PubMed

    Garcia, Gonçalo; Nanni, Sara; Figueira, Inês; Ivanov, Ines; McDougall, Gordon J; Stewart, Derek; Ferreira, Ricardo B; Pinto, Paula; Silva, Rui F M; Brites, Dora; Santos, Cláudia N

    2017-01-15

    Neuroinflammation is an integral part of the neurodegeneration process inherent to several aging dysfunctions. Within the central nervous system, microglia are the effective immune cells, responsible for neuroinflammatory responses. In this study, raspberries were subjected to in vitro digestion simulation to obtain the components that result from the gastrointestinal (GI) conditions, which would be bioaccessible and available for blood uptake. Both the original raspberry extract and the gastrointestinal bioaccessible (GIB) fraction protected neuronal and microglia cells against H2O2-induced oxidative stress and lipopolysaccharide (LPS)-induced inflammation, at low concentrations. Furthermore, this neuroprotective capacity was independent of intracellular ROS scavenging mechanisms. We show for the first time that raspberry metabolites present in the GIB fraction significantly inhibited microglial pro-inflammatory activation by LPS, through the inhibition of Iba1 expression, TNF-α release and NO production. Altogether, this study reveals that raspberry polyphenols may present a dietary route to the retardation or amelioration of neurodegenerative-related dysfunctions. PMID:27542476

  15. Aspirin’s Active Metabolite Salicylic Acid Targets High Mobility Group Box 1 to Modulate Inflammatory Responses

    PubMed Central

    Choi, Hyong Woo; Tian, Miaoying; Song, Fei; Venereau, Emilie; Preti, Alessandro; Park, Sang-Wook; Hamilton, Keith; Swapna, G V T; Manohar, Murli; Moreau, Magali; Agresti, Alessandra; Gorzanelli, Andrea; De Marchis, Francesco; Wang, Huang; Antonyak, Marc; Micikas, Robert J; Gentile, Daniel R; Cerione, Richard A; Schroeder, Frank C; Montelione, Gaetano T; Bianchi, Marco E; Klessig, Daniel F

    2015-01-01

    Salicylic acid (SA) and its derivatives have been used for millennia to reduce pain, fever and inflammation. In addition, prophylactic use of acetylsalicylic acid, commonly known as aspirin, reduces the risk of heart attack, stroke and certain cancers. Because aspirin is rapidly de-acetylated by esterases in human plasma, much of aspirin’s bioactivity can be attributed to its primary metabolite, SA. Here we demonstrate that human high mobility group box 1 (HMGB1) is a novel SA-binding protein. SA-binding sites on HMGB1 were identified in the HMG-box domains by nuclear magnetic resonance (NMR) spectroscopic studies and confirmed by mutational analysis. Extracellular HMGB1 is a damage-associated molecular pattern molecule (DAMP), with multiple redox states. SA suppresses both the chemoattractant activity of fully reduced HMGB1 and the increased expression of proinflammatory cytokine genes and cyclooxygenase 2 (COX-2) induced by disulfide HMGB1. Natural and synthetic SA derivatives with greater potency for inhibition of HMGB1 were identified, providing proof-of-concept that new molecules with high efficacy against sterile inflammation are attainable. An HMGB1 protein mutated in one of the SA-binding sites identified by NMR chemical shift perturbation studies retained chemoattractant activity, but lost binding of and inhibition by SA and its derivatives, thereby firmly establishing that SA binding to HMGB1 directly suppresses its proinflammatory activities. Identification of HMGB1 as a pharmacological target of SA/aspirin provides new insights into the mechanisms of action of one of the world’s longest and most used natural and synthetic drugs. It may also provide an explanation for the protective effects of low-dose aspirin usage. PMID:26101955

  16. Biodegradation of Triton X-100 and its primary metabolites by a bacterial community isolated from activated sludge.

    PubMed

    Wyrwas, Bogdan; Dymaczewski, Zbysław; Zgoła-Grześkowiak, Agnieszka; Szymański, Andrzej; Frańska, Magdalena; Kruszelnicka, Izabela; Ginter-Kramarczyk, Dobrochna; Cyplik, Paweł; Ławniczak, Łukasz; Chrzanowski, Łukasz

    2013-10-15

    A set of studies was carried using a continuous flow biodegradation unit in order to isolate a microbial community capable of efficient and complete utilization of octylphenol ethoxylates from activated sludge. Increasing concentrations of Triton X-100 (in the range of 1-1000 mg/l) were applied over a time period of 35 days in order to select microorganisms, which exhibit high tolerance towards this surfactant. The fate of the surfactant and its primary degradation products was assessed by HPLC/MS. It was observed that even small doses of the surfactant contributed to the disruption of the activated sludge, due to adsorption of primary Triton X-100 metabolites (octylphenol and short-chained ethoxylates) on the cells, although the long-chain octylphenol ethoxylates were efficiently degraded during the isolation process. The toxicity assessment of octylphenol as well as octylphenol di- and monoethoxylates towards activated sludge allowed for determination of EC50 values (8 and 55 mg/l, respectively). The identification of the residual microorganisms revealed the presence of Acinetobacter junii, Acinetobacter calcoaceticus, Aeromonas hydrophilia, Alcaligenes spp., Pseudomonas fluorescens and Sphingomonas capsulata. The isolated community exhibited a high resistance towards Triton X-100 and was capable of growth even at 10,000 mg/l, with the highest specific growth rate (0.47 h(-1)) observed at 4000 mg/l. Under aerobic conditions both octylphenol and the short-chained ethoxylates were completely degraded while no toxic effect towards the isolated bacterial community was observed. PMID:23770380

  17. Purification of human dihydro-orotate dehydrogenase and its inhibition by A77 1726, the active metabolite of leflunomide.

    PubMed Central

    Bruneau, J M; Yea, C M; Spinella-Jaegle, S; Fudali, C; Woodward, K; Robson, P A; Sautès, C; Westwood, R; Kuo, E A; Williamson, R A; Ruuth, E

    1998-01-01

    Leflunomide is currently in phase-III clinical trials for the treatment of rheumatoid arthritis. In this study, we have focused our efforts on the study of the mechanism of action of the active metabolite of leflunomide, A77 1726, in cells and tissue of human origin. The human high-affinity binding protein for radiolabelled A77 1726 was purified from solubilized U937 membranes by following the binding activity through the purification process and was characterized as the mitochondrial enzyme dihydro-orotate dehydrogenase (DHO-DH). The human and murine enzyme displayed identical pI and molecular mass values on SDS/PAGE (43 kDa), which contrasts notably with previous reports suggesting a molecular mass of 50 kDa for the human enzyme. DHO-DH activity was inhibited by A77 1726 and its analogue HR325 with similar potency in U937 and human spleen membrane preparations. HR325 was found to be anti-proliferative for phytohaemagglutinin-stimulated human peripheral blood mononuclear cells, at the same concentrations that caused accumulation of DHO and depletion of uridine. Supplementation of the cultures with exogenous uridine led to partial abrogation of the anti-proliferative effect. This is in line with our recent demonstration that the anti-proliferative effect in vitro of A77 1726 on lipopolysaccharide-stimulated mouse spleen cells was mediated by DHO-DH inhibition [Williamson, Yea, Robson, Curnock, Gadher, Hambleton, Woodward, Bruneau, Hambleton, Moss et al., (1995) J. Biol. Chem. 270, 22467-22472]. Thus, DHO-DH inhibition by A77 1726 and its analogues is responsible for the anti-proliferative effects in vitro of the compounds on human cells and is likely to be responsible for some of its effects in vivo. PMID:9820804

  18. Targeted metabolite analysis and biological activity of Pieris brassicae fed with Brassica rapa var. rapa.

    PubMed

    Pereira, David M; Noites, Alexandra; Valentão, Patricia; Ferreres, Federico; Pereira, José A; Vale-Silva, Luis; Pinto, Eugénia; Andrade, Paula B

    2009-01-28

    For the first time, an insect-plant system, Pieris brassicae fed with Brassica rapa var. rapa, was tested for its biological capacity, namely, antioxidant (DPPH*, *NO, and O(2)*- radicals) and antimicrobial (bacteria and fungi) activities. Samples from the insect's life cycle (larvae, excrements, exuviae, and butterfly) were always found to be more efficient than the host plant. Also, P. brassicae materials, as well as its host plant, were screened for phenolics and organic acids. The host plant revealed higher amounts of both compounds. Two phenolic acids, ferulic and sinapic, as well as kaempferol 3-Osophoroside, were common to insect (larvae and excrements) and plant materials, with excrements being considerably richer. Detection of sulfated compounds in excrements, absent in host plant, revealed that metabolic processes in this species involved sulfation. Additionally, deacylation and deglycosilation were observed. All matrices presented the same organic acids qualitative profile, with the exception of excrements. PMID:19115952

  19. Isolation, structure, and HIV-1-integrase inhibitory activity of structurally diverse fungal metabolites.

    PubMed

    Singh, Sheo B; Jayasuriya, Hiranthi; Dewey, Raymond; Polishook, Jon D; Dombrowski, Anne W; Zink, Deborah L; Guan, Ziqiang; Collado, Javier; Platas, Gonzalo; Pelaez, Fernando; Felock, Peter J; Hazuda, Daria J

    2003-12-01

    HIV-1 integrase is a critical enzyme for replication of HIV, and its inhibition is one of the most promising new drug strategies for anti-retroviral therapy, with potentially significant advantages over existing therapies. In this report, a series of HIV-1 inhibitors isolated from the organic extract of fermentations from terrestrial fungi is described. These fungal species, belonging to a variety of genera, were collected from throughout the world following the strict guidelines of Rio Convention on Biodiversity. The polyketide- and terpenoid-derived inhibitors are represented by two naphthoquinones, a biphenyl and two triphenyls, a benzophenone, four aromatics with or without catechol units, a linear aliphatic terpenoid, a diterpenoid, and a sesterterpenoid. These compounds inhibited the coupled and strand-transfer reaction of HIV-1 integrase with an IC(50) value of 0.5-120 micro M. The bioassay-directed isolation, structure elucidation, and HIV-1 inhibitory activity of these compounds are described. PMID:14714192

  20. Chemistry of the Nudibranch Aldisa andersoni: Structure and Biological Activity of Phorbazole Metabolites

    PubMed Central

    Nuzzo, Genoveffa; Ciavatta, Maria Letizia; Kiss, Robert; Mathieu, Véronique; Leclercqz, Helene; Manzo, Emiliano; Villani, Guido; Mollo, Ernesto; Lefranc, Florence; D’Souza, Lisette; Gavagnin, Margherita; Cimino, Guido

    2012-01-01

    The first chemical study of the Indo-Pacific dorid nudibranch Aldisa andersoni resulted in the isolation of five chlorinated phenyl-pyrrolyloxazoles belonging to the phorbazole series. Two new molecules, 9-chloro-phorbazole D and N1-methyl-phorbazole A, co-occurring with known phorbazoles A, B and D, have been characterized. Phorbazoles were found to be present mainly in the external part of the mollusc. The structures of the new compounds were determined by interpretation of spectroscopic data, mainly NMR and mass spectrometry and by comparison with the literature data. Evaluation of feeding-deterrence activity as well as in vitro growth inhibitory properties in human cancer cells was also carried out. PMID:23015775

  1. Colonic metabolites of berry polyphenols: the missing link to biological activity?

    PubMed

    Williamson, Gary; Clifford, Michael N

    2010-10-01

    The absorption of dietary phenols, polyphenols and tannins (PPT) is an essential step for biological activity and effects on health. Although a proportion of these dietary bioactive compounds are absorbed intact, depending on their chemical structure and the nature of any attached moiety (e.g. sugar, organic acid), substantial amounts of lower molecular weight catabolites are absorbed after biotransformation by the colon microflora. The main products in the colon are (a) benzoic acids (C6-C1), especially benzoic acid and protocatechuic acid; (b) phenylacetic acids (C6-C2), especially phenylacetic acid per se; (c) phenylpropionic acids (C6-C3), where the latter are almost entirely in the dihydro form, notably dihydrocaffeic acid, dihydroferulic acid, phenylpropionic acid and 3-(3'-hydroxyphenyl)-propionic acid. As a result of this biotransformation, some of these compounds can each reach mm concentrations in faecal water. Many of these catabolites are efficiently absorbed in the colon, appear in the blood and are ultimately excreted in the urine. In the case of certain polyphenols, such as anthocyanins, these catabolites are major products in vivo; protocatechuic acid is reported to represent a substantial amount of the ingested dose of cyanidin-3-O-glucoside. The major catabolites of berries, and especially blackcurrants, are predicted based on compositional data for polyphenols from berries and other sources. Since microbial catabolites may be present at many sites of the body in higher concentration than the parent compound, it is proposed that at least a part of the biological activities ascribed to berry polyphenols and other PPT are due to their colonic catabolites. PMID:20955650

  2. In vitro effects of brominated flame retardants and metabolites on CYP17 catalytic activity: A novel mechanism of action?

    SciTech Connect

    Canton, Rocio F. . E-mail: r.Fernandezcanton@iras.uu.nl; Sanderson, J. Thomas; Nijmeijer, Sandra; Bergman, Ake; Letcher, Robert J.; Berg, Martin van den

    2006-10-15

    Fire incidents have decreased significantly over the last 20 years due, in part, to regulations requiring addition of flame retardants (FRs) to consumer products. Five major classes of brominated flame retardants (BFRs) are hexabromocyclododecane isomers (HBCDs), tetrabromobisphenol-A (TBBPA) and three commercial mixtures of penta-, octa- and deca-polybrominated diphenyl ether (PBDE) congeners, which are used extensively as commercial FR additives. Furthermore, concentrations of PBDEs have been rapidly increasing during the 1999s in human breast milk and a number of endocrine effects have been reported. We used the H295R human adrenocortical carcinoma cell line to assess possible effects of some of these BFRs (PBDEs and several of their hydroxylated (OH) and methoxylated (CH{sub 3}O) metabolites or analogues), TBBPA and brominated phenols (BPs) on the combined 17{alpha}-hydroxylase and 17,20-lyase activities of CYP17. CYP17 enzyme catalyzes an important step in sex steroidogenesis and is responsible for the biosynthesis of dehydroepiandrosterone (DHEA) and androstenedione in the adrenals. In order to study possible interactions with BFRs, a novel enzymatic method was developed. The precursor substrate of CYP17, pregnenolone, was added to control and exposed H295R cells, and enzymatic production of DHEA was measured using a radioimmunoassay. In order to avoid pregnenolone metabolism via different pathways, specific chemical inhibitor compounds were used. None of the parent/precursor BFRs had a significant effect (P < 0.05) on CYP17 activity except for BDE-183, which showed significant inhibition of CYP17 activity at the highest concentration tested (10 {mu}M), with no signs of cytotoxicity as measured by mitochondrial toxicity tests (MTT). A strong inhibition of CYP17 activity was found for 6-OH-2,2',4,4'-tetrabromoDE (6-OH-BDE47) with a concentration-dependent decrease of almost 90% at 10 {mu}M, but with a concurrent decrease in cell viability at the higher

  3. Expression of a retinoic acid receptor (RAR)-like protein in the embryonic and adult nervous system of a protostome species.

    PubMed

    Carter, Christopher J; Rand, Christopher; Mohammad, Imtiaz; Lepp, Amanda; Vesprini, Nicholas; Wiebe, Olivia; Carlone, Robert; Spencer, Gaynor E

    2015-01-01

    The vitamin A metabolite, retinoic acid, is an important molecule in nervous system development and regeneration in vertebrates. Retinoic acid signaling in vertebrates is mediated by two classes of nuclear receptors, the retinoid X receptors (RXRs) and the retinoic acid receptors (RARs). Recently, evidence has emerged to suggest that many effects of retinoic acid are conserved between vertebrate and invertebrate nervous systems, even though the RARs were previously thought to be a vertebrate innovation and to not exist in non-chordates. We have cloned a full-length putative RAR from the CNS of the mollusc Lymnaea stagnalis (LymRAR). Immunoreactivity for the RAR protein was found in axons of adult neurons in the central nervous system and in growth cones of regenerating neurons in vitro. A vertebrate RAR antagonist blocked growth cone turning induced by exogenous all-trans retinoic acid, possibly suggesting a role for this receptor in axon guidance. We also provide immunostaining evidence for the presence of RAR protein in the developing, embryonic CNS, where it is also found in axonal processes. Using qPCR, we determined that LymRAR mRNA is detectable in the early veliger stage embryo and that mRNA levels increase significantly during embryonic development. Putative disruption of retinoid signaling in Lymnaea embryos using vertebrate RAR antagonists resulted in abnormal eye and shell development and in some instances completely halted development, resembling the effects of all-trans retinoic acid. This study provides evidence for RAR functioning in a protostome species. PMID:25504929

  4. All-trans retinoic acid potentiates cisplatin-induced kidney injury in rats: impact of retinoic acid signaling pathway.

    PubMed

    Elsayed, Abdelrahman M; Abdelghany, Tamer M; Akool, El-Sayed; Abdel-Aziz, Abdel-Aziz H; Abdel-Bakky, Mohamed S

    2016-03-01

    Cisplatin (cis-diammine dichloroplatinum (II), CDDP) is a widely used drug for treatment of various types of cancers. However, CDDP-induced nephrotoxicity remains the main dose-limiting side effect. Retinoids are a group of vitamin A-related compounds that exert their effects through retinoid receptors activation. In this study, we investigated the effect of CDDP treatment on retinoic acid receptor-α (RAR-α) and retinoid X receptor-α (RXR-α) expression. In addition, we investigated the possible modulatory effects of RAR agonist, all-trans retinoic acid (ATRA), on CDDP-induced nephrotoxicity. Rats were treated with saline, DMSO, CDDP, ATRA, or CDDP/ATRA. Twenty-four hours after the last ATRA injection, rats were killed; blood samples were collected; kidneys were dissected; and biochemical, immunohistochemical, and histological examinations were performed. Our results revealed that CDDP treatment significantly increased serum levels of creatinine and urea, with concomitant decrease in serum albumin. Moreover, reduced glutathione (GSH) content as well as superoxide dismutase (SOD) and catalase (CAT) activities were significantly reduced with concurrent increase in kidney malondialdehyde (MDA) content following CDDP treatment. Furthermore, CDDP markedly upregulated tubular RAR-α, RXR-α, fibrin, and inducible nitric oxide synthase (iNOS) protein expression. Although administration of ATRA to control rats did not produce marked alterations in kidney function parameters, administration of ATRA to CDDP-treated rats significantly exacerbated CDDP-induced nephrotoxicity. In addition, CDDP/ATRA co-treatment significantly increased RAR-α, RXR-α, fibrin, and iNOS protein expression compared to CDDP alone. In conclusion, we report, for the first time, the crucial role of retinoid receptors in CDDP-induced nephrotoxicity. Moreover, our findings indicate that co-administration of ATRA with CDDP, although beneficial on the therapeutic effects, their deleterious effects on

  5. Retinol dehydrogenase 10 but not retinol/sterol dehydrogenase(s) regulates the expression of retinoic acid-responsive genes in human transgenic skin raft culture.

    PubMed

    Lee, Seung-Ah; Belyaeva, Olga V; Wu, Lizhi; Kedishvili, Natalia Y

    2011-04-15

    Retinoic acid is essential for skin growth and differentiation, and its concentration in skin is controlled tightly. In humans, four different members of the short-chain dehydrogenase/reductase (SDR) superfamily of proteins were proposed to catalyze the rate-limiting step in the biosynthesis of retinoic acid (the oxidation of retinol to retinaldehyde). Epidermis contains at least three of these enzymes, but their relative importance for retinoic acid biosynthesis and regulation of gene expression during growth and differentiation of epidermis is not known. Here, we investigated the effect of the four human SDRs on retinoic acid biosynthesis, and their impact on growth and differentiation of keratinocytes using organotypic skin raft culture model of human epidermis. The results of this study demonstrate that ectopic expression of retinol dehydrogenase 10 (RDH10, SDR16C4) in skin rafts dramatically increases proliferation and inhibits differentiation of keratinocytes, consistent with the increased steady-state levels of retinoic acid and activation of retinoic acid-inducible genes in RDH10 rafts. In contrast, SDRs with dual retinol/sterol substrate specificity, namely retinol dehydrogenase 4 (RoDH4, SDR9C8), RoDH-like 3α-hydroxysteroid dehydrogenase (RL-HSD, SDR9C6), and RDH-like SDR (RDHL, SDR9C4) do not affect the expression of retinoic acid-inducible genes but alter the expression levels of several components of extracellular matrix. These results reveal essential differences in the metabolic contribution of RDH10 versus retinol/sterol dehydrogenases to retinoic acid biosynthesis and provide the first evidence that non-retinoid metabolic products of retinol/sterol dehydrogenases affect gene expression in human epidermis. PMID:21345790

  6. Retinol Dehydrogenase 10 but Not Retinol/Sterol Dehydrogenase(s) Regulates the Expression of Retinoic Acid-responsive Genes in Human Transgenic Skin Raft Culture*

    PubMed Central

    Lee, Seung-Ah; Belyaeva, Olga V.; Wu, Lizhi; Kedishvili, Natalia Y.

    2011-01-01

    Retinoic acid is essential for skin growth and differentiation, and its concentration in skin is controlled tightly. In humans, four different members of the short-chain dehydrogenase/reductase (SDR) superfamily of proteins were proposed to catalyze the rate-limiting step in the biosynthesis of retinoic acid (the oxidation of retinol to retinaldehyde). Epidermis contains at least three of these enzymes, but their relative importance for retinoic acid biosynthesis and regulation of gene expression during growth and differentiation of epidermis is not known. Here, we investigated the effect of the four human SDRs on retinoic acid biosynthesis, and their impact on growth and differentiation of keratinocytes using organotypic skin raft culture model of human epidermis. The results of this study demonstrate that ectopic expression of retinol dehydrogenase 10 (RDH10, SDR16C4) in skin rafts dramatically increases proliferation and inhibits differentiation of keratinocytes, consistent with the increased steady-state levels of retinoic acid and activation of retinoic acid-inducible genes in RDH10 rafts. In contrast, SDRs with dual retinol/sterol substrate specificity, namely retinol dehydrogenase 4 (RoDH4, SDR9C8), RoDH-like 3α-hydroxysteroid dehydrogenase (RL-HSD, SDR9C6), and RDH-like SDR (RDHL, SDR9C4) do not affect the expression of retinoic acid-inducible genes but alter the expression levels of several components of extracellular matrix. These results reveal essential differences in the metabolic contribution of RDH10 versus retinol/sterol dehydrogenases to retinoic acid biosynthesis and provide the first evidence that non-retinoid metabolic products of retinol/sterol dehydrogenases affect gene expression in human epidermis. PMID:21345790

  7. Antifungal Activity Against Plant Pathogens of Metabolites from the Endophytic Fungus Cladosporium cladosporioides

    PubMed Central

    Wang, Xiaoning; Radwan, Mohamed M.; Taráwneh, Amer H.; Gao, Jiangtao; Wedge, David E.; Rosa, Luiz H.; Cutler, Horace G.; Cutler, Stephen J.

    2013-01-01

    Bioassay-guided fractionation of Cladosporium cladosporioides (Fresen.) de Vries extracts led to the isolation of four compounds, including cladosporin, 1, isocladosporin, 2, 5′-hydroxyasperentin, 3, and cladosporin-8-methyl ether, 4. An additional compound 5′,6-diacetyl cladosporin, 5, was synthesized by acetylation of compound 3. Compounds 1-5 were evaluated for antifungal activity against plant pathogens. Phomopsis viticola was the most sensitive fungus to the tested compounds. At 30 μM, compound 1 exhibited 92.7%, 90.1%, 95.4% and 79.9% growth inhibition against Colletotrichum acutatum, Co. fragariae, Co. gloeosporioides and Phomopsis viticola, respectively. Compound 2 showed 50.4%, 60.2% and 83.0% growth inhibition at 30 μM against Co. fragariae, Co. gloeosporioides and P. viticola, respectively. Compounds 3 and 4 were isolated for the first time from Cladosporium cladosporioides. Moreover, the identification of essential structural features of the cladosporin nuclei has also been evaluated. These structures provide new templates for the potential treatment and management of plant diseases. PMID:23651409

  8. Determination of some selected secondary metabolites and their invitro antioxidant activity in commercially available Ethiopian tea (Camellia sinensis).

    PubMed

    Bizuayehu, Dereje; Atlabachew, Minaleshewa; Ali, Mirtachew Tihar

    2016-01-01

    Eight brands of tea (Camellia sinensis),which are cultivated and commercially available in Ethiopian market, were analyzed for estimation of their total secondary metabolites (polyphenols, flavonoids and tannins) content and free radical scavenging activity which is expressed on dry weight basis. In this present study, the total polyphenols, tannin and flavonoid contents were studied spectrophotometrically using Folin-Dennis, Folin-Dennis/protein precipitation and aluminium chloride methods respectively. The free radical scavenging activity was determined by using DPPH radical assay. Results of the analysis revealed that the total polyphenol content varied from 21.3 ± 0.24 to 31.6 ± 0.31 mg of gallic acid equivalent/g of dry matter. Total flavonoids content in the tea samples varied from 8.17 ± 0.68 to 23.2 ± 0.68 mg of catechin equivalent/g of dry weight and tannin content varied from 5.64 ± 0.39 7.45 ± 0.27 mg tannic acid equivalent/g of dry weight basis. The free radical scavenging activity among the tea brand samples ranged from 28.8 ± 1.86 to 80.0 ± 0.63 mg ascorbic acid equivalent/g and the half maximal inhibitory concentration (IC50%) values varied from 7.3 ± 1.35 to 64.0 ± 2.81 µg/mL of extract. The correlation between the antioxidant activity with total polyphenol content (R = 0.91325), with flavonoids (R = 0.80658) and with tannin (R = 0.73125) was calculated and maximum correlation value was found between polyphenol content and the free radical scavenging activity of the tea samples. The results in this study also revealed that green tea had the higher polyphenolic content and found to have the most promising antioxidant activity. This study further confirmed that Ethiopia tea is reach in phenolic compounds as compared to some overseas tea cultivars/varieties. PMID:27069832

  9. Biosynthesis and metabolism of retinoic acid: roles of CRBP and CRABP in retinoic acid: roles of CRBP and CRABP in retinoic acid homeostasis.

    PubMed

    Napoli, J L

    1993-02-01

    The enzymes that constitute the pathway of retinoic acid biosynthesis and metabolism may recognize retinoid binding proteins as effectors and substrates. Apocellular retinol-binding protein (CRBP) stimulates a bile-salt independent membrane-bound retinyl ester hydrolase resulting in the hydrolysis of endogenous retinyl esters and the formation of holoCRBP. HoloCRBP delivers retinol to a microsomal nicotin-amide-adenine dinucleotide phosphate-dependent dehydrogenase, protects it from artifactual oxidation and denies enzymes that cannot recognize the binding protein access to retinol. The retinal synthesized may be transferred from the microsomes to the cytosol by CRBP. A cytosolic retinal dehydrogenase has been purified that produces retinoic acid from retinal generated by microsomes in the presence of CRBP and from the complex CRBP-retinal itself. Thus, CRBP(type I) seems to channel retinoids through the reactions of retinoic acid synthesis via a series of protein-protein interactions. Cellular retinoic acid-binding protein (type I) facilitates retinoic acid metabolism by sequestering it and by acting as a low Km substrate, thereby also modulating the steady-state concentrations of retinoic acid. PMID:8381481

  10. Activation and silencing of secondary metabolites in Streptomyces albus and Streptomyces lividans after transformation with cosmids containing the thienamycin gene cluster from Streptomyces cattleya.

    PubMed

    Braña, Alfredo F; Rodríguez, Miriam; Pahari, Pallab; Rohr, Jurgen; García, Luis A; Blanco, Gloria

    2014-05-01

    Activation and silencing of antibiotic production was achieved in Streptomyces albus J1074 and Streptomyces lividans TK21 after introduction of genes within the thienamycin cluster from S. cattleya. Dramatic phenotypic and metabolic changes, involving activation of multiple silent secondary metabolites and silencing of others normally produced, were found in recombinant strains harbouring the thienamycin cluster in comparison to the parental strains. In S. albus, ultra-performance liquid chromatography purification and NMR structural elucidation revealed the identity of four structurally related activated compounds: the antibiotics paulomycins A, B and the paulomenols A and B. Four volatile compounds whose biosynthesis was switched off were identified by gas chromatography-mass spectrometry analyses and databases comparison as pyrazines; including tetramethylpyrazine, a compound with important clinical applications to our knowledge never reported to be produced by Streptomyces. In addition, this work revealed the potential of S. albus to produce many others secondary metabolites normally obtained from plants, including compounds of medical relevance as dihydro-β-agarofuran and of interest in perfume industry as β-patchoulene, suggesting that it might be an alternative model for their industrial production. In S. lividans, actinorhodins production was strongly activated in the recombinant strains whereas undecylprodigiosins were significantly reduced. Activation of cryptic metabolites in Streptomyces species might represent an alternative approach for pharmaceutical drug discovery. PMID:24633227

  11. INHIBITION OF RETINOID ACTIVITY BY COMPONENTS OF A PAPER MILL EFFLUENT

    EPA Science Inventory

    A cell line stably transfected with reporter genes activated by retinoic acid was used to test a paper mill effluent for the presence of retinoids or components that interfere with retinoic acid-stimulated gene transcription.

  12. Effects of chloro-s-triazine herbicides and metabolites on aromatase activity in various human cell lines and on vitellogenin production in male carp hepatocytes.

    PubMed Central

    Sanderson, J T; Letcher, R J; Heneweer, M; Giesy, J P; van den Berg, M

    2001-01-01

    We investigated a potential mechanism for the estrogenic properties of three chloro-s-triazine herbicides and six metabolites in vitro in several cell systems. We determined effects on human aromatase (CYP19), the enzyme that converts androgens to estrogens, in H295R (adrenocortical carcinoma), JEG-3 (placental choriocarcinoma), and MCF-7 (breast cancer) cells; we determined effects on estrogen receptor-mediated induction of vitellogenin in primary hepatocyte cultures of adult male carp (Cyprinus carpio). In addition to atrazine, simazine, and propazine, two metabolites--atrazine-desethyl and atrazine-desisopropyl--induced aromatase activity in H295R cells concentration-dependently (0.3-30 microM) and with potencies similar to those of the parent triazines. After a 24-hr exposure to 30 microM of the triazines, an apparent maximum induction of about 2- to 2.5-fold was achieved. The induction responses were confirmed by similar increases in CYP19 mRNA levels, determined by reverse-transcriptase polymerase chain reaction. In JEG-3 cells, where basal aromatase expression is about 15-fold greater than in H295R cells, the induction responses were similar but less pronounced; aromatase expression in MCF-7 cells was neither detectable nor inducible under our culture conditions. The fully dealkylated metabolite atrazine-desethyl-desisopropyl and the three hydroxylated metabolites (2-OH-atrazine-desethyl, -desisopropyl, and -desethyl-desisopropyl) did not induce aromatase activity. None of the triazine herbicides nor their metabolites induced vitellogenin production in male carp hepatocytes; nor did they antagonize the induction of vitellogenin by 100 nM (EC(50) 17beta-estradiol. These findings together with other reports indicate that the estrogenic effects associated with the triazine herbicides in vivo are not estrogen receptor-mediated, but may be explained partly by their ability to induce aromatase in vitro. PMID:11675267

  13. Substitution of Wheat for Corn in Beef Cattle Diets: Digestibility, Digestive Enzyme Activities, Serum Metabolite Contents and Ruminal Fermentation.

    PubMed

    Liu, Y F; Zhao, H B; Liu, X M; You, W; Cheng, H J; Wan, F C; Liu, G F; Tan, X W; Song, E L; Zhang, X L

    2016-10-01

    The objective of this study was to evaluate the effect of diets containing different amounts of wheat, as a partial or whole substitute for corn, on digestibility, digestive enzyme activities, serum metabolite contents and ruminal fermentation in beef cattle. Four Limousin×LuXi crossbred cattle with a body weight (400±10 kg), fitted with permanent ruminal, proximal duodenal and terminal ileal cannulas, were used in a 4×4 Latin square design with four treatments: Control (100% corn), 33% wheat (33% substitution for corn), 67% wheat (67% substitution for corn), and 100% wheat (100% substitution for corn) on a dry matter basis. The results showed that replacing corn with increasing amounts of wheat increased the apparent digestibility values of dry matter, organic matter, and crude protein (p<0.05). While the apparent digestibility of acid detergent fiber and neutral detergent fiber were lower with increasing amounts of wheat. Digestive enzyme activities of lipase, protease and amylase in the duodenum were higher with increasing wheat amounts (p<0.05), and showed similar results to those for the enzymes in the ileum except for amylase. Increased substitution of wheat for corn increased the serum alanine aminotransferase concentration (p<0.05). Ruminal pH was not different between those given only corn and those given 33% wheat. Increasing the substitution of wheat for corn increased the molar proportion of acetate and tended to increase the acetate-to-propionate ratio. Cattle fed 100% wheat tended to have the lowest ruminal NH3-N concentration compared with control (p<0.05), whereas no differences were observed among the cattle fed 33% and 67% wheat. These findings indicate that wheat can be effectively used to replace corn in moderate amounts to meet the energy and fiber requirements of beef cattle. PMID:26954111

  14. Cardiac Energy Dependence on Glucose Increases Metabolites Related to Glutathione and Activates Metabolic Genes Controlled by Mechanistic Target of Rapamycin

    PubMed Central

    Schisler, Jonathan C.; Grevengoed, Trisha J.; Pascual, Florencia; Cooper, Daniel E.; Ellis, Jessica M.; Paul, David S.; Willis, Monte S.; Patterson, Cam; Jia, Wei; Coleman, Rosalind A.

    2015-01-01

    Background Long chain acyl‐CoA synthetases (ACSL) catalyze long‐chain fatty acids (FA) conversion to acyl‐CoAs. Temporal ACSL1 inactivation in mouse hearts (Acsl1H−/−) impaired FA oxidation and dramatically increased glucose uptake, glucose oxidation, and mTOR activation, resulting in cardiac hypertrophy. We used unbiased metabolomics and gene expression analyses to elucidate the cardiac cellular response to increased glucose use in a genetic model of inactivated FA oxidation. Methods and Results Metabolomics analysis identified 60 metabolites altered in Acsl1H−/− hearts, including 6 related to glucose metabolism and 11 to cysteine and glutathione pathways. Concurrently, global cardiac transcriptional analysis revealed differential expression of 568 genes in Acsl1H−/− hearts, a subset of which we hypothesized were targets of mTOR; subsequently, we measured the transcriptional response of several genes after chronic mTOR inhibition via rapamycin treatment during the period in which cardiac hypertrophy develops. Hearts from Acsl1H−/− mice increased expression of several Hif1α‐responsive glycolytic genes regulated by mTOR; additionally, expression of Scl7a5, Gsta1/2, Gdf15, and amino acid‐responsive genes, Fgf21, Asns, Trib3, Mthfd2, were strikingly increased by mTOR activation. Conclusions The switch from FA to glucose use causes mTOR‐dependent alterations in cardiac metabolism. We identified cardiac mTOR‐regulated genes not previously identified in other cellular models, suggesting heart‐specific mTOR signaling. Increased glucose use also changed glutathione‐related pathways and compensation by mTOR. The hypertrophy, oxidative stress, and metabolic changes that occur within the heart when glucose supplants FA as a major energy source suggest that substrate switching to glucose is not entirely benign. PMID:25713290

  15. Antibacterial and antioxidant activity of the secondary metabolites from in vitro cultures of the Alice sundew (Drosera aliciae).

    PubMed

    Krolicka, Aleksandra; Szpitter, Anna; Maciag, Monika; Biskup, Edyta; Gilgenast, Ewelina; Wegrzyn, Grazyna; Lojkowska, Ewa

    2009-07-01

    The objective of the present study was to evaluate the antioxidant as well as the antibacterial properties of secondary metabolites obtained from Drosera aliciae (Alice sundew) plants grown in vitro and to examine the mechanism of their antimicrobial action. Bactericidal activity of extracts from D. aliciae, as well as pure ramentaceone (naphthoquinone), which is present in this plant, were examined against human pathogenic strains of micro-organisms that are both resistant and susceptible to antibiotics. A chloroform extract proved to be more effective than a methanol preparation against all of the tested strains, except for Pseudomonas aeruginosa isolates. The lowest minimal-bactericidal-concentration value was in the case of Staphylococcus aureus (25-50 mg fresh weight·ml(-1)). The influence of D. aliciae extracts and ramentaceone on the synthesis of DNA, RNA or proteins in cultures of Enterococcus faecalis was estimated by measurement of the incorporation of the radioactively labelled precursors [3H]thymidine, [3H]uridine or [3H]leucine respectively. The methanol extract of D. aliciae, except for a moderate effect on DNA synthesis, had no influence on RNA and protein synthesis. The chloroform preparation caused about a 75% decrease in [3H]uridine incorporation in comparison with the control after 60 min and a significant diminution in DNA and protein synthesis (44 and 30% respectively). Ramentaceone also decreased DNA and RNA synthesis, but less efficiently than did the chloroform extract, and it caused no changes in [3H]leucine incorporation. The methanol extract from D. aliciae proved to be an effective antioxidant in both the DPPH (2,2-diphenyl-10-picrylhydrazyl free radical) and the FRAP (ferric reducing antioxidant power) assay, with the activities exceeding those of well-known antioxidants, namely the flavonoids. The chloroform extract and ramentaceone showed no antioxidative properties. PMID:18782083

  16. Enhanced active metabolite generation and platelet inhibition with prasugrel compared to clopidogrel regardless of genotype in thienopyridine metabolic pathways.

    PubMed

    Braun, Oscar Ö; Angiolillo, Dominick J; Ferreiro, Jose L; Jakubowski, Joseph A; Winters, Kenneth J; Effron, Mark B; Duvvuru, Suman; Costigan, Timothy M; Sundseth, Scott; Walker, Joseph R; Saucedo, Jorge F; Kleiman, Neal S; Varenhorst, Christoph

    2013-12-01

    Clopidogrel response varies according to the presence of genetic polymorphisms. The CYP2C19*2 allele has been associated with impaired response; conflicting results have been reported for CYP2C19*17, ABCB1, and PON1 genotypes. We assessed the impact of CYP2C19, PON1, and ABCB1 polymorphisms on clopidogrel and prasugrel pharmacodynamic (PD) and pharmacokinetic (PK) parameters. Aspirin-treated patients (N=194) with coronary artery disease from two independent, prospective, randomised, multi-centre studies comparing clopidogrel (75 mg) and prasugrel (10 mg) were genotyped and classified by predicted CYP2C19 metaboliser phenotype (ultra metabolisers [UM] = *17 carriers; extensive metabolisers [EM] = *1/1 homozygotes; reduced metabolisers [RM] = *2 carriers). ABCB1 T/T and C/T polymorphisms and PON1 A/A, A/G and G/G polymorphisms were also genotyped. PD parameters were assessed using VerifyNow® P2Y12 and vasodilator stimulated phosphoprotein (VASP) expressed as platelet reactivity index (PRI) after 14 days of maintenance dosing. Clopidogrel and prasugrel active metabolite (AM) exposure was calculated in a cohort of 96 patients. For clopidogrel, genetic variants in CYP2C19, but not ABCB1 or PON1, affected PK and PD. For prasugrel, none of the measured genetic variants affected PK or PD. Compared with clopidogrel, platelet inhibition with prasugrel was greater even in the CYP2C19 UM phenotype. Prasugrel generated more AM and achieved greater platelet inhibition than clopidogrel irrespective of CYP2C19, ABCB1, and PON1 polymorphisms. The lack of effect from genetic variants on prasugrel AM generation or antiplatelet activity is consistent with previous studies in healthy volunteers and is consistent with improved efficacy in acute coronary syndrome patients managed with percutaneous coronary intervention. PMID:24009042

  17. Retinoic acid induces cells cultured from oral squamous cell carcinomas to become anti-angiogenic.

    PubMed Central

    Lingen, M. W.; Polverini, P. J.; Bouck, N. P.

    1996-01-01

    Retinoids have shown great promise as chemopreventive against the development of squamous cell carcinomas of the upper aerodigestive tract. However, the exact mechanism by which they block new tumors from arising is unknown. Here, we report that 13-cis- and all-trans-retinoic acid, used at clinically achievable doses of 10(-6) mol/L or less, can directly and specifically affect cell lines cultured from oral squamous cell carcinomas, inducing them to switch from an angiogenic to an anti-angiogenic phenotype. Although retinoic-acid-treated and untreated tumor cells make the same amount of interleukin-8, the major inducer of neovascularization produced by such tumor lines, they vary in production of inhibitory activity. Only the retinoic-acid-treated cells produce a potent angio-inhibitory activity that is able to block in vitro migration of endothelial cells toward tumor cell conditioned media and to halt neovascularization induced by such media in the rat cornea. Anti-angiogenic activity is induced in the tumor cells by low doses of retinoids in the absence of toxicity with a kinetics that suggest that it could be contributing to the effectiveness of the retinoids as chemopreventive agents. Images Figure 6 PMID:8686749

  18. All-trans and 9-cis retinoic acid alter rat hepatic stellate cell phenotype differentially

    PubMed Central

    Hellemans, K; Grinko, I; Rombouts, K; Schuppan, D; Geerts, A

    1999-01-01

    BACKGROUND—Hepatic stellate cells exert specific functions in the liver: storage of large amounts of retinyl esters, synthesis and breakdown of hepatic extracellular matrix, secretion of a variety of cytokines, and control of the diameter of the sinusoids.
AIMS—To examine the influence of all-trans retinoic acid (ATRA) and 9-cis retinoic acid (9RA) on extracellular matrix production and proliferation of activated hepatic stellate cells.
METHODS—Cells were isolated using collagenase/pronase, purified by centrifugation in nycodenz, and cultured for two weeks. At this time point the cells exhibited the activated phenotype. Cells were exposed to various concentrations of ATRA and 9RA. The expression of procollagens I, III, and IV, of fibronectin and of laminin were analysed by immunoprecipitation and northern hybridisation.
RESULTS—ATRA exerted a significant inhibitory effect on the synthesis of procollagens type I, III, and IV, fibronectin, and laminin, but did not influence stellate cell proliferation, whereas 9RA showed a clear but late effect on proliferation. 9RA increased procollagen I mRNA 1.9-fold, but did not affect the expression of other matrix proteins.
CONCLUSION—Results showed that ATRA and 9RA exert different, often contrary effects on activated stellate cells. These observations may explain prior divergent results obtained following retinoid administration to cultured stellate cells or in animals subjected to fibrogenic stimuli.


Keywords: hepatic stellate cells; retinoic acid; extracellular matrix proteins; proliferation PMID:10369717

  19. Secondary metabolites from Ganoderma.

    PubMed

    Baby, Sabulal; Johnson, Anil John; Govindan, Balaji

    2015-06-01

    Ganoderma is a genus of medicinal mushrooms. This review deals with secondary metabolites isolated from Ganoderma and their biological significance. Phytochemical studies over the last 40years led to the isolation of 431 secondary metabolites from various Ganoderma species. The major secondary compounds isolated are (a) C30 lanostanes (ganoderic acids), (b) C30 lanostanes (aldehydes, alcohols, esters, glycosides, lactones, ketones), (c) C27 lanostanes (lucidenic acids), (d) C27 lanostanes (alcohols, lactones, esters), (e) C24, C25 lanostanes (f) C30 pentacyclic triterpenes, (g) meroterpenoids, (h) farnesyl hydroquinones (meroterpenoids), (i) C15 sesquiterpenoids, (j) steroids, (k) alkaloids, (l) prenyl hydroquinone (m) benzofurans, (n) benzopyran-4-one derivatives and (o) benzenoid derivatives. Ganoderma lucidum is the species extensively studied for its secondary metabolites and biological activities. Ganoderma applanatum, Ganoderma colossum, Ganoderma sinense, Ganoderma cochlear, Ganoderma tsugae, Ganoderma amboinense, Ganoderma orbiforme, Ganoderma resinaceum, Ganoderma hainanense, Ganoderma concinna, Ganoderma pfeifferi, Ganoderma neo-japonicum, Ganoderma tropicum, Ganoderma australe, Ganoderma carnosum, Ganoderma fornicatum, Ganoderma lipsiense (synonym G. applanatum), Ganoderma mastoporum, Ganoderma theaecolum, Ganoderma boninense, Ganoderma capense and Ganoderma annulare are the other Ganoderma species subjected to phytochemical studies. Further phytochemical studies on Ganoderma could lead to the discovery of hitherto unknown biologically active secondary metabolites. PMID:25975187

  20. Retinoic acid: its biosynthesis and metabolism.

    PubMed

    Napoli, J L

    1999-01-01

    This article presents a model that integrates the functions of retinoid-binding proteins with retinoid metabolism. One of these proteins, the widely expressed (throughout retinoid target tissues and in all vertebrates) and highly conserved cellular retinol-binding protein (CRBP), sequesters retinol in an internal binding pocket that segregates it from the intracellular milieu. The CRBP-retinol complex appears to be the quantitatively major form of retinol in vivo, and may protect the promiscuous substrate from nonenzymatic degradation and/or non-specific enzymes. For example, at least seven types of dehydrogenases catalyze retinal synthesis from unbound retinol in vitro (NAD+ vs. NADP+ dependent, cytosolic vs. microsomal, short-chain dehydrogenases/reductases vs. medium-chain alcohol dehydrogenases). But only a fraction of these (some of the short-chain de-hydrogenases/reductases) have the fascinating additional ability of catalyzing retinal synthesis from CRBP-bound retinol as well. Similarly, CRBP and/or other retinoid-binding proteins function in the synthesis of retinal esters, the reduction of retinal generated from intestinal beta-carotene metabolism, and retinoic acid metabolism. The discussion details the evidence supporting an integrated model of retinoid-binding protein/metabolism. Also addressed are retinoid-androgen interactions and evidence incompatible with ethanol causing fetal alcohol syndrome by competing directly with retinol dehydrogenation to impair retinoic acid biosynthesis. PMID:10506831

  1. Network Analysis of Enzyme Activities and Metabolite Levels and Their Relationship to Biomass in a Large Panel of Arabidopsis Accessions[C][W][OA

    PubMed Central

    Sulpice, Ronan; Trenkamp, Sandra; Steinfath, Matthias; Usadel, Bjorn; Gibon, Yves; Witucka-Wall, Hanna; Pyl, Eva-Theresa; Tschoep, Hendrik; Steinhauser, Marie Caroline; Guenther, Manuela; Hoehne, Melanie; Rohwer, Johann M.; Altmann, Thomas; Fernie, Alisdair R.; Stitt, Mark

    2010-01-01

    Natural genetic diversity provides a powerful resource to investigate how networks respond to multiple simultaneous changes. In this work, we profile maximum catalytic activities of 37 enzymes from central metabolism and generate a matrix to investigate species-wide connectivity between metabolites, enzymes, and biomass. Most enzyme activities change in a highly coordinated manner, especially those in the Calvin-Benson cycle. Metabolites show coordinated changes in defined sectors of metabolism. Little connectivity was observed between maximum enzyme activities and metabolites, even after applying multivariate analysis methods. Measurements of posttranscriptional regulation will be required to relate these two functional levels. Individual enzyme activities correlate only weakly with biomass. However, when they are used to estimate protein abundances, and the latter are summed and expressed as a fraction of total protein, a significant positive correlation to biomass is observed. The correlation is additive to that obtained between starch and biomass. Thus, biomass is predicted by two independent integrative metabolic biomarkers: preferential investment in photosynthetic machinery and optimization of carbon use. PMID:20699391

  2. The Neurobiology of Retinoic Acid in Affective Disorders

    PubMed Central

    Bremner, J Douglas; McCaffery, Peter

    2009-01-01

    Current models of affective disorders implicate alterations in norepinephrine, serotonin, dopamine, and CRF/cortisol; however treatments targeted at these neurotransmitters or hormones have led to imperfect resolution of symptoms, suggesting that the neurobiology of affective disorders is incompletely understood. Until now retinoids have not been considered as possible contributors to affective disorders. Retinoids represent a family of compounds derived from Vitamin A that perform a large number of functions, many via the vitamin A product, retinoic acid. This signaling molecule binds to specific retinoic acid receptors in the brain which, like the glucocorticoid and thyroid hormone receptors, are part of the nuclear receptor superfamily and regulate gene transcription. Research in the field of retinoic acid in the CNS has focused on the developing brain, in part stimulated by the observation that isotretinoin (13-cis retinoic acid), an isomer of retinoic acid used in the treatment of acne, is highly teratogenic for the CNS. More recent work has suggested that retinoic acid may influence the adult brain; animal studies indicated that the administration of isotretinoin is associated with alterations in behavior as well as inhibition of neurogenesis in the hippocampus. Clinical evidence for an association between retinoids and depression includes case reports in the literature, studies of health care databases, and other sources. A preliminary PET study in human subjects showed that isotretinoin was associated with a decrease in orbitofrontal metabolism. Several studies have shown that the molecular components required for retinoic acid signaling are expressed in the adult brain ; the overlap of brain areas implicated in retinoic acid function and stress and depression suggest that retinoids could play a role in affective disorders. This report reviews the evidence in this area and describes several systems that may be targets of retinoic acid and which contribute

  3. All-trans retinoic acid mitigates methotrexate-induced liver injury in rats; relevance of retinoic acid signaling pathway.

    PubMed

    Ewees, Mohamed G; Abdelghany, Tamer M; Abdel-Aziz, Abdel-Aziz H; Abdel-Bakky, Mohamed S

    2015-09-01

    Methotrexate (MTX) is a widely used drug for treatment of rheumatic and autoimmune diseases as well as different types of cancer. One of the major side effects of MTX is hepatotoxicity. Retinoid receptors, including retinoid X receptor (RXR), and retinoic acid receptor (RAR) are vitamin A receptors that are highly expressed in the liver and regulate important physiological processes through regulation of different genes. In this study, we investigated the effect of MTX on RXR-α and RAR-α expression in the liver and the potential protective effects of all-trans retinoic acid (ATRA) in MTX-induced hepatotoxicity. Rats were randomly divided into five groups: The rates were treated with saline, DMSO, MTX (20 mg/kg/IP; single dose), ATRA (7.5 mg/kg/day, I.P), or MTX and ATRA. Rats were killed 24 h after the last ATRA injection. The liver tissues were dissected out, weighed, and subjected to histological, immunohistochemical, and biochemical examinations. Our results demonstrated that treatment with MTX resulted in significant decrease in reduced glutathione (GSH) content and superoxide dismutase (SOD) activity, with concomitant increase in ALT, AST, and MDA levels. In addition, MTX markedly downregulated the expression of both RXR-α and RAR-α, and changed the appearance of RXR-α to be very small speckled droplets. Treatment with ATRA significantly ameliorated MTX-induced effects on GSH, ALT, and MDA. Moreover, ATRA administration increased the expression and nuclear translocation of RXR-α in rat hepatocytes. In conclusion, our study revealed, for the first time, that retinoid receptors may play an important role in the MTX-induced hepatotoxicity. PMID:25971792

  4. Target interaction profiling of midostaurin and its metabolites in neoplastic mast cells predicts distinct effects on activation and growth

    PubMed Central

    Peter, Barbara; Winter, Georg E.; Blatt, Katharina; Bennett, Keiryn L.; Stefanzl, Gabriele; Rix, Uwe; Eisenwort, Gregor; Hadzijusufovic, Emir; Gridling, Manuela; Dutreix, Catherine; Hoermann, Gregor; Schwaab, Juliana; Radia, Deepti; Roesel, Johannes; Manley, Paul W.; Reiter, Andreas; Superti-Furga, Giulio; Valent, Peter

    2016-01-01

    Proteomic-based drug testing is an emerging approach to establish the clinical value and anti-neoplastic potential of multi-kinase inhibitors. The multikinase inhibitor midostaurin (PKC412) is a promising new agent used to treat patients with advanced systemic mastocytosis (SM). We examined the target interaction-profiles and the mast cell (MC)-targeting effects of two pharmacologically relevant midostaurin metabolites, CGP52421 and CGP62221. All three compounds, midostaurin and the two metabolites, suppressed IgE-dependent histamine secretion in basophils and MC with reasonable IC50 values. Midostaurin and CGP62221 also produced growth-inhibition and dephosphorylation of KIT in the MC leukemia cell line HMC-1.2, whereas the second metabolite, CGP52421, that accumulates in vivo, showed no substantial effects. Chemical proteomic profiling and drug-competition experiments revealed that midostaurin interacts with KIT and several additional kinase-targets. The key downstream-regulator FES was recognized by midostaurin and CGP62221, but not by CGP52421 in MC lysates, whereas the IgE-receptor-downstream target SYK was recognized by both metabolites. Together, our data show that the clinically relevant midostaurin metabolite CGP52421 inhibits IgE-dependent histamine release, but is a weak inhibitor of MC proliferation which may have clinical implications and may explain why mediator-related symptoms improve in SM patients even when disease progression occurs. PMID:26349526

  5. Target interaction profiling of midostaurin and its metabolites in neoplastic mast cells predicts distinct effects on activation and growth.

    PubMed

    Peter, B; Winter, G E; Blatt, K; Bennett, K L; Stefanzl, G; Rix, U; Eisenwort, G; Hadzijusufovic, E; Gridling, M; Dutreix, C; Hoermann, G; Schwaab, J; Radia, D; Roesel, J; Manley, P W; Reiter, A; Superti-Furga, G; Valent, P

    2016-02-01

    Proteomic-based drug testing is an emerging approach to establish the clinical value and anti-neoplastic potential of multikinase inhibitors. The multikinase inhibitor midostaurin (PKC412) is a promising new agent used to treat patients with advanced systemic mastocytosis (SM). We examined the target interaction profiles and the mast cell (MC)-targeting effects of two pharmacologically relevant midostaurin metabolites, CGP52421 and CGP62221. All three compounds, midostaurin and the two metabolites, suppressed IgE-dependent histamine secretion in basophils and MC with reasonable IC(50) values. Midostaurin and CGP62221 also produced growth inhibition and dephosphorylation of KIT in the MC leukemia cell line HMC-1.2, whereas the second metabolite, CGP52421, which accumulates in vivo, showed no substantial effects. Chemical proteomic profiling and drug competition experiments revealed that midostaurin interacts with KIT and several additional kinase targets. The key downstream regulator FES was recognized by midostaurin and CGP62221, but not by CGP52421 in MC lysates, whereas the IgE receptor downstream target SYK was recognized by both metabolites. Together, our data show that the clinically relevant midostaurin metabolite CGP52421 inhibits IgE-dependent histamine release, but is a weak inhibitor of MC proliferation, which may have clinical implications and may explain why mediator-related symptoms improve in SM patients even when disease progression occurs. PMID:26349526

  6. Retinoic acid metabolism blocking agents (RAMBAs) for treatment of cancer and dermatological diseases.

    PubMed

    Njar, Vincent C O; Gediya, Lalji; Purushottamachar, Puranik; Chopra, Pankaj; Vasaitis, Tadas Sean; Khandelwal, Aakanksha; Mehta, Jhalak; Huynh, Carlic; Belosay, Aashvini; Patel, Jyoti

    2006-07-01

    The naturally occurring retinoids and their synthetic analogs play a key role in differentiation, proliferation, and apoptosis, and their use/potential in oncology, dermatology and a variety of diseases are well documented. This review focuses on the role of all-trans-retinoic acid (ATRA), the principal endogenous metabolite of vitamin A (retinol) and its metabolism in oncology and dermatology. ATRA has been used successfully in differentiated therapy of acute promyelocytic leukemia, skin cancer, Kaposi's sarcoma, and cutaneous T-cell lymphoma, and also in the treatment of acne and psoriasis. However, its usefulness is limited by the rapid emergence of acquired ATRA resistance involving multifactoral mechanisms. A key mechanism of resistance involves ATRA-induced catabolism of ATRA. Thus, a novel strategy to overcome the limitation associated with exogenous ATRA therapy has been to modulate and/or increase the levels of endogenous ATRA by inhibiting the cytochrome P450-dependent ATRA-4-hydroxylase enzymes (particularly CYP26s) responsible for ATRA metabolism. These inhibitors are also referred to as retinoic acid metabolism blocking agents (RAMBAs). This review highlights development in the design, synthesis, and evaluation of RAMBAs. Major emphasis is given to liarozole, the most studied and only RAMBA in clinical use and also the new RAMBAs in development and with clinical potential. PMID:16530416

  7. Genomic antagonism between retinoic acid and estrogen signaling in breast cancer.

    PubMed

    Hua, Sujun; Kittler, Ralf; White, Kevin P

    2009-06-26

    Retinoic acid (RA) triggers antiproliferative effects in tumor cells, and therefore RA and its synthetic analogs have great potential as anticarcinogenic agents. Retinoic acid receptors (RARs) mediate RA effects by directly regulating gene expression. To define the genetic network regulated by RARs in breast cancer, we identified RAR genomic targets using chromatin immunoprecipitation and expression analysis. We found that RAR binding throughout the genome is highly coincident with estrogen receptor alpha (ERalpha) binding, resulting in a widespread crosstalk of RA and estrogen signaling to antagonistically regulate breast cancer-associated genes. ERalpha- and RAR-binding sites appear to be coevolved on a large scale throughout the human genome, often resulting in competitive binding activity at nearby or overlapping cis-regulatory elements. The highly coordinated intersection between these two critical nuclear hormone receptor signaling pathways provides a global mechanism for balancing gene expression output via local regulatory interactions dispersed throughout the genome. PMID:19563758

  8. The Expression of Bone Morphogenetic Protein 2 and Matrix Metalloproteinase 2 through Retinoic Acid Receptor Beta Induced by All-Trans Retinoic Acid in Cultured ARPE-19 Cells

    PubMed Central

    Gao, Zhenya; Huo, Lijun; Cui, Dongmei; Yang, Xiao; Zeng, Junwen

    2016-01-01

    Purpose All-trans retinoic acid (ATRA) plays an important role in ocular development. Previous studies found that retinoic acid could influence the metabolism of scleral remodeling by promoting retinal pigment epithelium (RPE) cells to secrete secondary signaling factors. The purpose of this study was to investigate whether retinoic acid affected secretion of bone morphogenetic protein 2 (BMP-2) and matrix metalloproteinase 2 (MMP-2) and to explore the signaling pathway of retinoic acid in cultured acute retinal pigment epithelial 19 (ARPE-19) cells. Methods The effects of ATRA (concentrations from 10−9 to 10−5 mol/l) on the expression of retinoic acid receptors (RARs) in ARPE-19 cells were examined at the mRNA and protein levels using reverse transcription-polymerase chain reaction (RT-PCR) and western blot assay, respectively. The effects of treating ARPE-19 cells with ATRA concentrations ranging from 10−9 to 10−5 mol/l for 24 h and 48 h or with 10-6mol/l ATRA at different times ranging from 6h to 72h were assessed using real-time quantitative PCR (qPCR) and enzyme-linked immunosorbent assay (ELISA). The contribution of RARβ-induced activation of ARPE-19 cells was confirmed using LE135, an antagonist of RARβ. Results RARβ mRNA levels significantly increased in the ARPE-19 cells treated with ATRA for 24h and 48h. These increases in RARβ mRNA levels were dose dependent (at concentrations of 10−9 to 10−5 mol/l) with a maximum effect observed at 10−6 mol/l. There were no significant changes in the mRNA levels of RARα and RARγ. Western blot assay revealed that RARβ protein levels were increased significantly in a time-dependent manner in ARPE-19 cells treated with 10−6 mol/l ATRA from 12 h to 72 h, with a marked increase observed at 24 h and 48 h. The upregulation of RARβ and the ATRA-induced secretion in ARPE-19 cells could be inhibited by the RARβ antagonist LE135. Conclusion ATRA induced upregulation of RARβ in ARPE-19 cells and stimulated

  9. Volatile Metabolites

    PubMed Central

    Rowan, Daryl D.

    2011-01-01

    Volatile organic compounds (volatiles) comprise a chemically diverse class of low molecular weight organic compounds having an appreciable vapor pressure under ambient conditions. Volatiles produced by plants attract pollinators and seed dispersers, and provide defense against pests and pathogens. For insects, volatiles may act as pheromones directing social behavior or as cues for finding hosts or prey. For humans, volatiles are important as flavorants and as possible disease biomarkers. The marine environment is also a major source of halogenated and sulfur-containing volatiles which participate in the global cycling of these elements. While volatile analysis commonly measures a rather restricted set of analytes, the diverse and extreme physical properties of volatiles provide unique analytical challenges. Volatiles constitute only a small proportion of the total number of metabolites produced by living organisms, however, because of their roles as signaling molecules (semiochemicals) both within and between organisms, accurately measuring and determining the roles of these compounds is crucial to an integrated understanding of living systems. This review summarizes recent developments in volatile research from a metabolomics perspective with a focus on the role of recent technical innovation in developing new areas of volatile research and expanding the range of ecological interactions which may be mediated by volatile organic metabolites. PMID:24957243

  10. Biochemical Characterization of the Active Anti-Hepatitis C Virus Metabolites of 2,6-Diaminopurine Ribonucleoside Prodrug Compared to Sofosbuvir and BMS-986094.

    PubMed

    Ehteshami, Maryam; Tao, Sijia; Ozturk, Tugba; Zhou, Longhu; Cho, Jong Hyun; Zhang, Hongwang; Amiralaei, Sheida; Shelton, Jadd R; Lu, Xiao; Khalil, Ahmed; Domaoal, Robert A; Stanton, Richard A; Suesserman, Justin E; Lin, Biing; Lee, Sam S; Amblard, Franck; Whitaker, Tony; Coats, Steven J; Schinazi, Raymond F

    2016-08-01

    Ribonucleoside analog inhibitors (rNAI) target the hepatitis C virus (HCV) RNA-dependent RNA polymerase nonstructural protein 5B (NS5B) and cause RNA chain termination. Here, we expand our studies on β-d-2'-C-methyl-2,6-diaminopurine-ribonucleotide (DAPN) phosphoramidate prodrug 1 (PD1) as a novel investigational inhibitor of HCV. DAPN-PD1 is metabolized intracellularly into two distinct bioactive nucleoside triphosphate (TP) analogs. The first metabolite, 2'-C-methyl-GTP, is a well-characterized inhibitor of NS5B polymerase, whereas the second metabolite, 2'-C-methyl-DAPN-TP, behaves as an adenosine base analog. In vitro assays suggest that both metabolites are inhibitors of NS5B-mediated RNA polymerization. Additional factors, such as rNAI-TP incorporation efficiencies, intracellular rNAI-TP levels, and competition with natural ribonucleotides, were examined in order to further characterize the potential role of each nucleotide metabolite in vivo Finally, we found that although both 2'-C-methyl-GTP and 2'-C-methyl-DAPN-TP were weak substrates for human mitochondrial RNA (mtRNA) polymerase (POLRMT) in vitro, DAPN-PD1 did not cause off-target inhibition of mtRNA transcription in Huh-7 cells. In contrast, administration of BMS-986094, which also generates 2'-C-methyl-GTP and previously has been associated with toxicity in humans, caused detectable inhibition of mtRNA transcription. Metabolism of BMS-986094 in Huh-7 cells leads to 87-fold higher levels of intracellular 2'-C-methyl-GTP than DAPN-PD1. Collectively, our data characterize DAPN-PD1 as a novel and potent antiviral agent that combines the delivery of two active metabolites. PMID:27216050

  11. Assessment of adrenocortical activity by non-invasive measurement of faecal cortisol metabolites in dromedary camels (Camelus dromedarius).

    PubMed

    Sid-Ahmed, Omer-Elfaroug; Sanhouri, Ahmed; Elwaseela, Badr-Eldin; Fadllalah, Imad; Mohammed, Galal-Eldin Elazhari; Möstl, Erich

    2013-08-01

    The aim of this study was to determine whether glucocorticoid production could be monitored non-invasively in dromedary camels by measuring faecal cortisol metabolites (FCMs). Five Sudanese dromedaries, two males and three females, were injected with a synthetic adrenocorticotropic hormone (ACTH) analogue. Blood samples were collected pre- and post-ACTH injection. Faeces were sampled after spontaneous defecation for five consecutive days (2 days before and 3 days after ACTH injection). Baseline plasma cortisol values ranged from 0.6 to 10.8 ng/ml in males and from 1.1 to 16.6 ng/ml in females, while peak values after ACTH injection were 10.9-41.9 in males and 10-42.2 ng/ml in females. Peak blood cortisol values were reached between 1.5 and 2.0 h after ACTH injection. The concentration of FCMs increased after ACTH injection in the faeces of both sexes, although steroid levels peaked earlier in males [24 h; (286.7-2,559.7 ng/g faeces)] than in females [36-48 h; (1,182.6-5,169.1 ng/g faeces)], reflecting increases of 3.1-8.3- and 4.3-8-fold above baseline levels. To detect chromatographic patterns of immunoreactive FCMs, faecal samples with high FCM concentrations from both sexes were pooled and subjected to reverse phase high performance liquid chromatography (RP-HPLC). RP-HPLC analysis revealed sex differences in the polarity of FCMs, with females showing more polar FCMs than males. We concluded that stimulation of adrenocortical activity by ACTH injection resulted in a measurable increase in blood cortisol that was reliably paralleled by increases in FCM levels. Thus, measurement of FCMs is a powerful tool for monitoring the adrenocortical responses of dromedaries to stressors in field conditions. PMID:23430659

  12. Species and gender differences in the formation of an active metabolite of a substituted 2,4-thiazolidinedione insulin sensitizer.

    PubMed

    Beconi, M; Mao, A; Creighton, M; Hop, C E C A; Chiu, S H L; Eydelloth, R; Franklin, R; Tang, F; Yu, N; Vincent, S

    2003-07-01

    1. The metabolism of a substituted 2,4-thiazolidinedione (P1) with dual PPARalpha/gamma activity was evaluated in male and female rats, dogs and monkeys. A para-hydroxylated metabolite (M1) with potent PPARgamma-selective agonist, was a major circulating drug-related component in female rats, dogs and monkeys, but not in male rats (M1-to-P1 exposure ratio of <1, 3-5, 5 and 5-11 in male rat, monkey, female rat, and dog, respectively). 2. M1 (%) formed in vitro (5, 53, 57-65, 67 and 67% in male rat, monkey, female rat, dog, and human liver microsomes, respectively), rank ordered with M1 (%) formed in vivo (24-45, 53-57, 78, 75-85%, for male rat, monkey, female rat and dog, respectively, after oral administration of P1). 3. The plasma clearance of M1 was higher in male rats (32 ml min(-1) kg(-1) compared with 6, 7 and 2 ml min(-1) kg(-1) in female rat, male monkey and male dogs, respectively). 4. The low amounts of M1 observed in male rats, with the appearance of products of the cleavage of the propyl group between the phenyl groups was probably due to the presence of the sex-specific CYP2C11, which cleaves P1 at the propyl bridge. None of the CYPs present in female rats cleaved P1 at this site and M1 was only produced by CYP2C6. In humans, only CYP2C8 and the polymorphic CYP2C19 produced M1. PMID:12893525

  13. Relation between clopidogrel active metabolite levels and different platelet aggregation methods in patients receiving clopidogrel and aspirin.

    PubMed

    Liang, Yan; Johnston, Marilyn; Hirsh, Jack; Pare, Guillaume; Li, Chunjian; Mehta, Shamir; Teo, Koon K; Sloane, Debi; Yi, Qilong; Zhu, Jun; Eikelboom, John W

    2012-11-01

    Clopidogrel is a prodrug that undergoes bioconversion via cytochrome P450 system to form an active metabolite (AM) that binds to the platelet ADP receptor. The antiplatelet effect of clopidogrel is commonly assessed by measuring the aggregatory response to 5 μM ADP by light transmission aggregation (LTA) or multiple electrode aggregometry (MEA) or by the vasodilator-stimulated phosphoprotein platelet reactivity index (VASP-PRI). To determine which of these three tests of platelet ADP receptor pathway inhibition most closely correlates with clopidogrel AM levels. We analyzed blood samples from 82 patients with coronary artery disease who were randomized to receive double-dose or standard dose clopidogrel for 2 weeks. We measured peak clopidogrel AM levels, platelet aggregation in response to ADP and VASP-PRI on days 1, and repeated all the measures on days 7 and 14. Linear regression analysis was used to examine the correlation between clopidogrel AM and LTA, MEA and VASP-PRI. Bland-Altman plots were used to explore the agreement between tests of the antiplatelet effects of clopidogrel. Clopidogrel AM on day 1 correlated most closely with VASP-PRI (r = -0.5767) and demonstrated weaker correlations with LTA (r = -0.4656) and MEA (r = -0.3384) (all p < 0.01). Intra-class correlation (ICC) between VASP-PRI and LTA was 0.6446; VASP-PRI and MEA was 0.4720; and LTA and MEA was 0.4693. Similar results were obtained on days 7 and 14. Commonly used pharmacodynamic measures of clopidogrel response are only moderately correlated with clopidogrel AM levels and may not be suitable to measure the adequacy of clopidogrel therapy. PMID:22797934

  14. Effect of bovine ABCG2 Y581S polymorphism on concentrations in milk of enrofloxacin and its active metabolite ciprofloxacin.

    PubMed

    Otero, J A; García-Mateos, D; de la Fuente, A; Prieto, J G; Álvarez, A I; Merino, G

    2016-07-01

    The ATP-binding cassette transporter G2 (ABCG2) is involved in the secretion of several drugs into milk. The bovine Y581S ABCG2 polymorphism increases the secretion into milk of the fluoroquinolone danofloxacin in Holstein cows. Danofloxacin and enrofloxacin are the fluoroquinolones most widely used in veterinary medicine. Both enrofloxacin (ENRO) and its active metabolite ciprofloxacin (CIPRO) reach milk at relatively high concentrations. The aim of this work was to study the effect of the bovine Y581S ABCG2 polymorphism on in vitro transport as well as on concentrations in plasma and in milk of ENRO and CIPRO. Experiments using cells overexpressing bovine ABCG2 showed the effects of ABCG2 on the transport of CIPRO, demonstrating more efficient in vitro transport of this antimicrobial by the S581 variant as compared with the Y581 variant. Animal studies administering 2.5mg/kg of ENRO subcutaneously to Y/Y 581 and Y/S 581 cows revealed that concentrations in plasma of ENRO and CIPRO were significantly lower in Y/S animals. Regardless of the genotype, the antimicrobial profile in milk after the administration of ENRO was predominantly of CIPRO. With respect to the genotype effects on the amounts of drugs present in milk, AUC0-24 values were more than 1.2 times higher in Y/S cows for ENRO and 2.2 times for CIPRO, indicating a greater capacity of Y581S to transfer these drugs into milk. These results emphasize the clinical relevance of this polymorphism as a factor affecting the concentrations in plasma and in milk of drugs of importance in veterinary medicine. PMID:27157572

  15. Microbial transformation of 20(S)-protopanaxadiol by Absidia corymbifera. Cytotoxic activity of the metabolites against human prostate cancer cells.

    PubMed

    Chen, Guangtong; Yang, Min; Nong, Shaojun; Yang, Xue; Ling, Yong; Wang, Donggeng; Wang, Xinyang; Zhang, Wei

    2013-01-01

    Biotransformation of 20(S)-protopanaxadiol (1) by the fungus Absidia corymbifera AS 3.3387 yielded five metabolites (2-6). On the basis of spectroscopic data analyses, the metabolites were identified as 26-hydroxyl-20(S)-protopanaxadiol (2), 23, 24-en-25-hydroxyl-20(S)-protopanaxadiol (3), 25-hydroxyl-20(S)-protopanaxadiol (4), 7β-hydroxyl-20(S)-protopanaxatriol (5), and 7-oxo-20(S)-protopanaxatriol (6), respectively. Among them, 5 and 6 are new compounds. These results indicated that A. corymbifera AS 3.3387 could catalyze the side-chain oxidation-reduction, 7β hydroxylation, and the specific C-7 dehydrogenation of derivatives of 20(S)-protopanaxadiol. The metabolites 2, 5, and 6 showed the more potent inhibitory effects against DU-145 and PC-3 cell lines than the substrate. PMID:23022533

  16. Retinoids induce integrin-independent lymphocyte adhesion through RAR-α nuclear receptor activity

    SciTech Connect

    Whelan, Jarrett T.; Wang, Lei; Chen, Jianming; Metts, Meagan E.; Nasser, Taj A.; McGoldrick, Liam J.; Bridges, Lance C.

    2014-11-28

    Highlights: • Transcription and translation are required for retinoid-induced lymphocyte adhesion. • RAR activation is sufficient to induced lymphocyte cell adhesion. • Vitamin D derivatives inhibit RAR-prompted lymphocyte adhesion. • Adhesion occurs through a novel binding site within ADAM disintegrin domains. • RARα is a key nuclear receptor for retinoid-dependent lymphocyte cell adhesion. - Abstract: Oxidative metabolites of vitamin A, in particular all-trans-retinoic acid (atRA), have emerged as key factors in immunity by specifying the localization of immune cells to the gut. Although it is appreciated that isomers of retinoic acid activate the retinoic acid receptor (RAR) and retinoid X receptor (RXR) family of nuclear receptors to elicit cellular changes, the molecular details of retinoic acid action remain poorly defined in immune processes. Here we employ a battery of agonists and antagonists to delineate the specific nuclear receptors utilized by retinoids to evoke lymphocyte cell adhesion to ADAM (adisintegrin and metalloprotease) protein family members. We report that RAR agonism is sufficient to promote immune cell adhesion in both immortal and primary immune cells. Interestingly, adhesion occurs independent of integrin function, and mutant studies demonstrate that atRA-induced adhesion to ADAM members required a distinct binding interface(s) as compared to integrin recognition. Anti-inflammatory corticosteroids as well as 1,25-(OH){sub 2}D{sub 3}, a vitamin D metabolite that prompts immune cell trafficking to the skin, potently inhibited the observed adhesion. Finally, our data establish that induced adhesion was specifically attributable to the RAR-α receptor isotype. The current study provides novel molecular resolution as to which nuclear receptors transduce retinoid exposure into immune cell adhesion.

  17. Chemical diversity of biologically active metabolites in the sclerotia of Inonotus obliquus and submerged culture strategies for up-regulating their production.

    PubMed

    Zheng, Weifa; Miao, Kangjie; Liu, Yubing; Zhao, Yanxia; Zhang, Meimei; Pan, Shenyuan; Dai, Yucheng

    2010-07-01

    Inonotus obliquus (Fr.) Pilat is a white rot fungus belonging to the family Hymenochaetaceae in the Basidiomycota. In nature, this fungus rarely forms a fruiting body but usually an irregular shape of sclerotial conk called 'Chaga'. Characteristically, I. obliquus produces massive melanins released to the surface of Chaga. As early as in the sixteenth century, Chaga was used as an effective folk medicine in Russia and Northern Europe to treat several human malicious tumors and other diseases in the absence of any unacceptable toxic side effects. Chemical investigations show that I. obliquus produces a diverse range of secondary metabolites including phenolic compounds, melanins, and lanostane-type triterpenoids. Among these are the active components for antioxidant, antitumoral, and antiviral activities and for improving human immunity against infection of pathogenic microbes. Geographically, however, this fungus is restricted to very cold habitats and grows very slowly, suggesting that Chaga is not a reliable source of these bioactive compounds. Attempts for culturing this fungus axenically all resulted in a reduced production of bioactive metabolites. This review examines the current progress in the discovery of chemical diversity of Chaga and their biological activities and the strategies to modulate the expression of desired pathways to diversify and up-regulate the production of bioactive metabolites by the fungus grown in submerged cultures for possible drug discovery. PMID:20532760

  18. Hesperidin metabolite hesperetin-7-O-glucuronide, but not hesperetin-3'-O-glucuronide, exerts hypotensive, vasodilatory, and anti-inflammatory activities.

    PubMed

    Yamamoto, Masaki; Jokura, Hiroko; Hashizume, Koujiro; Ominami, Hideo; Shibuya, Yusuke; Suzuki, Atsushi; Hase, Tadashi; Shimotoyodome, Akira

    2013-09-01

    Orally ingested hesperidin (HES) is hydrolyzed into hesperetin in the gastrointestinal tract and conjugated during absorption. Hesperetin conjugates are the main circulating metabolites in human and rat plasma. We previously reported that glucosyl hesperidin (GHES), a water-soluble HES derivative, prevents hypertension via improvement of endothelial dysfunction in spontaneously hypertensive rats (SHRs). Although these hesperetin conjugates seem to be responsible for hypotensive and endothelium-dependent vasodilatory activities of dietary GHES, little is known about the mechanisms of action of these conjugated metabolites. Therefore, the aim of the present study was to investigate the effects of hesperetin-7-O-β-d-glucuronide (HPT7G) and hesperetin-3'-O-β-d-glucuronide (HPT3'G), which are the predominant HES metabolites in rat plasma, on blood pressure and endothelial function. Intravenous administration of HPT7G (5 mg kg(-1)) decreased blood pressure in anesthetized SHRs. HPT7G enhanced endothelium-dependent vasodilation in response to acetylcholine, but had no effect on endothelium-independent vasodilation in response to sodium nitroprusside (SNP) in aortas isolated from SHRs. HPT7G decreased hydrogen peroxide-induced intracellular adhesion molecule-1 and monocyte chemoattractant protein-1 mRNA expression in rat aortic endothelial cells. In contrast, HPT3'G had little effect on these parameters. In conclusion, HPT7G exerted hypotensive, vasodilatory and anti-inflammatory activities, similar to hesperetin and these effects are associated, in part, with the activity of GHES and HES to improve hypertension and endothelial dysfunction. PMID:23831969

  19. Acute HCV/HIV Coinfection Is Associated with Cognitive Dysfunction and Cerebral Metabolite Disturbance, but Not Increased Microglial Cell Activation

    PubMed Central

    Garvey, Lucy J.; Pavese, Nicola; Ramlackhansingh, Anil; Thomson, Emma; Allsop, Joanna M.; Politis, Marios; Kulasegaram, Ranjababu; Main, Janice; Brooks, David J.; Taylor-Robinson, Simon D.; Winston, Alan

    2012-01-01

    Background Microglial cell activation and cerebral function impairment are described in both chronic hepatitis C viral (HCV) and Human-Immune-Deficiency viral (HIV) infections. The aim of this study was to investigate the effect of acute HCV infection upon cerebral function and microglial cell activation in HIV-infected individuals. Methods A case-control study was conducted. Subjects with acute HCV and chronic HIV coinfection (aHCV) were compared to matched controls with chronic HIV monoinfection (HIVmono). aHCV was defined as a new positive plasma HCV RNA within 12 months of a negative RNA test. Subjects underwent neuro-cognitive testing (NCT), cerebral proton magnetic resonance spectroscopy (1H-MRS) and positron emission tomography (PET) using a 11C-radiolabeled ligand (PK11195), which is highly specific for translocator protein 18 kDA receptors on activated microglial cells. Differences between cases and controls were assessed using linear regression modelling. Results Twenty-four aHCV cases completed NCT and 1H-MRS, 8 underwent PET. Of 57 HIVmono controls completing NCT, 12 underwent 1H-MRS and 8 PET. Subjects with aHCV demonstrated on NCT, significantly poorer executive function (mean (SD) error rate 26.50(17.87) versus 19.09(8.12), p = 0.001) and on 1H-MRS increased myo-inositol/creatine ratios (mI/Cr, a marker of cerebral inflammation) in the basal ganglia (ratio of 0.71(0.22) versus 0.55(0.23), p = 0.03), compared to subjects with HIVmono. On PET imaging, no difference in 11C-PK11195 binding potential (BP) was observed between study groups (p>0.10 all cerebral locations), however lower BPs were associated with combination antiretroviral therapy (cART) use in the parietal (p = 0.01) and frontal (p = 0.03) cerebral locations. Discussion Poorer cognitive performance and disturbance of cerebral metabolites are observed in subjects with aHC,V compared to subjects with HIVmono. Higher 11C-PK11195 BP was not observed in subjects with aHCV, but was

  20. Vitamin A and immune function: retinoic acid modulates population dynamics in antigen receptor and CD38-stimulated splenic B cells.

    PubMed

    Chen, Qiuyan; Ross, A Catharine

    2005-10-01

    Vitamin A and its active metabolite, all-trans retinoic acid (RA), regulate the antibody response in vivo, although the underlying mechanisms are not well understood. We have investigated the regulation by RA of B cell population dynamics and Ig gene expression in purified splenic mouse B cells stimulated through the B cell antigen receptor (BCR) and/or CD38, a BCR coreceptor. After ligation of the BCR and/or CD38, B cells became more heterogeneous in size. RA substantially restrained this change, concomitant with inhibition of cell proliferation. To examine B cell heterogeneity more closely, we categorized stimulated B cells by size (forward angle light scatter) and determined cell division dynamics, germ-line Ig heavy chain gene transcription and surface IgG1 (sIgG1) expression. Flow cytometric analysis of carboxyfluorescein diacetate succinimidyl ester-labeled B cells costained for sIgG1 showed that the more proliferative groups of B cells were smaller, whereas cells expressing more sIgG1 were larger. RA enriched the latter population, whereas cell division frequency in general and the number of smaller B cells that had undergone division cycles were reduced. Although RA significantly inhibited Ig germ-line transcript levels in the total B cell population, CD19(-)IgG1(+) B cells, which represent a more differentiated phenotype, were enriched. Furthermore, pax-5 mRNA was decreased and activation-induced cytidine deaminase mRNA was increased in RA-treated stimulated B cells. Thus, RA regulated factors known to be required for Ig class switch recombination and modulated the population dynamics of ligation-stimulated B cells, while promoting the progression of a fraction of B cells into differentiated sIgG-expressing cells. PMID:16093312

  1. Retinoic Acid Induces Embryonic Stem Cell Differentiation by Altering Both Encoding RNA and microRNA Expression

    PubMed Central

    Yu, Mengying; Wu, Haibo; Ai, Zhiying; Wu, Yongyan; Liu, Hongliang; Du, Juan; Guo, Zekun; Zhang, Yong

    2015-01-01

    Retinoic acid (RA) is a vitamin A metabolite that is essential for early embryonic development and promotes stem cell neural lineage specification; however, little is known regarding the impact of RA on mRNA transcription and microRNA levels on embryonic stem cell differentiation. Here, we present mRNA microarray and microRNA high-output sequencing to clarify how RA regulates gene expression. Using mRNA microarray analysis, we showed that RA repressed pluripotency-associated genes while activating ectoderm markers in mouse embryonic stem cells (mESCs). Moreover, RA modulated the DNA methylation of mESCs by altering the expression of epigenetic-associated genes such as Dnmt3b and Dnmt3l. Furthermore, H3K4me2, a pluripotent histone modification, was repressed by RA stimulation. From microRNA sequence data, we identified two downregulated microRNAs, namely, miR-200b and miR-200c, which regulated the pluripotency of stem cells. We found that miR-200b or miR-200c deficiency suppressed the expression of pluripotent genes, including Oct4 and Nanog, and activated the expression of the ectodermal marker gene Nestin. These results demonstrate that retinoid induces mESCs to differentiate by regulating miR-200b/200c. Our findings provide the landscapes of mRNA and microRNA gene networks and indicate the crucial role of miR-200b/200c in the RA-induced differentiation of mESCs. PMID:26162091

  2. The relationship of nitrogen and C/N ratio with secondary metabolites levels and antioxidant activities in three varieties of Malaysian kacip Fatimah (Labisia pumila Blume).

    PubMed

    Ibrahim, Mohd Hafiz; Jaafar, Hawa Z E

    2011-01-01

    Kacip Fatimah (Labisia pumila Blume), one of the most famous and widely used herbs, especially in Southeast Asia, is found to have interesting bioactive compounds and displays health promoting properties. In this study, the antioxidant activities of the methanol extracts of leaves, stems and roots of three varieties of L. pumila (var. alata, pumila and lanceolata) were evaluated in an effort to compare and validate the medicinal potential of this indigenous Malaysian herb species. The antioxidant activity determined by the 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay, as well as the total amount of phenolics and flavonoids were the highest in the leaves, followed by the stems and roots in all the varieties. A similar trend was displayed by the ferric reducing antioxidant potential (FRAP) activity, suggesting that the L. pumila varieties possess high foliar antioxidant properties. At low FRAP activity concentrations, the values of the leaves' inhibition activity in the three varieties were significantly higher than those of the stems and roots, with var. alata exhibiting higher antioxidant activities and total contents of phenolics and flavonoids compared to the varieties pumila and lanceolata. The high production of secondary metabolites and antioxidant activities in var. alata were firmly related to low nitrogen content and high C/N ratio in plant parts. The study also demonstrated a positive correlation between secondary metabolite content and antioxidant activities, and revealed that the consumption of L. pumila could exert several beneficial effects by virtue of its antioxidant activity. PMID:21716173

  3. Characterization of DNA Binding and Retinoic Acid Binding Properties of Retinoic Acid Receptor

    NASA Astrophysics Data System (ADS)

    Yang, Na; Schule, Roland; Mangelsdorf, David J.; Evans, Ronald M.

    1991-05-01

    High-level expression of the full-length human retinoic acid receptor (RAR) α and the DNA binding domain of the RAR in Escherichia coli was achieved by using a T7 RNA polymerase-directed expression system. After induction, full-length RAR protein was produced at an estimated level of 20% of the total bacterial proteins. Both intact RAR molecules and the DNA binding domain bind to the cognate DNA response element with high specificity in the absence of retinoic acid. However, this binding is enhanced to a great extent upon the addition of eukaryotic cell extracts. The factor responsible for this enhancement is heat-sensitive and forms a complex with RAR that binds to DNA and exhibits a distinct migration pattern in the gel-mobility-shift assay. The interaction site of the factor with RAR is localized in the 70-amino acid DNA binding region of RAR. The hormone binding ability of the RARα protein was assayed by a charcoal absorption assay and the RAR protein was found to bind to retinoic acid with a K_d of 2.1 x 10-10 M.

  4. Comparison of the Function and Expression of CYP26A1 and CYP26B1, the two Retinoic Acid Hydroxylases

    PubMed Central

    Topletz, Ariel R.; Thatcher, Jayne E.; Zelter, Alex; Lutz, Justin D.; Tay, Suzanne; Nelson, Wendel L.; Isoherranen, Nina

    2011-01-01

    All-trans-retinoic acid (atRA) is an important signaling molecule in all chordates. The cytochrome P450 enzymes CYP26 are believed to partially regulate cellular concentrations of atRA via oxidative metabolism and hence affect retinoid homeostasis and signaling. CYP26A1 and CYP26B1 are atRA hydroxylases that catalyze formation of similar metabolites in cell systems. However, they have only 40% sequence similarity suggesting differences between the two enzymes. The aim of this study was to determine whether CYP26A1 and CYP26B1 have similar catalytic activity, form different metabolites from atRA and are expressed in different tissues in adults. The mRNA expression of CYP26A1 and CYP26B1 correlated between human tissues except for human cerebellum in which CYP26B1 was the predominant CYP26 and liver in which CYP26A1 dominated. Quantification of CYP26A1 and CYP26B1 protein in human tissues was in agreement with the mRNA expression and showed correlation between the two isoforms. Qualitatively, recombinant CYP26A1 and CYP26B1 formed the same primary and sequential metabolites from atRA. Quantitatively, CYP26B1 had a lower Km (19nM) and Vmax (0.8pmol/min/pmol) than CYP26A1 (Km=50nM and Vmax=10pmol/min/pmol) for formation of 4-OH-RA. The major atRA metabolites 4-OH-RA, 18-OH-RA and 4-oxo-RA were all substrates of CYP26A1 and CYP26B1, and CYP26A1 had a 2–10 fold higher catalytic activity towards all substrates tested. This study shows that CYP26A1 and CYP26B1 are qualitatively similar RA hydroxylases with overlapping expression profiles. CYP26A1 has higher catalytic activity than CYP26B1 and seems to be responsible for metabolism of atRA in tissues that function as a barrier for atRA exposure. PMID:22020119

  5. Distribution of topical ocular nepafenac and its active metabolite amfenac to the posterior segment of the eye.

    PubMed

    Chastain, James E; Sanders, Mark E; Curtis, Michael A; Chemuturi, Nagendra V; Gadd, Martha E; Kapin, Michael A; Markwardt, Kerry L; Dahlin, David C

    2016-04-01

    Nepafenac ophthalmic suspensions, 0.1% (NEVANAC(®)) and 0.3% (ILEVRO™), are topical nonsteroidal anti-inflammatory drug (NSAID) products approved in the United States, Europe and various other countries to treat pain and inflammation associated with cataract surgery. NEVANAC is also approved in Europe for the reduction in the risk of postoperative macular edema (ME) associated with cataract surgery in diabetic patients. The efficacy against ME suggests that topical administration leads to distribution of nepafenac or its active metabolite amfenac to the posterior segment of the eye. This article evaluates the ocular distribution of nepafenac and amfenac and the extent of local delivery to the posterior segment of the eye, following topical ocular instillation in animal models. Nepafenac ophthalmic suspension was instilled unilaterally in New Zealand White rabbits as either a single dose (0.1%; one drop) or as multiple doses (0.3%, one drop, once-daily for 4 days, or 0.1% one drop, three-times daily for 3 days and one morning dose on day 4). Nepafenac (0.3%) was also instilled unilaterally in cynomolgus monkeys as multiple doses (one drop, three-times daily for 7 days). Nepafenac and amfenac concentrations in harvested ocular tissues were measured using high-performance liquid chromatography/mass spectrometry. Locally-distributed compound concentrations were determined as the difference in levels between dosed and undosed eyes. In single-dosed rabbit eyes, peak concentrations of locally-distributed nepafenac and amfenac showed a trend of sclera > choroid > retina. Nepafenac peak levels in sub-samples posterior to the eye equator and inclusive of the posterior pole (E-PP) were 55.1, 4.03 and 2.72 nM, respectively, at 0.25 or 0.50 h, with corresponding amfenac peak levels of 41.9, 3.10 and 0.705 nM at 1 or 4 h. By comparison, peak levels in sclera, choroid and retina sub-samples in a band between the ora serrata and the equator (OS-E) were 13- to 40-fold

  6. Glucuronic acid and the ethanol metabolite ethyl-glucuronide cause toll-like receptor 4 activation and enhanced pain.

    PubMed

    Lewis, Susannah S; Hutchinson, Mark R; Zhang, Yingning; Hund, Dana K; Maier, Steven F; Rice, Kenner C; Watkins, Linda R

    2013-05-01

    We have previously observed that the non-opioid morphine metabolite, morphine-3-glucuronide, enhances pain via a toll-like receptor 4 (TLR4) dependent mechanism. The present studies were undertaken to determine whether TLR4-dependent pain enhancement generalizes to other classes of glucuronide metabolites. In silico modeling predicted that glucuronic acid alone and ethyl glucuronide, a minor but long-lasting ethanol metabolite, would dock to the same MD-2 portion of the TLR4 receptor complex previously characterized as the docking site for morphine-3-glucuronide. Glucuronic acid, ethyl glucuronide and ethanol all caused an increase in TLR4-dependent reporter protein expression in a cell line transfected with TLR4 and associated co-signaling molecules. Glucuronic acid-, ethyl glucuronide-, and ethanol-induced increases in TLR4 signaling were blocked by the TLR4 antagonists LPS-RS and (+)-naloxone. Glucuronic acid and ethyl glucuronide both caused allodynia following intrathecal injection in rats, which was blocked by intrathecal co-administration of the TLR4 antagonist LPS-RS. The finding that ethyl glucuronide can cause TLR4-dependent pain could have implications for human conditions such as hangover headache and alcohol withdrawal hyperalgesia, as well as suggesting that other classes of glucuronide metabolites could have similar effects. PMID:23348028

  7. Glucuronic acid and the ethanol metabolite ethyl-glucuronide cause Toll-like receptor 4 activation and enhanced pain

    PubMed Central

    Lewis, Susannah S.; Hutchinson, Mark R.; Zhang, Yingning; Hund, Dana K.; Maier, Steven F.; Rice, Kenner C.; Watkins, Linda R.

    2013-01-01

    We have previously observed that the non-opioid morphine metabolite, morphine-3-glucuronide, enhances pain via a toll-like receptor 4 (TLR4) dependent mechanism. The present studies were undertaken to determine whether TLR4-dependent pain enhancement generalizes to other classes of glucuronide metabolites. In silico modeling predicted that glucuronic acid alone and ethyl glucuronide, a minor but long-lasting ethanol metabolite, would dock to the same MD-2 portion of the TLR4 receptor complex previously characterized as the docking site for morphine-3-glucuronide. Glucuronic acid, ethyl glucuronide and ethanol all caused an increase in TLR4-dependent reporter protein expression in a cell line transfected with TLR4 and associated co-signaling molecules. Glucuronic acid-, ethyl glucuronide-, and ethanol-induced increases in TLR4 signaling were blocked by the TLR4 antagonists LPS-RS and (+)-naloxone. Glucuronic acid and ethyl glucuronide both caused allodynia following intrathecal injection in rats, which was blocked by intrathecal co-administration of the TLR4 antagonist LPS-RS. The finding that ethyl glucuronide can cause TLR4-dependent pain could have implications for human conditions such as hangover headache and alcohol withdrawal hyperalgesia, as well as suggesting that other classes of glucuronide metabolites could have similar effects. PMID:23348028

  8. Enhancement of fludarabine sensitivity by all-trans-retinoic acid in chronic lymphocytic leukemia cells

    PubMed Central

    Fernández-Calotti, Paula X.; Lopez-Guerra, Mónica; Colomer, Dolors; Pastor-Anglada, Marçal

    2012-01-01

    Background A subset of patients with fludarabine-resistant chronic lymphocytic leukemia has previously been shown to express elevated intracellular levels of the concentrative high-affinity fludarabine transporter hCNT3, without any detectable related activity